Stem cell therapy: a clinical trial of stroke.
Bhasin Ashu,Srivastava M V Padma,Mohanty Sujata,Bhatia Rohit,Kumaran Senthil S,Bose Sushmita
Clinical neurology and neurosurgery
BACKGROUND:The alarming disability burden and a high prevalence rate of stroke in India has encouraged the researchers to develop regenerative therapies to reduce clinical deficits. This study evaluates safety, feasibility and efficacy of autologous mononuclear and mesenchymal cell transplantation in stroke patients evaluated on clinical scores and functional imaging (fMRI and DTI). METHODS:Forty (n=40) stroke patients were recruited with the inclusion criteria as: 3 months to 2 years of index event, power of hand muscles of at least 2; Brunnstrom stage: 2-5; conscious and comprehendible. Fugl Meyer (FM), modified Barthel Index (mBI), Medical Research Council (MRC) grade for strength, Ashworth tone scale and functional imaging was used for assessments at baseline, 8 weeks and 24 weeks. 50-60 million cells in 250 ml saline were infused intravenously over 2-3 h. RESULTS:The safety test profile was normal with no mortality or cell related adverse reactions in stem cell patients. Among outcome parameters, only modified Barthel Index (mBI) showed statistical significant improvement (p<0.05) in the stem cell group. An increased number of cluster activation in Brodmann areas BA 4, BA 6 was observed post stem cell infusion indicating neural plasticity. CONCLUSION:Autologous intravenous stem cell therapy is safe and feasible. Stem cells act as "scaffolds" for neural transplantation and may aid in repair mechanisms in stroke.
A long-term follow-up study of intravenous autologous mesenchymal stem cell transplantation in patients with ischemic stroke.
Lee Jin Soo,Hong Ji Man,Moon Gyeong Joon,Lee Phil Hyu,Ahn Young Hwan,Bang Oh Young,
Stem cells (Dayton, Ohio)
We previously evaluated the short-term follow-up preliminary data of mesenchymal stem cells (MSCs) transplantation in patients with ischemic stroke. The present study was conducted to evaluate the long-term safety and efficacy of i.v. MSCs transplantation in a larger population. To accomplish this, we performed an open-label, observer-blinded clinical trial of 85 patients with severe middle cerebral artery territory infarct. Patients were randomly allocated to one of two groups, those who received i.v. autologous ex vivo cultured MSCs (MSC group) or those who did not (control group), and followed for up to 5 years. Mortality of any cause, long-term side effects, and new-onset comorbidities were monitored. Of the 52 patients who were finally included in this study, 16 were the MSC group and 36 were the control group. Four (25%) patients in the MSC group and 21 (58.3%) in the control group died during the follow-up period, and the cumulative surviving portion at 260 weeks was 0.72 in the MSC group and 0.34 in the control group (log-rank; p = .058). Significant side effects were not observed following MSC treatment. The occurrence of comorbidities including seizures and recurrent vascular episodes did not differ between groups. When compared with the control group, the follow-up modified Rankin Scale (mRS) score was decreased, whereas the number of patients with a mRS of 0-3 increased in the MSC group (p = .046). Clinical improvement in the MSC group was associated with serum levels of stromal cell-derived factor-1 and the degree of involvement of the subventricular region of the lateral ventricle. Intravenous autologous MSCs transplantation was safe for stroke patients during long-term follow-up. This therapy may improve recovery after stroke depending on the specific characteristics of the patients.
Autologous mesenchymal stem cells in chronic stroke.
Bhasin Ashu,Srivastava M V Padma,Kumaran S Senthil,Mohanty Sujata,Bhatia Rohit,Bose Sushmita,Gaikwad Shailesh,Garg Ajay,Airan Balram
Cerebrovascular diseases extra
BACKGROUND:Cell transplantation is a 'hype and hope' in the current scenario. It is in the early stage of development with promises to restore function in chronic diseases. Mesenchymal stem cell (MSC) transplantation in stroke patients has shown significant improvement by reducing clinical and functional deficits. They are feasible and multipotent and have homing characteristics. This study evaluates the safety, feasibility and efficacy of autologous MSC transplantation in patients with chronic stroke using clinical scores and functional imaging (blood oxygen level-dependent and diffusion tensor imaging techniques). METHODS:Twelve chronic stroke patients were recruited; inclusion criteria were stroke lasting 3 months to 1 year, motor strength of hand muscles of at least 2, and NIHSS of 4-15, and patients had to be conscious and able to comprehend. Fugl Meyer (FM), modified Barthel index (mBI), MRC, Ashworth tone grade scale scores and functional imaging scans were assessed at baseline, and after 8 and 24 weeks. Bone marrow was aspirated under aseptic conditions and expansion of MSC took 3 weeks with animal serum-free media (Stem Pro SFM). Six patients were administered a mean of 50-60 × 10(6) cells i.v. followed by 8 weeks of physiotherapy. Six patients served as controls. This was a non-randomized experimental controlled trial. RESULTS:Clinical and radiological scanning was normal for the stem cell group patients. There was no mortality or cell-related adverse reaction. The laboratory tests on days 1, 3, 5 and 7 were also normal in the MSC group till the last follow-up. The FM and mBI showed a modest increase in the stem cell group compared to controls. There was an increased number of cluster activation of Brodmann areas BA 4 and BA 6 after stem cell infusion compared to controls, indicating neural plasticity. CONCLUSION:MSC therapy aiming to restore function in stroke is safe and feasible. Further randomized controlled trials are needed to evaluate its efficacy.
Stem cell-derived extracellular vesicle therapy for acute brain insults and neurodegenerative diseases.
Stem cell-based therapy is a promising approach for treating a variety of disorders, including acute brain insults and neurodegenerative diseases. Stem cells such as mesenchymal stem cells (MSCs) secrete extracellular vesicles (EVs), circular membrane fragments (30 nm-1 μm) that are shed from the cell surface, carrying several therapeutic molecules such as proteins and microRNAs. Because EV-based therapy is superior to cell therapy in terms of scalable production, biodistribution, and safety profiles, it can be used to treat brain diseases as an alternative to stem cell therapy. This review presents evidences evaluating the role of stem cell-derived EVs in stroke, traumatic brain injury, and degenerative brain diseases, such as Alzheimer's disease and Parkinson' disease. In addition, stem cell-derived EVs have better profiles in biocompatibility, immunogenicity, and safety than those of small chemical and macromolecules. The advantages and disadvantages of EVs compared with other strategies are discussed. Even though EVs obtained from native stem cells have potential in the treatment of brain diseases, the successful clinical application is limited by the short half-life, limited targeting, rapid clearance after application, and insufficient payload. We discuss the strategies to enhance the efficacy of EV therapeutics. Finally, EV therapies have yet to be approved by the regulatory authorities. Major issues are discussed together with relevant advances in the clinical application of EV therapeutics. [BMB Reports 2022; 55(1): 20-29].
Mesenchymal Stromal Cells (MSCs): A Promising Tool for Cell-Based Angiogenic Therapy.
Ulpiano Cristiana,da Silva Cláudia L,Monteiro Gabriel A
Current gene therapy
The Mesenchymal stromal cells (MSCs) are a diverse subset of adult multipotent precursors, known for their potential therapeutic properties in regenerative medicine mainly sustained by paracrine effects through secretion of a variety of biologically active molecules. MSC secretome includes a wide range of soluble protein factors, composed of growth factors and cytokines, and vesicular components, which transfer proteins and genetic material modulating the host microenvironment. In particular, MSC-derived secretome mediates the different steps of the angiogenic process, inducing endothelial cell functions in vitro and promoting angiogenesis in vivo. As a result, MSCs have been widely explored as a promising cell-based therapy in diseases caused by insufficient angiogenesis. Numerous studies of myocardial infarction, ischemic stroke, and critical limb ischemia in animals have shown that human MSCs can enhance angiogenesis and accelerate tissue regeneration. This extensive preclinical work encouraged the study of these remarkable cells for the treatment of these disorders in human clinical settings. The present review provides a comprehensive overview of the pro-angiogenic potential of MSCs and paracrine effectors of their secretome. In addition, bioengineering strategies, including ex vivo preconditioning and genetic modification approaches, to enhance MSC innate angiogenic properties, and thereby therapeutic potency, will be presented. Finally, an update on completed preclinical and clinical studies with MSCs for the treatment of ischemia-related diseases will be discussed.
Neuroprotective effect of stroke pretreated MSCs against cerebral ischemia/reperfusion injury in rats.
Tang Wenxue,Lv Xin,Huang Jinxiu,Wang Baiyong,Lin Leqing,Shen Yueliang,Yao Yanmei
BACKGROUND:Mesenchymal stem cells (MSCs) have been shown to enhance neurological recovery after stroke. In this study, a middle cerebral artery occlusion (MCAO) rat model was designed to assess the neuroprotective effects of stroke pretreated MSCs on cerebral ischemia/reperfusion (I/R) injury. METHOD:The MSCs were isolated and cultured in medium with 10% FBS, normal control serum (NS), or stroke serum (SS). Then these MSCs were injected into each group rats (n=6) 1 day after MCAO and continue feeding for 28 days. A battery of behavioral tests, TTC staining, hematoxylin and eosin staining, ELISA, and TUNEL assay were used to assess neural injury. Moreover, in order to detected the enhancement of neuronal regeneration and angiogenesis, immunofluorescence and western blotting were performed to assess the expression of trophic factor and growth factor. RESULT:After treatment, the behaviour was improvement significantly. And the infarct area, brain lesion, and apoptosis cells were significantly decreased in the SS-MSCs group than others. It also modulated the inflammation by attenuating inflammatory cytokines. In addition, the number of neurogenesis positive cells, the expression of trophic factors and growth factors were significantly higher in the SS-MSCs group than others. At the same time, FBS-MSCs and NS-MSCs showed differences in the expression of trophic factors and growth factors, but the results were not as well as SS-MSCs. CONCLUSION:SS-MSCs administration after the reperfusion led to neuroprotection by improving pathological changes, behavioral improvement, neurogenesis, suppression of apoptosis and inflammatory as well as angiogenesis.
Safety and Efficacy of Intraventricular Delivery of Bone Marrow-Derived Mesenchymal Stem Cells in Hemorrhagic Stroke Model.
Huang Peng,Freeman William D,Edenfield Brandy H,Brott Thomas G,Meschia James F,Zubair Abba C
External ventricular drain (EVD) is used clinically to relieve intracranial pressure and occasionally to deliver medications following intracerebral hemorrhage (ICH). Mesenchymal stem cell (MSC) therapy has been shown to be neuroprotective and can induce neuroregeneration in stroke models. We evaluated the safety and efficacy of delivering MSCs intraventricularly in a rat hemorrhagic stroke model. Using autologous blood, hemorrhagic stroke was induced at specific coordinates in the right basal ganglia. After 30 minutes, rats were treated with either bone marrow-derived MSCs or a phosphate-buffered saline placebo via direct intraventricular infusion. Three dosages (2 × 10/kg, 5 × 10/kg, and 1 × 10/kg) of MSCs were administered. Forelimb use asymmetry test was employed to evaluate functional improvement after cell therapy. At the end of the experiment, peripheral blood samples and organs were harvested; biochemistry, cytokine, and growth factor analysis and histology evaluations were performed to explore cell toxicity and cell fate, and the effects of MSC therapy on injury volume, anti-inflammation, and neurogenesis. Intraventricular administration of MSCs in ICH rat model showed improved behavior and alleviated brain damage. Additionally, treated ICH rats showed significantly reduced expression of IL-1α, IL-6, and IFN-γ. No obvious cell toxicity was noticed through blood chemistry and histology evaluation. None of the infused MSCs were detected at the end of the experiment. EVD is safe and effective to use as a method of delivering MSCs to treat ICH. Intraventricularly delivered MSCs have anti-inflammatory properties and a capacity to induce neurogenesis and improve function following ICH injury.
Considering risk factors for the effectiveness of translational therapies in brain stroke.
Gutiérrez-Vargas Johanna Andrea,Cardona-Gómez Gloria Patricia
Journal of the neurological sciences
Multiple studies on cerebral ischemia have been performed in animal models to propose different strategies of neuroprotection that mitigate either the early or late consequences of the disease. These therapies have been successful in reducing the volume of infarction, the proinflammatory cascade, and the amount of free radicals, as well as reversing markers of neurodegeneration, among other events. However, when those strategies are translated to clinical studies, their effectiveness is not reproduced. This review will focus on highlighting some of the main limitations of the animal models of stroke that lead to unsuccessful translational therapies and the common risk factors in humans that should be carefully considered in the experimental design of future studies to generate a more realistic spatiotemporal physiopathology and improve therapeutic efficacy in cerebral ischemia.
Promoting Brain Repair and Regeneration After Stroke: a Plea for Cell-Based Therapies.
Dabrowski Ania,Robinson Thomas J,Felling Ryan J
Current neurology and neuroscience reports
PURPOSE OF REVIEW:After decades of hype, cell-based therapies are emerging into the clinical arena for the purposes of promoting recovery after stroke. In this review, we discuss the most recent science behind the role of cell-based therapies in ischemic stroke and the efforts to translate these therapies into human clinical trials. RECENT FINDINGS:Preclinical data support numerous beneficial effects of cell-based therapies in both small and large animal models of ischemic stroke. These benefits are driven by multifaceted mechanisms promoting brain repair through immunomodulation, trophic support, circuit reorganization, and cell replacement. Cell-based therapies offer tremendous potential for improving outcomes after stroke through multimodal support of brain repair. Based on recent clinical trials, cell-based therapies appear both feasible and safe in all phases of stroke. Ongoing translational research and clinical trials will further refine these therapies and have the potential to transform the approach to stroke recovery and rehabilitation.
Safety and efficacy of multipotent adult progenitor cells in acute ischaemic stroke (MASTERS): a randomised, double-blind, placebo-controlled, phase 2 trial.
Hess David C,Wechsler Lawrence R,Clark Wayne M,Savitz Sean I,Ford Gary A,Chiu David,Yavagal Dileep R,Uchino Ken,Liebeskind David S,Auchus Alexander P,Sen Souvik,Sila Cathy A,Vest Jeffrey D,Mays Robert W
The Lancet. Neurology
BACKGROUND:Multipotent adult progenitor cells are a bone marrow-derived, allogeneic, cell therapy product that modulates the immune system, and represents a promising therapy for acute stroke. We aimed to identify the highest, well-tolerated, and safest single dose of multipotent adult progenitor cells, and if they were efficacious as a treatment for stroke recovery. METHODS:We did a phase 2, randomised, double-blind, placebo-controlled, dose-escalation trial of intravenous multipotent adult progenitor cells in 33 centres in the UK and the USA. We used a computer-generated randomisation sequence and interactive voice and web response system to assign patients aged 18-83 years with moderately severe acute ischaemic stroke and a National Institutes of Health Stroke Scale (NIHSS) score of 8-20 to treatment with intravenous multipotent adult progenitor cells (400 million or 1200 million cells) or placebo between 24 h and 48 h after symptom onset. Patients were ineligible if there was a change in NIHSS of four or more points during at least a 6 h period between screening and randomisation, had brainstem or lacunar infarct, a substantial comorbid disease, an inability to undergo an MRI scan, or had a history of splenectomy. In group 1, patients were enrolled and randomly assigned in a 3:1 ratio to receive 400 million cells or placebo and assessed for safety through 7 days. In group 2, patients were randomly assigned in a 3:1 ratio to receive 1200 million cells or placebo and assessed for safety through the first 7 days. In group 3, patients were enrolled, randomly assigned, and stratified by baseline NIHSS score to receive 1200 million cells or placebo in a 1:1 ratio within 24-48 h. Patients, investigators, and clinicians were masked to treatment assignment. The primary safety outcome was dose-limiting toxicity effects. The primary efficacy endpoint was global stroke recovery, which combines dichotomised results from the modified Rankin scale, change in NIHSS score from baseline, and Barthel index at day 90. Analysis was by intention to treat (ITT) including all patients in groups 2 and 3 who received the investigational agent or placebo. This study is registered with ClinicalTrials.gov, number NCT01436487. FINDINGS:The study was done between Oct 24, 2011, and Dec 7, 2015. After safety assessments in eight patients in group 1, 129 patients were randomly assigned (67 to receive multipotent adult progenitor cells and 62 to receive placebo) in groups 2 and 3 (1200 million cells). The ITT populations consisted of 65 patients who received multipotent adult progenitor cells and 61 patients who received placebo. There were no dose-limiting toxicity events in either group. There were no infusional or allergic reactions and no difference in treatment-emergent adverse events between the groups (64 [99%] of 65 patients in the multipotent adult progenitor cell group vs 59 [97%] of 61 in the placebo group). There was no difference between the multipotent adult progenitor cell group and placebo groups in global stroke recovery at day 90 (odds ratio 1·08 [95% CI 0·55-2·09], p=0·83). INTERPRETATION:Administration of multipotent adult progenitor cells was safe and well tolerated in patients with acute ischaemic stroke. Although no significant improvement was observed at 90 days in neurological outcomes with multipotent adult progenitor cells treatment, further clinical trials evaluating the efficacy of the intervention in an earlier time window after stroke (<36 h) are planned. FUNDING:Athersys Inc.
Intra-arterial bone marrow mononuclear cells in ischemic stroke: a pilot clinical trial.
Moniche Francisco,Gonzalez Alejandro,Gonzalez-Marcos Jose-Ramon,Carmona Magdalena,Piñero Pilar,Espigado Ildefonso,Garcia-Solis David,Cayuela Aurelio,Montaner Joan,Boada Cristina,Rosell Anna,Jimenez Maria-Dolores,Mayol Antonio,Gil-Peralta Alberto
BACKGROUND AND PURPOSE:Bone marrow mononuclear cell (BM-MNC) intra-arterial transplantation improves recovery in experimental models of ischemic stroke. We aimed to assess the safety, feasibility, and biological effects of autologous BM-MNC transplantation in patients with stroke. METHODS:A single-blind (outcomes assessor) controlled Phase I/II trial was conducted in patients with middle cerebral artery stroke. Autologous BM-MNCs were injected intra-arterially between 5 and 9 days after stroke. Follow-up was done for up to 6 months and blood samples were collected for biological markers. The primary outcome was safety and feasibility of the procedure. The secondary outcome was improvement in neurological function. RESULTS:Ten cases (BM-MNC-treated) and 10 control subjects (BM-MNC-nontreated) were consecutively included. Mean National Institutes of Health Stroke Scale before the procedure was 15.6. Mean BM-MNCs injected were 1.59×10(8). There was no death, stroke recurrence, or tumor formation during follow-up, although 2 cases had an isolate partial seizure at 3 months. After transplantation, higher plasma levels of beta nerve growth factor (β-nerve growth factor) were found compared with control subjects (P=0.02). There were no significant differences in neurological function at 180 days. A trend to positive correlation between number of CD34+ cells injected and Barthel Index was found (r=0.56, P=0.09). CONCLUSIONS:Intra-arterial BM-MNC transplantation in subacute ischemic stroke is feasible and seems to be safe. Larger randomized trials are needed to confirm the safety and elucidate the efficacy of BM-MNC transplantation. CLINICAL TRIAL REGISTRATION URL:www.clinicaltrials.gov. Unique identifier: NCT00761982.
Hypoxia/Reoxygenation-Preconditioned Human Bone Marrow-Derived Mesenchymal Stromal Cells Rescue Ischemic Rat Cortical Neurons by Enhancing Trophic Factor Release.
Kim Young Seo,Noh Min Young,Cho Kyung Ah,Kim Hyemi,Kwon Min-Soo,Kim Kyung Suk,Kim Juhan,Koh Seong-Ho,Kim Seung Hyun
Bone marrow-derived mesenchymal stromal cells (BM-MSCs) represent a promising tool for stem cell-based therapies. However, the majority of MSCs fail to reach the injury site and have only minimal therapeutic effect. In this study, we assessed whether hypoxia/reoxygenation (H/R) preconditioning of human BM-MSCs could increase their functional capacity and beneficial effect on ischemic rat cortical neurons. Human BM-MSCs were cultured under hypoxia (1% O2) and with long-term reoxygenation for various times to identify the optimal conditions for increasing their viability and proliferation. The effects of H/R preconditioning on the BM-MSCs were assessed by analyzing the expression of prosurvival genes, trophic factors, and cell migration assays. The functionally improved BM-MSCs were cocultured with ischemic rat cortical neurons to compare with normoxic cultured BM-MSCs. Although the cell viability and proliferation of BM-MSCs were reduced after 1 day of hypoxic culture (1% O2), when this was followed by 5-day reoxygenation, the BM-MSCs recovered and multiplied extensively. The immunophenotype and trilineage differentiation of BM-MSCs were also maintained under this H/R preconditioning. In addition, the preconditioning enhanced the expression of prosurvival genes, the messenger RNA (mRNA) levels of various trophic factors and migration capacity. Finally, coculture with the H/R-preconditioned BM-MSCs promoted the survival of ischemic rat cortical neurons. H/R preconditioning of BM-MSCs increases prosurvival signals, trophic factor release, and cell migration and appears to increase their ability to rescue ischemic cortical neurons. This optimized H/R preconditioning procedure could provide the basis for a new strategy for stem cell therapy in ischemic stroke patients.
The effect of hypoxia preconditioning on the neural and stemness genes expression profiling in human umbilical cord blood mesenchymal stem cells.
Kheirandish Maryam,Gavgani Somaie Piri,Samiee Shahram
Transfusion and apheresis science : official journal of the World Apheresis Association : official journal of the European Society for Haemapheresis
In recent years, human umbilical cord blood-derived mesenchymal stem cell (hUB-MSCs) has been regarded as an alternative source for stem cell therapy. In this study, we evaluated the effect of hypoxia preconditioning (HPC) on the expression of Nt-3, GFAP, Nestin, Oct-4 and Nanog genes and proliferative capacity of hUB-MSCs in comparison with normoxic conditions. HPC+Hypoxia protocol includes cultured hUB-MSCs for 15min at 2.5% O and after that reoxygenation for 30min at 21% O (HPC), and then hypoxia preconditioned hUB-MSCs subjected to 2.5% O for 72h (Hypoxia). Conclusively, the results showed that hypoxic preconditioning is an effective strategy for enhancing proliferation capacity of hUB-MSCs, and also can trigger expression of some of the neural genes. In addition, the concept of involvement of oxygen tension in the expression of some of the neural genes of hUB-MSCs would be a good sign of enhanced neural differentiation potential in vitro.
Preconditioning influences mesenchymal stem cell properties in vitro and in vivo.
Hu Chenxia,Li Lanjuan
Journal of cellular and molecular medicine
Various diseases and toxic factors easily impair cellular and organic functions in mammals. Organ transplantation is used to rescue organ function, but is limited by scarce resources. Mesenchymal stem cell (MSC)-based therapy carries promising potential in regenerative medicine because of the self-renewal and multilineage potency of MSCs; however, MSCs may lose biological functions after isolation and cultivation for a long time in vitro. Moreover, after they are injected in vivo and migrate into the damaged tissues or organs, they encounter a harsh environment coupled with death signals due to the inadequate tensegrity structure between the cells and matrix. Preconditioning, genetic modification and optimization of MSC culture conditions are key strategies to improve MSC functions in vitro and in vivo, and all of these procedures will contribute to improving MSC transplantation efficacy in tissue engineering and regenerative medicine. Preconditioning with various physical, chemical and biological factors is possible to preserve the stemness of MSCs for further application in studies and clinical tests. In this review, we mainly focus on preconditioning and the corresponding mechanisms for improving MSC activities in vitro and in vivo; we provide a glimpse into the promotion of MSC-based cell therapy development for regenerative medicine. As a promising consequence, MSC transplantation can be applied for the treatment of some terminal diseases and can prolong the survival time of patients in the near future.
Mesenchymal Stem Cell Therapy for Ischemic Tissues.
Yong Kar Wey,Choi Jane Ru,Mohammadi Mehdi,Mitha Alim P,Sanati-Nezhad Amir,Sen Arindom
Stem cells international
Ischemic diseases such as myocardial infarction, ischemic stroke, and critical limb ischemia are immense public health challenges. Current pharmacotherapy and surgical approaches are insufficient to completely heal ischemic diseases and are associated with a considerable risk of adverse effects. Alternatively, human mesenchymal stem cells (hMSCs) have been shown to exhibit immunomodulation, angiogenesis, and paracrine secretion of bioactive factors that can attenuate inflammation and promote tissue regeneration, making them a promising cell source for ischemic disease therapy. This review summarizes the pathogenesis of ischemic diseases, discusses the potential therapeutic effects and mechanisms of hMSCs for these diseases, and provides an overview of challenges of using hMSCs clinically for treating ischemic diseases.
Mesenchymal stem cells in cardiac regeneration: a detailed progress report of the last 6 years (2010-2015).
Singh Aastha,Singh Abhishek,Sen Dwaipayan
Stem cell research & therapy
Mesenchymal stem cells have been used for cardiovascular regenerative therapy for decades. These cells have been established as one of the potential therapeutic agents, following several tests in animal models and clinical trials. In the process, various sources of mesenchymal stem cells have been identified which help in cardiac regeneration by either revitalizing the cardiac stem cells or revascularizing the arteries and veins of the heart. Although mesenchymal cell therapy has achieved considerable admiration, some challenges still remain that need to be overcome in order to establish it as a successful technique. This in-depth review is an attempt to summarize the major sources of mesenchymal stem cells involved in myocardial regeneration, the significant mechanisms involved in the process with a focus on studies (human and animal) conducted in the last 6 years and the challenges that remain to be addressed.
Mesenchymal stem cells: immunobiology and role in immunomodulation and tissue regeneration.
Kode Jyoti A,Mukherjee Shayanti,Joglekar Mugdha V,Hardikar Anandwardhan A
Mesenchymal stem cells (MSC) are multipotent cells that differentiate into osteoblasts, myocytes, chondrocytes and adipocytes as well as insulin-producing cells. The mechanism underlying their in vivo differentiation is not clear and is thought to be caused by spontaneous cell fusion or factors present in the microenvironment. However, their ease of isolation, high 'ex-vivo' expansion potential and ability to differentiate into multiple lineages make them attractive tools for potential use in cell therapy. MSC have been isolated from several tissues, including bone/bone marrow, fat, Wharton's jelly, umbilical cord blood, placenta and pancreas. The 'immunosuppressive' property of human MSC makes them an important candidate for cellular therapy in allogeneic settings. Use of allogeneic MSC for repair of large defects may be an alternative to autologous and allogeneic tissue-grafting procedures. An allogeneic approach would enable MSC to be isolated from any donor, expanded and cryopreserved, providing a readily available source of progenitors for cell replacement therapy. Their immunomodulatory properties have raised the possibility of establishing allogeneic MSC banks for tissue regeneration. These facts are strongly reflected in the current exponential growth in stem cell research in the pharmaceutical and biotechnology communities. Current knowledge regarding the immunobiology and clinical application of MSC needs to be strengthened further to establish MSC as a safe and effective therapeutic tool in regenerative medicine. This paper discusses human MSC with particular reference to the expression of their surface markers, their role as immunomodulators and their multilineage differentiation potential and possible use in tissue regeneration and repair.
The PI3k/Akt pathway is associated with angiogenesis, oxidative stress and survival of mesenchymal stem cells in pathophysiologic condition in ischemia.
Samakova A,Gazova A,Sabova N,Valaskova S,Jurikova M,Kyselovic J
Ischemic diseases are characterized by reduced blood supply to a tissue or an organ due to obstruction of blood vessels. The most serious and most common ischemic diseases include ischemic heart disease, ischemic stroke, and critical limb ischemia. Revascularization is the first choice of therapy, but the cell therapy is being introduced as a possible way of treatment for no-option patients. One of the possibilities of cell therapy is the use of mesenchymal stem cells (MSCs). MSCs are easily isolated from bone marrow and can be defined as non-hematopoietic multipotent adult stem cells population with a defined capacity for self-renewal and differentiation into cell types of all three germ layers depending on their origin. Since 1974, when Friedenstein and coworkers (Friedenstein et al. 1974) first time isolated and characterized MSCs, MSC-based therapy has been shown to be safe and effective. Nevertheless, many scientists and clinical researchers want to improve the success of MSCs in regenerative therapy. The secret of successful cell therapy may lie, along with the homing, in secretion of biologically active molecules including cytokines, growth factors, and chemokines known as MSCs secretome. One of the intracellular signalling mechanism includes the activity of phosphatidylinositol-3-kinase (phosphoinositide 3-kinase) (PI3K) - protein kinase B (serine-threonine protein kinase Akt) (Akt) pathway. This PI3K/Akt pathway plays key roles in many cell types in regulating cell proliferation, differentiation, apoptosis, and migration. Pre-conditioning of MSCs could improve efficacy of signalling mechanism.
Mesenchymal stem cells and their use in therapy: what has been achieved?
