Hypoxia-Elicited Mesenchymal Stem Cell-Derived Small Extracellular Vesicles Alleviate Myocardial Infarction by Promoting Angiogenesis through the miR-214/Sufu Pathway.
Stem cells international
Objective:Myocardial infarction is a leading cause of mortality worldwide. Angiogenesis in the infarct border zone is vital for heart function restoration after myocardial infarction. Hypoxia-induced MSC modification is a safe and effective approach for angiogenesis in clinical therapy; however, the mechanism still requires further investigation. In our study, we preconditioned human umbilical cord mesenchymal stem cells (huMSCs) with hypoxia and isolated the small extracellular vesicles (sEVs) to promote cardiac repair. We also investigated the potential mechanisms. Method:huMSCs were preconditioned with hypoxia (1% O and 5% CO at 37°C for 48 hours), and their sEVs were isolated using the Total Exosome Isolation reagent kit. To explore the role of miR-214 in MSC-derived sEVs, sEVs with low miR-214 expression were prepared by transfecting miR-214 inhibitor into huMSCs before hypoxia pretreatment. Scratch assays and tube formation assays were performed in sEVs cocultured with HUVECs to assess the proangiogenic capability of MSC-sEVs and MSC-sEVs. Rat myocardial infarction models were used to investigate the ability of miR-214-differentially expressed sEVs in cardiac repair. Echocardiography, Masson's staining, and immunohistochemical staining for CD31 were performed to assess cardiac function, the ratio of myocardial fibrosis, and the capillary density after sEV implantation. The potential mechanism by which MSC-sEVs enhance angiogenesis was explored by RT-qPCR and western blotting. Results:Tube formation and scratch assays demonstrated that the proangiogenic capability of huMSC-derived sEVs was enhanced by hypoxia pretreatment. Echocardiography and Masson's staining showed greater improvements in heart function and less ventricular remodeling after MSC-sEV transplantation. The angiogenic capability was reduced following miR-214 knockdown in MSC-sEVs. Furthermore, Sufu, a target of miR-214, was decreased, and hedgehog signaling was activated in HUVECs. Conclusion:We found that hypoxia induced miR-214 expression both in huMSCs and their sEVs. Transplantation of MSC-sEVs into a myocardial infarction model improved cardiac repair by increasing angiogenesis. Mechanistically, MSC-sEVs promote HUVEC tube formation and migration by transferring miR-214 into recipient cells, inhibiting Sufu expression, and activating the hedgehog pathway. Hypoxia-induced vesicle modification is a feasible way to restore heart function after myocardial infarction.
10.1155/2023/1662182
The Effects of Hypoxia-Preconditioned Dental Stem Cell-Derived Secretome on Tissue Regeneration.
Tissue engineering. Part B, Reviews
Mesenchymal stroma cells derived from oral tissues are known as dental stem cells (DSCs). Owing to their unique therapeutic niche and clinical accessibility, DSCs serve as a promising treatment option for bone defects and oral tissue regeneration. DSCs exist in a hypoxic microenvironment , which is far lower than the current 20% oxygen concentration used in culture. It has been widely reported that the application of an oxygen concentration less than 5% in the culture of DSCs is beneficial for preserving stemness and promoting proliferation, migration, and paracrine activity. The paracrine function of DSCs involves the secretome, which includes conditioned media (CM) and soluble bioactive molecules, as well as extracellular vesicles extracted from CM. Hypoxia can play a role in immunomodulation and angiogenesis by altering the protein or nucleic acid components in the secretory group, which enhances the therapeutic potential of DSCs. This review summarizes the biological characteristics of DSCs, the influence of hypoxia on DSCs, the impact of hypoxia on the secretory group of DSCs, and the latest progress on the use of DSCs secretory group in tissue regeneration based on hypoxia pretreatment. We highlighted the multifunctional biological effect of hypoxia culture on tissue regeneration and provided a summary of the current mechanism of hypoxia in the pretreatment of DSCs.
10.1089/ten.TEB.2024.0054
Novel neutrophil extracellular trap-related mechanisms in diabetic wounds inspire a promising treatment strategy with hypoxia-challenged small extracellular vesicles.