Fernández Vallone V B,Romaniuk M A,Choi H,Labovsky V,Otaegui J,Chasseing N A
Differentiation; research in biological diversity
The considerable therapeutic potential of human multipotent mesenchymal stromal cells or mesenchymal stem cells (MSCs) has generated increasing interest in a wide variety of biomedical disciplines. Nevertheless, researchers report studies on MSCs using different methods of isolation and expansion, as well as different approaches to characterize them; therefore, it is increasingly difficult to compare and contrast study outcomes. To begin to address this issue, the Mesenchymal and Tissue Stem Cell Committee of the International Society for Cellular Therapy proposed minimal criteria to define human MSCs. First, MSCs must be plastic-adherent when maintained in standard culture conditions (α minimal essential medium plus 20% fetal bovine serum). Second, MSCs must express CD105, CD73 and CD90, and MSCs must lack expression of CD45, CD34, CD14 or CD11b, CD79α or CD19 and HLA-DR surface molecules. Third, MSCs must differentiate into osteoblasts, adipocytes and chondroblasts in vitro. MSCs are isolated from many adult tissues, in particular from bone marrow and adipose tissue. Along with their capacity to differentiate and transdifferentiate into cells of different lineages, these cells have also generated great interest for their ability to display immunomodulatory capacities. Indeed, a major breakthrough was the finding that MSCs are able to induce peripheral tolerance, suggesting that they may be used as therapeutic tools in immune-mediated disorders. Although no significant adverse events have been reported in clinical trials to date, all interventional therapies have some inherent risks. Potential risks for undesirable events, such as tumor development, that might occur while using these stem cells for therapy must be taken into account and contrasted against the potential benefits to patients.
Neurorestoration after stroke.
Azad Tej D,Veeravagu Anand,Steinberg Gary K
Recent advancements in stem cell biology and neuromodulation have ushered in a battery of new neurorestorative therapies for ischemic stroke. While the understanding of stroke pathophysiology has matured, the ability to restore patients' quality of life remains inadequate. New therapeutic approaches, including cell transplantation and neurostimulation, focus on reestablishing the circuits disrupted by ischemia through multidimensional mechanisms to improve neuroplasticity and remodeling. The authors provide a broad overview of stroke pathophysiology and existing therapies to highlight the scientific and clinical implications of neurorestorative therapies for stroke.
Clinical Trials With Mesenchymal Stem Cells: An Update.
Squillaro Tiziana,Peluso Gianfranco,Galderisi Umberto
In the last year, the promising features of mesenchymal stem cells (MSCs), including their regenerative properties and ability to differentiate into diverse cell lineages, have generated great interest among researchers whose work has offered intriguing perspectives on cell-based therapies for various diseases. Currently the most commonly used adult stem cells in regenerative medicine, MSCs, can be isolated from several tissues, exhibit a strong capacity for replication in vitro, and can differentiate into osteoblasts, chondrocytes, and adipocytes. However, heterogeneous procedures for isolating and cultivating MSCs among laboratories have prompted the International Society for Cellular Therapy (ISCT) to issue criteria for identifying unique populations of these cells. Consequently, the isolation of MSCs according to ISCT criteria has produced heterogeneous, nonclonal cultures of stromal cells containing stem cells with different multipotent properties, committed progenitors, and differentiated cells. Though the nature and functions of MSCs remain unclear, nonclonal stromal cultures obtained from bone marrow and other tissues currently serve as sources of putative MSCs for therapeutic purposes, and several findings underscore their effectiveness in treating different diseases. To date, 493 MSC-based clinical trials, either complete or ongoing, appear in the database of the US National Institutes of Health. In the present article, we provide a comprehensive review of MSC-based clinical trials conducted worldwide that scrutinizes biological properties of MSCs, elucidates recent clinical findings and clinical trial phases of investigation, highlights therapeutic effects of MSCs, and identifies principal criticisms of the use of these cells. In particular, we analyze clinical trials using MSCs for representative diseases, including hematological disease, graft-versus-host disease, organ transplantation, diabetes, inflammatory diseases, and diseases in the liver, kidney, and lung, as well as cardiovascular, bone and cartilage, neurological, and autoimmune diseases.
Shattering barriers toward clinically meaningful MSC therapies.
Levy Oren,Kuai Rui,Siren Erika M J,Bhere Deepak,Milton Yuka,Nissar Nabeel,De Biasio Michael,Heinelt Martina,Reeve Brock,Abdi Reza,Alturki Meshael,Fallatah Mohanad,Almalik Abdulaziz,Alhasan Ali H,Shah Khalid,Karp Jeffrey M
More than 1050 clinical trials are registered at FDA.gov that explore multipotent mesenchymal stromal cells (MSCs) for nearly every clinical application imaginable, including neurodegenerative and cardiac disorders, perianal fistulas, graft-versus-host disease, COVID-19, and cancer. Several companies have or are in the process of commercializing MSC-based therapies. However, most of the clinical-stage MSC therapies have been unable to meet primary efficacy end points. The innate therapeutic functions of MSCs administered to humans are not as robust as demonstrated in preclinical studies, and in general, the translation of cell-based therapy is impaired by a myriad of steps that introduce heterogeneity. In this review, we discuss the major clinical challenges with MSC therapies, the details of these challenges, and the potential bioengineering approaches that leverage the unique biology of MSCs to overcome the challenges and achieve more potent and versatile therapies.
Gingiva-Derived Mesenchymal Stem Cells: Potential Application in Tissue Engineering and Regenerative Medicine - A Comprehensive Review.
Kim Dane,Lee Alisa E,Xu Qilin,Zhang Qunzhou,Le Anh D
Frontiers in immunology
A unique subpopulation of mesenchymal stem cells (MSCs) has been isolated and characterized from human gingival tissues (GMSCs). Similar to MSCs derived from other sources of tissues, e.g. bone marrow, adipose or umbilical cord, GMSCs also possess multipotent differentiation capacities and potent immunomodulatory effects on both innate and adaptive immune cells through the secretion of various types of bioactive factors with immunosuppressive and anti-inflammatory functions. Uniquely, GMSCs are highly proliferative and have the propensity to differentiate into neural cell lineages due to the neural crest-origin. These properties have endowed GMSCs with potent regenerative and therapeutic potentials in various preclinical models of human disorders, particularly, some inflammatory and autoimmune diseases, skin diseases, oral and maxillofacial disorders, and peripheral nerve injuries. All types of cells release extracellular vesicles (EVs), including exosomes, that play critical roles in cell-cell communication through their cargos containing a variety of bioactive molecules, such as proteins, nucleic acids, and lipids. Like EVs released by other sources of MSCs, GMSC-derived EVs have been shown to possess similar biological functions and therapeutic effects on several preclinical diseases models as GMSCs, thus representing a promising cell-free platform for regenerative therapy. Taken together, due to the easily accessibility and less morbidity of harvesting gingival tissues as well as the potent immunomodulatory and anti-inflammatory functions, GMSCs represent a unique source of MSCs of a neural crest-origin for potential application in tissue engineering and regenerative therapy.
Impurity of stem cell graft by murine embryonic fibroblasts - implications for cell-based therapy of the central nervous system.
Molcanyi Marek,Mehrjardi Narges Zare,Schäfer Ute,Haj-Yasein Nadia Nabil,Brockmann Michael,Penner Marina,Riess Peter,Reinshagen Clemens,Rieger Bernhard,Hannes Tobias,Hescheler Jürgen,Bosche Bert
Frontiers in cellular neuroscience
Stem cells have been demonstrated to possess a therapeutic potential in experimental models of various central nervous system disorders, including stroke. The types of implanted cells appear to play a crucial role. Previously, groups of the stem cell network NRW implemented a feeder-based cell line within the scope of their projects, examining the implantation of stem cells after ischemic stroke and traumatic brain injury. Retrospective evaluation indicated the presence of spindle-shaped cells in several grafts implanted in injured animals, which indicated potential contamination by co-cultured feeder cells (murine embryonic fibroblasts - MEFs). Because feeder-based cell lines have been previously exposed to a justified criticism with regard to contamination by animal glycans, we aimed to evaluate the effects of stem cell/MEF co-transplantation. MEFs accounted for 5.3 ± 2.8% of all cells in the primary FACS-evaluated co-culture. Depending on the culture conditions and subsequent purification procedure, the MEF-fraction ranged from 0.9 to 9.9% of the cell suspensions in vitro. MEF survival and related formation of extracellular substances in vivo were observed after implantation into the uninjured rat brain. Impurity of the stem cell graft by MEFs interferes with translational strategies, which represents a threat to the potential recipient and may affect the graft microenvironment. The implications of these findings are critically discussed.
Pitfalls and fallacies interfering with correct identification of embryonic stem cells implanted into the brain after experimental traumatic injury.
Molcanyi Marek,Bosche Bert,Kraitsy Klaus,Patz Silke,Zivcak Jozef,Riess Peter,El Majdoub Faycal,Hescheler Jürgen,Goldbrunner Roland,Schäfer Ute
Journal of neuroscience methods
Cell-therapy was proposed to be a promising tool in case of death or impairment of specific cell types. Correct identification of implanted cells became crucial when evaluating the success of transplantation therapy. Various methods of cell labeling have been employed in previously published studies. The use of intrinsic signaling of green fluorescent protein (GFP) has led to a well known controversy in the field of cardiovascular research. We encountered similar methodological pitfalls after transplantation of GFP-transfected embryonic stem cells into rat brains following traumatic brain injury (TBI). As the identification of implanted graft by intrinsic autofluorescence failed, anti-GFP labeling coupled to fluorescent and conventional antibodies was needed to visualize the implanted cells. Furthermore, different cell types with strong intrinsic autofluorescence were found at the sites of injury and transplantation, thus mimicking the implanted stem cells. GFP-positive stem cells were correctly localized, using advanced histological techniques. The activation of microglia/macrophages, accompanying the transplantation post TBI, was shown to be a significant source of artefacts, interfering with correct identification of implanted stem cells. Dependent on the strategy of stem cell tracking, the phagocytosis of implanted cells as observed in this study, might also impede the interpretation of results. Critical appraisal of previously published data as well as a review of different histological techniques provide tools for a more accurate identification of transplanted stem cells.
Extract derived from rat brains in the acute phase following traumatic brain injury impairs survival of undifferentiated stem cells and induces rapid differentiation of surviving cells.
Bentz Kristine,Molcanyi Marek,Schneider Annette,Riess Peter,Maegele Marc,Bosche Bert,Hampl Jürgen A,Hescheler Jürgen,Patz Silke,Schäfer Ute
Cellular physiology and biochemistry : international journal of experimental cellular physiology, biochemistry, and pharmacology
Dramatic cerebral responses following brain injury (TBI) comprise inflammation, cell death, and modulation of trophic factor release. These cerebral modulations might induce and/or attenuate acute neuronal damage. Here, we investigated the effect of tissue extract derived from healthy (HBE) or injured rat brain (TBE) on the differentiation of cultured embryonic stem cells in vitro. Rats were sacrificed at t = 45 minutes following lateral fluid-percussion injury and extracts of cerebral tissue were prepared from 4-6 healthy or injured rat brain hemispheres. Murine embryonic stem cells (CGR8) cultured in serum-free medium were then conditioned for a week with HBE or TBE. Omission of serum from the culture medium induced neural differentiation of CGR8 stem cells, as indicated by a significant time dependent down-regulation of oct-4 with a concomitant upregulation of nestin after 7 days. In parallel cell loss was observed that seemed to be largely due to apoptotic cell death. In TBE treated cells, on the other hand, a significant amplification of apoptotic cell death, enhancement of nestin and MAP2 expression and marked morphological changes such as axonal-like outgrowth was observed within 3 days of conditioning. Treatment of stem cells with HBE resulted in less pronounced neuronal differentiation processes. Axonal-like outgrowth was not observed. Our data suggest that during the early acute phase of traumatic injury the cerebral environment is disposed to detrimental as well as potent protective signals that seem to rapidly induce neurogenic processes.
Iron and intracerebral hemorrhage: from mechanism to translation.
Xiong Xiao-Yi,Wang Jian,Qian Zhong-Ming,Yang Qing-Wu
Translational stroke research
Intracerebral hemorrhage (ICH) is a leading cause of morbidity and mortality around the world. Currently, there is no effective medical treatment available to improve functional outcomes in patients with ICH due to its unknown mechanisms of damage. Increasing evidence has shown that the metabolic products of erythrocytes are the key contributor of ICH-induced secondary brain injury. Iron, an important metabolic product that accumulates in the brain parenchyma, has a detrimental effect on secondary injury following ICH. Because the damage mechanism of iron during ICH-induced secondary injury is clear, iron removal therapy research on animal models is effective. Although many animal and clinical studies have been conducted, the exact metabolic pathways of iron and the mechanisms of iron removal treatments are still not clear. This review summarizes recent progress concerning the iron metabolism mechanisms underlying ICH-induced injury. We focus on iron, brain iron metabolism, the role of iron in oxidative injury, and iron removal therapy following ICH, and we suggest that further studies focus on brain iron metabolism after ICH and the mechanism for iron removal therapy.
Effect of hemopexin treatment on outcome after intracerebral hemorrhage in mice.
Chen-Roetling Jing,Li Yang,Cao Yang,Yan Zhe,Lu Xiangping,Regan Raymond F
Heme release from hemoglobin may contribute to secondary injury after intracerebral hemorrhage (ICH). The primary endogenous defense against heme toxicity is hemopexin, a 57 kDa glycoprotein that is depleted in the CNS after hemorrhagic stroke. We hypothesized that systemic administration of exogenous hemopexin would reduce perihematomal injury and improve outcome after experimental ICH. Intraperitoneal treatment with purified human plasma hemopexin beginning 2 h after striatal ICH induction and repeated daily for the following two days reduced blood-brain barrier disruption and cell death at 3 days. However, it had no effect on neurological deficits at 4 or 7 days or striatal cell viability at 8 days. Continuous daily hemopexin administration had no effect on striatal heme content at 3 or 7 days, and did not attenuate neurological deficits, inflammatory cell infiltration, or perihematomal cell viability at 8 days. These results suggest that systemic hemopexin treatment reduces early injury after ICH, but this effect is not sustained, perhaps due to an imbalance between striatal tissue heme and hemopexin content at later time points. Future studies should investigate its effect when administered by methods that more efficiently target CNS delivery.
Animal models of spontaneous intracerebral hemorrhage.
Alharbi Bader Murshed,Tso Michael K,Macdonald R Loch
OBJECTIVES:Intracerebral hemorrhage (ICH) is a type of stroke that results in significant mortality and morbidity. Currently there is no definitive treatment for this disease. The paucity of animal models that reflect the heterogeneity of this spontaneous human disease could be the reason. METHODS:In this review, we searched the literature for animal models of spontaneous ICH and found eight relevant papers. RESULTS:Two were related to hypertension and six were related to cerebral amyloid angiopathy (CAA). One model used double transgenic mice overexpressing human renin and angiotensinogen which caused the mice to be hypertensive. Induction of ICH, however required addition of a high salt diet and nitric oxide synthase inhibition. Another mouse model of hypertension employed subcutaneous angiotensin II infusion and nitric oxide synthase inhibition plus acute injections of angiotensin to further elevate blood pressure. Five CAA models were in transgenic mice overexpressing amyloid precursor protein. One relied on the natural development of CAA in squirrel monkeys. CONCLUSIONS:While all of the spontaneous ICH models have some advantages, the disadvantages include the sporadic time of onset of ICH and variability in size and location of ICH. Since there are no known efficacious treatments for ICH, it is not known if findings in the animal models will find treatments that are effective in humans.
Complex Clearance Mechanisms After Intraventricular Hemorrhage and rt-PA Treatment-a Review on Clinical Trials.
Bosche Bert,Mergenthaler Philipp,Doeppner Thorsten R,Hescheler Jürgen,Molcanyi Marek
Translational stroke research
Intracerebral hemorrhage in combination with intraventricular hemorrhage (IVH) is a severe type of stroke frequently leading to prolonged clinical care, continuous disability, shunt dependency, and high mortality. The molecular mechanisms induced by IVH are complex and not fully understood. Moreover, the treatment options for IVH are limited. Intraventricular recombinant tissue plasminogen activator (rt-PA) dissolves the blood clot in the ventricular system; however, whether the clinical outcome is thereby positively affected is still being debated. The mechanistic cascade induced by intraventricular rt-PA therapy may cure and harm in parallel. Despite the fact that intraventricular blood clots are thereby dissolved, blood derivatives enter the parenchyma and may still adversely affect functional structures of the brain: Smaller blood clots may obstruct the perivascular (Virchow-Robin) space and thereby the glymphatic system with detrimental consequences for cerebrospinal fluid (CSF)/interstitial fluid (ISF) flow. These clots, blood cells but also blood derivatives in the perivascular space, destabilize the blood-brain barrier from the brain parenchyma side, thereby also functionally weakening the neurovascular unit. This may lead to further accommodation of serum proteins in the ISF and particularly in the perivascular space further contributing to the adverse effects on the neuronal microenvironment. Finally, the arterial (Pacchionian) granulations have to cope with ISF containing this "blood, cell, and protein cocktail," resulting in obstruction and insufficient function of the arterial granulations, followed by a malresorptive hydrocephalus. Particularly in light of currently improved knowledge on the physiologic and pathophysiologic clearance of cerebrospinal fluid and interstitial fluid, a critical discussion and reevaluation of our current therapeutic strategies to treat intraventricular hemorrhages are needed to successfully treat patients suffering from this severe type of stroke. In this review, we therefore summarize and discuss recent clinical trials and future directions for the field of IVH with respect to the currently increased understanding of the glymphatic system and the neurovascular unit pathophysiology.
TOOTH (The Open study Of dental pulp stem cell Therapy in Humans): Study protocol for evaluating safety and feasibility of autologous human adult dental pulp stem cell therapy in patients with chronic disability after stroke.
Nagpal Anjali,Kremer Karlea L,Hamilton-Bruce Monica A,Kaidonis Xenia,Milton Austin G,Levi Christopher,Shi Songtao,Carey Leeanne,Hillier Susan,Rose Miranda,Zacest Andrew,Takhar Parabjit,Koblar Simon A
International journal of stroke : official journal of the International Stroke Society
RATIONALE:Stroke represents a significant global disease burden. As of 2015, there is no chemical or biological therapy proven to actively enhance neurological recovery during the chronic phase post-stroke. Globally, cell-based therapy in stroke is at the stage of clinical translation and may improve neurological function through various mechanisms such as neural replacement, neuroprotection, angiogenesis, immuno-modulation, and neuroplasticity. Preclinical evidence in a rodent model of middle cerebral artery ischemic stroke as reported in four independent studies indicates improvement in neurobehavioral function with adult human dental pulp stem cell therapy. Human adult dental pulp stem cells present an exciting potential therapeutic option for improving post-stroke disability. AIMS:TOOTH (The Open study Of dental pulp stem cell Therapy in Humans) will investigate the use of autologous stem cell therapy for stroke survivors with chronic disability, with the following objectives: (a) determine the maximum tolerable dose of autologous dental pulp stem cell therapy; (b) define that dental pulp stem cell therapy at the maximum tolerable dose is safe and feasible in chronic stroke; and (c) estimate the parameters of efficacy required to design a future Phase 2/3 clinical trial. METHODS AND DESIGN:TOOTH is a Phase 1, open-label, single-blinded clinical trial with a pragmatic design that comprises three stages: Stage 1 will involve the selection of 27 participants with middle cerebral artery ischemic stroke and the commencement of autologous dental pulp stem cell isolation, growth, and testing in sequential cohorts (n = 3). Stage 2 will involve the transplantation of dental pulp stem cell in each cohort of participants with an ascending dose and subsequent observation for a 6-month period for any dental pulp stem cell-related adverse events. Stage 3 will investigate the neurosurgical intervention of the maximum tolerable dose of autologous dental pulp stem cell followed by 9 weeks of intensive task-specific rehabilitation. Advanced magnetic resonance and positron emission tomography neuro-imaging, and clinical assessment will be employed to probe any change afforded by stem cell therapy in combination with rehabilitation. SAMPLE SIZE ESTIMATES:Nine participants will step-wise progress in Stage 2 to a dose of up to 10 million dental pulp stem cell, employing a cumulative 3 + 3 statistical design with low starting stem cell dose and subsequent dose escalation, assuming that an acceptable probability of dose-limiting complications is between 1 in 6 (17%) and 1 in 3 (33%) of patients. In Stage 3, another 18 participants will receive an intracranial injection with the maximum tolerable dose of dental pulp stem cell. OUTCOMES:The primary outcomes to be measured are safety and feasibility of intracranial administration of autologous human adult DPSC in patients with chronic stroke and determination of the maximum tolerable dose in human subjects. Secondary outcomes include estimation of the measures of effectiveness required to design a future Phase 2/3 clinical trial.
Transplantation of human dental pulp stem cells ameliorates brain damage following acute cerebral ischemia.
Nito Chikako,Sowa Kota,Nakajima Masataka,Sakamoto Yuki,Suda Satoshi,Nishiyama Yasuhiro,Nakamura-Takahashi Aki,Nitahara-Kasahara Yuko,Ueda Masayuki,Okada Takashi,Kimura Kazumi
Biomedicine & pharmacotherapy = Biomedecine & pharmacotherapie
AIMS:Numerous experimental studies have shown that cellular therapy, including human dental pulp stem cells (DPSCs), is an attractive strategy for ischemic brain injury. Herein, we examined the effects of intravenous DPSC administration after transient middle cerebral artery occlusion in rats. METHODS:Male Sprague-Dawley rats received a transient 90 min middle cerebral artery occlusion. DPSCs (1 × 10 cells) or vehicle were administered via the femoral vein at 0 h or 3 h after ischemia-reperfusion. PKH26, a red fluorescent cell linker, was used to track the transplanted cells in the brain. Infarct volume, neurological deficits, and immunological analyses were performed at 24 h and 72 h after reperfusion. RESULTS:PKH26-positive cells were observed more frequently in the ipsilateral than the contralateral hemisphere. DPSCs transplanted at 0 h after reperfusion significantly reduced infarct volume and reversed motor deficits at 24 h and 72 h recovery. DPSCs transplanted at 3 h after reperfusion also significantly reduced infarct volume and improved motor function compared with vehicle groups at 24 h and 72 h recovery. Further, DPSC transplantation significantly inhibited microglial activation and pro-inflammatory cytokine expression compared with controls at 72 h after reperfusion. Moreover, DPSCs attenuated neuronal degeneration in the cortical ischemic boundary area. CONCLUSIONS:Systemic delivery of human DPSCs after reperfusion reduced ischemic damage and improved functional recovery in a rodent ischemia model, with a clinically relevant therapeutic window. The neuroprotective action of DPSCs may relate to the modulation of neuroinflammation during the acute phase of stroke.
Postnatal human dental pulp stem cells (DPSCs) in vitro and in vivo.
Gronthos S,Mankani M,Brahim J,Robey P G,Shi S
Proceedings of the National Academy of Sciences of the United States of America
Dentinal repair in the postnatal organism occurs through the activity of specialized cells, odontoblasts, that are thought to be maintained by an as yet undefined precursor population associated with pulp tissue. In this study, we isolated a clonogenic, rapidly proliferative population of cells from adult human dental pulp. These DPSCs were then compared with human bone marrow stromal cells (BMSCs), known precursors of osteoblasts. Although they share a similar immunophenotype in vitro, functional studies showed that DPSCs produced only sporadic, but densely calcified nodules, and did not form adipocytes, whereas BMSCs routinely calcified throughout the adherent cell layer with clusters of lipid-laden adipocytes. When DPSCs were transplanted into immunocompromised mice, they generated a dentin-like structure lined with human odontoblast-like cells that surrounded a pulp-like interstitial tissue. In contrast, BMSCs formed lamellar bone containing osteocytes and surface-lining osteoblasts, surrounding a fibrous vascular tissue with active hematopoiesis and adipocytes. This study isolates postnatal human DPSCs that have the ability to form a dentin/pulp-like complex.
Multipotent mesenchymal stem cells with immunosuppressive activity can be easily isolated from dental pulp.
Pierdomenico Laura,Bonsi Laura,Calvitti Mario,Rondelli Damiano,Arpinati Mario,Chirumbolo Gabriella,Becchetti Ennio,Marchionni Cosetta,Alviano Francesco,Fossati Valentina,Staffolani Nicola,Franchina Michele,Grossi Alberto,Bagnara Gian Paolo
BACKGROUND:Bone marrow mesenchymal stem cells (MSCs) are currently being investigated in preclinical and clinical settings because of their multipotent differentiative capacity or, alternatively, their immunosuppressive function. The aim of this study was to evaluate dental pulp (DP) as a potential source of MSCs instead of bone marrow (BM). METHODS:Flow cytometric analysis showed that DP-MSCs and BM-MSCs were equally SH2, SH3, SH4, CD29 and CD 166 positive. The in vitro proliferative kinetics of MSCs were measured by 3H-thymidine incorporation uptake. The immunosuppressive function of MSCs was then tested by coculturing PHA-stimulated allogeneic T cells with or without MSCs for 3 days. RESULTS:BM-MSCs could be differentiated in vitro into osteogenic, chondrogenic and adipogenic lineages. DP-MSCs showed osteogenic and adipocytic differentiation, but did not differentiate into chondrocytes. Although DP-MSCs grow rapidly in vitro between day 3 and day 8 of culture and then decrease their proliferation by day 15, BM-MSCs have a stable and continuous proliferation over the same period of time. The addition of DP-MSCs or BM-MSCs resulted in 91 +/- 4% and 75 +/- 3% inhibition of T cell response, respectively, assessed by a 3H-thymidine assay. CONCLUSIONS:Dental pulp is an easily accessible and efficient source of MSCs, with different kinetics and differentiation potentialities from MSCs as isolated from the bone marrow. The rapid proliferative capacity together with the immunoregulatory characteristics of DP-MSCs may prompt future studies aimed at using these cells in the treatment or prevention of T-cell alloreactivity in hematopoietic or solid organ allogeneic transplantation.
Impact of Dental Pulp Stem Cells Overexpressing Hepatocyte Growth Factor after Cerebral Ischemia/Reperfusion in Rats.
Sowa Kota,Nito Chikako,Nakajima Masataka,Suda Satoshi,Nishiyama Yasuhiro,Sakamoto Yuki,Nitahara-Kasahara Yuko,Nakamura-Takahashi Aki,Ueda Masayuki,Kimura Kazumi,Okada Takashi
Molecular therapy. Methods & clinical development
Hepatocyte growth factor (HGF) has neuroprotective effects against ischemia-induced injuries. Dental pulp stem cell (DPSC) transplantation attenuates tissue injury in the brain of rats with post-transient middle cerebral artery occlusion. We sought to determine whether DPSCs that overexpress HGF can enhance their therapeutic effects on brain damage post-ischemia/reperfusion injury. Treatment with DPSCs overexpressing HGF reduced infarct volumes compared to unmodified DPSC treatment at 3 and 7 days post-transient middle cerebral artery occlusion. The use of unmodified DPSCs and DPSCs overexpressing HGF was associated with improved motor function compared to that with administration of vehicle at 7 days post-transient middle cerebral artery occlusion. DPSCs overexpressing HGF significantly inhibited microglial activation and pro-inflammatory cytokine production along with suppression of neuronal degeneration. Post-reperfusion, DPSCs overexpressing HGF attenuated the decreases in tight junction proteins, maintained blood-brain barrier integrity, and increased microvessel density in peri-infarct areas. The administration of DPSCs overexpressing HGF during the acute phase of stroke increased their neuroprotective effects by modulating inflammation and blood-brain barrier permeability, thereby promoting improvements in post-ischemia/reperfusion brain injury.
Human Dental Pulp Stem Cells Are More Effective Than Human Bone Marrow-Derived Mesenchymal Stem Cells in Cerebral Ischemic Injury.
Song Miyeoun,Lee Jae-Hyung,Bae Jinhyun,Bu Youngmin,Kim Eun-Cheol
We compared the therapeutic effects and mechanism of transplanted human dental pulp stem cells (hDPSCs) and human bone marrow-derived mesenchymal stem cells (hBM-MSCs) in a rat stroke model and an in vitro model of ischemia. Rats were intravenously injected with hDPSCs or hBM-MSCs 24 h after middle cerebral artery occlusion (MCAo), and both groups showed improved functional recovery and reduced infarct volume versus control rats, but the hDPSC group showed greater reduction in infarct volume than the hBM-MSC group. The positive area for the endothelial cell marker was greater in the lesion boundary areas in the hDPSC group than in the hBM-MSC group. Administration of hDPSCs to rats with stroke significantly decreased reactive gliosis, as evidenced by the attenuation of MCAo-induced GFAP+/nestin+ and GFAP+/Musashi-1+ cells, compared with hBM-MSCs. In vivo findings were confirmed by in vitro data illustrating that hDPSCs showed superior neuroprotective, migratory, and in vitro angiogenic effects in oxygen-glucose deprivation (OGD)-injured human astrocytes (hAs) versus hBM-MSCs. Comprehensive comparative bioinformatics analyses from hDPSC- and hBM-MSC-treated in vitro OGD-injured hAs were examined by RNA sequencing technology. In gene ontology and KEGG pathway analyses, significant pathways in the hDPSC-treated group were the MAPK and TGF-β signaling pathways. Thus, hDPSCs may be a better cell therapy source for ischemic stroke than hBM-MSCs.
Dental-Pulp Stem Cells as a Therapeutic Strategy for Ischemic Stroke.
Regenerative medicine aims to restore human functions by regenerating organs and tissues using stem cells or living tissues for the treatment of organ and tissue defects or dysfunction. Clinical trials investigating the treatment of cerebral infarction using mesenchymal stem cells, a type of somatic stem cell therapy, are underway. The development and production of regenerative medicines using somatic stem cells is expected to contribute to the treatment of cerebral infarction, a central nervous system disease for which there is no effective treatment. Numerous experimental studies have shown that cellular therapy, including the use of human dental pulp stem cells, is an attractive strategy for patients with ischemic brain injury. This review describes the basic research, therapeutic mechanism, clinical trials, and future prospects for dental pulp stem cell therapy, which is being investigated in Japan in first-in-human clinical trials for the treatment of patients with acute cerebral ischemia.
Dental Pulp Stem Cells: An Attractive Alternative for Cell Therapy in Ischemic Stroke.
Lan Xiaoyan,Sun Zhengwu,Chu Chengyan,Boltze Johannes,Li Shen
Frontiers in neurology
Ischemic stroke is a major cause of disability and mortality worldwide, but effective restorative treatments are very limited at present. Regenerative medicine research revealed that stem cells are promising therapeutic options. Dental pulp stem cells (DPSCs) are autologously applicable cells that origin from the neural crest and exhibit neuro-ectodermal features next to multilineage differentiation potentials. DPSCs are of increasing interest since they are relatively easy to obtain, exhibit a strong proliferation ability, and can be cryopreserved for a long time without losing their multi-directional differentiation capacity. Besides, use of DPSCs can avoid fundamental problems such as immune rejection, ethical controversy, and teratogenicity. Therefore, DPSCs provide a tempting prospect for stroke treatment.
Mesenchymal stem cells inhibit and stimulate mixed lymphocyte cultures and mitogenic responses independently of the major histocompatibility complex.
Le Blanc K,Tammik L,Sundberg B,Haynesworth S E,Ringdén O
Scandinavian journal of immunology
We aimed to study the effects of mesenchymal stem cells (MSCs) on alloreactivity and effects of T-cell activation on human peripheral blood lymphocytes (PBLs) in vitro. MSCs were expanded from the bone marrow of healthy subjects. MSCs isolated from second to third passage were positive for CD166, CD105, CD44, CD29, SH-3 and SH-4, but negative for CD34 and CD45. MSCs cultured in osteogenic, adipogenic or chondrogenic media differentiated, respectively, into osteocytes, adipocytes or chondrocytes. MSC added to PBL cultures had various effects, ranging from slight inhibition to stimulation of DNA synthesis. The stimulation index (SI = (PBL + MSC)/PBL) varied between 0.2 and 7.3. The SI was not affected by the MSC dose or by the addition of allogeneic or autologous MSCs to the lymphocytes. Suppression of proliferative activity was observed in all experiments after the addition of 10,000-40,000 MSCs to mixed lymphocyte cultures (MLCs). Lymphocyte proliferation was 10-90%, compared with a control MLC run in parallel without MSCs. In contrast, the addition of fewer MSCs (10-1000 cells) led to a less consistent suppression or a marked lymphocyte proliferation in several experiments, ranging from 40 to 190% of the maximal lymphocyte proliferation in control MLCs. The ability to inhibit or stimulate T-cell alloresponses appeared to be independent of the major histocompatibility complex, as results were similar using 'third party' MSCs or MSCs that were autologous to the responder or stimulating PBLs. The strongest inhibitory effect was seen if MSCs were added at the beginning of the 6 day culture, and the effect declined if MSCs were added on day 3 or 5. Marked inhibitory effects of allogeneic and autologous MSCs (15,000) were also noted after mitogenic lymphocyte stimulation by phytohaemagglutinin (median lymphocyte proliferation of 30% of controls), Concanavalin A (56%) and protein A (65%). Little, if any, inhibition occurred after stimulation with pokeweed mitogen. Low numbers of MSCs (150 cells) were unable to inhibit mitogen-induced T-cell responses. MSCs have significant immune modulatory effects on MLCs and after mitogenic stimulation of PBL. High numbers of MSCs suppress alloreactive T cells, whereas very low numbers clearly stimulated lymphocyte proliferation in some experiments. The effect of a larger number of MSCs on MLCs seems more dependent on cell dose than histocompatibility and could result from an 'overload' of a stimulatory mechanism.
Bone marrow mesenchymal stem cells inhibit the response of naive and memory antigen-specific T cells to their cognate peptide.
Krampera Mauro,Glennie Sarah,Dyson Julian,Scott Diane,Laylor Ruthline,Simpson Elizabeth,Dazzi Francesco
Mesenchymal stem cells (MSCs) have been recently shown to inhibit T-cell proliferation to polyclonal stimuli. We characterized the effect of MSCs of bone marrow origin on the T-cell response of naive and memory T cells to their cognate antigenic epitopes. The immune response to murine male transplantation antigens, HY, was selected because the peptide identity and major histocompatibility complex (MHC) restriction of the immunodominant epitopes are known. C57BL/6 female mice immunized with male cells were the source of memory T cells, whereas C6 mice transgenic for HY-specific T-cell receptor provided naive T cells. Responder cells were stimulated in vitro with male spleen cells or HY peptides in the presence or absence of MSCs. MSCs inhibited HY-specific naive and memory T cells in a dose-dependent fashion and affected cell proliferation, cytotoxicity, and the number of interferon gamma (IFN-gamma)-producing HY peptide-specific T cells. However, the MSC inhibitory effect did not selectively target antigen-reactive T cells. When MSCs were added to the T-cell cultures in a Transwell system or MSCs were replaced by MSC culture supernatant, the inhibitory activity was abrogated. T-cell reactivity was also restored if MSCs were removed from the cultures. The expression of MHC molecules and the presence in culture of antigen-presenting cells (APCs) or of CD4(+)/CD25(+) regulatory T cells were not required for MSCs to inhibit. We conclude that MSCs inhibit naive and memory T-cell responses to their cognate antigens. Overall our data suggest that MSCs physically hinder T cells from the contact with APCs in a noncognate fashion.
Human bone marrow stromal cells suppress T-lymphocyte proliferation induced by cellular or nonspecific mitogenic stimuli.
Di Nicola Massimo,Carlo-Stella Carmelo,Magni Michele,Milanesi Marco,Longoni Paolo D,Matteucci Paola,Grisanti Salvatore,Gianni Alessandro M
CD2(+) T lymphocytes obtained from either the donor of bone marrow stromal cells (BMSCs) or a third party were cultured in mixed lymphocyte reactions (MLRs) with either allogeneic dendritic cells (DCs) or peripheral blood lymphocytes (PBLs). When autologous or allogeneic BMSCs were added back to T cells stimulated by DCs or PBLs, a significant and dose-dependent reduction of T-cell proliferation, ranging from 60% +/- 5% to 98% +/- 1%, was evident. Similarly, addition of BMSCs to T cells stimulated by polyclonal activators resulted in a 65% +/- 5% (P =.0001) suppression of proliferation. BMSC- induced T-cell suppression was still evident when BMSCs were added in culture as late as 5 days after starting of MLRs. BMSC-inhibited T lymphocytes were not apoptotic and efficiently proliferated on restimulation. BMSCs significantly suppressed both CD4(+) and CD8(+) T cells (65% +/- 5%, [P =.0005] and 75% +/- 15% [P =.0005], respectively). Transwell experiments, in which cell-cell contact between BMSCs and effector cells was prevented, resulted in a significant inhibition of T-lymphocyte proliferation, suggesting that soluble factors were involved in this phenomenon. By using neutralizing monoclonal antibodies, transforming growth factor beta1 and hepatocyte growth factor were identified as the mediators of BMSC effects. In conclusion, our data demonstrate that (1) autologous or allogeneic BMSCs strongly suppress T-lymphocyte proliferation, (2) this phenomenon that is triggered by both cellular as well as nonspecific mitogenic stimuli has no immunologic restriction, and (3) T-cell inhibition is not due to induction of apoptosis and is likely due to the production of soluble factors.
Concise review: mesenchymal stem cells: their phenotype, differentiation capacity, immunological features, and potential for homing.
Chamberlain Giselle,Fox James,Ashton Brian,Middleton Jim
Stem cells (Dayton, Ohio)
MSCs are nonhematopoietic stromal cells that are capable of differentiating into, and contribute to the regeneration of, mesenchymal tissues such as bone, cartilage, muscle, ligament, tendon, and adipose. MSCs are rare in bone marrow, representing approximately 1 in 10,000 nucleated cells. Although not immortal, they have the ability to expand manyfold in culture while retaining their growth and multilineage potential. MSCs are identified by the expression of many molecules including CD105 (SH2) and CD73 (SH3/4) and are negative for the hematopoietic markers CD34, CD45, and CD14. The properties of MSCs make these cells potentially ideal candidates for tissue engineering. It has been shown that MSCs, when transplanted systemically, are able to migrate to sites of injury in animals, suggesting that MSCs possess migratory capacity. However, the mechanisms underlying the migration of these cells remain unclear. Chemokine receptors and their ligands and adhesion molecules play an important role in tissue-specific homing of leukocytes and have also been implicated in trafficking of hematopoietic precursors into and through tissue. Several studies have reported the functional expression of various chemokine receptors and adhesion molecules on human MSCs. Harnessing the migratory potential of MSCs by modulating their chemokine-chemokine receptor interactions may be a powerful way to increase their ability to correct inherited disorders of mesenchymal tissues or facilitate tissue repair in vivo. The current review describes what is known about MSCs and their capacity to home to tissues together with the associated molecular mechanisms involving chemokine receptors and adhesion molecules.
Mesenchymal stem cells: biology and potential clinical uses.
Deans R J,Moseley A B
There has been an increasing interest in recent years in the stromal cell system functioning in the support of hematopoiesis. The stromal cell system has been proposed to consist of marrow mesenchymal stem cells that are capable of self-renewal and differentiation into various connective tissue lineages. Recent efforts demonstrated that the multiple mesenchymal lineages can be clonally derived from a single mesenchymal stem cell, supporting the proposed paradigm. Dexter demonstrated in 1982 that an adherent stromal-like culture was able to support maintenance of hematopoietic stem as well as early B lymphopoeisis. Recent data from in vitro models demonstrating the essential role of stromal support in hematopoiesis shaped the view that cell-cell interactions in the marrow microenvironment are critical for normal hematopoietic function and differentiation. Maintenance of the hematopoietic stem cell population has been used to increase the efficiency of hematopoietic stem cell gene transfer. High-dose chemotherapy and frequently cause stromal damage with resulting hematopoietic defects. Data from preclinical transplantation studies suggested that stromal cell infusions not only prevent the occurrence of graft failure, but they have an immunomodulatory effect. Preclinical and early clinical safety studies are paving the way for further applications of mesenchymal stem cells in the field of transplantation with respect to hematopoietic support, immunoregulation, and graft facilitation.
Immunosuppression for in vivo research: state-of-the-art protocols and experimental approaches.
Diehl Rita,Ferrara Fabienne,Müller Claudia,Dreyer Antje Y,McLeod Damian D,Fricke Stephan,Boltze Johannes
Cellular & molecular immunology
Almost every experimental treatment strategy using non-autologous cell, tissue or organ transplantation is tested in small and large animal models before clinical translation. Because these strategies require immunosuppression in most cases, immunosuppressive protocols are a key element in transplantation experiments. However, standard immunosuppressive protocols are often applied without detailed knowledge regarding their efficacy within the particular experimental setting and in the chosen model species. Optimization of such protocols is pertinent to the translation of experimental results to human patients and thus warrants further investigation. This review summarizes current knowledge regarding immunosuppressive drug classes as well as their dosages and application regimens with consideration of species-specific drug metabolization and side effects. It also summarizes contemporary knowledge of novel immunomodulatory strategies, such as the use of mesenchymal stem cells or antibodies. Thus, this review is intended to serve as a state-of-the-art compendium for researchers to refine applied experimental immunosuppression and immunomodulation strategies to enhance the predictive value of preclinical transplantation studies.
Transplantation of cryopreserved human umbilical cord blood mononuclear cells does not induce sustained recovery after experimental stroke in spontaneously hypertensive rats.
Weise Gesa,Lorenz Marlene,Pösel Claudia,Maria Riegelsberger Ute,Störbeck Veronika,Kamprad Manja,Kranz Alexander,Wagner Daniel-Christoph,Boltze Johannes
Journal of cerebral blood flow and metabolism : official journal of the International Society of Cerebral Blood Flow and Metabolism
Previous studies have highlighted the enormous potential of cell-based therapies for stroke not only to prevent ischemic brain damage, but also to amplify endogenous repair processes. Considering its widespread availability and low immunogenicity human umbilical cord blood (HUCB) is a particularly attractive stem cell source. Our goal was to investigate the neurorestorative potential of cryopreserved HUCB mononuclear cells (MNC) after permanent middle cerebral artery occlusion (MCAO) in spontaneously hypertensive rats (SHR). Human umbilical cord blood MNC or vehicle solution was administered intravenously 24 hours after MCAO. Experimental groups were as follows: (1) quantitative polymerase chain reaction (PCR) of host-derived growth factors up to 48 hours after stroke; (2) immunohistochemical analysis of astroglial scarring; (3) magnetic resonance imaging (MRI) and weekly behavioral tests for 2 months after stroke. Long-term functional outcome and lesion development on MRI were not beneficially influenced by HUCB MNC therapy. Furthermore, HUCB MNC treatment did not change local growth factor levels and glial scarring extent. In summary, we could not demonstrate neurorestorative properties of HUCB MNC after stroke in SHR. Our results advise caution regarding a prompt translation of cord blood therapy into clinical stroke trials as long as deepened knowledge about its precise modes of action is missing.
Lesional and perilesional tissue characterization by automated image processing in a novel gyrencephalic animal model of peracute intracerebral hemorrhage.
Boltze Johannes,Ferrara Fabienne,Hainsworth Atticus H,Bridges Leslie R,Zille Marietta,Lobsien Donald,Barthel Henryk,McLeod Damian D,Gräßer Felix,Pietsch Sören,Schatzl Ann-Kathrin,Dreyer Antje Y,Nitzsche Björn
Journal of cerebral blood flow and metabolism : official journal of the International Society of Cerebral Blood Flow and Metabolism
Intracerebral hemorrhage (ICH) is an important stroke subtype, but preclinical research is limited by a lack of translational animal models. Large animal models are useful to comparatively investigate key pathophysiological parameters in human ICH. To (i) establish an acute model of moderate ICH in adult sheep and (ii) an advanced neuroimage processing pipeline for automatic brain tissue and hemorrhagic lesion determination; 14 adult sheep were assigned for stereotactically induced ICH into cerebral white matter under physiological monitoring. Six hours after ICH neuroimaging using 1.5T MRI including structural as well as perfusion and diffusion, weighted imaging was performed before scarification and subsequent neuropathological investigation including immunohistological staining. Controlled, stereotactic application of autologous blood caused a space-occupying intracerebral hematoma of moderate severity, predominantly affecting white matter at 5 h post-injection. Neuroimage post-processing including lesion probability maps enabled automatic quantification of structural alterations including perilesional diffusion and perfusion restrictions. Neuropathological and immunohistological investigation confirmed perilesional vacuolation, axonal damage, and perivascular blood as seen after human ICH. The model and imaging platform reflects key aspects of human ICH and enables future translational research on hematoma expansion/evacuation, white matter changes, hematoma evacuation, and other aspects.
Deep Learning to Decipher the Progression and Morphology of Axonal Degeneration.
Palumbo Alex,Grüning Philipp,Landt Svenja Kim,Heckmann Lara Eleen,Bartram Luisa,Pabst Alessa,Flory Charlotte,Ikhsan Maulana,Pietsch Sören,Schulz Reinhard,Kren Christopher,Koop Norbert,Boltze Johannes,Madany Mamlouk Amir,Zille Marietta
Axonal degeneration (AxD) is a pathological hallmark of many neurodegenerative diseases. Deciphering the morphological patterns of AxD will help to understand the underlying mechanisms and develop effective therapies. Here, we evaluated the progression of AxD in cortical neurons using a novel microfluidic device together with a deep learning tool that we developed for the enhanced-throughput analysis of AxD on microscopic images. The trained convolutional neural network (CNN) sensitively and specifically segmented the features of AxD including axons, axonal swellings, and axonal fragments. Its performance exceeded that of the human evaluators. In an in vitro model of AxD in hemorrhagic stroke induced by the hemolysis product hemin, we detected a time-dependent degeneration of axons leading to a decrease in axon area, while axonal swelling and fragment areas increased. Axonal swellings preceded axon fragmentation, suggesting that swellings may be reliable predictors of AxD. Using a recurrent neural network (RNN), we identified four morphological patterns of AxD (granular, retraction, swelling, and transport degeneration). These findings indicate a morphological heterogeneity of AxD in hemorrhagic stroke. Our EntireAxon platform enables the systematic analysis of axons and AxD in time-lapse microscopy and unravels a so-far unknown intricacy in which AxD can occur in a disease context.
Neuronal Death After Hemorrhagic Stroke In Vitro and In Vivo Shares Features of Ferroptosis and Necroptosis.
Zille Marietta,Karuppagounder Saravanan S,Chen Yingxin,Gough Peter J,Bertin John,Finger Joshua,Milner Teresa A,Jonas Elizabeth A,Ratan Rajiv R
BACKGROUND AND PURPOSE:Intracerebral hemorrhage leads to disability or death with few established treatments. Adverse outcomes after intracerebral hemorrhage result from irreversible damage to neurons resulting from primary and secondary injury. Secondary injury has been attributed to hemoglobin and its oxidized product hemin from lysed red blood cells. The aim of this study was to identify the underlying cell death mechanisms attributable to secondary injury by hemoglobin and hemin to broaden treatment options. METHODS:We investigated cell death mechanisms in cultured neurons exposed to hemoglobin or hemin. Chemical inhibitors implicated in all known cell death pathways were used. Identified cell death mechanisms were confirmed using molecular markers and electron microscopy. RESULTS:Chemical inhibitors of ferroptosis and necroptosis protected against hemoglobin- and hemin-induced toxicity. By contrast, inhibitors of caspase-dependent apoptosis, protein or mRNA synthesis, autophagy, mitophagy, or parthanatos had no effect. Accordingly, molecular markers of ferroptosis and necroptosis were increased after intracerebral hemorrhage in vitro and in vivo. Electron microscopy showed that hemin induced a necrotic phenotype. Necroptosis and ferroptosis inhibitors each abrogated death by >80% and had similar therapeutic windows in vitro. CONCLUSIONS:Experimental intracerebral hemorrhage shares features of ferroptotic and necroptotic cell death, but not caspase-dependent apoptosis or autophagy. We propose that ferroptosis or necroptotic signaling induced by lysed blood is sufficient to reach a threshold of death that leads to neuronal necrosis and that inhibition of either of these pathways can bring cells below that threshold to survival.
Novel targets, treatments, and advanced models for intracerebral haemorrhage.
Zille Marietta,Farr Tracy D,Keep Richard F,Römer Christine,Xi Guohua,Boltze Johannes
Intracerebral haemorrhage (ICH) is the second most common type of stroke and a major cause of mortality and disability worldwide. Despite advances in surgical interventions and acute ICH management, there is currently no effective therapy to improve functional outcomes in patients. Recently, there has been tremendous progress uncovering new pathophysiological mechanisms underlying ICH that may pave the way for the development of therapeutic interventions. Here, we highlight emerging targets, but also existing gaps in preclinical animal modelling that prevent their exploitation. We particularly focus on (1) ICH aetiology, (2) the haematoma, (3) inflammation, and (4) post-ICH pathology. It is important to recognize that beyond neurons and the brain, other cell types and organs are crucially involved in ICH pathophysiology and successful interventions likely will need to address the entire organism. This review will spur the development of successful therapeutic interventions for ICH and advanced animal models that better reflect its aetiology and pathophysiology.
Integrin α4 Overexpression on Rat Mesenchymal Stem Cells Enhances Transmigration and Reduces Cerebral Embolism After Intracarotid Injection.
Cui Li-Li,Nitzsche Franziska,Pryazhnikov Evgeny,Tibeykina Marina,Tolppanen Laura,Rytkönen Jussi,Huhtala Tuulia,Mu Jing-Wei,Khiroug Leonard,Boltze Johannes,Jolkkonen Jukka
BACKGROUND AND PURPOSE:Very late antigen-4 (integrin α4β1)/vascular cell adhesion molecule-1 mediates leukocyte trafficking and transendothelial migration after stroke. Mesenchymal stem cells (MSCs) typically express integrin β1 but insufficient ITGA4 (integrin α4), which limits their homing after intravascular transplantation. We tested whether ITGA4 overexpression on MSCs increases cerebral homing after intracarotid transplantation and reduces MSC-borne cerebral embolism. METHODS:Rat MSCs were lentivirally transduced to overexpress ITGA4. In vitro transendothelial migration was assessed using a Boyden chamber assay. Male Wistar rats intracarotidly received 0.5×10 control or modified MSCs 24 hours after sham or stroke surgery. In vivo behavior of MSCs in the cerebral vasculature was observed by intravital microscopy and single-photon emission computed tomography for up to 72 hours. RESULTS:Transendothelial migration of ITGA4-overexpressing MSCs was increased in vitro. MSCs were passively entrapped in microvessels in vivo and occasionally formed large cell aggregates causing local blood flow interruptions. MSCs were rarely found in perivascular niches or parenchyma at 72 hours post-transplantation, but ITGA4 overexpression significantly decreased cell aggregation and ameliorated the evoked cerebral embolism in stroke rats. CONCLUSIONS:ITGA4 overexpression on MSCs enhances transendothelial migration in vitro, but not in vivo, although it improves safety after intracarotid transplantation into stroke rats.
Pulmonary passage is a major obstacle for intravenous stem cell delivery: the pulmonary first-pass effect.
Fischer Uwe M,Harting Matthew T,Jimenez Fernando,Monzon-Posadas Werner O,Xue Hasen,Savitz Sean I,Laine Glen A,Cox Charles S
Stem cells and development
Intravenous (IV) stem cell delivery for regenerative tissue therapy has been increasingly used in both experimental and clinical trials. However, recent data suggest that the majority of administered stem cells are initially trapped in the lungs. We sought to investigate variables that may affect this pulmonary first-pass effect. In anesthetized Sprague-Dawley rats, silicone tubing catheters were placed in the left internal jugular vein and common carotid artery. We investigated four different cell types: mesenchymal stromal cells (MSC), multipotent adult progenitor cells (MAPCs), bone marrow-derived mononuclear cells (BMMC), and neural stem cells (NSC). Cells were co-labeled with Qtracker 655 (for flow cytometry) and Qtracker 800 (for infrared imaging) and infused intravenously with continual arterial sample collection. Samples were analyzed via flow cytometry to detect labeled cells reaching the arterial circulation. Following sampling and exsanguination, heart, lungs, spleen, kidney, and liver were harvested and placed on an infrared imaging system to identify the presence of labeled cells. The majority of MSCs were trapped inside the lungs following intravenous infusion. NSC and MAPC pulmonary passage was 2-fold and BMMC passage was 30-fold increased as compared to MSCs. Inhibition of MSC CD49d significantly increased MSC pulmonary passage. Infusion via two boluses increased pulmonary MSC passage as compared to single bolus administration. Infrared imaging revealed stem cells evenly distributed over all lung fields. Larger stem and progenitor cells are initially trapped inside the lungs following intravenous administration with a therapeutically questionable number of cells reaching the arterial system acutely.
Intravenous Human Umbilical Cord-Derived Mesenchymal Stromal Cell Administration in Models of Moderate and Severe Intracerebral Hemorrhage.
Mello Tanira Giara,Rosado-de-Castro Paulo Henrique,Campos Raquel Maria Pereira,Vasques Juliana Ferreira,Rangel-Junior William Simões,Mattos Raphael Santos de Almeida Rezende de,Puig-Pijuan Teresa,Foerster Bernd Uwe,Gutfilen Bianca,Souza Sergio Augusto Lopes,Boltze Johannes,Paiva Fernando Fernandes,Mendez-Otero Rosalia,Pimentel-Coelho Pedro Moreno
Stem cells and development
Intracerebral hemorrhage (ICH) is as a life-threatening condition that can occur in young adults, often causing long-term disability. Recent preclinical data suggest mesenchymal stromal cell (MSC)-based therapies as promising options to minimize brain damage after ICH. However, therapeutic evidence and mechanistic insights are still limited, particularly when compared with other disorders such as ischemic stroke. Herein, we employed a model of collagenase-induced ICH in young adult rats to investigate the potential therapeutic effects of an intravenous injection of human umbilical cord Wharton's jelly-derived MSCs (hUC-MSCs). Two doses of collagenase were used to cause moderate or severe hemorrhages. Magnetic resonance imaging showed that animals treated with hUC-MSCs after moderate ICH had smaller residual hematoma volumes than vehicle-treated rats, whereas the cell therapy failed to decrease the hematoma volume in animals with a severe ICH. Functional assessments (rotarod and elevated body swing tests) were performed for up to 21 days after ICH. Enduring neurological impairments were seen only in animals subjected to severe ICH, but the cell therapy did not induce statistically significant improvements in the functional recovery. The biodistribution of Technetium-99m-labeled hUC-MSCs was also evaluated, showing that most cells were found in organs such as the spleen and lungs 24 h after transplantation. Nevertheless, it was possible to detect a weak signal in the brain, which was higher in the ipsilateral hemisphere of rats subjected to a severe ICH. These data indicate that hUC-MSCs have moderately beneficial effects in cases of less severe brain hemorrhages in rats by decreasing the residual hematoma volume, and that optimization of the therapy is still necessary.
Mesenchymal Stem Cell-Derived Neuron-Like Cell Transplantation Combined with Electroacupuncture Improves Synaptic Plasticity in Rats with Intracerebral Hemorrhage via mTOR/p70S6K Signaling.
Yang Guoqiang,Zhu Jiayi,Zhan Guwen,Fan Guangbi,Deng Li,Tang Huajun,Jiang Xiaoqian,Chen Bo,Yang Chaoxian
Stem cells international
Previous studies have shown that the combination of mesenchymal stem cell (MSC) transplantation and electroacupuncture (EA) stimulation is a neuroprotective strategy for treating intracerebral hemorrhage (ICH). However, the underlying mechanisms by which the combined treatment promotes neuroprotection remain unclear. This study was designed to investigate the effects of the combined treatment on synaptic plasticity and elucidate their underlying mechanisms. Therefore, rat ICH models were established by injecting collagenase and heparin, and the animals were randomly divided into model control (MC), EA stimulation (EA), MSC-derived neuron-like cell transplantation (MSC-dNLCs), and MSC-dNLC transplantation combined with EA stimulation (MSC-dNLCs+EA) groups. We observed the ultrastructure of the brain and measured the brain water content (BWC) and the levels of the microtubule-associated protein 2 (MAP2), galactocerebrosidase (GALC), and glial fibrillary acidic protein (GFAP) proteins. We also measured the levels of the phosphorylated mammalian target of rapamycin (mTOR) and 70 kDa ribosomal protein S6 kinase (p70S6K) proteins, as well as the expression of synapse-related proteins. The BWC increased in rats after ICH and decreased significantly in ICH rats treated with MSC-dNLC transplantation, EA stimulation, or combined therapy. Meanwhile, after ICH, the number of blood vessels increased more evidently, but only the combined treatment reduced the number of blood vessels among rats receiving the three treatments. Moreover, the levels of MAP2, GALC, postsynaptic density 95 (PSD95), and synaptophysin (SYP) proteins, as well as the levels of the phosphorylated mTOR and p70S6k proteins, increased in the MSC-dNLCs+EA group compared with those in the MSC-dNLCs and EA groups. Compared with the MC group, GFAP expression was significantly reduced in the MSC-dNLCs, EA, and MSC-dNLCs+EA groups, but the differences among the three treatment groups were not significant. In addition, the number of synapses increased only in the MSC-dNLCs+EA group compared to the MC group. Based on these data, the combination of MSC-dNLC transplantation and EA stimulation exerts a synergistic effect on improving the consequences of ICH by relieving cerebral edema and glial scarring, promoting the survival of neurons and oligodendrocytes, and activating mTOR/p70S6K signaling to enhance synaptic plasticity.
Neural differentiation of brain-derived neurotrophic factor-expressing human umbilical cord blood-derived mesenchymal stem cells in culture via TrkB-mediated ERK and β-catenin phosphorylation and following transplantation into the developing brain.