Bioactive materials
Neutrophil extracellular traps (NETs) have been considered a significant unfavorable factor for wound healing in diabetes, but the mechanisms remain unclear. The therapeutic application of small extracellular vesicles (sEVs) derived from mesenchymal stem cells (MSCs) has received considerable attention for their properties. Hypoxic preconditioning is reported to enhance the therapeutic potential of MSC-derived sEVs in regenerative medicine. Therefore, the aim of this study is to illustrate the detailed mechanism of NETs in impairment of diabetic wound healing and develop a promising NET-targeting treatment based on hypoxic pretreated MSC-derived sEVs (Hypo-sEVs). Excessive NETs were found in diabetic wounds and in high glucose (HG)-induced neutrophils. Further research showed that high concentration of NETs impaired the function of fibroblasts through activating endoplasmic reticulum (ER) stress. Hypo-sEVs efficiently promoted diabetic wound healing and reduced the excessive NET formation by transferring miR-17-5p. Bioinformatic analysis and RNA interference experiment revealed that miR-17-5p in Hypo-sEVs obstructed the NET formation by targeting TLR4/ROS/MAPK pathway. Additionally, miR-17-5p overexpression decreased NET formation and overcame NET-induced impairment in fibroblasts, similar to the effects of Hypo-sEVs. Overall, we identify a previously unrecognized NET-related mechanism in diabetic wounds and provide a promising NET-targeting strategy for wound treatment.
10.1016/j.bioactmat.2023.04.007
Hypoxia-pretreated mesenchymal stem cell-derived exosomes-loaded low-temperature extrusion 3D-printed implants for neural regeneration after traumatic brain injury in canines.
Frontiers in bioengineering and biotechnology
Regenerating brain defects after traumatic brain injury (TBI) still remains a significant difficulty, which has motivated interest in 3D printing to design superior replacements for brain implantation. Collagen has been applied to deliver cells or certain neurotrophic factors for neuroregeneration. However, its fast degradation rate and poor mechanical strength prevent it from being an excellent implant material after TBI. In the present study, we prepared 3D-printed collagen/silk fibroin/hypoxia-pretreated human umbilical cord mesenchymal stem cells (HUCMSCs)-derived exosomes scaffolds (3D-CS-HMExos), which possessed favorable physical properties suitable biocompatibility and biodegradability and were attractive candidates for TBI treatment. Furthermore, inspired by exosomal alterations resulting from cells in different external microenvironments, exosomes were engineered through hypoxia stimulation of mesenchymal stem cells and were proposed as an alternative therapy for promoting neuroregeneration after TBI. We designed hypoxia-preconditioned (Hypo) exosomes derived from HUCMSCs (Hypo-MExos) and proposed them as a selective therapy to promote neuroregeneration after TBI. For the current study, 3D-CS-HMExos were prepared for implantation into the injured brains of beagle dogs. The addition of hypoxia-induced exosomes further exhibited better biocompatibility and neuroregeneration ability. Our results revealed that 3D-CS-HMExos could significantly promote neuroregeneration and angiogenesis due to the doping of hypoxia-induced exosomes. In addition, the 3D-CS-HMExos further inhibited nerve cell apoptosis and proinflammatory factor (TNF-α and IL-6) expression and promoted the expression of an anti-inflammatory factor (IL-10), ultimately enhancing the motor functional recovery of TBI. We proposed that the 3D-CS-loaded encapsulated hypoxia-induced exosomes allowed an adaptable environment for neuroregeneration, inhibition of inflammatory factors and promotion of motor function recovery in TBI beagle dogs. These beneficial effects implied that 3D-CS-HMExos implants could serve as a favorable strategy for defect cavity repair after TBI.
10.3389/fbioe.2022.1025138
MiR-126-3p-Enriched Extracellular Vesicles from Hypoxia-Preconditioned VSC 4.1 Neurons Attenuate Ischaemia-Reperfusion-Induced Pain Hypersensitivity by Regulating the PIK3R2-Mediated Pathway.