Lim Jung Yeon,Park Sang In,Kim Seong Muk,Jun Jin Ae,Oh Ji Hyeon,Ryu Chung Hun,Jeong Chang Hyun,Park Sun Hwa,Park Soon A,Oh Wonil,Chang Jong Wook,Jeun Sin-Soo
The ability of mesenchymal stem cells (MSCs) to differentiate into neural cells makes them potential replacement therapeutic candidates in neurological diseases. Presently, overexpression of brain-derived neurotrophic factor (BDNF), which is crucial in the regulation of neural progenitor cell differentiation and maturation during development, was sufficient to convert the mesodermal cell fate of human umbilical cord blood-derived MSCs (hUCB-MSCs) into a neuronal fate in culture, in the absence of specialized induction chemicals. BDNF overexpressing hUCB-MSCs (MSCs-BDNF) yielded an increased number of neuron-like cells and, surprisingly, increased the expression of neuronal phenotype markers in a time-dependent manner compared with control hUCB-MSCs. In addition, MSCs-BDNF exhibited a decreased labeling for MSCs-related antigens such as CD44, CD73, and CD90, and decreased potential to differentiate into mesodermal lineages. Phosphorylation of the receptor tyrosine kinase B (TrkB), which is a receptor of BDNF, was increased significantly in MSC-BDNF. BDNF overexpression also increased the phosphorylation of β-catenin and extracellular signal-regulated kinases (ERKs). Inhibition of TrkB availability by treatment with the TrkB-specific inhibitor K252a blocked the BDNF-stimulated phosphorylation of β-catenin and ERKs, indicating the involvement of both the β-catenin and ERKs signals in the BDNF-stimulated and TrkB-mediated neural differentiation of hUCB-MSCs. Reduction of β-catenin availability using small interfering RNA-mediated gene silencing inhibited ERKs phosphorylation. However, β-catenin activation was maintained. In addition, inhibition of β-catenin and ERKs expression levels abrogated the BDNF-stimulated upregulation of neuronal phenotype markers. Furthermore, MSC-BDNF survived and migrated more extensively when grafted into the lateral ventricles of neonatal mouse brain, and differentiated significantly into neurons in the olfactory bulb and periventricular astrocytes. These results indicate that BDNF induces the neural differentiation of hUCB-MSCs in culture via the TrkB-mediated phosphorylation of ERKs and β-catenin and following transplantation into the developing brain.
Senescence of mesenchymal stem cells (Review).
Li Yi,Wu Qiong,Wang Yujia,Li Li,Bu Hong,Bao Ji
International journal of molecular medicine
Mesenchymal stem cells (MSCs) have been used in cell-based therapy for various diseases, due to their immunomodulatory and inflammatory effects. However, the function of MSCs is known to decline with age, a process that is called senescence. To date, the process of MSC senescence remains unknown as in-depth understanding of the mechanisms involved in cellular senescence is lacking. First, senescent MSCs are so heterogeneous that not all of them express the same phenotypic markers. In addition, the genes and signaling pathways which regulate this process in MSCs are still unknown. Thus, an understanding of the molecular processes controlling MSC senescence is crucial to determining the drivers and effectors of age-associated MSC dysfunction. Moreover, the proper use of MSCs for clinical application requires a general understanding of the MSC aging process. Furthermore, such knowledge is essential for the development of therapeutic interventions that can slow or reverse age-related degenerative changes to enhance repair processes and maintain healthy function in aging tissues. To further clarify the properties of senescent cells, as well as to present significant findings from studies on the mechanisms of cellular aging, we summarize these biological features in the senescence of MSCs in this scenario. This review summarizes recent advances in our understanding of the markers and differentiation potential indicating MSC senescence, as well as factors affecting MSC senescence with particular emphasis on the roles of oxidative stress, intrinsic changes in telomere shortening, histone deacetylase and DNA methyltransferase, genes and signaling pathways and immunological properties.
The axon-glia unit in white matter stroke: mechanisms of damage and recovery.
Rosenzweig Shira,Carmichael S Thomas
Approximately one quarter of all strokes in humans occur in white matter, and the progressive nature of white matter lesions often results in severe physical and mental disability. Unlike cortical grey matter stroke, the pathology of white matter stroke revolves around disrupted connectivity and injured axons and glial cells, rather than neuronal cell bodies. Consequently, the mechanisms behind ischemic damage to white matter elements, the regenerative responses of glial cells and their signaling pathways, all differ significantly from those in grey matter. Development of effective therapies for white matter stroke would require an enhanced understanding of the complex cellular and molecular interactions within the white matter, leading to the identification of new therapeutic targets. This review will address the unique properties of the axon-glia unit during white matter stroke, describe the challenging process of promoting effective white matter repair, and discuss recently-identified signaling pathways which may hold potential targets for repair in this disease. This article is part of a Special Issue entitled SI: Cell Interactions In Stroke.
Allogeneic bone marrow stromal cells promote glial-axonal remodeling without immunologic sensitization after stroke in rats.
Li Yi,McIntosh Kevin,Chen Jieli,Zhang Chunling,Gao Qi,Borneman Jade,Raginski Kim,Mitchell James,Shen Lihong,Zhang Jing,Lu Dunyue,Chopp Michael
We evaluated the effects of allogeneic bone marrow stromal cell treatment of stroke on functional outcome, glial-axonal architecture, and immune reaction. Female Wistar rats were subjected to 2 h of middle cerebral artery occlusion. Rats were injected intravenously with PBS, male allogeneic ACI--or syngeneic Wistar--bone marrow stromal cells at 24 h after ischemia and sacrificed at 28 days. Significant functional recovery was found in both cell-treated groups compared to stroke rats that did not receive BMSCs, but no difference was detected between allogeneic and syngeneic cell-treated rats. No evidence of T cell priming or humoral antibody production to marrow stromal cells was found in recipient rats after treatment with allogeneic cells. Similar numbers of Y-chromosome+ cells were detected in the female rat brains in both groups. Significantly increased thickness of individual axons and myelin, and areas of the corpus callosum and the numbers of white matter bundles in the striatum were detected in the ischemic boundary zone of cell-treated rats compared to stroked rats. The areas of the contralateral corpus callosum significantly increased after cell treatment compared to normal rats. Processes of astrocytes remodeled from hypertrophic star-like to tadpole-like shape and oriented parallel to the ischemic regions after cell treatment. Axonal projections emanating from individual parenchymal neurons exhibited an overall orientation parallel to elongated radial processes of reactive astrocytes of the cell-treated rats. Allogeneic and syngeneic bone marrow stromal cell treatment after stroke in rats improved neurological recovery and enhanced reactive oligodendrocyte and astrocyte related axonal remodeling with no indication of immunologic sensitization in adult rat brain.
Umbilical cord-derived mesenchymal stem cells with forced expression of hepatocyte growth factor enhance remyelination and functional recovery in a rat intracerebral hemorrhage model.
Liu An Min,Lu Gang,Tsang Kam Sze,Li Guo,Wu Yan,Huang Zheng Song,Ng Ho Keung,Kung Hsiang Fu,Poon Wai Sang
BACKGROUND:Spontaneous intracerebral hemorrhage (ICH) carries a high mortality rate, with survivors commonly left with permanent neurological deficits. Mesenchymal stem cell (MSC) transplantation promotes functional recovery in experimental ICH, and treatment with hepatocyte growth factor (HGF) is beneficial in ischemic stroke. OBJECTIVE:We hypothesize that transplantation of MSCs with previous transduction of HGF has an additive effect in promoting neurological recovery through myelin and axonal regeneration. METHODS:HGF transduction to human umbilical cord-derived MSCs using lentiviral plasmid pWPI-HGF-GFP was prepared. One week after a collagenase-induced ICH, 80 male Sprague-Dawley rats were divided into 3 groups for stereotactic injection of phosphate-buffered saline (group I), MSC transplant (group II), and HGF-transduced MSC transplant (group III), respectively, into the left ventricle. The animals were assessed weekly for 5 weeks using the Rotarod motor function test, at which time they were killed for Luxol fast blue myelin staining and appropriate immunohistochemistry and Western blotting. RESULTS:Animals receiving transplanted HGF-transduced MSCs (group III) exhibited significantly better motor function recovery than animals treated with MSCs alone (group II), which in turn performed better than the phosphate-buffered saline controls at 2 weeks after transplantation. Luxol fast blue staining of myelin displayed significantly less demyelination and significantly higher reactivity in myelin basic protein and growth-associated protein-43 in immunohistochemistry and Western blotting and significantly reduced myelin-associated glycoprotein activity in group III animals. CONCLUSION:Animals transplanted with HGF-transduced MSCs 1 week after experimental ICH were shown to achieve a better neurological recovery. This improved neurological recovery from ICH is attributed to nerve fiber remyelination and axonal regeneration.
Ultrastructural Characteristics of Neuronal Death and White Matter Injury in Mouse Brain Tissues After Intracerebral Hemorrhage: Coexistence of Ferroptosis, Autophagy, and Necrosis.
Li Qian,Weiland Abigail,Chen Xuemei,Lan Xi,Han Xiaoning,Durham Frederick,Liu Xi,Wan Jieru,Ziai Wendy C,Hanley Daniel F,Wang Jian
Frontiers in neurology
Although intracerebral hemorrhage (ICH) is a devastating disease worldwide, the pathologic changes in ultrastructure during the acute and chronic phases of ICH are poorly described. In this study, transmission electron microscopy was used to examine the ultrastructure of ICH-induced pathology. ICH was induced in mice by an intrastriatal injection of collagenase. Pathologic changes were observed in the acute (3 days), subacute (6 days), and chronic (28 days) phases. Compared with sham animals, we observed various types of cell death in the injured striatum during the acute phase of ICH, including necrosis, ferroptosis, and autophagy. Different degrees of axon degeneration in the striatum were seen in the acute phase, and axonal demyelination was observed in the ipsilateral striatum and corpus callosum at late time points. In addition, phagocytes, resident microglia, and infiltrating monocyte-macrophages were present around red blood cells and degenerating neurons and were observed to engulf red blood cells and other debris. Many synapses appeared abnormal or were lost. This systematic analysis of the pathologic changes in ultrastructure after ICH in mice provides information that will be valuable for future ICH pathology studies.
Injury mechanisms in acute intracerebral hemorrhage.
Wilkinson D Andrew,Pandey Aditya S,Thompson B Gregory,Keep Richard F,Hua Ya,Xi Guohua
Intracerebral hemorrhage (ICH) is the most common hemorrhagic stroke subtype, and rates are increasing with an aging population. Despite an increase in research and trials of therapies for ICH, mortality remains high and no interventional therapy has been demonstrated to improve outcomes. We review known mechanisms of injury, recent clinical trial results, and newly discovered signaling pathways involved in hematoma clearance. Enthusiasm remains high for methods of minimally invasive clot removal as well as pharmacologic strategies to improve recovery after ICH, both of which are currently being evaluated in clinical trials. This article is part of the Special Issue entitled 'Cerebral Ischemia'.
Intracerebral hematoma extends via perivascular spaces and perineurium.
Yin Jia,Lü Tian-Ming,Qiu Guang,Huang Rui-Yu,Fang Min,Wang Yuan-Yuan,Xiao Duan,Liu Xiao-Jia
The Tohoku journal of experimental medicine
Intracerebral hemorrhage (ICH) is a devastating disorder associated with high morbidity and mortality. ICH results in the formation of hematoma that affects not only the primary site of injury but also the remote regions. In fact, hematoma can extend via perivascular spaces (also called Virchow-Robin spaces, VRS) and perineurium in an animal model of ICH. In the present study, we used magnetic resonance imaging (MRI) with susceptibility-weighted imaging (SWI) to investigate the characteristics of the perivascular and perineural extensions of hematomas in patients with ICH. A total of 20 ICH patients without secondary subarachnoid and secondary intraventricular hemorrhages were recruited. Brain MRI scans, including SWI, T1, and T2-weighted images, were performed between 17 h to 7 days after the onset of ICH. MRI with SWI revealed that paramagnetic substances spread along the VRS or the perineurium. Such distribution could cause the formation of cerebral microbleeds (CMBs). However, the distribution of remote hemorrhagic lesions varied, depending on the size and location of the original hematoma. The unenhanced CT scans of the 20 patients did not show any hyperdensity around the blood vessels and nerve tracts outside the hematoma. These results indicate the perivascular and perineural extensions of hematomas in patients with ICH, which is formed by the leakage of the original hematoma via the VRS or perineurium. We also provide a new explanation for the series of pathological processes involved in ICH, including the remote effects of hematoma and the formation of CMBs in patients with ICH.
The Role of MSCs and Cell Fusion in Tissue Regeneration.
Dörnen Jessica,Dittmar Thomas
International journal of molecular sciences
Regenerative medicine is concerned with the investigation of therapeutic agents that can be used to promote the process of regeneration after injury or in different diseases. Mesenchymal stem/stromal cells (MSCs) and their secretome-including extracellular vesicles (EVs) are of great interest, due to their role in tissue regeneration, immunomodulatory capacity and low immunogenicity. So far, clinical studies are not very conclusive as they show conflicting efficacies regarding the use of MSCs. An additional process possibly involved in regeneration might be cell fusion. This process occurs in both a physiological and a pathophysiological context and can be affected by immune response due to inflammation. In this review the role of MSCs and cell fusion in tissue regeneration is discussed.
Targeting the plasticity of mesenchymal stromal cells to reroute the course of acute myeloid leukemia.
Borella Giulia,Da Ros Ambra,Borile Giulia,Porcù Elena,Tregnago Claudia,Benetton Maddalena,Marchetti Anna,Bisio Valeria,Montini Barbara,Michielotto Barbara,Cani Alice,Leszl Anna,Campodoni Elisabetta,Sandri Monica,Montesi Monica,Bresolin Silvia,Cairo Stefano,Buldini Barbara,Locatelli Franco,Pigazzi Martina
Bone marrow (BM) microenvironment contributes to the regulation of normal hematopoiesis through a finely tuned balance of self-renewal and differentiation processes, cell-cell interaction, and secretion of cytokines that during leukemogenesis are altered and favor tumor cell growth. In pediatric acute myeloid leukemia (AML), chemotherapy is the standard of care, but >30% of patients still relapse. The need to accelerate the evaluation of innovative medicines prompted us to investigate the role of mesenchymal stromal cells (MSCs) in the leukemic niche to define its contribution to the mechanism of leukemia drug escape. We generated a humanized 3-dimensional (3D) niche with AML cells and MSCs derived from either patients (AML-MSCs) or healthy donors. We observed that AML cells establish physical connections with MSCs, mediating a reprogrammed transcriptome inducing aberrant cell proliferation and differentiation and severely compromising their immunomodulatory capability. We confirmed that AML cells modulate h-MSCs transcriptional profile promoting functions similar to the AML-MSCs when cocultured in vitro, thus facilitating leukemia progression. Conversely, MSCs derived from BM of patients at time of disease remission showed recovered healthy features at transcriptional and functional levels, including the secretome. We proved that AML blasts alter MSCs activities in the BM niche, favoring disease development and progression. We discovered that a novel AML-MSC selective CaV1.2 channel blocker drug, lercanidipine, is able to impair leukemia progression in 3D both in vitro and when implanted in vivo if used in combination with chemotherapy, supporting the hypothesis that synergistic effects can be obtained by dual targeting approaches.
The Fate of Erythrocytes after Cerebral Hemorrhage.
Xia Fan,Keep Richard F,Ye Fenghui,Holste Katherine G,Wan Shu,Xi Guohua,Hua Ya
Translational stroke research
After a cerebral hemorrhage (intracerebral, subarachnoid, and intraventricular), extravasated blood contributes to both initial brain injury, via physical disruption and mass effect, and secondary injury, through the release of potentially neurotoxic and pro-inflammatory factors such as hemoglobin, iron, and peroxiredoxin-2. Erythrocytes are a major blood component and are a source of such damaging factors. Erythrolysis after cerebral hemorrhage releases potential neurotoxins, contributing to brain injury and edema. Alternatively, erythrocyte phagocytosis via microglia or macrophages may limit the spill of neurotoxins therefore limiting subsequent brain injury. The aim of this review is to discuss the process of phagocytosis of erythrocytes by microglia or macrophages after cerebral hemorrhage, the effect of erythrolysis on brain injury, novel mechanisms of erythrocyte and phagocyte egress from the brain, and exciting new targets in this pathway to attenuate brain injury. Understanding the fate of erythrocytes after cerebral hemorrhage may uncover additional potential interventions for clinical translational research.
Imaging of Spontaneous Intracerebral Hemorrhage.
Jain Abhi,Malhotra Ajay,Payabvash Seyedmehdi
Neuroimaging clinics of North America
Primary or nontraumatic spontaneous intracerebral hemorrhage (ICH) comprises approximately 15% to 20% of all stroke. ICH has a mortality of approximately 40% within the first month, and 75% mortality and morbidity rate within the first year. Despite reduction in overall stroke incidence, hemorrhagic stroke incidence has remained steady since 1980. Neuroimaging is critical in detection of ICH, determining the underlying cause, identification of patients at risk of hematoma expansion, and directing the treatment strategy. This article discusses the neuroimaging methods of ICH, imaging markers for clinical outcome prediction, and future research directions with attention to the latest evidence-based guidelines.
MSC Secretome as a Promising Tool for Neuroprotection and Neuroregeneration in a Model of Intracerebral Hemorrhage.
Karagyaur Maxim,Dzhauari Stalik,Basalova Nataliya,Aleksandrushkina Natalia,Sagaradze Georgy,Danilova Natalia,Malkov Pavel,Popov Vladimir,Skryabina Mariya,Efimenko Anastasia,Tkachuk Vsevolod
Multipotent mesenchymal stromal cells (MSCs) are considered to be critical contributors to injured tissue repair and regeneration, and MSC-based therapeutic approaches have been applied to many peripheral and central neurologic disorders. It has been demonstrated that the beneficial effects of MSC are mainly mediated by the components of their secretome. In the current study, we have explored the neuroprotective potential of the MSC secretome in a rat model of intracerebral hemorrhage and shown that a 10-fold concentrated secretome of human MSC and its combination with the brain-derived neurotrophic factor (BDNF) provided a better survival and neurological outcome of rats within 14 days of intracerebral hemorrhage compared to the negative (non-treated) and positive (BDNF) control groups. We found that it was due to the ability of MSC secretome to stimulate neuron survival under conditions of glutamate-induced neurotoxicity. However, the lesion volume did not shrink in these rats, and this also correlated with prominent microglia activation. We hypothesize that this could be caused by the species-specificity of the used MSC secretome and provide evidence to confirm this. Thus, we have found that allogenic rat MSC secretome was more effective than xenogenic human MSC secretome in the rat intracerebral hemorrhage model: it reduced the volume of the lesion and promoted excellent survival and neurological outcome of the treated rats.
Ghrelin attenuates secondary brain injury following intracerebral hemorrhage by inhibiting NLRP3 inflammasome activation and promoting Nrf2/ARE signaling pathway in mice.
Cheng Yijun,Chen Bin,Xie Wanqun,Chen Zhenghong,Yang Guoyuan,Cai Yu,Shang Hanbing,Zhao Weiguo
Ghrelin, a brain-gut peptide, has been proven to exert neuroprotection in different kinds of neurological diseases; however, its role and the potential molecular mechanisms in secondary brain injury (SBI) after intracerebral hemorrhage (ICH) are still unknown. In this study, we investigate whether treatment with ghrelin may attenuate SBI in a murine ICH model, and if so, whether the neuroprotective effects are due to the inhibition of nucleotide-binding oligomerization domain-like receptor pyrin domain-containing 3 (NLRP3) inflammasome activation and promotion of nuclear factor-E2-related factor 2 (Nrf2)/antioxidative response element (ARE) signaling pathway. Stereotactically intrastriatal infusion of autologous blood was performed to mimic ICH. Ghrelin was given intraperitoneally immediately following ICH and again 1 h later. Results showed that ghrelin attenuated neurobehavioral deficits, brain edema, hematoma volume, and perihematomal cell death post-ICH. Ghrelin inhibited the NLRP3 inflammasome activation and subsequently suppressed the neuroinflammatory response as evidenced by reduced microglia activation, neutrophil infiltration, and pro-inflammatory mediators release after ICH. Additionally, ghrelin alleviated ICH-induced oxidative stress according to the chemiluminescence of luminol and lucigenin, malondialdehyde (MDA) content, and total superoxide dismutase (SOD) activity assays. These changes were accompanied by upregulation of Nrf2 expression, Nrf2 nuclear accumulation, and enhanced Nrf2 DNA binding activity, as well as by increased expressions of Nrf2 downstream target antioxidative genes, including NAD(P)H quinine oxidoreductase-1 (NQO1), glutathione cysteine ligase regulatory subunit (GCLC), and glutathione cysteine ligase modulatory subunit (GCLM). Together, our data suggested that ghrelin protected against ICH-induced SBI by inhibiting NLRP3 inflammasome activation and promoting Nrf2/ARE signaling pathway.
BMAL1 attenuates intracerebral hemorrhage-induced secondary brain injury in rats by regulating the Nrf2 signaling pathway.
Annals of translational medicine
BACKGROUND:Intracerebral hemorrhage (ICH) is a severe cerebrovascular disease with high morbidity and mortality rates. Oxidative stress and inflammation are important pathological mechanisms of secondary brain injury (SBI) after ICH. Brain and muscle Arnt-like protein 1 (BMAL1), which forms the core component of the circadian clock, was previously shown to be involved in many diseases and to participate in oxidative stress and inflammatory responses. However, the role of BMAL1 in SBI following ICH is unknown. In addition, treatments targeting miR-155 and its downstream signaling pathway may exert a beneficial effect on SBI after ICH, while miR-155 may regulate Bmal1 mRNA stability and translation. Nevertheless, researchers have not clearly determined whetheantagomir-155 upregulates BMAL1 expression and subsequently attenuates ICH-induced brain injury in rats. METHODS:After establishing an ICH rat model by injecting autologous blood, the time course of changes in levels of the BMAL1 protein after ICH was analyzed. Subsequently, this study was designed to investigate the potential role and mechanisms of BMAL1 in SBI following ICH using lentiviral overexpression and antagomir-155 treatments. RESULTS:BMAL1 protein levels were significantly decreased in the ICH group compared to the sham group. Genetic overexpression of BMAL1 alleviated oxidative stress, inflammation, brain edema, blood-brain barrier injury, neuronal death, and neurological dysfunction induced by ICH. On the other hand, we observed that inhibiting miRNA-155 using antagomir-155 promoted the expression of BMAL1 and further activated the nuclear factor erythroid 2-related factor 2 (Nrf2) signaling pathway to attenuate brain injury after ICH. CONCLUSIONS:These results reveal that BMAL1 serves as a neuroprotective agent in ICH and upregulation of BMAL1 attenuates ICH-induced SBI. Therefore, BMAL1 may be a promising therapeutic target for SBI following ICH.
Overexpression of CX3CR1 in Adipose-Derived Stem Cells Promotes Cell Migration and Functional Recovery After Experimental Intracerebral Hemorrhage.
Li Gaigai,Yu Haihan,Liu Na,Zhang Ping,Tang Yingxin,Hu Yang,Zhang Ye,Pan Chao,Deng Hong,Wang Jiahui,Li Qi,Tang Zhouping
Frontiers in neuroscience
Stem cell therapy has emerged as a new promising therapeutic strategy for intracerebral hemorrhage (ICH). However, the efficiency of stem cell therapy is partially limited by low retention and engraftment of the delivered cells. Therefore, it's necessary to improve the migration ability of stem cells to the injured area in order to save the costs and duration of cell preparation. This study aimed to investigate whether overexpression of CX3CR1, the specific receptor of chemokine fractalkine (FKN), in adipose-derived stem cells (ADSCs) can stimulate the cell migration to the injured area in the brain, improve functional recovery and protect against cell death following experimental ICH. ADSCs were isolated from subcutaneous adipose tissues of rats. ICH was induced by means of an injection of collagenase type VII. ELISA showed that the expression levels of fractalkine/FKN were increased at early time points, with a peak at day 3 after ICH. And it was found that different passages of ADSCs could express the chemokine receptor CX3CR1. Besides, the chemotactic movements of ADSCs toward fractalkine have been verified by transwell migration assay. ADSCs overexpressing CX3CR1 were established through lentivirus transfection. We found that after overexpression of CX3CR1 receptor, the migration ability of ADSCs was increased both and . In addition, reduced cell death and improved sensory and motor functions were seen in the mice ICH model. Thus, ADSCs overexpression CX3CR1 might be taken as a promising therapeutic strategy for the treatment of ICH.
A Role for Endoplasmic Reticulum Stress in Intracerebral Hemorrhage.
Mohammed Thangameeran Shaik Ismail,Tsai Sheng-Tzung,Hung Hsiang-Yi,Hu Wei-Fen,Pang Cheng-Yoong,Chen Shin-Yuan,Liew Hock-Kean
The endoplasmic reticulum (ER) is an intracellular organelle that performs multiple functions, such as lipid biosynthesis, protein folding, and maintaining intracellular calcium homeostasis. Thus, conditions wherein the ER is unable to fold proteins is defined as ER stress, and an inbuilt quality control mechanism, called the unfolded protein response (UPR), is activated during ER stress, which serves as a recovery system that inhibits protein synthesis. Further, based on the severity of ER stress, the response could involve both proapoptotic and antiapoptotic phases. Intracerebral hemorrhage (ICH) is the second most common subtype of cerebral stroke and many lines of evidence have suggested a role for the ER in major neurological disorders. The injury mechanism during ICH includes hematoma formation, which in turn leads to inflammation, elevated intracranial pressure, and edema. A proper understanding of the injury mechanism(s) is required to effectively treat ICH and closing the gap between our current understanding of ER stress mechanisms and ICH injury can lead to valuable advances in the clinical management of ICH.
Potential of stem cell therapy in intracerebral hemorrhage.
Huang Abel Po-Hao,Hsu Yi-Hua,Wu Meng-Shiue,Tsai Hsin-Han,Su Chia-Yi,Ling Thai-Yen,Hsu Shan-Hui,Lai Dar-Ming
Molecular biology reports
Spontaneous intracerebral hemorrhage (ICH) is a common disease associated with high mortality and morbidity. The treatment of patients with ICH includes medical and surgical interventions. New areas of surgical intervention have been focused on the evacuation of hematoma through minimally invasive neurosurgery. In contrast, there have been no significant advances in the development of medical interventions for functional recovery after ICH. Stem cells exert multiple therapeutic functions and have emerged as a promising treatment strategy. Herein, we summarized the pathophysiology of ICH and its treatment targets, and we introduced the therapeutic mechanisms of stem cells (e.g. neutrotrophy and neuroregeneration). Moreover, we reviewed and summarized the experimental designs of the preclinical studies, including the types of cells and the timing and routes of stem cell administration. We further listed and reviewed the completed/published and ongoing clinical trials supporting the safety and efficacy of stem cell therapy in ICH. The limitations of translating preclinical studies into clinical trials and the objectives of future studies were discussed. In conclusion, current literatures showed that stem cell therapy is a promising treatment in ICH and further translation research on judiciously selected group of patients is warranted before it can be extensively applied in clinical practice.
Potential effects of mesenchymal stem cell derived extracellular vesicles and exosomal miRNAs in neurological disorders.
Nakano Masako,Fujimiya Mineko
Neural regeneration research
Mesenchymal stem cells are multipotent cells that possess anti-inflammatory, anti-apoptotic and immunomodulatory properties. The effects of existing drugs for neurodegenerative disorders such as Alzheimer's disease are limited, thus mesenchymal stem cell therapy has been anticipated as a means of ameliorating neuronal dysfunction. Since mesenchymal stem cells are known to scarcely differentiate into neuronal cells in damaged brain after transplantation, paracrine factors secreted from mesenchymal stem cells have been suggested to exert therapeutic effects. Extracellular vesicles and exosomes are small vesicles released from mesenchymal stem cells that contain various molecules, including proteins, mRNAs and microRNAs. In recent years, administration of exosomes/extracellular vesicles in models of neurological disorders has been shown to improve neuronal dysfunctions, via exosomal transfer into damaged cells. In addition, various microRNAs derived from mesenchymal stem cells that regulate various genes and reduce neuropathological changes in various neurological disorders have been identified. This review summarizes the effects of exosomes/extracellular vesicles and exosomal microRNAs derived from mesenchymal stem cells on models of stroke, subarachnoid and intracerebral hemorrhage, traumatic brain injury, and cognitive impairments, including Alzheimer's disease.
Extracellular vesicles derived from bone marrow mesenchymal stem cells alleviate neuroinflammation after diabetic intracerebral hemorrhage via the miR-183-5p/PDCD4/NLRP3 pathway.
Ding H,Jia Y,Lv H,Chang W,Liu F,Wang D
Journal of endocrinological investigation
OBJECTIVES:Intracerebral hemorrhage (ICH) induced by diabetes results in further brain injury and nerve cell death. Bone marrow mesenchymal stem cell (BMSC) transplantation contributes to attenuating neurological deficits after ICH. This study investigated the mechanism of extracellular vesicles (EVs) derived from BMSCs in reducing neuroinflammation after diabetic ICH. METHODS:BMSC-EVs were isolated and identified. The rat model of db/db-ICH was established and the model rats were administered with EVs. miR-183-5p expression in brain tissues of db/db-ICH rats was detected. The brain injury of db/db-ICH rats was evaluated by measuring neurobehavioral score, brain water content and inflammatory factors. BV2 cells were cultured in vitro to establish high-glucose (HG)-Hemin-BV2 cell model. The levels of reactive oxygen species (ROS) and inflammatory factors in BV2 cells were measured, and BV2 cell viability and apoptosis were assessed. The targeting relationship between miR-183-5p and PDCD4 was predicted and verified. The activation of PDCD4/NLRP3 pathway in rat brain tissues and BV2 cells was detected. RESULTS:miR-183-5p expression was reduced in db/db-ICH rats brain tissues. BMSC-EVs ameliorated cranial nerve function, decreased brain water content and repressed inflammatory response by carrying miR-183-5p. BMSC-EVs mitigated HG-Hemin-BV2 cell injury, reduced ROS level and suppressed inflammatory response. miR-183-5p targeted PDCD4. PDCD4 promoted BV2 cell inflammation by activating the NLRP3 pathway. BMSC-EVs inhibited HG-Hemin-BV2 cell inflammation through the miR-183-5p/PDCD4/NLRP3 pathway, and inhibition of miR-183-5p reversed the protective effect of EVs. CONCLUSION:BMSC-EVs carried miR-183-5p into db/db-ICH rat brain tissues and repressed the NLRP3 pathway by targeting PDCD4, thus alleviating neuroinflammation after diabetic ICH.
MicroRNA-21 Overexpression Promotes the Neuroprotective Efficacy of Mesenchymal Stem Cells for Treatment of Intracerebral Hemorrhage.
Zhang Heyu,Wang Yanzhe,Lv Qing,Gao Jun,Hu Liuting,He Zhiyi
Frontiers in neurology
Intracerebral hemorrhage (ICH) has high morbidity and mortality, with no effective treatment at present. One possible therapeutic strategy involves the use of mesenchymal stem cells (MSCs), which have shown promise in experimental models and have great potential for treating nervous illnesses in humans. However, many deficiencies in MSC treatment still need to be addressed, including their poor survival rate post-transplantation. Previously, we reported that the microRNA-21 (miR-21) is downregulated in ICH patients' blood and brain tissue. In this study, we aimed to examine its role and therapeutic efficacy in ICH using miR-21-overexpressing MSCs. We found that this microRNA can enhance MSC survival and recovery of neurological function in ICH rats. Its mechanism of action involves reduced neuronal apoptosis. In addition, we demonstrated that miR-21 can be transported to neurons through exosomes derived from MSCs and that it can target transient receptor potential melastatin 7 ( to alleviate neuronal injury following ICH. We also observed that the NF-κB pathway is involved in the regulation of miR-21 in neural cells. In conclusion, miR-21 significantly enhances the survival of MSCs in ICH, and miR-21-overexpressing MSCs clearly improved neurological function in ICH rats. Transplantation of miR-21-overexpressing MSCs may, therefore, provide an effective strategy for neuroprotection and treatment of cerebrovascular diseases.
Hypoxic preconditioning rejuvenates mesenchymal stem cells and enhances neuroprotection following intracerebral hemorrhage via the miR-326-mediated autophagy.
Liu Jianyang,He Jialin,Ge Lite,Xiao Han,Huang Yan,Zeng Liuwang,Jiang Zheng,Lu Ming,Hu Zhiping
Stem cell research & therapy
BACKGROUND:Intracerebral hemorrhage (ICH) is a major public health concern, and mesenchymal stem cells (MSCs) hold great potential for treating ICH. However, the quantity and quality of MSCs decline in the cerebral niche, limiting the potential efficacy of MSCs. Hypoxic preconditioning is suggested to enhance the survival of MSCs and augment the therapeutic efficacy of MSCs in ICH. MicroRNAs (miRNAs) are known to mediate cellular senescence. However, the precise mechanism by which miRNAs regulate the senescence of hypoxic MSCs remains to be further studied. In the present study, we evaluated whether hypoxic preconditioning enhances the survival and therapeutic effects of olfactory mucosa MSC (OM-MSC) survival and therapeutic effects in ICH and investigated the mechanisms by which miRNA ameliorates hypoxic OM-MSC senescence. METHODS:In the in vivo model, ICH was induced in mice by administration of collagenase IV. At 24 h post-ICH, 5 × 10 normoxia or hypoxia OM-MSCs or saline was administered intracerebrally. The behavioral outcome, neuronal apoptosis, and OM-MSC survival were evaluated. In the in vitro model, OM-MSCs were exposed to hemin. Cellular senescence was examined by evaluating the expressions of P16INK4A, P21, P53, and by β-galactosidase staining. Microarray and bioinformatic analyses were performed to investigate the differences in the miRNA expression profiles between the normoxia and hypoxia OM-MSCs. Autophagy was confirmed using the protein expression levels of LC3, P62, and Beclin-1. RESULTS:In the in vivo model, transplanted OM-MSCs with hypoxic preconditioning exhibited increased survival and tissue-protective capability. In the in vitro model, hypoxia preconditioning decreased the senescence of OM-MSCs exposed to hemin. Bioinformatic analysis identified that microRNA-326 (miR-326) expression was significantly increased in the hypoxia OM-MSCs compared with that of normoxia OM-MSCs. Upregulation of miR-326 alleviated normoxia OM-MSC senescence, whereas miR-326 downregulation increased hypoxia OM-MSC senescence. Furthermore, we showed that miR-326 alleviated cellular senescence by upregulating autophagy. Mechanistically, miR-326 promoted the autophagy of OM-MSCs via the PI3K signaling pathway by targeting polypyrimidine tract-binding protein 1 (PTBP1). CONCLUSIONS:Our study shows that hypoxic preconditioning delays OM-MSC senescence and augments the therapeutic efficacy of OM-MSCs in ICH by upregulating the miR-326/PTBP1/PI3K-mediated autophagy.
Hypoxia-preconditioned mesenchymal stem cells attenuate microglial pyroptosis after intracerebral hemorrhage.
Annals of translational medicine
BACKGROUND:Microglia plays a vital role in neuroinflammation, contributing to the pathogenesis of intracerebral hemorrhage (ICH)-induced brain injury. Mesenchymal stem cells (MSCs) hold great potential for treating ICH. We previously revealed that MSCs ameliorate the microglial pyroptosis caused by an ischemic stroke. However, whether MSCs can modulate microglial pyroptosis after ICH remains unknown. This study aimed to investigate the neuroprotective effects of hypoxia-preconditioned olfactory mucosa MSCs (OM-MSCs) on ICH and the possible mechanisms. METHODS:ICH was induced in mice via administration of collagenase IV. At 6 h post-ICH, 2-4×10 normoxic/hypoxic OM-MSCs or saline were intracerebrally administered. To evaluate the neuroprotective effects, the behavioral outcome, apoptosis, and neuronal injury were measured. Microglia activation and pro-inflammatory cytokines were applied to detect neuroinflammation. Microglial pyroptosis was determined by western blotting, immunofluorescence staining, and transmission electron microscopy (TEM). RESULTS:The two OM-MSC-transplanted groups exhibited significantly improved functional recovery and reduced neuronal injury, especially the hypoxic OM-MSCs group. Hypoxic OM-MSCs attenuated microglial activation as well as the levels of interleukin-1β (IL-1β) and tumor necrosis factor-α (TNF-α). Moreover, we found that hypoxia-preconditioned OM-MSCs ameliorated pyroptosis by diminishing the levels of pyroptosis-associated proteins in peri-hematoma brain tissues, decreasing the expression of the microglial nod-like receptor family protein 3 (NLRP3) and caspase-1, and reducing the membrane pores on microglia post-ICH. CONCLUSIONS:Our study showed that hypoxic preconditioning augments the therapeutic efficacy of OM-MSCs, and hypoxia-preconditioned OM-MSCs alleviate microglial pyroptosis in the ICH model.
Exosomes Derived from MicroRNA-146a-5p-Enriched Bone Marrow Mesenchymal Stem Cells Alleviate Intracerebral Hemorrhage by Inhibiting Neuronal Apoptosis and Microglial M1 Polarization.
Drug design, development and therapy
INTRODUCTION:Intracerebral hemorrhage (ICH) is a devastating type of stroke with high mortality, and the effective therapies for ICH remain to be explored. Exosomes (Exos) have been found to play important roles in cell communication by transferring molecules, including microRNAs (miRNAs/miRs). MiRNAs are critical regulators of genes involved in many various biological processes and have been demonstrated to aggravate or alleviate brain damages induced by ICH. The aim of the present study was to investigate the effect of Exos derived from miR-146a-5p-enriched bone marrow mesenchymal stem cells (BMSCs-miR-146a-5p-Exos) on experimental ICH. METHODS:ICH was induced in adult male Sprague-Dawley rats by an intrastriatal injection of collagenase type IV. At 24 h after surgery, Exos were administrated. For detecting apoptotic cells, TUNEL staining was performed using an in situ Cell Death Detection Kit. Fluoro-Jade B staining was performed to detect degenerating neurons. Immunofluorescence assay was performed to detect the expression of myeloperoxidase (MPO) and OX-42. The binding of miR-146a-5p and its target genes was confirmed by luciferase reporter assay. RESULTS:At 24 h after surgery, BMSCs-miR-146a-5p-Exos administration significantly improved neurological function, reduced apoptotic and degenerative neurons, and inhibited inflammatory response. Furthermore, miR-146a-5p-enriched Exos obviously inhibited the M1 polarization of microglia after ICH in rats, accompanied by the reduced expression of pro-inflammatory mediators releasing by M1 microglia including inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2) and monocyte chemoattractant protein-1 (MCP-1). Finally, we observed that miR-146a-5p directly targeted interleukin-1 receptor-associated kinase1 (IRAK1) and nuclear factor of activated T cells 5 (NFAT5), which contributed to the inflammation response and the polarization of M1 microglia/macrophages. CONCLUSION:We demonstrated that miR-146a-5p-riched BMSCs-Exos could offer neuroprotection and functional improvements after ICH through reducing neuronal apoptosis, and inflammation associated with the inhibition of microglial M1 polarization by downregulating the expression of IRAK1 and NFAT5.
Electroacupuncture Enhance Therapeutic Efficacy of Mesenchymal Stem Cells Transplantation in Rats With Intracerebral Hemorrhage.
Deng Li,Zhou Ling,Zhu Yan,Fan Guangbi,Tang Huajun,Zheng Yujie,Gao Xiaoqing,Guo Kan,Zhou Peng,Yang Chaoxian
Stem cell reviews and reports
BACKGROUND:Previous studies have showed the beneficial effects of mesenchymal stem cells (MSCs) on experimental intracerebral hemorrhage (ICH) animal. Enhancement of the treatment efficacy of MSCs in ICH is essential, considering the diseases association with high rates of disability and mortality. Some auxiliary methods to enhance the beneficial efficacy of MSCs have been introduced. However, the effect of electroacupuncture (EA) on the therapeutic efficacy of MSCs transplantation in hemorrhagic stroke and its potential mechanism is not explored. METHODS:ICH rat models were established using collagenase and heparin. 48 h after ICH induction, the rats were randomly divided into model control (MC), MSCs transplantation (MSCs), EA stimulation (EA) and MSCs transplantation combined with EA stimulation (MSCs + EA) groups. We used mNSS test and gait analysis to assess neurological function of rats, and PET/CT to evaluate the volume of hemorrhage focus and level of glucose uptake. The concentrations of MDA, SOD, NSE, S100B and MBP in serum or plasma were examined with ELISA. Neural differentiation of MSCs, and the expressions of Bcl-2, Bax, Arg-1 and iNOS proteins around hematoma were detected by immunofluorescence and immunohistochemistry staining respectively. Western blot was carried out to analyze the expression levels of COX4, OGDH, PDH-E1α, Bcl-2 and Bax proteins. TUNEL staining was used to estimate cell apoptosis and transmission electron microscopy (TEM) was used to observe the ultrastructure and number of mitochondria. RESULTS:Our data showed that EA promoted neuron-like differentiation of transplanted MSCs and the expressions of BDNF and NGF proteins in ICH rats. The score of mNSS and the gait analysis showed that the recovery of the neurological function in the MSCs + EA group was better than that in the MSCs and EA groups. EA improved the structure of brain tissue, and alleviated brain injury further after MSCs transplantation in ICH rats. When compared with the MSCs and EA groups, the level of glucose uptake and numbers of mitochondria and Arg-1 positive cells in MSCs + EA group increased significantly, but the numbers of apoptotic cells and iNOS positive cells and volume of hemorrhage focus reduced. The expressional levels of COX4, OGDH, PDH-E1α and Bcl-2 proteins increased, while the expressional level of Bax protein decreased compared with those in the MSCs and EA groups. CONCLUSIONS:Our results reveal that EA improve therapeutic efficacy of MSCs transplantation in ICH rats.
Effects of GDNF-Transfected Marrow Stromal Cells on Rats with Intracerebral Hemorrhage.
Deng Li,Gao Xiaoqing,Fan Guangbi,Yang Chaoxian
Journal of stroke and cerebrovascular diseases : the official journal of National Stroke Association
OBJECTIVE:The present study aimed to investigate the effects of Mesenchymal stem cells/glial cell line derived neurotrophic factor (MSCs/GDNF) transplantation on nerve reconstruction in rats with intracerebral hemorrhage. METHODS:GDNF transduction to MSCs was using adenovirus vector pAdEasy-1-pAdTrack-CMV prepared. Intracerebral hemorrhage (ICH) was induced by injection of collagenase and heparin into the caudate putamen. At the third day after a collagenase-induced ICH, adult male SD rats were randomly divided into saline group, MSCs group and MSCs/GDNF group. Immunofluorescence and RT-PCR were performed to detect the differentiation of MSCs or MSCs with an adenovirus vector encoding GDNF gene in vivo and in vitro. RESULT:After 6 hours of induction, both MSCs and MSCs/GDNF expressed neuro or glial specific markers and synaptic-associated proteins (SYN, GAP-43, PSD-95); additionally, they secreted bioactive compounds (BDNF, NGF-β). MSCs/GDNF transplantation, compared to MSCs and saline solution injection, significantly improved neurological functions after ICH. The grafted MSCs or MSCs/GDNF survived in the striatum after 2 weeks of transplantation and expressed the neural cell-specific biomarkers NSE, MAP2, and GFAP. CONCLUSION:These findings demonstrate that MSCs/GDNF transplantation contributes to improved neurological function in experimental ICH rats. The mechanisms are possibly due to neuronal replacement and enhanced neurotrophic factor secretion.
Effect of Pretreatment with the NADPH Oxidase Inhibitor Apocynin on the Therapeutic Efficacy of Human Placenta-Derived Mesenchymal Stem Cells in Intracerebral Hemorrhage.
Min Saehong,Kim Ok Joon,Bae Jinkun,Chung Tae Nyoung
International journal of molecular sciences
Several studies have demonstrated the beneficial effect of mesenchymal stem cells (MSCs) on intracerebral hemorrhage (ICH). Enhancement of the therapeutic efficacy of MSCs in ICH is necessary, considering the diseases high association with mortality and morbidity. Various preconditioning methods to enhance the beneficial properties of MSCs have been introduced. We suggested apocynin, a well-known nicotinamide adenine dinucleotide phosphate (NADPH) oxidase inhibitor, as a novel preconditioning regimen to enhance the therapeutic efficacy of MSCs in ICH. Rat ICH models were made using bacterial collagenase. 24 h after ICH induction, the rats were randomly divided into apocynin-preconditioned MSC-treated (Apo-MSC), naïve MSC-treated and control groups. Hematoma volume, brain edema, and degenerating neuron count were compared at 48 h after the ICH induction. The expression of tight junction proteins (occludin, zona occludens [ZO]-1) were also compared. Hematoma size, hemispheric enlargement and degenerating neuron count were significantly lower in the Apo-MSC group than in the naïve MSC group ( = 0.004, 0.013 and 0.043, respectively), while the expression of occludin was higher ( = 0.024). Apocynin treatment enhances the therapeutic efficacy of MSCs in ICH in the acute stage, through the improvement of the beneficial properties of MSCs, such as neuroprotection and the reinforcement of endovascular integrity of cerebral vasculature.
Effect of MicroRNA-126a-3p on Bone Marrow Mesenchymal Stem Cells Repairing Blood-brain Barrier and Nerve Injury after Intracerebral Hemorrhage.
Wang Chunyan,Cao Jingwei,Duan Shurong,Xu Ran,Yu Hongli,Huo Xin,Qian Yuanyuan
Journal of stroke and cerebrovascular diseases : the official journal of National Stroke Association
OBJECTIVE:Intracerebral hemorrhage (ICH) is a disease that threatens human health due to its high morbidity and mortality. On behalf of finding the better methods in the treatment of ICH, researchers pay more attention to a new technology which is finding effective genes to modify stem cells. METHODS:In this study, we isolated, cultured and identified bone marrow mesenchymal stem cells (MSCs) in vitro. Further, the MSCs (transfected with lentivirus expressing microRNA-126a-3p (miR-126)) were injected into the type Ⅶ collagenase-induced ICH rats to investigate the recovery effects of blood-brain barrier (BBB) and nerve damage in vivo. RESULTS:The MSCs surface marker molecules (CD29: 98.5%; CD90: 96.5%) were highly expressed, and the blood cell surface molecule was negatively expressed (CD45: 2%). Meanwhile, it was verified that miR-126 facilitated the differentiation of MSCs into vascular endothelial cells, owing to the rise of markers (CD31 and VE-cadherin). The modified neurological severity score, modified limb placing test score, brain water content and evans blue content were reduced after transplanted miR-126-modified MSCs. It was found that miR-126 accelerated the differentiation of MSCs into vascular endothelial cells via immunohistochemical staining in vivo. HE staining indicated the area of edema was obviously decreased compared with that in ICH + vector-MSCs group. MiR-126-modified MSCs alleviated the cell apoptosis in brain tissues by TUNEL assay. In addition, the mRNA and protein expression of protease activated receptor-1 and matrix metalloproteinase-9 were diminished, whilst the expression of zonula occludens-1 (ZO-1) and claudin-5 were enhanced in ICH+miR-126-MSCs group. Immunofluorescence assay revealed that miR-126-modified MSCs decreased the disruption of tight junction (ZO-1 and claudin-5). CONCLUSIONS:All data illustrate that miR-126-modified MSCs repair BBB and nerve injury after ICH.
Mesenchymal stromal cell injection protects against oxidative stress in Escherichia coli-induced acute lung injury in mice.
Shalaby Sally M,El-Shal Amal S,Abd-Allah Somia H,Selim Assmaa O,Selim Sally A,Gouda Zienab A,Abd El Motteleb Dalia M,Zanfaly Hala E,El-Assar Heba M,Abdelazim Shymaa
BACKGROUND AIMS:Stem cells may be a promising therapy for acute respiratory distress syndrome. Recent in vivo and in vitro studies suggested that the mesenchymal stromal cells (MSCs) have anti-oxidative stress properties. We hypothesized that intravenous injection of bone marrow-derived mesenchymal stem cells (MSCs) could attenuate Escherichia coli-induced acute lung injury (ALI) in mice by controlling the oxidative stress status. METHODS:Eighty mice were randomly divided into four groups: group 1 (control group) received 25 μL of saline as a vehicle; group 2 contained E coli-induced ALI mice; group 3 included mice that received MSCs before induction of ALI; group 4 included mice that received MSCs after induction of ALI. Lung samples were isolated and assayed for oxidative stress variables and histopathologic analysis. Total anti-oxidant capacity was measured in broncho-alveolar lavage. RESULTS:Pre- and post-injury MSC injection increased survival, reduced pulmonary edema and attenuated lung injuries in ALI mice. Histologically, MSCs exhibited a considerable degree of preservation of the pulmonary alveolar architecture. An increase of anti-oxidant enzyme activities and a decrease of myeloperoxidase activity and malondialdehyde levels in the MSC recipient groups versus the ALI group were found. Furthermore, the total anti-oxidant capacity and reduced glutathione levels were significantly increased in MSCs recipient groups versus the ALI group. Weak +ve inducible nitric oxide synthase immuno-expression in groups that received MSCs was detected. Pre-injury MSC injection showed better effects than did post-injury MSC injection. CONCLUSIONS:Systemic bone marrow-derived MSC injection was effective in modulating the oxidative stress status in E coli-induced acute lung injury in mice.
Programmed Death Ligand 1 Plays a Neuroprotective Role in Experimental Autoimmune Neuritis by Controlling Peripheral Nervous System Inflammation of Rats.
Ding Yanan,Han Ranran,Jiang Wei,Xiao Jinting,Liu Haijie,Chen Xiuju,Li Xiaowen,Hao Junwei
Journal of immunology (Baltimore, Md. : 1950)
Programmed death 1 (PD-1; CD279), a member of the CD28 family, is an inhibitory receptor on T cells and is responsible for T cell dysfunction in infectious diseases and cancers. The ligand for PD-1, programmed death ligand 1 (PD-L1; also known as B7-H1, CD274), is a member of the B7 family. The engagement of PD-1 with programmed death ligand can downregulate autoreactive T cells that participate in multiple autoimmune diseases. Experimental autoimmune neuritis (EAN) is an animal model of Guillain-Barré syndrome, and the pathogenesis of EAN is mediated principally through T cells and macrophages. In this study, we investigated the effects of PD-L1 in EAN rats. For preventative and therapeutic management, we administered PD-L1, which successfully decreased the severity of EAN; it alleviated the neurologic course of EAN, as well as inhibited the infiltration of inflammatory cells and demyelination of sciatic nerves. Our data revealed that PD-L1 treatment inhibited lymphocyte proliferation and altered T cell differentiation by inducing decreases in IFN-γCD4 Th1 cells and IL-17CD4 Th17 cells and increases in IL-4CD4 Th2 cells and Foxp3CD4 regulatory T cells. The expression levels of p-STAT3 and Foxp3 were significantly different in PD-L1-treated groups compared with the control group. Additionally, PD-L1 regulated the expression of Foxp3 and p-STAT3 in EAN, probably by inhibiting PI3K/AKT/mTOR signaling expression. In summary, PD-L1 is a potentially useful agent for the treatment of EAN because of its anti-inflammatory and neuroprotective effects.
Programmed death-1 pathway in host tissues ameliorates Th17/Th1-mediated experimental chronic graft-versus-host disease.
Fujiwara Hideaki,Maeda Yoshinobu,Kobayashi Koichiro,Nishimori Hisakazu,Matsuoka Ken-Ichi,Fujii Nobuharu,Kondo Eisei,Tanaka Takehiro,Chen Lieping,Azuma Miyuki,Yagita Hideo,Tanimoto Mitsune
Journal of immunology (Baltimore, Md. : 1950)
Chronic graft-versus-host disease (GVHD) is a major cause of late death and morbidity after allogeneic hematopoietic cell transplantation, but its pathogenesis remains unclear. We investigated the role of the programmed death-1 (PD-1) pathway in chronic GVHD using a well-defined mouse model of B10.D2 (H-2(d)) donor to BALB/c (H-2(d)) recipients. PD-1 expression on allogeneic donor T cells was upregulated continuously in chronic GVHD development, whereas PD-L1 expression in host tissues was transiently upregulated and declined to basal levels in the late posttransplant period. Blockade of the PD-1 pathway by anti-PD-1, anti-PD-L1, or anti-PD-L2 mAbs exacerbated clinical and pathologic chronic GVHD. Chimeric mice revealed that PD-L1 expression in host tissues suppressed expansion of IL-17(+)IFN-γ(+) T cells, and that PD-L1 expression on hematopoietic cells plays a role in the development of regulatory T cells only during the early transplantation period but does not affect the severity of chronic GVHD. Administration of the synthetic retinoid Am80 overcame the IL-17(+)IFN-γ(+) T cell expansion caused by PD-L1 deficiency, resulting in reduced chronic GVHD damage in PD-L1(-/-) recipients. Stimulation of the PD-1 pathway also alleviated chronic GVHD. These results suggest that the PD-1 pathway contributes to the suppression of Th17/Th1-mediated chronic GVHD and may represent a new target for the prevention or treatment of chronic GVHD.
Mesenchymal stromal cells induce regulatory T cells via epigenetic conversion of human conventional CD4 T cells in vitro.
Azevedo Rita I,Minskaia Ekaterina,Fernandes-Platzgummer Ana,Vieira Ana I S,da Silva Cláudia L,Cabral Joaquim M S,Lacerda João F
Stem cells (Dayton, Ohio)
Regulatory T cells (Treg) play a critical role in immune tolerance. The scarcity of Treg therapy clinical trials in humans has been largely due to the difficulty in obtaining sufficient Treg numbers. We performed a preclinical investigation on the potential of mesenchymal stromal cells (MSCs) to expand Treg in vitro to support future clinical trials. Human peripheral blood mononuclear cells from healthy donors were cocultured with allogeneic bone marrow-derived MSCs expanded under xenogeneic-free conditions. Our data show an increase in the counts and frequency of CD4 CD25 Foxp3 CD127 Treg cells (4- and 6-fold, respectively) after a 14-day coculture. However, natural Treg do not proliferate in coculture with MSCs. When purified conventional CD4 T cells (Tcon) are cocultured with MSCs, only cells that acquire a Treg-like phenotype proliferate. These MSC-induced Treg-like cells also resemble Treg functionally, since they suppress autologous Tcon proliferation. Importantly, the DNA methylation profile of MSC-induced Treg-like cells more closely resembles that of natural Treg than of Tcon, indicating that this population is stable. The expression of PD-1 is higher in Treg-like cells than in Tcon, whereas the frequency of PDL-1 increases in MSCs after coculture. TGF-β levels are also significantly increased MSC cocultures. Overall, our data suggest that Treg enrichment by MSCs results from Tcon conversion into Treg-like cells, rather than to expansion of natural Treg, possibly through mechanisms involving TGF-β and/or PD-1/PDL-1 expression. This MSC-induced Treg population closely resembles natural Treg in terms of phenotype, suppressive ability, and methylation profile.
PD-L1 (Programmed Death Ligand 1) Protects Against Experimental Intracerebral Hemorrhage-Induced Brain Injury.
Han Ranran,Luo Jiaying,Shi Yanchao,Yao Yang,Hao Junwei
BACKGROUND AND PURPOSE:Intracerebral hemorrhage (ICH) is a neurologically destructive stroke, for which no valid treatment is available. This preclinical study examined the therapeutic effect of PD-L1 (programmed death ligand 1), a B7 family member and a ligand for both PD-1 (programmed death 1) and B7-1 (CD80), in a murine ICH model. METHODS:ICH was induced by injecting autologous blood into 252 male C57BL/6 and Rag1 mice. One hour later, ICH mice were randomly assigned to receive an intraperitoneal injection of vehicle, PD-L1, or anti-PD-L1 antibody. Neurological function was assessed along with brain edema, brain infiltration of immune cells, blood-brain barrier integrity, neuron death, and mTOR (mammalian target of rapamycin) pathway products. RESULTS:PD-L1 significantly attenuated neurological deficits, reduced brain edema, and decreased hemorrhage volume in ICH mice. PD-L1 specifically downsized the number of brain-infiltrating CD4 T cells and the percentages of Th1 and Th17 cells but increased the percentages of Th2 and regulatory T cells. In the PD-L1-treated group, we observed an amelioration of the inflammatory milieu, decreased cell death, and enhanced blood-brain barrier integrity. PD-L1 also inhibited the mTOR pathway. The administration of anti-PD-L1 antibody produced the opposite effects to those of PD-L1 in ICH mice. CONCLUSIONS:PD-L1 provided protection from the damaging consequences of ICH.
Mechanism of White Matter Injury and Promising Therapeutic Strategies of MSCs After Intracerebral Hemorrhage.
Li Jing,Xiao Linglong,He Dian,Luo Yunhao,Sun Haitao
Frontiers in aging neuroscience
Intracerebral hemorrhage (ICH) is the most fatal subtype of stroke with high disability and high mortality rates, and there is no effective treatment. The predilection site of ICH is in the area of the basal ganglia and internal capsule (IC), where exist abundant white matter (WM) fiber tracts, such as the corticospinal tract (CST) in the IC. Proximal or distal white matter injury (WMI) caused by intracerebral parenchymal hemorrhage is closely associated with poor prognosis after ICH, especially motor and sensory dysfunction. The pathophysiological mechanisms involved in WMI are quite complex and still far from clear. In recent years, the neuroprotection and repairment capacity of mesenchymal stem cells (MSCs) has been widely investigated after ICH. MSCs exert many unique biological effects, including self-recovery by producing growth factors and cytokines, regenerative repair, immunomodulation, and neuroprotection against oxidative stress, providing a promising cellular therapeutic approach for the treatment of WMI. Taken together, our goal is to discuss the characteristics of WMI following ICH, including the mechanism and potential promising therapeutic targets of MSCs, aiming at providing new clues for future therapeutic strategies.
Treatment with Mesenchymal-Derived Extracellular Vesicles Reduces Injury-Related Pathology in Pyramidal Neurons of Monkey Perilesional Ventral Premotor Cortex.