Molecular neurobiology
Recent evidence suggests that hypoxia preconditioning can alter the microRNA (miRNA) profile of extracellular vesicles (EVs) and has better neuroprotective effects when enriched miRs are delivered to recipients. However, the roles of exosomal miRNAs in regulating ischaemia-reperfusion (IR)-induced pain hypersensitivity are largely unknown. Thus, we isolated EVs from normoxia-conditioned neurons (Nor-VSC EVs) and Hypo-VSC EVs by ultracentrifugation. After the initial screening by a microarray analysis and quantitative RT-PCR (qRT-PCR), miR-126-3p, which was detected as the most altered miR in the Hypo-VSC EVs, was further confirmed by applying GW4869 to inhibit exosomal secretion. Moreover, transfection with a miR-126 mimic obviously increased miR-126-3p expression in Nor-VSC EVs, whereas a miR-126 inhibitor prevented the increase in miR-126-3p in Hypo-VSC EVs. A rat model of pain was established by performing 8-min occlusion of the aorta. Following IR, compared with the Nor-VSC EVs- or antagomir-126-injected rats, the Hypo-VSC EVs-injected rats displayed improved pain hypersensitivity demonstrated as higher PWT and PWL values. Mechanistically, PIK3R2 is a target of miR-126-3p and might be a modulator of the phosphoinositide 3-kinase (PI3K)/Akt pathway as the PIK3R2 and PI3K immunoreactivities in each group were changed in opposite directions. Compared with the controls, higher protein levels of PI3K and phosphorylated Akt but lower levels of phosphorylated nuclear factor-κ B (NF-κB), tumour necrosis factor (TNF)-α and interleukin (IL)-1β were detected in the spinal cords of the Hypo-VSC EVs-injected rats, and these effects were impaired by an injection of Hypo-VSC EVs combined with antagomir-126. Collectively, the miR-126-3p-enriched Hypo-VSC EVs attenuated IR-induced pain hypersensitivity by restoring miR-126-3p expression in the injured spinal cord and subsequently modulating PIK3R2-mediated PI3K/Akt and NF-κB signalling pathways.
10.1007/s12035-020-02159-y
Exosomes derived from hypoxic mesenchymal stem cells restore ovarian function by enhancing angiogenesis.
Stem cell research & therapy
BACKGROUND:hucMSC-exosomes can be engineered to strengthen their therapeutic potential, and the present study aimed to explore whether hypoxic preconditioning can enhance the angiogenic potential of hucMSC-exosomes in an experimental model of POF. METHODS:Primary hucMSCs and ROMECs were isolated from fresh tissue samples and assessed through a series of experiments. Exosomes were isolated from hucMSCs under normoxic or hypoxic conditions (norm-Exos and hypo-Exos, respectively) and then characterized using classic experimental methods. Based on a series of angiogenesis-related assays, we found that hypo-Exos significantly promoted ROMEC proliferation, migration, and tube formation and increased angiogenesis-promoting molecules in vitro. Histology, immunohistochemistry, and immunofluorescence experiments in a rat model of POF demonstrated that hypoxia pretreatment strengthens the therapeutic angiogenic effect of hucMSC-exosomes in vivo. Subsequently, high-throughput miRNA sequencing, qRT‑PCR analysis, and western blotting were employed to identify the potential molecular mechanism. RESULTS:We found that hypo-Exos enhance endothelial function and angiogenesis via the transfer of miR-205-5p in vitro and in vivo. Finally, based on the results of bioinformatics analysis, dual luciferase reporter assays, and gain- and loss-of-function studies, we found evidence indicating that exosomal miR-205-5p enhances angiogenesis by targeting the PTEN/PI3K/AKT/mTOR signalling pathway. These results indicated for the first time that exosomes derived from hypoxia-conditioned hucMSCs strongly enhance angiogenesis via the transfer of miR-205-5p by targeting the PTEN/PI3K/AKT/mTOR signalling pathway. CONCLUSIONS:Our findings provide a theoretical basis and demonstrate the potential application of a novel cell-free approach with stem cell-derived products in the treatment of POF.
10.1186/s13287-024-04111-6
Exosomes from myoblasts induced by hypoxic preconditioning improved ventricular conduction by increasing Cx43 expression in hypothermia ischemia reperfusion hearts.
Cytotechnology
Myocardial ischemia-reperfusion arrhythmia after cardiac surgery is common and seriously affects quality of life. Remote ischemic preconditioning can reduce the myocardial damage caused by severe ischemia. However, the underlying mechanism is not well understood. This study aimed to investigate the effects of exosomes derived from C2C12 mouse myoblasts after hypoxic preconditioning (HP) on ventricular conduction in hypothermic ischemia-reperfusion hearts. Myocardial ischemia-reperfusion model rats were established using the Langendorff cardiac perfusion system. Exosomes derived from normoxic (ExoA) and hypoxia-preconditioned (ExoB) C2C12 cells were injected into the jugular vein of the model rats. The time to heartbeat restoration, arrhythmia type and duration, and heart rate were recorded after myocardial ischemia-reperfusion. Conduction velocity on the surface of left ventricle was measured using a microelectrode array after 30 min of balanced perfusion, 15 min of reperfusion, and 30 min of reperfusion. Immunohistochemistry and western blotting were performed to determine the distribution and relative expression of connexin 43 (Cx43). ExoB contained more exosomes than ExoA, showing that HP stimulated the release of exosomes. The IR + ExoB group showed faster recovery of ventricular myocardial activity, a lower arrhythmia score, faster conduction velocity, and better electrical conductivity than the IR group. ExoB increased the expression of Cx43 and reduced its lateralization in the ventricular muscle. Our study showed that exosomes induced by hypoxic preconditioning can improve ventricular myocardial conduction and reperfusion arrhythmia in isolated hearts after hypothermic ischemia-reperfusion. Graphical abstract:
10.1007/s10616-024-00634-1
Exosomes from hypoxia-treated human adipose-derived mesenchymal stem cells enhance angiogenesis through VEGF/VEGF-R.