Medalla Maria,Chang Wayne,Calderazzo Samantha M,Go Veronica,Tsolias Alexandra,Goodliffe Joseph W,Pathak Dhruba,De Alba Diego,Pessina Monica,Rosene Douglas L,Buller Benjamin,Moore Tara L
The Journal of neuroscience : the official journal of the Society for Neuroscience
Functional recovery after cortical injury, such as stroke, is associated with neural circuit reorganization, but the underlying mechanisms and efficacy of therapeutic interventions promoting neural plasticity in primates are not well understood. Bone marrow mesenchymal stem cell-derived extracellular vesicles (MSC-EVs), which mediate cell-to-cell inflammatory and trophic signaling, are thought be viable therapeutic targets. We recently showed, in aged female rhesus monkeys, that systemic administration of MSC-EVs enhances recovery of function after injury of the primary motor cortex, likely through enhancing plasticity in perilesional motor and premotor cortices. Here, using whole-cell patch-clamp recording and intracellular filling in acute slices of ventral premotor cortex (vPMC) from rhesus monkeys () of either sex, we demonstrate that MSC-EVs reduce injury-related physiological and morphologic changes in perilesional layer 3 pyramidal neurons. At 14-16 weeks after injury, vPMC neurons from both vehicle- and EV-treated lesioned monkeys exhibited significant hyperexcitability and predominance of inhibitory synaptic currents, compared with neurons from nonlesioned control brains. However, compared with vehicle-treated monkeys, neurons from EV-treated monkeys showed lower firing rates, greater spike frequency adaptation, and excitatory:inhibitory ratio. Further, EV treatment was associated with greater apical dendritic branching complexity, spine density, and inhibition, indicative of enhanced dendritic plasticity and filtering of signals integrated at the soma. Importantly, the degree of EV-mediated reduction of injury-related pathology in vPMC was significantly correlated with measures of behavioral recovery. These data show that EV treatment dampens injury-related hyperexcitability and restores excitatory:inhibitory balance in vPMC, thereby normalizing activity within cortical networks for motor function. Neuronal plasticity can facilitate recovery of function after cortical injury, but the underlying mechanisms and efficacy of therapeutic interventions promoting this plasticity in primates are not well understood. Our recent work has shown that intravenous infusions of mesenchymal-derived extracellular vesicles (EVs) that are involved in cell-to-cell inflammatory and trophic signaling can enhance recovery of motor function after injury in monkey primary motor cortex. This study shows that this EV-mediated enhancement of recovery is associated with amelioration of injury-related hyperexcitability and restoration of excitatory-inhibitory balance in perilesional ventral premotor cortex. These findings demonstrate the efficacy of mesenchymal EVs as a therapeutic to reduce injury-related pathologic changes in the physiology and structure of premotor pyramidal neurons and support recovery of function.
The inhibitory effect of mesenchymal stem cell on blood-brain barrier disruption following intracerebral hemorrhage in rats: contribution of TSG-6.
Chen Min,Li Xifeng,Zhang Xin,He Xuying,Lai Lingfeng,Liu Yanchao,Zhu Guohui,Li Wei,Li Hui,Fang Qinrui,Wang Zequn,Duan Chuanzhi
Journal of neuroinflammation
BACKGROUND:Mesenchymal stem cells (MSCs) are well known having beneficial effects on intracerebral hemorrhage (ICH) in previous studies. The therapeutic mechanisms are mainly to investigate proliferation, differentiation, and immunomodulation. However, few studies have used MSCs to treat blood-brain barrier (BBB) leakage after ICH. The influence of MSCs on the BBB and its related mechanisms were investigated when MSCs were transplanted into rat ICH model in this study. METHODS:Adult male Sprague-Dawley (SD) rats were randomly divided into sham-operated group, PBS-treated (ICH + PBS) group, and MSC-treated (ICH + MSC) group. ICH was induced by injection of IV collagenase into the rats' brains. MSCs were transplanted intravenously into the rats 2 h after ICH induction in MSC-treated group. The following factors were compared: inflammation, apoptosis, behavioral changes, inducible nitric oxide synthase (iNOS), matrix metalloproteinase 9 (MMP-9), peroxynitrite (ONOO(-)), endothelial integrity, brain edema content, BBB leakage, TNF-α stimulated gene/protein 6 (TSG-6), and nuclear factor-κB (NF-κB) signaling pathway. RESULTS:In the ICH + MSC group, MSCs decreased the levels of proinflammatory cytokines and apoptosis, downregulated the density of microglia/macrophages and neutrophil infiltration at the ICH site, reduced the levels of iNOS and MMP-9, attenuated ONOO(-) formation, and increased the levels of zonula occludens-1 (ZO-1) and claudin-5. MSCs also improved the degree of brain edema and BBB leakage. The protective effect of MSCs on the BBB in ICH rats was possibly invoked by increased expression of TSG-6, which may have suppressed activation of the NF-κB signaling pathway. The levels of iNOS and ONOO(-), which played an important role in BBB disruption, decreased due to the inhibitory effects of TSG-6 on the NF-κB signaling pathway. CONCLUSIONS:Our results demonstrated that intravenous transplantation of MSCs decreased the levels of ONOO(-) and degree of BBB leakage and improved neurological recovery in a rat ICH model. This strategy may provide a new insight for future therapies that aim to prevent breakdown of the BBB in patients with ICH and eventually offer therapeutic options for ICH.
Bone marrow stromal cells attenuate sepsis via prostaglandin E(2)-dependent reprogramming of host macrophages to increase their interleukin-10 production.
Németh Krisztián,Leelahavanichkul Asada,Yuen Peter S T,Mayer Balázs,Parmelee Alissa,Doi Kent,Robey Pamela G,Leelahavanichkul Kantima,Koller Beverly H,Brown Jared M,Hu Xuzhen,Jelinek Ivett,Star Robert A,Mezey Eva
Sepsis causes over 200,000 deaths yearly in the US; better treatments are urgently needed. Administering bone marrow stromal cells (BMSCs -- also known as mesenchymal stem cells) to mice before or shortly after inducing sepsis by cecal ligation and puncture reduced mortality and improved organ function. The beneficial effect of BMSCs was eliminated by macrophage depletion or pretreatment with antibodies specific for interleukin-10 (IL-10) or IL-10 receptor. Monocytes and/or macrophages from septic lungs made more IL-10 when prepared from mice treated with BMSCs versus untreated mice. Lipopolysaccharide (LPS)-stimulated macrophages produced more IL-10 when cultured with BMSCs, but this effect was eliminated if the BMSCs lacked the genes encoding Toll-like receptor 4, myeloid differentiation primary response gene-88, tumor necrosis factor (TNF) receptor-1a or cyclooxygenase-2. Our results suggest that BMSCs (activated by LPS or TNF-alpha) reprogram macrophages by releasing prostaglandin E(2) that acts on the macrophages through the prostaglandin EP2 and EP4 receptors. Because BMSCs have been successfully given to humans and can easily be cultured and might be used without human leukocyte antigen matching, we suggest that cultured, banked human BMSCs may be effective in treating sepsis in high-risk patient groups.
Mesenchymal stem cells as trophic mediators.
Caplan Arnold I,Dennis James E
Journal of cellular biochemistry
Adult marrow-derived Mesenchymal Stem Cells (MSCs) are capable of dividing and their progeny are further capable of differentiating into one of several mesenchymal phenotypes such as osteoblasts, chondrocytes, myocytes, marrow stromal cells, tendon-ligament fibroblasts, and adipocytes. In addition, these MSCs secrete a variety of cytokines and growth factors that have both paracrine and autocrine activities. These secreted bioactive factors suppress the local immune system, inhibit fibrosis (scar formation) and apoptosis, enhance angiogenesis, and stimulate mitosis and differentiation of tissue-intrinsic reparative or stem cells. These effects, which are referred to as trophic effects, are distinct from the direct differentiation of MSCs into repair tissue. Several studies which tested the use of MSCs in models of infarct (injured heart), stroke (brain), or meniscus regeneration models are reviewed within the context of MSC-mediated trophic effects in tissue repair.
Therapeutic benefit of intravenous administration of bone marrow stromal cells after cerebral ischemia in rats.
Chen J,Li Y,Wang L,Zhang Z,Lu D,Lu M,Chopp M
BACKGROUND AND PURPOSE:We tested the hypothesis that intravenous infusion of bone marrow derived-marrow stromal cells (MSCs) enter the brain and reduce neurological functional deficits after stroke in rats. METHODS:Rats (n=32) were subjected to 2 hours of middle cerebral artery occlusion (MCAO). Test groups consisted of MCAO alone (group 1, n=6); intravenous infusion of 1x10(6) MSCs at 24 hours after MCAO (group 2, n=6); or infusion of 3x10(6) MSCs (group 3, n=7). Rats in groups 1 to 3 were euthanized at 14 days after MCAO. Group 4 consisted of MCAO alone (n=6) and group 5, intravenous infusion of 3x10(6) MSCs at 7 days after MCAO (n=7). Rats in groups 4 and 5 were euthanized at 35 days after MCAO. For cellular identification, MSCs were prelabeled with bromodeoxyuridine. Behavioral tests (rotarod, adhesive-removal, and modified Neurological Severity Score [NSS]) were performed before and at 1, 7, 14, 21, 28, and 35 days after MCAO. Immunohistochemistry was used to identify MSCs or cells derived from MSCs in brain and other organs. RESULTS Significant recovery of somatosensory behavior and Neurological Severity Score (P<0.05) were found in animals infused with 3x10(6) MSCs at 1 day or 7 days compared with control animals. MSCs survive and are localized to the ipsilateral ischemic hemisphere, and a few cells express protein marker phenotypic neural cells. CONCLUSIONS:MSCs delivered to ischemic brain tissue through an intravenous route provide therapeutic benefit after stroke. MSCs may provide a powerful autoplastic therapy for stroke.
Human umbilical cord mesenchymal stem cells transplantation promotes cutaneous wound healing of severe burned rats.
Liu Lingying,Yu Yonghui,Hou Yusen,Chai Jiake,Duan Hongjie,Chu Wanli,Zhang Haijun,Hu Quan,Du Jundong
BACKGROUND:Severe burns are a common and highly lethal trauma. The key step for severe burn therapy is to promote the wound healing as early as possible, and reports indicate that mesenchymal stem cell (MSC) therapy contributes to facilitate wound healing. In this study, we investigated effect of human umbilical cord MSCs (hUC-MSCs) could on wound healing in a rat model of severe burn and its potential mechanism. METHODS:Adult male Wistar rats were randomly divided into sham, burn, and burn transplanted hUC-MSCs. GFP labeled hUC-MSCs or PBS was intravenous injected into respective groups. The rate of wound closure was evaluated by Image Pro Plus. GFP-labeled hUC-MSCs were tracked by in vivo bioluminescence imaging (BLI), and human-specific DNA expression in wounds was detected by PCR. Inflammatory cells, neutrophils, macrophages, capillaries and collagen types I/III in wounds were evaluated by histochemical staining. Wound blood flow was evaluated by laser Doppler blood flow meter. The levels of proinflammatory and anti-inflammatory factors, VEGF, collagen types I/III in wounds were analyzed using an ELISA. RESULTS:We found that wound healing was significantly accelerated in the hUC-MSC therapy group. The hUC-MSCs migrated into wound and remarkably decreased the quantity of infiltrated inflammatory cells and levels of IL-1, IL-6, TNF-α and increased levels of IL-10 and TSG-6 in wounds. Additionally, the neovascularization and levels of VEGF in wounds in the hUC-MSC therapy group were markedly higher than those in other control groups. The ratio of collagen types I and III in the hUC-MSC therapy group were markedly higher than that in the burn group at indicated time after transplantation. CONCLUSION:The study suggests that hUC-MSCs transplantation can effectively improve wound healing in severe burned rat model. Moreover, these data might provide the theoretical foundation for the further clinical application of hUC-MSC in burn areas.
Human adipose-derived stem cell adipogenesis induces paracrine regulation of the invasive ability of MCF-7 human breast cancer cells
Zhao Yang,Gao Jianhua,Lu Feng
Experimental and therapeutic medicine
The aim of this study was to determine the effects of paracrine regulation on the invasive ability of MCF-7 human breast cancer cells through human adipose-derived stem cell (hADSC) adipogenesis. hADSC differentiation of the third and fourth passages of cells was induced in different induction media: osteogenic, adipogenic and chondrogenic. Transwell migration assays in the differently conditioned media, flow cytometry, enzyme-linked immunosorbent assay and western blot analysis for selected cytokines were performed. The flow cytometric analysis demonstrated positive expression of CD29, CD44 and CD105, while expression of CD34 and CD45 was not identified. The transwell migration assay showed that the invasive ability of MCF-7 cells was significantly enhanced during hADSC adipogenesis. hADSCs exerted a significantly positive effect on the invasive activity of MCF-7 cells during adipo-genesis. The results indicate that the high expression levels of activating protein 2 (aP2) in MCF-7 and adipocytes induced for 12 days may be associated with cell growth, invasion and metastasis. Peroxisome proliferator-activated receptor γ may be involved in fatty syntheses during adipogenic initiation and following adipogenic differentiation, possibly acting as a protection factor resulting in cell maturation and differentiation. This study also demonstrated that the expression of vascular endothelial growth factor was repressed by hADSCs, while that of matrix metalloproteinase-2 and urokinase-type plasminogen activator was increased to a significant level.
Anti-inflammatory and immunomodulatory mechanisms of mesenchymal stem cell transplantation in experimental traumatic brain injury.
Zhang Run,Liu Yi,Yan Ke,Chen Lei,Chen Xiang-Rong,Li Peng,Chen Fan-Fan,Jiang Xiao-Dan
Journal of neuroinflammation
BACKGROUND:Previous studies have shown beneficial effects of mesenchymal stem cell (MSC) transplantation in central nervous system (CNS) injuries, including traumatic brain injury (TBI). Potential repair mechanisms involve transdifferentiation to replace damaged neural cells and production of growth factors by MSCs. However, few studies have simultaneously focused on the effects of MSCs on immune cells and inflammation-associated cytokines in CNS injury, especially in an experimental TBI model. In this study, we investigated the anti-inflammatory and immunomodulatory properties of MSCs in TBI-induced neuroinflammation by systemic transplantation of MSCs into a rat TBI model. METHODS/RESULTS:MSCs were transplanted intravenously into rats 2 h after TBI. Modified neurologic severity score (mNSS) tests were performed to measure behavioral outcomes. The effect of MSC treatment on neuroinflammation was analyzed by immunohistochemical analysis of astrocytes, microglia/macrophages, neutrophils and T lymphocytes and by measuring cytokine levels [interleukin (IL)-1α, IL-1β, IL-4, IL-6, IL-10, IL-17, tumor necrosis factor-α, interferon-γ, RANTES, macrophage chemotactic protein-1, macrophage inflammatory protein 2 and transforming growth factor-β1] in brain homogenates. The immunosuppression-related factors TNF-α stimulated gene/protein 6 (TSG-6) and nuclear factor-κB (NF-κB) were examined by reverse transcription-polymerase chain reaction and Western blotting. Intravenous MSC transplantation after TBI was associated with a lower density of microglia/macrophages and peripheral infiltrating leukocytes at the injury site, reduced levels of proinflammatory cytokines and increased anti-inflammatory cytokines, possibly mediated by enhanced expression of TSG-6, which may suppress activation of the NF-κB signaling pathway. CONCLUSIONS:The results of this study suggest that MSCs have the ability to modulate inflammation-associated immune cells and cytokines in TBI-induced cerebral inflammatory responses. This study thus offers a new insight into the mechanisms responsible for the immunomodulatory effect of MSC transplantation, with implications for functional neurological recovery after TBI.
Interleukin-33 reduces neuronal damage and white matter injury via selective microglia M2 polarization after intracerebral hemorrhage in rats.
Chen Zhouqing,Xu Na,Dai Xuejiao,Zhao Chongshun,Wu Xin,Shankar Sandhya,Huang Huachen,Wang Zhong
Brain research bulletin
Interleukin-33 (IL-33) is closely related to the regulation of immunological cells, and its receptor ST2 is a member of the interleukin-1 (IL-1) receptor family. Inflammatory responses play critical roles in neuronal damage and white matter injury (WMI) post intracerebral hemorrhage (ICH). In this study, we tried to explore the role of IL-33 in neuronal damage and WMI after ICH and the underlying mechanisms. The in vivo ICH model was performed by autologous whole blood injection into the right basal ganglia in rats. Immunoblotting, immunofluorescence, brain water content measurement, FJB staining, and TUNEL staining were applied in this study. IL-33 expression was increased in whole brain tissues post-ICH, mainly rapidly increased in ipsilateral astrocyte and microglia, but stayed at a low level in neurons. Intracerebroventricular infusion of IL-33 after ICH attenuated short-term and long-term neurological deficits, WMI, neuronal degeneration, cell death and promoted the transformation of microglia phenotype from M1 to M2 in brain tissues after ICH. These results suggest that IL-33 reduces neuronal damage and WMI by promoting microglia M2 polarization after ICH, thereby improving the outcomes of neurological function.
IL-33, an interleukin-1-like cytokine that signals via the IL-1 receptor-related protein ST2 and induces T helper type 2-associated cytokines.
Schmitz Jochen,Owyang Alexander,Oldham Elizabeth,Song Yaoli,Murphy Erin,McClanahan Terril K,Zurawski Gerard,Moshrefi Mehrdad,Qin Jinzhong,Li Xiaoxia,Gorman Daniel M,Bazan J Fernando,Kastelein Robert A
Cytokines of the interleukin-1 (IL-1) family, such as IL-1 alpha/beta and IL-18, have important functions in host defense, immune regulation, and inflammation. Insight into their biological functions has led to novel therapeutic approaches to treat human inflammatory diseases. Within the IL-1 family, IL-1 alpha/beta, IL-1Ra, and IL-18 have been matched to their respective receptor complexes and have been shown to have distinct biological functions. The most prominent orphan IL-1 receptor is ST 2. This receptor has been described as a negative regulator of Toll-like receptor-IL-1 receptor signaling, but it also functions as an important effector molecule of T helper type 2 responses. We report a member of the IL-1 family, IL-33, which mediates its biological effects via IL-1 receptor ST 2, activates NF-kappaB and MAP kinases, and drives production of T(H)2-associated cytokines from in vitro polarized T(H)2 cells. In vivo, IL-33 induces the expression of IL-4, IL-5, and IL-13 and leads to severe pathological changes in mucosal organs.
IL-33 improves wound healing through enhanced M2 macrophage polarization in diabetic mice.
He Rongguo,Yin Hui,Yuan Baohong,Liu Tao,Luo Li,Huang Ping,Dai Liangcheng,Zeng Kang
IL-33 is a newly discovered member of the IL-1 family and has been identified as a potent inducer of Th2 type immunity. Emerging evidence imply that IL-33 may also act as an alarm to alert the immune system when released by epithelial barrier tissues during trauma or infection. In this study, we further investigate the potential efficacy of IL-33 on dermal wound healing in streptozotocin-induced diabetic mice. A full-thickness skin wound was generated on the back of diabetic mice and treated with IL-33 or vehicle topically. Our data showed that IL-33 delivery contributed to diabetic wound closure with wounds gaping narrower and exhibiting elevated re-epithelialization. IL-33 promoted the new extracellular matrix (ECM) deposition and angiogenesis formation, which indicates an important role of IL-33 on matrix synthesis and neovascularization. Meanwhile, IL-33 accelerated the development of M2 macrophages in wound sites in vivo, and amplified IL-13-induced polarization of bone marrow-derived macrophages toward a M2 phenotype in vitro. Furthermore, IL-33-amplified M2 macrophages augmented the proliferation of fibroblasts and ECM deposition. All together, these results strongly suggest manipulation of IL-33-mediated signal might be a potential therapeutic approach for diabetic skin wounds.
Mesenchymal Stem Cell Therapy in Intracerebral Haemorrhagic Stroke.
Bedini Gloria,Bersano Anna,Zanier Elisa R,Pischiutta Francesca,Parati Eugenio A
Current medicinal chemistry
BACKGROUND:Spontaneous intracerebral haemorrhage (ICH) is a relatively common fatal disease, with an overall global incidence estimated at 24.6 per 100,000 person- years. Given the high degree of morbidity and mortality associated with ICH, therapies that may have neuroprotective effects are of increasing interest to clinicians. In this last context, cell therapies offer the promise of improving the disease course which cannot be addressed adequately by existing treatments. OBJECTIVE:The aim of this review is to evaluate the protective effects and molecular mechanisms of mesenchymal stem cells (MSCs) on haemorrhagic brain following ICH. We also discuss possible emerging therapeutic approaches worth of further research. METHODS AND RESULTS:The available literature on the therapeutic potential of MSCs in ICH animal models clearly demonstrated that MSCs enhance the functional recovery and reduce the volume of the infarct size exerting anti-inflammatory and angiogenic properties. However, the quality of the original articles investigating the efficacy of stem cell therapies in ICH animal models is still poor and the lack of ICH clinical trial does not permit to reach any relevant conclusions. CONCLUSION:Further studies have to be implemented in order to achieve standardized methods of MSCs isolation, characterization and administration to improve ICH treatments with MSCs or MSC-derived products.
BM-MSC Transplantation Alleviates Intracerebral Hemorrhage-Induced Brain Injury, Promotes Astrocytes Vimentin Expression, and Enhances Astrocytes Antioxidation via the Cx43/Nrf2/HO-1 Axis.
Chen Xiao,Liang Huaibin,Xi Zhiyu,Yang Yong,Shan Huimin,Wang Baofeng,Zhong Zhihong,Xu Canxin,Yang Guo-Yuan,Sun Qingfang,Sun Yuhao,Bian Liuguan
Frontiers in cell and developmental biology
Intracerebral hemorrhage (ICH) is a particularly severe form of stroke, and reactive astrogliosis is a common response following injury to the central nervous system (CNS). Mesenchymal stem cells (MSCs) are reported to promote neurogenesis and alleviate the late side effects in injured brain regions. Gap junctions (Gjs) are abundant in the brain, where the richest connexin (Cx) is Cx43, most prominently expressed in astrocytes. Nuclear factor erythroid 2-related factor 2 (Nrf2) is an essential transcription factor regulating antioxidant reactions. Here, we aimed to explore whether bone marrow MSCs (BM-MSCs) could alleviate brain injury and protect astrocytes from apoptosis, by regulating Cx43 and Nrf2. We validated the effect of BM-MSC transplantation in an ICH model and and detected changes using immunofluorescence, as well as protein and mRNA expression of glial fibrillary acidic protein (GFAP), vimentin (VIM), Cx43, Nrf2, and heme oxygenase-1 (HO-1). Our results showed that BM-MSC transplantation attenuated brain injury after ICH and upregulated VIM expression and . Additionally, Cx43 upregulation and Nrf2 nuclear translocation were observed in astrocytes cocultured with BM-MSC. Knockdown of Cx43 by siRNA restrained Nrf2 nuclear translocation. Cx43 and Nrf2 had a connection as determined by immunofluorescence and coimmunoprecipitation. We demonstrated that astrocytes undergo astroglial-mesenchymal phenotype switching and have anti-apoptotic abilities after BM-MSC transplantation, where Cx43 upregulation triggers Nrf2 nuclear translocation and promotes its phase II enzyme expression. The Cx43/Nrf2 interaction of astrocytes after BM-MSC transplantation may provide an important therapeutic target in the management of ICH.
Intranasal administration of human MSC for ischemic brain injury in the mouse: in vitro and in vivo neuroregenerative functions.
Donega Vanessa,Nijboer Cora H,Braccioli Luca,Slaper-Cortenbach Ineke,Kavelaars Annemieke,van Bel Frank,Heijnen Cobi J
Intranasal treatment with C57BL/6 MSCs reduces lesion volume and improves motor and cognitive behavior in the neonatal hypoxic-ischemic (HI) mouse model. In this study, we investigated the potential of human MSCs (hMSCs) to treat HI brain injury in the neonatal mouse. Assessing the regenerative capacity of hMSCs is crucial for translation of our knowledge to the clinic. We determined the neuroregenerative potential of hMSCs in vitro and in vivo by intranasal administration 10 d post-HI in neonatal mice. HI was induced in P9 mouse pups. 1×10(6) or 2×10(6) hMSCs were administered intranasally 10 d post-HI. Motor behavior and lesion volume were measured 28 d post-HI. The in vitro capacity of hMSCs to induce differentiation of mouse neural stem cell (mNSC) was determined using a transwell co-culture differentiation assay. To determine which chemotactic factors may play a role in mediating migration of MSCs to the lesion, we performed a PCR array on 84 chemotactic factors 10 days following sham-operation, and at 10 and 17 days post-HI. Our results show that 2×10(6) hMSCs decrease lesion volume, improve motor behavior, and reduce scar formation and microglia activity. Moreover, we demonstrate that the differentiation assay reflects the neuroregenerative potential of hMSCs in vivo, as hMSCs induce mNSCs to differentiate into neurons in vitro. We also provide evidence that the chemotactic factor CXCL10 may play an important role in hMSC migration to the lesion site. This is suggested by our finding that CXCL10 is significantly upregulated at 10 days following HI, but not at 17 days after HI, a time when MSCs no longer reach the lesion when given intranasally. The results described in this work also tempt us to contemplate hMSCs not only as a potential treatment option for neonatal encephalopathy, but also for a plethora of degenerative and traumatic injuries of the nervous system.
Bone marrow mesenchymal stem cells transplantation alleviates brain injury after intracerebral hemorrhage in mice through the Hippo signaling pathway.
Chen Xiao,Xu Can-Xin,Liang Huaibin,Xi Zhiyu,Pan Jiaji,Yang Yong,Sun Qingfang,Yang Guoyuan,Sun Yuhao,Bian Liuguan
Intracerebral hemorrhage (ICH) is a common acute nervous system disease with high mortality and severe disability. Mesenchymal stem cells (MSCs) have been reported to promote neurogenesis and to alleviate side effects in areas of brain injury areas. The Hippo pathway regulates diverse cellular processes, including cell survival, proliferation, differentiation, and organ size. Here, we found that transplantation of bone marrow MSCs (BM-MSCs) into the brains of mice could alleviate ICH-mediated injury and protect astrocytes from apoptosis by regulating mammalian sterile 20-like kinase (MST)1 and Yes-associated protein (YAP). Knocking down of MST1 by si-RNA triggered YAP nuclear translocation. We further demonstrated that astrocytes undergo astroglial-mesenchymal phenotype switching and become capable of proliferating after BM-MSC transplantation via the Hippo signaling pathway. Together, our identification of the Hippo pathway in mediating the beneficial effects of BM-MSCs may provide a novel therapeutic target in the treatment and management of ICH.
Differentiation and neurological benefit of the mesenchymal stem cells transplanted into the rat brain following intracerebral hemorrhage.
Zhang Huabiao,Huang Zhiyong,Xu Yuming,Zhang Suming
Spontaneous intracerebral hemorrhage (ICH) is often a fatal event. In a patient who survives the initial ictus, the resulting hematoma within brain parenchyma can trigger a series of events that lead to secondary insults and severe neurological deficits. Great efforts have been focused on searching for new approaches to help patients recover neurological function after ICH. Previous studies indicate that mesenchymal stem cells (MSCs) grafted into the ischemic rat brain can improve neurological function. However, there is no report regarding whether MSCs can be used in the same way to improve the neurological function after ICH. We generated the ICH model by injecting collagenase VII into rat brain. Subsequently, 5-bromo-2-deoxyuridine (BrdU)-labeled mesenchymal stem cells were delivered into the brain through carotid artery, cervical vein or lateral ventricle. The distribution and differentiation of MSCs were investigated by methods of immunohistochemistry. We found that MSCs were able to differentiate into neural cells in vitro as well as in the rat brain after ICH. The injected MSCs were able to migrate into hippocampus, blooding foci and ipsilateral cortex. In the hippocampus, MSCs differentiated into neurons; but in surrounding bleeding foci, they differentiated into neurons and astrocytes. In the ipsilateral cortex, MSCs differentiated into neurons, astrocytes and oligodendrocytes. Notably, the motor function of the rats in the carotid artery (CA) group and the lateral ventricle (LV) group improved significantly. Collectively, our study indicates that MSCs are able to differentiate into neural cells in the rat brain after ICH and can significantly improve motor function.
Molecular targeting of malignant glioma cells with an EphA2-specific immunotoxin delivered by human bone marrow-derived mesenchymal stem cells.
Sun Xin-Lin,Xu Zhi-Min,Ke Yi-Quan,Hu Chang-Chen,Wang Shi-Yong,Ling Geng-Qiang,Yan Zhong-Jie,Liu Yi-Jing,Song Zhen-Hua,Jiang Xiao-Dan,Xu Ru-Xiang
Immunotoxins have shown great promise as an alternative treatment for brain malignancies such as gliomas, but their failure to penetrate into the tumor mass remains a major problem. Mesenchymal stem cells exhibit tropism to tumor tissue and may serve as a cellular vehicle for the delivery and local production of antitumor agents. In this study, we used human bone marrow-derived mesenchymal stem cells (hMSCs) as a vehicle for the targeted delivery of EphrinA1-PE38, a very specific immunotoxin against the EphA2 receptor that is overexpressed in gliomas. hMSCs were transduced with adenovirus to express secretable EphrinA1-PE38. Our invitro assays confirmed the expression, release and selective killing effect of the immunotoxin produced by hMSCs. Furthermore, the intratumoral injection of engineered hMSCs was effective at inhibiting tumor growth in a malignant glioma tumor model. These results indicate that gene therapy utilizing EphrinA1-PE38-secreting hMSCs may provide a novel approach for the local treatment of malignant gliomas.
Targeted intracerebral delivery of the anti-inflammatory cytokine IL13 promotes alternative activation of both microglia and macrophages after stroke.
Hamzei Taj Somayyeh,Le Blon Debbie,Hoornaert Chloé,Daans Jasmijn,Quarta Alessandra,Praet Jelle,Van der Linden Annemie,Ponsaerts Peter,Hoehn Mathias
Journal of neuroinflammation
BACKGROUND:Subtle adjustment of the activation status of CNS resident microglia and peripheral macrophages, to promote their neuroprotective and neuroregenerative functions, may facilitate research towards curing neurodegenerative disorders. In the present study, we investigated whether targeted intracerebral delivery of the anti-inflammatory cytokine interleukin (IL)13, by means of transplanting IL13-expressing mesenchymal stem cells (IL13-MSCs), can promote a phenotypic switch in both microglia and macrophages during the pro-inflammatory phase in a mouse model of ischemic stroke. METHODS:We used the CXCR1 CCR2 transgenic mouse model to separately recognize brain-resident microglia from infiltrated macrophages. Quantitative immunohistochemical analyses were applied to characterize polarization phenotypes of both cell types. RESULTS:Distinct behaviors of both cell populations were noted dependent on the anatomical site of the lesion. Immunohistochemistry revealed that mice grafted with IL13-MSCs, in contrast to non-grafted and MSC-grafted control mice, were able to drive recruited microglia and macrophages into an alternative activation state, as visualized by a significant increase of Arg-1 and a noticeable decrease of MHC-II expression at day 14 after ischemic stroke. Interestingly, both Arg-1 and MHC-II were expressed more abundantly in macrophages than in microglia, further confirming the distinct behavior of both cell populations. CONCLUSIONS:The current data highlight the importance of controlled and localized delivery of the anti-inflammatory cytokine IL13 for modulation of both microglia and macrophage responses after ischemic stroke, thereby providing pre-clinical rationale for the application of L13-MSCs in future investigations of neurodegenerative disorders.
Activated Mesenchymal Stem Cells Induce Recovery Following Stroke Via Regulation of Inflammation and Oligodendrogenesis.
Tobin Matthew K,Stephen Terilyn K L,Lopez Kyra L,Pergande Melissa R,Bartholomew Amelia M,Cologna Stephanie M,Lazarov Orly
Journal of the American Heart Association
Background Brain repair mechanisms fail to promote recovery after stroke, and approaches to induce brain regeneration are scarce. Mesenchymal stem cells (MSC) are thought to be a promising therapeutic option. However, their efficacy is not fully elucidated, and the mechanism underlying their effect is not known. Methods and Results The middle cerebral artery occlusion model was utilized to determine the efficacy of interferon-γ-activated mesenchymal stem cells (aMSCγ) as an acute therapy for stroke. Here we show that treatment with aMSCγ is a more potent therapy for stroke than naive MSC. aMSCγ treatment results in significant functional recovery assessed by the modified neurological severity score and open-field analysis compared with vehicle-treated animals. aMSCγ-treated animals showed significant reductions in infarct size and inhibition of microglial activation. The aMSCγ treatment suppressed the hypoxia-induced microglial proinflammatory phenotype more effectively than treatment with naive MSC. Importantly, treatment with aMSCγ induced recruitment and differentiation of oligodendrocyte progenitor cells to myelin-producing oligodendrocytes in vivo. To elucidate the mechanism underlying high efficacy of aMSCγ therapy, we examined the secretome of aMSCγ and compared it to that of naive MSC. Intriguingly, we found that aMSCγ but not nMSC upregulated neuron-glia antigen 2, an important extracellular signal and a hallmark protein of oligodendrocyte progenitor cells. Conclusions These results suggest that activation of MSC with interferon-γ induces a potent proregenerative, promyelinating, and anti-inflammatory phenotype of these cells, which increases the potency of aMSCγ as an effective therapy for ischemic stroke.
Intravenous administration of human adipose-derived stem cells ameliorates motor and cognitive function for intracerebral hemorrhage mouse model.
Kuramoto Yoji,Takagi Toshinori,Tatebayashi Kotaro,Beppu Mikiya,Doe Nobutaka,Fujita Mitsugu,Yoshimura Shinichi
Even today, intracerebral hemorrhage (ICH) is a major cause of death and disabilities. Rehabilitation is preferentially applied for functional recovery although its effect is limited. Recent studies have suggested that intravenous administration of mesenchymal stem cells would improve the post-ICH neurological deficits. Human adipose-derived stem cells (hADSCs) have been established in our laboratory. We aimed to evaluate the therapeutic efficacy of the hADSCs on the post-ICH neurological deficits using a clinical-relevant ICH mouse model. We also evaluated immune responses to clarify the underlying mechanisms. The hADSCs expressed MSC markers at high levels. The hADSCs administration into the ICH-bearing mice improved the neurological deficits during the subacute phases, which was shown by neurobehavioral experiments. Besides, the hADSC administration decreased the number of CD11CD45 cells and increased the proportion of CD86 and Ly6C cells in the ICH lesions. In summary, intravenous administration of hADSCs during the acute phase improved ICH-induced neurological deficits during the subacute phase because of the suppression of acute inflammation mediated by CD11CD45 subpopulations. Our data suggest that hADSCs can be served as a novel strategy for ICH treatment.
Bone marrow-derived mesenchymal stem cells maintain the resting phenotype of microglia and inhibit microglial activation.
Yan Ke,Zhang Run,Sun Chengmei,Chen Lei,Li Peng,Liu Yi,Peng Lingmei,Sun Haitao,Qin Kun,Chen Fanfan,Huang Weiyi,Chen Yuxin,Lv Bingke,Du Mouxuan,Zou Yuxi,Cai Yingqian,Qin Lingsha,Tang Yanping,Jiang Xiaodan
Many studies have shown that microglia in the activated state may be neurotoxic. It has been proven that uncontrolled or over-activated microglia play an important role in many neurodegenerative disorders. Bone marrow-derived mesenchymal stem cells (BMSCs) have been shown in many animal models to have a therapeutic effect on neural damage. Such a therapeutic effect is attributed to the fact that BMSCs have the ability to differentiate into neurons and to produce trophic factors, but there is little information available in the literature concerning whether BMSCs play a therapeutic role by affecting microglial activity. In this study, we triggered an inflammatory response situation in vitro by stimulating microglia with the bacterial endotoxin lipopolysaccharide (LPS), and then culturing these microglia with BMSC-conditioned medium (BMSC-CM). We found that BMSC-CM significantly inhibited proliferation and secretion of pro-inflammatory factors by activated microglia. Furthermore, we found that the phagocytic capacity of microglia was also inhibited by BMSC-CM. Finally, we investigated whether the induction of apoptosis and the production of nitric oxide (NO) were involved in the inhibition of microglial activation. We found that BMSC-CM significantly induced apoptosis of microglia, while no apoptosis was apparent in the LPS-stimulated microglia. Our study also provides evidence that NO participates in the inhibitory effect of BMSCs. Our experimental results provide evidence that BMSCs have the ability to maintain the resting phenotype of microglia or to control microglial activation through their production of several factors, indicating that BMSCs could be a promising therapeutic tool for treatment of diseases associated with microglial activation.
Transplantation of hypoxia preconditioned bone marrow mesenchymal stem cells enhances angiogenesis and neurogenesis after cerebral ischemia in rats.
Wei Ling,Fraser Jamie L,Lu Zhong-Yang,Hu Xinyang,Yu Shan Ping
Neurobiology of disease
Hypoxic preconditioning of stem cells and neural progenitor cells has been tested for promoting cell survival after transplantation. The present investigation examined the hypothesis that hypoxic preconditioning of bone marrow mesenchymal stem cells (BMSCs) could not only enhance their survival but also reinforce regenerative properties of these cells. BMSCs from eGFP engineered rats or pre-labeled with BrdU were pre-treated with normoxia (20% O(2), N-BMSCs) or sub-lethal hypoxia (0.5% O(2). H-BMSCs). The hypoxia exposure up-regulated HIF-1α and trophic/growth factors in BMSCs, including brain-derived neurotrophic factor (BDNF), glial cell-derived neurotrophic factor (GDNF), vascular endothelial growth factor (VEGF) and its receptor FIK-1, erythropoietin (EPO) and its receptor EPOR, stromal derived factor-1 (SDF-1) and its CXC chemokine receptor 4 (CXCR4). Meanwhile, many pro-inflammatory cytokines/chemokines were down-regulated in H-BMSCs. N-BMSCs or H-BMSCs were intravenously injected into adult rats 24h after 90-min middle cerebral artery occlusion. Comparing to N-BMSCs, transplantation of H-BMSCs showed greater effect of suppressing microglia activity in the brain. Significantly more NeuN-positive and Glut1-positive cells were seen in the ischemic core and peri-infarct regions of the animals received H-BMSC transplantation than that received N-BMSCs. Some NeuN-positive and Glut-1-positive cells showed eGFP or BrdU immunoflourescent reactivity, suggesting differentiation from exogenous BMSCs into neuronal and vascular endothelial cells. In Rotarod test performed 15days after stroke, animals received H-BMSCs showed better locomotion recovery compared with stroke control and N-BMSC groups. We suggest that hypoxic preconditioning of transplanted cells is an effective means of promoting their regenerative capability and therapeutic potential for the treatment of ischemic stroke.
Human Placenta-Derived Mesenchymal Stem Cells Reduce Mortality and Hematoma Size in a Rat Intracerebral Hemorrhage Model in an Acute Phase.
Choi Bo Young,Kim Ok Joon,Min Sae-Hong,Jeong Jeong Hyun,Suh Sang Won,Chung Tae Nyoung
Stem cells international
Intracerebral hemorrhage (ICH) is a critical disease, highly associated with mortality and morbidity. Several studies have demonstrated the beneficial effect of mesenchymal stem cells (MSCs) on ICH, mostly focused on their mid-to-long-term effect. Acute hematoma expansion is one of the most important prognostic factors of ICH. We hypothesized that MSCs would decrease mortality and hematoma size in acute ICH, based on the findings of a few recent researches reporting their effect on blood-brain barrier and endothelial integrity. Rat ICH models were made using bacterial collagenase. One hour after ICH induction, the rats were randomly divided into MSC-treated and control groups. Mortality, hematoma volume, ventricular enlargement, brain edema, and degenerating neuron count were compared at 24 hours after ICH induction. Expression of tight junction proteins (ZO-1, occludin) and coagulation factor VII mRNA was also compared. Mortality rate (50% versus 8.3%), hematoma size, ventricular size, hemispheric enlargement, and degenerating neuron count were significantly lower in the MSC-treated group ( = 0.034, 0.038, 0.001, 0.022, and <0.001, resp.), while the expression of ZO-1 and occludin was higher ( = 0.007 and 0.012). Administration of MSCs may prevent hematoma expansion in the hyperacute stage of ICH and decrease acute mortality by enhancing the endothelial integrity of cerebral vasculature.
Protracted Morphological Changes in the Corticospinal Tract Within the Cervical Spinal Cord After Intracerebral Hemorrhage in the Right Striatum of Mice.
Ng Anson Cho Kiu,Yao Min,Cheng Stephen Yin,Li Jing,Huang Jian-Dong,Wu Wutian,Leung Gilberto Ka Kit,Sun Haitao
Frontiers in neuroscience
Intracerebral hemorrhage (ICH) is associated with high morbidity and mortality rates. Currently, there is no promising treatment that improves prognosis significantly. While a thorough investigation of the pathological process within the primary site of injury in the brain has been conducted by the research field, the focus was mainly on gray matter injury, which partly accounted for the failure of discovery of clinically efficacious treatments. It is not until recent years that white matter (WM) injury in the brain after subcortical ICH was examined. As WM tracts form networks between different regions, damage to fibers should impair brain connectivity, resulting in functional impairment. Although WM changes have been demonstrated in the brain after ICH, alterations distant from the initial injury site down in the spinal cord are unclear. This longitudinal study, for the first time, revealed prolonged morphological changes of the contralesional dorsal corticospinal tract (CST) in the spinal cord 5 weeks after experimental ICH in mice by confocal microscopy and transmission electron microscopy, implying that the structural integrity of the CST was compromised extensively after ICH. Given the important role of CST in motor function, future translational studies targeting motor recovery should delineate the treatment effects on CST integrity.
Modified behavioural tests to detect white matter injury- induced motor deficits after intracerebral haemorrhage in mice.
Chen Weixiang,Xia Min,Guo Chao,Jia Zhengcai,Wang Jie,Li Chengcheng,Li Mingxi,Tang Xiaoqin,Hu Rong,Chen Yujie,Liu Xin,Feng Hua
Motor function deficit induced by white matter injury (WMI) is one of the most severe complications of intracerebral haemorrhage (ICH). The degree of WMI is closely related to the prognosis of patients after ICH. However, the current behavioural assessment of motor function used in the ICH mouse model is mainly based on that for ischaemic stroke and lacks the behavioural methods that accurately respond to WMI. Here, a series of easy-to-implement behavioural tests were performed to detect motor deficits in mice after ICH. The results showed that the grip strength test and the modified pole test not only can better distinguish the degree of motor dysfunction between different volumes of blood ICH models than the Basso Mouse Scale and the beam walking test but can also accurately reflect the severity of WMI characterized by demyelination, axonal swelling and the latency of motor-evoked potential delay induced by ICH. In addition, after ICH, the results of grip tests and modified pole tests, rather than the Basso Mouse Scale and the beam walking test, were worse than those observed after intraventricular haemorrhage (IVH), which was used as a model of brain haemorrhage in non-white matter areas. These results indicate that the grip strength test and the modified pole test have advantages in detecting the degree of motor deficit induced by white matter injury after ICH in mice.
Long term depression in the CA1 field is associated with a transient decrease in pre- and postsynaptic PKC substrate phosphorylation.
Ramakers G M,Heinen K,Gispen W H,de Graan P N
The Journal of biological chemistry
Induction of homosynaptic long term depression (LTD) in the CA1 field of the hippocampus is thought to require activation of N-methyl-d-aspartate receptors, an elevation of postsynaptic Ca(2+) levels, and a subsequent increase in phosphatase activity. To investigate the spatial and temporal changes in protein phosphatase activity following LTD induction, we determined the in situ phosphorylation state of a pre- (GAP-43/B-50) and postsynaptic (RC3) protein kinase C substrate during N-methyl-d-aspartate receptor-dependent LTD in the CA1 field of rat hippocampal slices. We show that LTD is associated with a transient (<30 min) and D-AP5-sensitive reduction in GAP-43/B-50 and RC3 phosphorylation and that LTD is prevented by the phosphatase inhibitors okadaic acid and cyclosporin A. Our data provide strong evidence for a transient increase in pre- and postsynaptic phosphatase activity during LTD. Since the in situ phosphorylation of the calmodulin-binding proteins GAP-43/B-50 and RC3 changes during both LTD and long term potentiation, these proteins may form part of the link between the Ca(2+) signal and Ca(2+)/calmodulin-dependent processes implicated in long term potentiation and LTD.
Synaptic localization of growth-associated protein 43 in cultured hippocampal neurons during synaptogenesis.
Morita Shoko,Miyata Seiji
Cell biochemistry and function
Growth-associated protein 43 (GAP-43), a novel axonal phosphoprotein, is originally identified as a growth-cone-specific protein of developing neurons in vitro. The expression of GAP-43 is also shown to be up-regulated concomitant with increased synaptic plasticity in the brains in vivo, but how GAP-43 is concerned with synaptic plasticity is not well understood. In the present study, therefore, we aimed to elucidate subcellular localization of GAP-43 as culture development of rat hippocampal neurons. Western blotting showed that the expression of GAP-43 in the cerebral and hippocampal tissues was prominently high at postnatal days 14 and 21 or the active period of synaptogenesis. Double-labelling immunohistochemistry with an axonal marker Tau revealed that the immunoreactivity of GAP-43 was seen throughout axons of cultured hippocampal neurons but stronger at axonal puncta of developing neurons than axonal processes. Double-labelling immunohistochemistry with presynaptic terminal markers of synapsin and synaptotagmin revealed that the immunoreactivity of GAP-43 was observed mostly at weak synapsin- and synaptotagmin-positive puncta rather than strong ones. The quantitative analysis of immunofluorescent intensity showed a clear inverse correlation between GAP-43 and either synapsin or synaptotagmin expression. These data indicate that GAP-43 is highly expressed at immature growing axonal terminals and its expression is decreased along with the maturation of synaptogenesis.
GAP-43 is essential for the neurotrophic effects of BDNF and positive AMPA receptor modulator S18986.
Gupta S K,Mishra R,Kusum S,Spedding M,Meiri K F,Gressens P,Mani S
Cell death and differentiation
Positive alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) receptor modulators include benzamide compounds that allosterically modulate AMPA glutamate receptors. These small molecules that cross the blood-brain barrier have been shown to act as a neuroprotectant by increasing the levels of endogenous brain-derived neurotrophic factor (BDNF). Positive AMPA receptor modulators have also been shown to increase the levels of growth-associated protein-43 (GAP-43). GAP-43 plays a major role in many aspects of neuronal function in vertebrates. The goal of this study was to determine whether GAP-43 was important in mediating the actions of positive AMPA receptor modulator (S18986) and BDNF. Using cortical cultures from GAP-43 knockout and control mice, we show that (1) GAP-43 is upregulated in response to S18986 and BDNF in control cultures; (2) this upregulation of GAP-43 is essential for mediating the neuroprotective effects of S18986 and BDNF; (3) administration of S18986 and BDNF leads to an increase in the expression of the glutamate transporters GLT-1 and GLAST that are key to limiting excitotoxic cell death and this increase in GLT-1 and GLAST expression is completely blocked in the absence of GAP-43. Taken together this study concludes that GAP-43 is an important mediator of the neurotrophic effects of S18986 and BDNF on neuronal survival and plasticity, and is essential for the success of positive AMPA receptor modulator-BDNF-based neurotrophin therapy.
Electroacupuncture stimulates the proliferation and differentiation of endogenous neural stem cells in a rat model of ischemic stroke.
Tan Feng,Wang Jian,Liu Jing Xian,Wang Chen,Li Miaodan,Gu Yong
Experimental and therapeutic medicine
Electroacupuncture (EA) may stimulate neurogenesis in animal models of ischemic stroke; however, the associated mechanisms are not clear. The present study aimed to evaluate the neurogenesis efficacy of EA on ischemic stroke and the underlying associated mechanisms. A model of middle cerebral artery occlusion (MCAO) was employed as the rat model of brain ischemia and reperfusion. EA treatment at the GV20 (Baihui) and GV14 (Dazhui) acupoints was conducted for 30 min daily following MCAO. Immunofluorescence was performed to measure the number of bromodeoxyuridine (BrdU)/nestin- or BrdU/doublecortin (DCX)-positive cells in the sham, MCAO and MCAO + EA groups. Results indicated that EA stimulation significantly decreased the neurological score and neuronal loss in rats in the MCAO group (both P<0.05). Furthermore, immunostaining assays indicated that BrdU/nestin- and BrdU/DCX-positive cells in EA-treated rats were significantly increased (P<0.05) when compared with the rats in the MCAO group, indicating EA may induce the proliferation and differentiation of endogenous neural stem cells (eNSCs) during cerebral ischemia-reperfusion. In addition, EA treatment significantly enhanced the protein expression levels of plasticity-related gene 5 (PRG5), a critical neurogenesis factor, and significantly decreased the protein expression levels of three neurogenesis inhibiting molecules, NogoA, lysophosphatidic acid and RhoA (all P<0.05). These results suggested that EA promotes the proliferation and differentiation of eNSCs, likely through modulating PRG5/RhoA signaling.
Electroacupuncture ameliorates memory impairments by enhancing oligodendrocyte regeneration in a mouse model of prolonged cerebral hypoperfusion.
Ahn Sung Min,Kim Yu Ri,Kim Ha Neui,Shin Yong-Il,Shin Hwa Kyoung,Choi Byung Tae
We modeled prolonged cerebral hypoperfusion in mice using bilateral common carotid artery stenosis (BCAS) and electroacupuncture (EA) stimulation was applied at two acupoints, Baihui (GV20) and Dazhui (GV14). In behavioral tests of memory, BCAS produced impairments in spatial and short-term memory in mice that were attenuated by therapeutic EA stimulation. Therapeutic use of EA in BCAS also enhanced oligodendrocyte (OL) differentiation from oligodendrocyte precursor cells (OPCs), in association with white matter improvements in the corpus callosum (CC). In PCR analyses of growth factor gene expression, significant positive changes in 3 genes were observed following EA stimulation in BCAS, and here we highlight alterations in neurotrophin-4/5 (NT4/5). We confirmed EA-mediated positive changes in the expression of NT4/5 and its receptor, tyrosine receptor kinase B (TrkB). Treatment of naïve and BCAS + EA animals with a selective TrkB antagonist, ANA-12, produced losses of myelin and cognitive function that were ameliorated by EA therapy. Moreover, following BCAS we observed an EA-dependent increase in phospho-activated CREB (a downstream mediator of NT4/5-TrkB signaling) in OPCs and OLs of the CC. Our results suggest that EA stimulation promotes the recovery of memory function following white matter injury via a mechanism that promotes oligodendrocyte regeneration and involves NT4/5-TrkB signaling.
Therapeutic Potential of a Combination of Electroacupuncture and TrkB-Expressing Mesenchymal Stem Cells for Ischemic Stroke.
Ahn Sung Min,Kim Yu Ri,Shin Yong-Il,Ha Ki Tae,Lee Seo-Yeon,Shin Hwa Kyoung,Choi Byung Tae
We prepared and grafted tropomyosin receptor kinase B (TrkB) gene-transfected mesenchymal stem cells (TrkB-MSCs) into the ischemic penumbra and investigated whether electroacupuncture (EA) treatment could promote functional recovery from ischemic stroke. For the behavioral test, TrkB-MSCs+EA resulted in significantly improved motor function compared to that obtained with MSCs+EA or TrkB-MSCs alone. At 30 days after middle cerebral artery occlusion (MCAO), the largest number of grafted MSCs was detected in the TrkB-MSC+EA group. Some differentiation into immature neuroblasts and astrocytes was detected; however, only a few mature neuron-like cells were found. Compared to other treatments, TrkB-MSCs+EA upregulated the expression of mature brain-derived neurotrophic factor (BDNF) and neurotrophin-4/5 (NT4) and induced the activation of TrkB receptor and its transcription factor cAMP response element-binding protein (CREB). At 60 days after MCAO, EA highly promoted the differentiation of TrkB-MSCs into mature neuron-like cells compared to the effect in MSCs. A selective TrkB antagonist, ANA-12, reverted the effect of TrkB-MSCs+EA in motor function recovery and survival of grafted MSCs. Our results suggest that EA combined with grafted TrkB-MSCs promotes the expression of BDNF and NT4, induces the differentiation of TrkB-MSCs, and improves motor function. TrkB-MSCs could serve as effective therapeutic agents for ischemic stroke if used in combination with BDNF/NT4-inducing therapeutic approaches.
Concise review: hitting the right spot with mesenchymal stromal cells.
Tolar Jakub,Le Blanc Katarina,Keating Armand,Blazar Bruce R
Stem cells (Dayton, Ohio)
Mesenchymal stromal cells or mesenchymal stem cells (MSCs) have captured considerable scientific and public interest because of their potential to limit physical and immune injury, to produce bioactive molecules and to regenerate tissues. MSCs are phenotypically heterogeneous and distinct subpopulations within MSC cultures are presumed to contribute to tissue repair and the modulation of allogeneic immune responses. As the first example of efficacy, clinical trials for prevention and treatment of graft-versus-host disease after hematopoietic cell transplantation show that MSCs can effectively treat human disease. The view of the mechanisms whereby MSCs function as immunomodulatory and reparative cells has evolved simultaneously. Initially, donor MSCs were thought to replace damaged cells in injured tissues of the recipient. More recently, however, it has become increasingly clear that even transient MSC engraftment may exert favorable effects through the secretion of cytokines and other paracrine factors, which engage and recruit recipient cells in productive tissue repair. Thus, an important reason to investigate MSCs in mechanistic preclinical models and in clinical trials with well-defined end points and controls is to better understand the therapeutic potential of these multifunctional cells. Here, we review the controversies and recent insights into MSC biology, the regulation of alloresponses by MSCs in preclinical models, as well as clinical experience with MSC infusions (Table 1) and the challenges of manufacturing a ready supply of highly defined transplantable MSCs.
Comparative study of regenerative effects of mesenchymal stem cells derived from placental amnion, chorion and umbilical cord on dermal wounds.
Ertl Juliane,Pichlsberger Melanie,Tuca Alexandru-Cristian,Wurzer Paul,Fuchs Jakob,Geyer Stefan H,Maurer-Gesek Barbara,Weninger Wolfgang J,Pfeiffer Dagmar,Bubalo Vladimir,Parvizi Daryousch,Kamolz Lars-Peter,Lang Ingrid
OBJECTIVE:Mesenchymal stem/stromal cells derived from human term placentas (PMSCs) are novel therapeutic agents and more topical than ever. Here we evaluated the effects of three types of PMSCs on wound healing in an in vivo mouse model: Amnion-derived MSCs (AMSCs), blood vessel-derived MSCs (BV-MSCs) from the chorionic plate and Wharton's jelly-derived MSCs (WJ-MSCs) from the umbilical cord. METHODS:We topically applied PMSCs onto skin wounds in mice using the dermal substitute Matriderm as carrier and evaluated wound healing parameters. In addition, we investigated the effects of all PMSC types under co-application with placental endothelial cells (PLECs). After 8 days, we compared the percent of wound closure and the angiogenic potential between all groups. RESULTS:AMSCs, BV-MSCs and WJ-MSCs significantly induced a faster healing and a higher number of blood vessels in the wound when compared to controls (Matriderm-alone). PLECs did not further improve the advantageous effects of PMSC-treatment. Quantitative data and 3D analysis by high resolution episcopic microscopy confirmed a lower density of vessels in Matriderm/PMSCs/PLECs co-application compared to Matriderm/PMSCs treatment. CONCLUSION:Results indicate that all three PMSC types exert similar beneficial effects on wound closure and neovascularization in our mouse model. PRACTICE:Using Matriderm as carrier for PMSCs propagates rapid cell migration towards the wound area that allows a fast and clinically practicable method for stem cell application. IMPLICATIONS:These promising effects warrant further investigation in clinical trials.
Comparison of Matrigel and Matriderm as a carrier for human amnion-derived mesenchymal stem cells in wound healing.
Tuca Alexandru-Cristian,Ertl Juliane,Hingerl Kerstin,Pichlsberger Melanie,Fuchs Jakob,Wurzer Paul,Pfeiffer Dagmar,Bubalo Vladimir,Parvizi Daryousch,Kamolz Lars-Peter,Lang Ingrid
Amnion-derived mesenchymal stem cells (AMSC) are a promising tool in regenerative medicine. Here we evaluated the utility of Matrigel and Matriderm as carrier for the topical application of AMSC to mice skin wounds. In both application forms, AMSC promoted neovascularization of the wound area. Matrigel proved as excellent matrix for AMSC and immigrating mouse cells, but the solid Matriderm enabled a more adequate positioning of AMSC into the wound. Although AMSC did not attach to Matriderm, they reliably induced wound reduction. Thus, a combined administration of AMSC/Matriderm could be beneficial to potentiate the encouraging effects on wound healing.
Mesenchymal stromal cells from the human placenta promote neovascularization in a mouse model in vivo.
Kinzer M,Hingerl K,König J,Reinisch A,Strunk D,Huppertz B,Lang I
Cell transplantation is a promising strategy in regenerative medicine for revascularization of ischemic tissues. Based on our observation that placental mesenchymal stromal cells (PMSC) enhance endothelial cell viability in vitro via secretion of angiogenic factors, we asked whether PMSC support vascular growth in vivo. PMSC were isolated from amnion and placental endothelial cells (PLEC) from chorion and either separately or co-transplanted subcutaneously into immune-deficient mice. Co-transplantation resulted in a higher number of perfused human vessels (CD31+/vimentin+) containing mouse glycophorin A+ erythrocytes. Results indicate positive effects of PMSC on neovascularization in vivo, making them attractive candidates to create autologous PMSC/PLEC pairs for research and transplantation.
Placental mesenchymal stromal cells derived from blood vessels or avascular tissues: what is the better choice to support endothelial cell function?