Han Yudi,Ren Jing,Bai Yun,Pei Xuetao,Han Yan
The international journal of biochemistry & cell biology
BACKGROUND:We previously reported that co-transplantation of exosomes from hypoxia-preconditioned adipose mesenchymal stem cells (ADSCs) improves the neoangiogenesis and survival of the grafted tissue. This study aimed to investigate the molecular mechanism of this protective effect. METHODS:Exosomes were collected from normoxia-treated (nADSC-Exo) or hypoxia--treated (hypADSC-Exo) human ADSCs, and their pro-angiogenic capacity was evaluated in human umbilical vein endothelial cells (HUVECs) and a nude mouse model of subcutaneous fat grafting. Protein array was used to compare the exosome-derived proteins between nADSC-Exo and hypADSC-Exo. RESULTS:Compared with the nADSC-Exo group and untreated control, hypADSC-Exo treatment significantly promoted proliferation, migration and tube-formation capability of HUVECs. Protein array revealed that the levels of vascular endothelial growth factor (VEGF), epidermal growth factor (EGF), fibroblast growth factor (FGF) and their receptors (VEGF-R2, VEGF-R3), and monocyte chemoattractant protein 2 (MCP-2), monocyte chemoattractant protein 4 (MCP-4) were significantly higher in the hypADSC-Exo than in the nADSC-Exo. In the nude mice model of fat grafting, immunofluorescence of CD31 showed that hypADSC-Exo dramatically improved neovascularization around the graft. Furthermore, compared with nADSC-Exo and control groups, cotransplantation of hypADSC-Exo significantly increased the protein expression of EGF, FGF, VEGF/VEGF-R, angiopoietin-1(Ang-1) and tyrosine kinase with immunoglobulin-like and EGF-like domains 1(Tie-1, an angiopoietin receptor) in the grafted tissue at 30 days after transplantation. Immunohistochemical analysis demonstrated that hypADSC-Exo treatment significantly increased VEGF-R expression in the grafted tissue. CONCLUSIONS:Exosomes from hypoxia-treated human ADSCs possess a higher capacity to enhance angiogenesis in fat grafting, at least partially, via regulating VEGF/VEGF-R signaling.
10.1016/j.biocel.2019.01.017
Hypoxia-preconditioned WJ-MSC spheroid-derived exosomes delivering miR-210 for renal cell restoration in hypoxia-reoxygenation injury.
Stem cell research & therapy
BACKGROUND:Recent advancements in mesenchymal stem cell (MSC) technology have paved the way for innovative treatment options for various diseases. These stem cells play a crucial role in tissue regeneration and repair, releasing local anti-inflammatory and healing signals. However, challenges such as homing issues and tumorigenicity have led to exploring MSC-exosomes as a promising alternative. MSC-exosomes have shown therapeutic potential in conditions like renal ischemia-reperfusion injury, but low production yields hinder their clinical use. METHODS:To address this limitation, we examined hypoxic preconditioning of Wharton jelly-derived MSCs (WJ-MSCs) 3D-cultured in spheroids on isolated exosome yields and miR-21 expression. We then evaluated their capacity to load miR-210 into HEK-293 cells and mitigate ROS production, consequently enhancing their survival and migration under hypoxia-reoxygenation conditions. RESULTS:MiR-210 overexpression was significantly induced by optimized culture and preconditioning conditions, which also improved the production yield of exosomes from grown MSCs. The exosomes enriched with miR-210 demonstrated a protective effect by improving survival, reducing apoptosis and ROS accumulation in damaged renal cells, and ultimately promoting cell migration. CONCLUSION:The present study underscores the possibility of employing advanced techniques to maximize the therapeutic attributes of exosomes produced from WJ-MSC spheroid for improved recovery outcomes in ischemia-reperfusion injuries.
10.1186/s13287-024-03845-7
Hypoxic preconditioning BMSCs-exosomes inhibit cardiomyocyte apoptosis after acute myocardial infarction by upregulating microRNA-24.