König Julia,Weiss Gregor,Rossi Daniele,Wankhammer Karin,Reinisch Andreas,Kinzer Manuela,Huppertz Berthold,Pfeiffer Dagmar,Parolini Ornella,Lang Ingrid
Stem cells and development
Mesenchymal stromal cells (MSCs) are promising tools for therapeutic revascularization of ischemic tissues and for support of vessel formation in engineered tissue constructs. Recently, we could show that avascular-derived MSCs from placental amnion release soluble factors that exhibit survival-enhancing effects on endothelial cells (ECs). We hypothesize that MSCs derived from placental blood vessels might have even more potent angiogenic effects. Therefore, we isolated and characterized MSCs from placental chorionic blood vessels (bv-MSCs) and tested their angiogenic potential in comparison to amnion-derived avascular MSCs (av-MSCs). bv-MSCs express a very similar surface marker profile compared with av-MSCs and could be differentiated toward the adipogenic and osteogenic lineages. bv-MSCs exert immunosuppressive properties on peripheral blood mononuclear cells, suggesting that they are suitable for cell transplantation settings. Conditioned medium (Cdm) from av-MSCs and bv-MSCs significantly enhanced EC viability, whereas only Cdm from bv-MSCs significantly increased EC migration and network formation (Matrigel assay). Angiogenesis array analysis of av- and bv-MSC-Cdm revealed a similar secretion pattern of angiogenic factors, including angiogenin, interleukins-6 and -8, and tissue inhibitors of matrix metalloproteinase-1 and 2. Enzyme-linked immunosorbent assay analysis showed that, in contrast to av-MSCs, bv-MSCs secreted vascular endothelial growth factor. In direct coculture with bv-MSCs, ECs showed a significantly increased formation of vessel-like structures compared with av-MSCs. With regard to therapeutic treatment, bv-MSCs and particularly their Cdm might be valuable to stimulate angiogenesis especially in ischemic tissues. av-MSCs and their Cdm could be beneficial in conditions when it is required to promote the survival and stabilization of blood vessels without the risk of unmeant angiogenesis.
Amnion-derived mesenchymal stem cells improve viability of endothelial cells exposed to shear stress in ePTFE grafts.
Pfeiffer Dagmar,Wankhammer Karin,Stefanitsch Christina,Hingerl Kerstin,Huppertz Berthold,Dohr Gottfried,Desoye Gernot,Lang Ingrid
The International journal of artificial organs
PURPOSE::Blood vessel reconstruction is an increasing need of patients suffering from cardiovascular diseases. For the development of microvascular prostheses, efficient endothelialization is mandatory to prevent graft occlusion. Here, we assessed the impact of amnion-derived mesenchymal stem/stromal cells (hAMSC), known for their important angiogenic potential, on the integrity and stability of endothelial cells exposed to shear stress in vascular grafts. METHODS::Human placental endothelial cells (hPEC) were cultured at the inner surface of an expanded polytetrafluoroethylene (ePTFE) graft positioned within a bioreactor and exposed to a minimal shear stress of 0.015 dyne/cm or a physiological shear stress of 0.92 dyne/cm. hAMSC attached to the outer graft surface were able to interact with human placental endothelial cells by paracrine factors. RESULTS::Microscopical analysis and evaluation of glucose/lactate metabolism evidenced successful cell seeding of the graft: hPEC formed a stable monolayer, hAMSC showed a continuous growth during 72 h incubation. hAMSC improved the viability of hPEC exposed to 0.015 dyne/cm as shown by a decreased lactate dehydrogenase release of 13% after 72 h compared to hPEC single culture. The viability-enhancing effect of hAMSC on hPEC was further improved by 13% under physiological shear stress. Angiogenesis array analysis revealed that hPEC exposed to physiological shear stress and hAMSC co-culture reduced the secretion of angiogenin, GRO, MCP-1, and TIMP-2. CONCLUSION::hAMSC exerted best survival-enhancing effects on hPEC under exposure to physiological shear stress and modulated endothelial function by paracrine factors. Our data support further studies on the development of grafts functionalized with hAMSC-derived secretomes to enable fast clinical application.
Mesenchymal Stem Cells Transplantation in Intracerebral Hemorrhage: Application and Challenges.
Gong Yu-Hua,Hao Shi-Lei,Wang Bo-Chu
Frontiers in cellular neuroscience
Intracerebral hemorrhage (ICH) is one of the leading causes of death and long-term disability worldwide. Mesenchymal stem cell (MSC) therapies have demonstrated improved outcomes for treating ICH-induced neuronal defects, and the neural network reconstruction and neurological function recovery were enhanced in rodent ICH models through the mechanisms of neurogenesis, angiogenesis, anti-inflammation, and anti-apoptosis. However, many key issues associated with the survival, differentiation, and safety of grafted MSCs after ICH remain to be resolved, which hinder the clinical translation of MSC therapy. Herein, we reviewed an overview of the research status of MSC transplantation after ICH in different species including rodents, swine, monkey, and human, and the challenges for MSC-mediated ICH recovery from pathological microenvironment have been summarized. Furthermore, some efficient strategies for the outcome improvement of MSC transplantation were proposed.
The potential of stem cell therapy for stroke: is PISCES the sign?
Smith Helen K,Gavins Felicity N E
FASEB journal : official publication of the Federation of American Societies for Experimental Biology
Substantial developments in the field of stem cell research point toward novel therapies for the treatment of diseases such as stroke. This review covers the establishment of tissue damage in stroke and the status of current therapies. We evaluate stem cell therapy with respect to other treatments, including clinical, preclinical, and failed, and provide a comprehensive account of stem cell clinical trials for stroke therapy currently underway. Finally, we describe mechanisms through which stem cells improve outcome in experimental stroke as well as potential pitfalls this basic research has identified.
Stem cell mediation of functional recovery after stroke in the rat.
Ramos-Cabrer Pedro,Justicia Carles,Wiedermann Dirk,Hoehn Mathias
BACKGROUND:Regenerative strategies of stem cell grafting have been demonstrated to be effective in animal models of stroke. In those studies, the effectiveness of stem cells promoting functional recovery was assessed by behavioral testing. These behavioral studies do, however, not provide access to the understanding of the mechanisms underlying the observed functional outcome improvement. METHODOLOGY/PRINCIPAL FINDINGS:In order to address the underlying mechanisms of stem cell mediated functional improvement, this functional improvement after stroke in the rat was investigated for six months after stroke by use of fMRI, somatosensory evoked potentials by electrophysiology, and sensorimotor behavior testing. Stem cells were grafted ipsilateral to the ischemic lesion. Rigorous exclusion of spontaneous recovery as confounding factor permitted to observe graft-related functional improvement beginning after 7 weeks and continuously increasing during the 6-month observation period. The major findings were i) functional improvement causally related to the stem cells grafting; ii) tissue replacement can be excluded as dominant factor for stem cell mediated functional improvement; iii) functional improvement occurs by exclusive restitution of the function in the original representation field, without clear contributions from reorganization processes, and iv) stem cells were not detectable any longer after six months. CONCLUSIONS/SIGNIFICANCE:A delayed functional improvement due to stem cell implantation has been documented by electrophysiology, fMRI and behavioral testing. This functional improvement occurred without cells acting as a tissue replacement for the necrotic tissue after the ischemic event. Combination of disappearance of grafted cells after six months on histological sections with persistent functional recovery was interpreted as paracrine effects by the grafted stem cells being the dominant mechanism of cell activity underlying the observed functional restitution of the original activation sites. Future studies will have to investigate whether the stem cell mediated improvement reactivates the original representation target field by using original connectivity pathways or by generating/activating new ones for the stimulus.
Systemic transplantation of human adipose stem cells attenuated cerebral inflammation and degeneration in a hemorrhagic stroke model.
Kim Jeong-Min,Lee Soon-Tae,Chu Kon,Jung Keun-Hwa,Song Eun-Cheol,Kim Se-Jeong,Sinn Dong-In,Kim Jin-Hee,Park Dong-Kyu,Kang Kyung-Mook,Hyung Hong Nan,Park Hee-Kwon,Won Chong-Hyun,Kim Kyu-Han,Kim Manho,Kun Lee Sang,Roh Jae-Kyu
Adipose-derived stem cells (ASCs) are readily accessible multipotent mesenchymal stem cells and are known to secrete multiple growth factors, and thereby to have cytoprotective effects in various injury models. In the present study, the authors investigated the neuroprotective effect of ASCs in an intracerebral hemorrhage (ICH) model. ICH was induced via the stereotaxic infusion of collagenase, and human ASCs (three million cells per animal) isolated from human fresh fat tissue, were intravenously administered at 24 h post-ICH induction. Acute brain inflammation markers, namely, cell numbers positively stained for terminal transferase dUTP nick end labeling (TUNEL), myeloperoxidase (MPO), or OX-42, and brain water content were checked at 3 days post-ICH. In addition, the authors quantified brain degeneration by measuring hemispheric atrophy and perihematomal glial thickness at 6 weeks post-ICH, and determined modified limb placing behavioral scores weekly over 5 weeks post-ICH. The results showed that brain water content, TUNEL+, and MPO+ cell numbers were significantly reduced in the ASC-transplanted rats. ASC transplantation attenuated neurological deficits from 4 to 5 weeks post-ICH, and reduced both the brain atrophy and the glial proliferation at 6 weeks. Transplanted ASCs were found to densely populate perihematomal areas at 6 weeks, and to express endothelial markers (von Willebrand factor and endothelial barrier antigen), but not neuronal or glial markers. In summary, ASCs transplantation in the ICH model reduced both acute cerebral inflammation and chronic brain degeneration, and promoted long-term functional recovery.
Stem Cell Therapy: A Promising Therapeutic Method for Intracerebral Hemorrhage.
Gao Liansheng,Xu Weilin,Li Tao,Chen Jingyin,Shao Anwen,Yan Feng,Chen Gao
Spontaneous intracerebral hemorrhage (ICH) is one type of the most devastating cerebrovascular diseases worldwide, which causes high morbidity and mortality. However, efficient treatment is still lacking. Stem cell therapy has shown good neuroprotective and neurorestorative effect in ICH and is a promising treatment. In this study, our aim was to review the therapeutic effects, strategies, related mechanisms and safety issues of various types of stem cell for ICH treatment. Numerous studies had demonstrated the therapeutic effects of diverse stem cell types in ICH. The potential mechanisms include tissue repair and replacement, neurotrophy, promotion of neurogenesis and angiogenesis, anti-apoptosis, immunoregulation and anti-inflammation and so forth. The microenvironment of the central nervous system (CNS) can also influence the effects of stem cell therapy. The detailed therapeutic strategies for ICH treatment such as cell type, the number of cells, time window, and the routes of medication delivery, varied greatly among different studies and had not been determined. Moreover, the safety issues of stem cell therapy for ICH should not be ignored. Stem cell therapy showed good therapeutic effect in ICH, making it a promising treatment. However, safety should be carefully evaluated, and more clinical trials are required before stem cell therapy can be extensively applied to clinical use.
Transplantation of adipose-derived stem cells is associated with neural differentiation and functional improvement in a rat model of intracerebral hemorrhage.
Chen Juan,Tang Ying-Xin,Liu Yong-Ming,Chen Ji,Hu Xiao-Qing,Liu Na,Wang Shu-Xin,Zhang Yu,Zeng Wen-Gao,Ni Hou-Jie,Zhao Bin,Chen Yan-Fang,Tang Zhou-Ping
CNS neuroscience & therapeutics
AIMS:To examine whether transplantation of adipose-derived stem cells (ADSCs) induces neural differentiation and improves neural function in a rat intracerebral hemorrhage (ICH) model. METHODS:Adipose-derived stem cells cells were isolated from inguinal fat pad of rat. ICH was induced by injection of collagenase type IV into the right basal ganglia of rat. Forty-eight hours after ICH, ADSCs cells (10 μL of 2-4 × 10(7) cells/mL) were injected into the right lateral cerebral ventricle. The differentiation of ADSCs was detected in vitro and in vivo. The neural function was evaluated with Zea Longa 5-grade scale at day 1, 3, 7, 14, or 28. RESULTS:Our data demonstrated that ADSCs differentiated into cells that shared the similarities of neurons or astrocytes in vitro. Transplantation of ADSCs decreased cell apoptosis and the transplanted ADSCs were able to differentiate into neuron-like and astrocyte-like cells around the hematoma, accompanied with upregulation of vascular endothelial growth factor expression and improvement of neural function. CONCLUSIONS:Our data suggest that transplantation of ADSCs could be a therapeutic approach for ICH stroke.
Clinical and preclinical translation of cell-based therapies using adipose tissue-derived cells.
Gimble Jeffrey M,Guilak Farshid,Bunnell Bruce A
Stem cell research & therapy
Adipose tissue is now recognized as an accessible, abundant, and reliable site for the isolation of adult stem cells suitable for tissue engineering and regenerative medicine applications. The past decade has witnessed an explosion of preclinical data relating to the isolation, characterization, cryopreservation, differentiation, and transplantation of freshly isolated stromal vascular fraction cells and adherent, culture-expanded, adipose-derived stromal/stem cells in vitro and in animal models. This body of work has provided evidence supporting clinical translational applications of adipose-derived cells in safety and efficacy trials. The present article reviews the case reports and phase I-III clinical evidence using autologous adipose-derived cells that have been published, to date, in the fields of gastroenterology, neurology, orthopedics, reconstructive surgery, and related clinical disciplines. Future directions and challenges facing the field are discussed and evaluated.
Effects of human umbilical cord mesenchymal stem cell transplantation combined with minimally invasive hematoma aspiration on intracerebral hemorrhage in rats.
Zhang Qinghua,Shang Xiao,Hao Maolin,Zheng Maoyong,Li Yanxia,Liang Zhigang,Cui Yuanxiao,Liu Zhenhua
American journal of translational research
This study is to investigate the effects of human umbilical cord-mesenchymal stem cells (HUC-MSCs) transplantation combined with minimally invasive hematoma aspiration on neural functional recovery and p53 gene expression in rats with intracerebral hemorrhage (ICH). Collagenase type-IV was injected to the caudate nucleus of the rats to make ICH models. One hundred and twenty Sprague-Dawley rats with successful modeling were randomly divided into 4 groups, including the ICH group, hematoma aspiration group, HUC-MSCs transplantation group and HUC-MSCs transplantation combined with hematoma aspiration group (combination group). Neural functional status of the rats was assessed by modified neurological severity score (mNSS). Expression of p53 in the cerebral tissues surrounding ICH was detected by immunohistochemical assays. The scores of mNSS and the expression of p53 gene in the hematoma aspiration group, the HUC-MSCs transplantation group and the combination group were significantly lower than those in the ICH group at each indicated time point (p < 0.05). Intriguingly, mNSS scores and p53 expression in the combination group were significantly lower than those in the hematoma aspiration group on day 7, 14 and 30 (p < 0.05), and significantly lower than those in the HUC-MSCs transplantation group on day 14 and 30 (p < 0.05). HUC-MSCs transplantation combined with minimally invasive hematoma aspiration is more effective than either therapy alone in rats with ICH and could distinctly reduce the damage of nerve cells.
Cell therapy for cerebral hemorrhage: Five year follow-up report.
Chang Zhitian,Mao Gengsheng,Sun Lizhong,Ao Qiang,Gu Yongquan,Liu Ying
Experimental and therapeutic medicine
The aim of the study was to examine treatment of cerebral hemorrhages with bone-marrow or human umbilical cord-derived mesenchymal stem cells (BMSCs or Hu-MSCs) and conventional surgical approaches, and determine and compare the effectiveness, feasibility, safety and reproducibility of each method. A retrospective analysis was performed on a cohort of cell-treated cerebral hemorrhage patients from October 1, 2007 to October 1, 2009. A total of 24 patients, all of whom received conventional surgical treatment, were classified as follows: i) The control group consisted of 8 patients who received only hematoma removal surgery, ii) the autologous group consisted of 7 patients who received additional autologous bone marrow mononuclear cell transplantation, and iii) the allograft group consisted of 9 patients who received additional umbilical cord mononuclear cell transplantation. After conventional hematoma removal surgery and X-ray supervision within 24 h and at 7 days, neurological disability and function tests were completed 3, 6, 12, 36 and 60 months later. The T-cell marker plasma levels were analyzed after 60 months. The results showed that, at approximately 3.5 months after graft the hematomas in all the groups were completely reabsorbed as observed on computed tomography scans. However, the functional outcomes in the cell-transplanted groups were better than in the control group after 5 years. While the National Institutes of Health Stroke Scale, modified Rankin score and modified Barthel index scores were simliar in the cell-transplanted groups, patients in the allograft group had better outcomes than those in the autologous graft group starting at 3 months and until the end of the follow-up period. The serum levels of T-cell markers CD4, CD56 and human leukocyte antigen-DR in the allograft group showed no signs of immunogenic graft complications and there were no significant differences in T-cell subtypes among the patient groups. The results of the present study suggest that, treatment of cerebral hemorrhage patients can be safely and effectively accomplished using Hu-MSC grafting and larger clinical trials should be considered in the future.
Intracerebral and Intravenous Transplantation Represents a Favorable Approach for Application of Human Umbilical Cord Mesenchymal Stromal Cells in Intracerebral Hemorrhage Rats.
Xie Jiang,Wang Bin,Wang Lian,Dong Fang,Bai Gang,Liu Yongjun
Medical science monitor : international medical journal of experimental and clinical research
BACKGROUND Intracerebral hemorrhage (ICH) is one severe subtype of stroke, with a very complex pathology. Stem cell-based therapy holds promising potential in the treatment of neurological disorders. Human umbilical cord-derived mesenchymal stem cells (UC-MSCs) have a therapeutic effect in recovery from brain damage following ICH. The aim of this study was to identify an effective and convenient way of using UC-MSCs in the ICH rat model. MATERIAL AND METHODS CM-DiI-labeled human UC-MSCs were transplanted intracerebrally or intravenously into collagenase VII-induced ICH rat models. Neurological function was evaluated before ICH and at 0, 7, 14, 21, and 28 days after treatment. ICH rats were sacrificed to evaluate the injury volume. Neurogenesis and angiogenesis and vascular areas were investigated using microtubule-associated protein 2 (MAP2), glial fibrillary acidic protein (GFAP), and 4',6-diamidino-2-phenylindole (DAPI) immunohistochemistry at two weeks after transplantation. RESULTS The intracerebral and intravenous administration of UC-MSCs both resulted in significant improvement in neurological function and decrease in injury volume of ICH rats. Transplanted UC-MSCs were chemotactic in vivo and showed a predominant distribution around the ICH region. In addition, UC-MSCs could integrate into the cerebral vasculature in both groups. CONCLUSIONS Both intracerebral and intravenous administration of UC-MSCs could have a favorable effect on recovery of neurological function in ICH rats, although the fundamental mechanisms may be different between the two groups. Our data suggest that intravenous implantation of UC-MSCs could serve as a favorable approach for cell-based therapy in central nervous system (CNS) diseases according to clinical needs.
Bone Marrow Mesenchymal Stem Cell Transplantation Increases GAP-43 Expression via ERK1/2 and PI3K/Akt Pathways in Intracerebral Hemorrhage.
Cui Jianzhong,Cui Changmeng,Cui Ying,Li Ran,Sheng Huaxin,Jiang Xiaohua,Tian Yanxia,Wang Kaijie,Gao Junling
Cellular physiology and biochemistry : international journal of experimental cellular physiology, biochemistry, and pharmacology
BACKGROUND/AIMS:Intracerebral hemorrhage (ICH) occurs in hypertensive patients and results in high rates of mortality and disability. This study determined whether bone marrow mesenchymal stem cell (BMSC) transplantation affects axonal regeneration and examined the underlying mechanisms after the administration of PD98059 (p-ERK1/2 inhibitor) or/ and LY294002 (PI3K inhibitor). The hypothesis that was intended to be tested was that BMSC transplantation regulates the expression of growth-associated protein-43 (GAP-43) via the ERK1/2 and PI3K/Akt signaling pathways. METHODS:Seventy-five male rats (250-280 g) were subjected to intracerebral blood injection and then randomly received a vehicle, BMSCs, PD98059 or LY294002 treatment. Neurological deficits were evaluated prior to injury and at 1, 3 and 7 days post-injury. The expression of GAP-43, Akt, p-Akt, ERK1/2, and p-ERK1/2 proteins was measured by western blot analysis. RESULTS:BMSC transplantation attenuated neurological deficits 3-7 days post-ICH. The expression of GAP-43 was increased 3 days following BMSC transplantation. However, this increase was inhibited by either PD98059 or LY294002 treatment. Treatment with both PD98059 and LY294002 was more effective than was treatment with an individual compound. CONCLUSION:BMSC transplantation could attenuate neurological deficits and activate axonal regeneration in this rat ICH model. The protective effects might be associated with increased GAP-43 expression by activating both the ERK1/2 and PI3K/Akt signaling pathways.
Neuroprotection by PlGF gene-modified human mesenchymal stem cells after cerebral ischaemia.
Liu H,Honmou O,Harada K,Nakamura K,Houkin K,Hamada H,Kocsis J D
Brain : a journal of neurology
Intravenous delivery of mesenchymal stem cells (MSCs) prepared from adult bone marrow reduces infarction size and ameliorates functional deficits in rat cerebral ischaemia models. Placental growth factor (PlGF) is angiogenic to impaired non-neural tissue. To test the hypothesis that PlGF contributes to the therapeutic benefits of MSC delivery in cerebral ischaemia, we compared the efficacy of systemic delivery of human MSCs (hMSCs) and hMSCs transfected with a fibre-mutant F/RGD adenovirus vector with a PlGF gene (PlGF-hMSCs). A permanent middle cerebral artery occlusion (MCAO) was induced by intraluminal vascular occlusion with a microfilament. hMSCs and PlGF-hMSCs were intravenously injected into the rats 3 h after MCAO. Lesion size was assessed at 3 and 6 h, and 1, 3, 4 and 7 days using MR imaging and histology. Functional outcome was assessed using the limb placement test and the treadmill stress test. Both hMSCs and PlGF-hMSCs reduced lesion volume, induced angiogenesis and elicited functional improvement compared with the control sham group, but the effect was greater in the PlGF-hMSC group. Enzyme-linked immunosorbent assay of the infarcted hemisphere revealed an increase in PlGF in both hMSC groups, but a greater increase in the PlGF-hMSC group. These data support the hypothesis that PlGF contributes to neuroprotection and angiogenesis in cerebral ischaemia, and cellular delivery of PlGF to the brain can be achieved by intravenous delivery of hMSCs.
Multilineage potential of stable human mesenchymal stem cell line derived from fetal marrow.
Nagai Atsushi,Kim Woo K,Lee Hong J,Jeong Han S,Kim Kwang S,Hong Seok H,Park In H,Kim Seung U
Human bone marrow contains two major cell types, hematopoietic stem cells (HSCs) and mesenchymal stem cells (MSCs). MSCs possess self-renewal capacity and pluripotency defined by their ability to differentiate into osteoblasts, chondrocytes, adipocytes and muscle cells. MSCs are also known to differentiate into neurons and glial cells in vitro, and in vivo following transplantation into the brain of animal models of neurological disorders including ischemia and intracerebral hemorrhage (ICH) stroke. In order to obtain sufficient number and homogeneous population of human MSCs, we have clonally isolated permanent and stable human MSC lines by transfecting primary cell cultures of fetal human bone marrow MSCs with a retroviral vector encoding v-myc gene. One of the cell lines, HM3.B10 (B10), was found to differentiate into neural cell types including neural stem cells, neurons, astrocytes and oligodendrocytes in vitro as shown by expression of genetic markers for neural stem cells (nestin and Musashi1), neurons (neurofilament protein, synapsin and MAP2), astrocytes (glial fibrillary acidic protein, GFAP) and oligodendrocytes (myelin basic protein, MBP) as determined by RT-PCR assay. In addition, B10 cells were found to differentiate into neural cell types as shown by immunocytochical demonstration of nestin (for neural stem cells), neurofilament protein and beta-tubulin III (neurons) GFAP (astrocytes), and galactocerebroside (oligodendrocytes). Following brain transplantation in mouse ICH stroke model, B10 human MSCs integrate into host brain, survive, differentiate into neurons and astrocytes and induce behavioral improvement in the ICH animals. B10 human MSC cell line is not only a useful tool for the studies of organogenesis and specifically for the neurogenesis, but also provides a valuable source of cells for cell therapy studies in animal models of stroke and other neurological disorders.
Plasticity of cultured mesenchymal stem cells: switch from nestin-positive to excitable neuron-like phenotype.
Wislet-Gendebien Sabine,Hans Grégory,Leprince Pierre,Rigo Jean-Michel,Moonen Gustave,Rogister Bernard
Stem cells (Dayton, Ohio)
Bone marrow mesenchymal stem cells (MSCs) can differentiate into several types of mesenchymal cells, including osteocytes, chondrocytes, and adipocytes, but, under appropriate experimental conditions, can also differentiate into nonmesenchymal cells--for instance, neural cells. These observations have raised interest in the possible use of MSCs in cell therapy strategies for various neurological disorders. In the study reported here, we addressed the question of in vitro differentiation of MSCs into functional neurons. First, we demonstrate that when they are co-cultured with cerebellar granule neurons, adult MSCs can express neuronal markers. Two factors are needed for the emergence of neuronal differentiation of the MSCs: the first one is nestin expression by MSCs (nestin is a marker for the responsive character of MSCs to extrinsic signals), and the second one is a direct cell-cell interaction between neural cells and MSCs that allows the integration of these extrinsic signals. Three different approaches suggest that neural phenotypes arise from MSCs by a differentiation rather than a cell fusion process, although this last phenomenon can also coexist. The expression of several genes--including sox, pax, notch, delta, frizzled, and erbB--was analyzed by quantitative reverse transcription polymerase chain reaction (RT-PCR) in order to further characterize the nestin-positive phenotype compared to the nestin-negative one. An overexpression of sox2, sox10, pax6, fzd, erbB2, and erbB4 is found in nestin-positive MSCs. Finally, electrophysiological analyses demonstrate that MSC-derived neuron-like cells can fire single-action potentials and respond to several neurotransmitters such as GABA, glycine, and glutamate. We conclude that nestin-positive MSCs can differentiate in vitro into excitable neuron-like cells.
After Intracerebral Hemorrhage, Oligodendrocyte Precursors Proliferate and Differentiate Inside White-Matter Tracts in the Rat Striatum.
Joseph Michael J E,Caliaperumal Jayalakshmi,Schlichter Lyanne C
Translational stroke research
Damage to myelinated axons contributes to neurological deficits after acute CNS injury, including ischemic and hemorrhagic stroke. Potential treatments to promote re-myelination will require fully differentiated oligodendrocytes, but almost nothing is known about their fate following intracerebral hemorrhage (ICH). Using a rat model of ICH in the striatum, we quantified survival, proliferation, and differentiation of oligodendrocyte precursor cells (OPCs) (at 1, 3, 7, 14, and 28 days) in the peri-hematoma region, surrounding striatum, and contralateral striatum. In the peri-hematoma, the density of Olig2(+) cells increased dramatically over the first 7 days, and this coincided with disorganization and fragmentation of myelinated axon bundles. Very little proliferation (Ki67(+)) of Olig2(+) cells was seen in the anterior subventricular zone from 1 to 28 days. However, by 3 days, many were proliferating in the peri-hematoma region, suggesting that local proliferation expands their population. By 14 days, the density of Olig2(+) cells declined in the peri-hematoma region, and, by 28 days, it reached the low level seen in the contralateral striatum. At these later times, many surviving axons were aligned into white-matter bundles, which appeared less swollen or fragmented. Oligodendrocyte cell maturation was prevalent over the 28-day period. Densities of immature OPCs (NG2(+)Olig2(+)) and mature (CC-1(+)Olig2(+)) oligodendrocytes in the peri-hematoma increased dramatically over the first week. Regardless of the maturation state, they increased preferentially inside the white-matter bundles. These results provide evidence that endogenous oligodendrocyte precursors proliferate and differentiate in the peri-hematoma region and have the potential to re-myelinate axon tracts after hemorrhagic stroke.
Axonal dysfunction in internal capsule is closely associated with early motor deficits after intracerebral hemorrhage in mice.
Hijioka Masanori,Anan Junpei,Matsushita Hideaki,Ishibashi Hayato,Kurauchi Yuki,Hisatsune Akinori,Seki Takahiro,Katsuki Hiroshi
Previously we showed that expansion of intracerebral hemorrhage (ICH) into the internal capsule greatly aggravated neurological symptoms in mice. Here we examined ICH-associated events in the internal capsule with relation to neurological dysfunction. Corticospinal axons labeled by biotinylated dextran amine exhibited fragmented appearance after ICH induced by local injection of collagenase into the internal capsule. Fragmentation of axonal structures was confirmed by neurofilament-H immunostaining, which was evident from 6h after induction of ICH. We also observed accumulation of amyloid precursor protein, which indicated compromised axonal transport, from 3h after induction of ICH. The early defect in axonal transport was accompanied by a robust decline in motor performance. Local application of an axonal transport inhibitor colchicine to the internal capsule induced a prompt decline in motor performance, suggesting that compromised axonal transport is closely associated with early neurological dysfunction in ICH. Arrest of axonal transport and fragmentation of axonal structures were also induced by local injection of thrombin, but not by thrombin receptor activator peptide-6, a protease-activated receptor-1 agonist. These results suggest that receptor-independent actions of thrombin mediate disruption of structure and function of axons by hemorrhage expansion into the internal capsule, which leads to severe neurological dysfunction.