Zhang C-S,Shao K,Liu C-W,Li C-J,Yu B-T
European review for medical and pharmacological sciences
OBJECTIVE:To elucidate the regulatory effect of hypoxic preconditioning bone marrow mesenchymal stem cells (BMSCs)-exosomes on cardiomyocyte apoptosis in acute myocardial infarction (AMI) rats. MATERIALS AND METHODS:BMSCs-derived exosomes were extracted by Exoquick method. Expressions of exosome surface markers were determined by Western blot. The AMI model in rats was established by LAD ligation. Rats were randomly assigned into sham group, AMI group, AMI+H-exo group and AMI+N-exo group. MicroRNA-24 expression in rat myocardium was detected at different time points. Subsequently, hypoxic preconditioning or normoxic preconditioning BMSCs-exosomes were intramyocardially injected into rats. Infarct size was calculated through TTC (triphenyltetrazolium chloride) staining. Cardiomyocyte apoptosis was accessed with Terminal Deoxynucleotidyl Transferase dUTP Nick-end Labeling (TUNEL). Heart function of AMI rats was evaluated by echocardiography. Protein expressions of apoptotic genes in rat myocardium were detected by Western blot. RESULTS:The mRNA level of microRNA-24 was higher in H-exo group than N-exo group. Injection of hypoxic preconditioning BMSCs-exosomes markedly upregulated microRNA-24 level, reduced infarct size and improved cardiac function in AMI rats. Protein expressions of Bax, caspase-3 and cleaved-caspase-3 were downregulated by BMSCs-exosomes treatment. H9c2 cells showed upregulated microRNA-24 level and decreased apoptotic rate after incubation with hypoxic preconditioning BMSCs-exosomes. The above cellular performances were partially reversed by transfection of microRNA-24 inhibitor. CONCLUSIONS:Hypoxic preconditioning BMSCs-exosomes inhibit cardiomyocyte apoptosis in AMI rats by upregulating microRNA-24.
10.26355/eurrev_201908_18560
Exosomes Secreted from Hypoxia-Preconditioned Mesenchymal Stem Cells Prevent Steroid-Induced Osteonecrosis of the Femoral Head by Promoting Angiogenesis in Rats.
BioMed research international
Recent studies have suggested that exosomes exert similar therapeutic effects to those of mesenchymal stem cells (MSCs) in regenerative medicine and MSCs-derived exosomes exhibit therapeutic effects on steroid-induced osteonecrosis of the femoral head (ONFH). Furthermore, reparative functions of exosomes from MSCs are enhanced by hypoxia treatment of the cells. However, there are no related reports about whether exosomes derived from hypoxia-preconditioned MSCs could show better therapeutic effects on steroid-induced ONFH. In vitro, we investigated the effects of hypoxia precondition on exosomes derived from bone marrow mesenchymal stem cells (BMMSCs) from rats and the proangiogenic ability of exosomes derived from hypoxia-preconditioned BMMSCs. In vivo, we investigated the role of exosomes from hypoxia-preconditioned BMMSCs on angiogenesis and protecting osteonecrosis in a rat ONFH model. We found that the potential of the proangiogenic ability of exosomes derived from hypoxia-preconditioned BMMSCs was higher than exosomes derived from BMMSCs cultured under normoxia. Exosomes derived from hypoxia-preconditioned BMMSCs significantly promoted proliferation, migration, vascular endothelial growth factor (VEGF) expression, and tube formation of human umbilical vein endothelial cells (HUVECs) compared with exosomes derived from BMMSCs cultured under normoxia. Administration of exosomes derived from hypoxia-preconditioned BMMSCs significantly prevented bone loss and increased vessel volume in the femoral head compared with exosomes derived from BMMSCs cultured under normoxia. Taken together, our data suggest that exosomes derived from hypoxia-preconditioned BMMSCs exert better therapeutic effects on steroid-induced ONFH by promoting angiogenesis and preventing bone loss.
10.1155/2021/6655225
Advances in application of hypoxia-preconditioned mesenchymal stem cell-derived exosomes.
Frontiers in cell and developmental biology
Mesenchymal stem cells (MSCs) primarily secrete physiologically functional exosomes via paracrine effects that act on various adjacent and distant cells, thus exerting their therapeutic effects. In recent years, hypoxic preconditioning, as a novel MSC culture mode, has emerged as a research hotspot. Many previous studies have shown the role and underlying regulatory mechanisms of hypoxic preconditioning in various diseases, which has provided sufficient reference materials for the MSC research field. Therefore, this review summarizes the progress in application of hypoxia-preconditioned MSC-derived exosomes that substantially increases and improves the biological activity of specific molecules, such as microRNA.
10.3389/fcell.2024.1446050