Rebamipide-loaded chitosan nanoparticles accelerate prostatic wound healing by inhibiting M1 macrophage-mediated inflammation via the NF-κB signaling pathway.
Sun Menghao,Deng Zheng,Shi Fei,Zhou Zheng,Jiang Chenyi,Xu Zhilu,Cui Xiaoming,Li Wentong,Jing Yifeng,Han Bangmin,Zhang Weifen,Xia Shujie
A large proportion of benign prostatic hyperplasia (BPH) patients suffer from lower urinary tract symptoms after surgery due to the presence of prostatic urothelium wounds. Rebamipide (RBM) exerts wound healing promotion and anti-inflammatory effects on various tissues, including the urothelium. However, intravesical administration of RBM is hindered due to its low solubility and resulting unsustainable drug concentrations in the bladder. In this study, RBM-loaded chitosan nanoparticles (RBM/CTS NPs) were prepared using the ionic cross-linking method. Physicochemical characteristics and the wound healing promotion effect, as well as in vitro influence on macrophages were evaluated. The results show that RBM/CTS NPs are spherical with uniform size distribution, while slower and sustained in vitro release of RBM is presented. In vivo, faster wound healing and improved re-epithelialization progress were observed after treatment with RBM/CTS NPs in a model of thulium laser resection of the prostate (TmLRP). The degree of local inflammatory response decreased, as confirmed by decreasing numbers of pro-inflammatory M1 phenotype macrophages and levels of IL-1β, IL-6, IL-12 and TNF-α in the urine of canines. We also found that RBM/CTS NPs suppress macrophage M1 polarization induced by lipopolysaccharide and interferon-γ and inhibit the activation of the NF-κB signaling pathway. Therefore, as a novel therapeutic strategy, intravesical administration of RBM/CTS NPs can effectively avoid drug intolerance and drug wastage, accelerating the postoperative wound repairing of the prostatic urethra by suppressing macrophage M1 phenotype polarization.
Keratinocyte autophagy enables the activation of keratinocytes and fibroblastsand facilitates wound healing.
Qiang Lei,Yang Seungwon,Cui Yan-Hong,He Yu-Ying
Macroautophagy/autophagy is a cellular catabolic process that is implicated in several physiological and pathological processes. However, the role of epidermal autophagy in wound healing remains unknown. Here, using mice with genetic ablation of the essential (autophagy related 5) or (autophagy related 7) in their epidermis to inhibit autophagy, we show that keratinocyte autophagy regulates wound healing in mice. Wounding induces the expression of autophagy genes in mouse skin. Epidermis-specific autophagy deficiency inhibits wound closure, re-epithelialization, keratinocyte proliferation and differentiation, dermal granulation tissue formation, and infiltration of immune cells including macrophages, neutrophils, and mast cells, while it does not affect angiogenesis. Using cytokine array screening, we found that autophagy deficiency inhibits the transcription and production of the cytokine CCL2/MCP-1 by TNF. At the molecular level, TNF induces autophagic flux and the expression of autophagy genes through NFKB in epidermal keratinocytes. TNF promotes transcription through the autophagy-AMPK-BRAF-MAPK1/3/ERK-activator protein 1 (AP1) pathway. Indeed, treating mice with recombinant CCL2 can reverse the effect of autophagy deficiency in keratinocytes. At the cellular level, we found that induction via autophagy in keratinocytes is required not only for keratinocyte migration and proliferation but also for dermal fibroblast activation. Our findings demonstrate a critical role of epidermal autophagy in wound healing and elucidate a critical molecular machinery coordinating keratinocyte-fibroblast interaction in skin repair.: ACTA2/α-SMA: actin alpha 2, smooth muscle; ACTB: β-actin; ADGRE1: adhesion G protein-coupled receptor E1; AMPK: AMP-activated protein kinase; AP1: activator protein 1; AP1-RE: AP1 response element; ATG: autophagy-related; ATG16L1: autophagy related 16 like 1; BECN1: beclin 1; BRAF: B-Raf proto-oncogene, serine/threonine kinase; C5: complement C5; CCL2/MCP-1: C-C motif chemokine ligand 2; CCL3: C-C motif chemokine ligand 3; CK: cytokeratin; cKO: conditional knockout; CRTC1: CREB-regulated transcription coactivator 1; CXCL1: C-X-C motif chemokine ligand 1; CXCL2: C-X-C motif chemokine ligand 2; ECM: extracellular matrix; EGF: epidermal growth factor; FGF7: fibroblast growth factor 7; GABARAPL2: GABA type A receptor associated protein like 2; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; HBEGF: heparin binding EGF like growth factor; HPRT1: hypoxanthine phosphoribosyltransferase 1; IHC: immunohistochemical; IL1B: interleukin 1 beta; KRT10: keratin 10; KRT14: keratin 14; MAP1LC3B/LC3B-I/II: microtubule-associated protein 1 light chain 3 beta; MAPK1/3/ERK: mitogen-activated protein kinase 1/3; MKI67/Ki-67: marker of proliferation; MPO: myeloperoxidase; NFKB: NF-kappa B, nuclear factor kappa-light-chain-enhancer of activated B cells; NFKB-RE: NFKB response element; PDGF: platelet-derived growth factor; PECAM1: platelet and endothelial cell adhesion molecule 1; PRKAA1: protein kinase AMP-activated catalytic subunit alpha 1; RELA/p65: RELA proto-oncogene, NFKB subunit; shCON: small hairpin ; siNC: negative control; siRNA: small interfering RNA; SP1: sp1 transcription factor; SQSTM1/p62: sequestosome 1; TGFA: transforming growth factor alpha; TGFB1: transforming growth factor beta 1; TIMP1: TIMP metallopeptidase inhibitor 1; TNF/TNF-alpha: tumor necrosis factor; TREM1: triggering receptor expressed on myeloid cells 1; WT: wild-type.
MicroRNA-146a Deficiency Delays Wound Healing in Normal and Diabetic Mice.
Bi Xinling,Zhou Li,Liu Yanfang,Gu Jun,Mi Qing-Sheng
Advances in wound care
MiRNAs are important regulators of inflammation and wound healing. However, the mechanisms through which miRNAs regulate wound healing under normal and diabetic conditions are poorly understood. We aimed to determine the effects of miR-146a on the pathogenesis of wound healing in normal and streptozotocin (STZ)-induced diabetic mice. Wild-type (WT) and miR-146a knockout (KO) mice were induced to develop diabetes with STZ. Next, skin and corneal wounds were produced and measured. Percent wound closure and histology were evaluated. Inflammation at wound sites was analyzed using flow cytometry, reverse-transcription PCR, and western blot. Healing of wounded skin was significantly delayed in miR-146a KO compared with WT mice. However, corneal epithelial wound healing did not differ significantly in the mice with normal blood glucose, whereas corneal and skin wound healing was significantly delayed in KO mice with diabetes. Neutrophil infiltration increased in skin wounds of KO compared with normal mice. The potential mechanisms were associated with dysregulated interleukin 1β, tumor necrosis factor alpha (TNF-α), IRAK1 (interleukin-1 receptor-associated kinase 1), TRAF6 (TNF receptor-associated factor 6), and nuclear factor kappa B (NF-κB) signaling induced by miR-146a KO. Skin wound healing was delayed in miR-146a KO mice and enhanced inflammatory responses were mediated by the NF-κB signaling pathway. Deficiency in miR-146a delayed skin wound healing by enhancing inflammatory responses in normal and diabetic mice. Therefore, miR-146a may be a potential target for modulation to accelerate skin wound healing.
Brazilian red propolis improves cutaneous wound healing suppressing inflammation-associated transcription factor NFκB.
Corrêa Flavia Regina Sobreira,Schanuel Fernanda Seabra,Moura-Nunes Nathalia,Monte-Alto-Costa Andréa,Daleprane Julio Beltrame
Biomedicine & pharmacotherapy = Biomedecine & pharmacotherapie
The use of natural products in wound healing has been extensively studied in the context of complementary and alternative medicine. Propolis, a natural product, is a polyphenol-rich resin used for this purpose. This study aimed to investigate the effect of Brazilian Red Propolis Extract (BRPE) on inflammation and wound healing in mice, using a tissue repair model. The BRPE polyphenol content was analyzed by liquid chromatography coupled to mass spectrometry (LC/MS). A full-thickness excision lesion was created, and mice were treated orally with daily doses of vehicle solution (water-alcohol solution containing 2% of ethanol, control group) or 100mg/kg of BRPE (P100 group) during nine consecutive days. BRPE chemical composition analysis showed that this complex matrix contains several phenolic compounds such as phenolic acids, phenolic terpenes and flavonoids (especially catechins, flavonols, chalcones, isoflavones, isoflavans, pterocarpans and bioflavonoids). After BRPE administration, it was observed that, when compared to the control group, P100 group presented faster wound closure (p<0.001); less neutrophils per mm (p<0.05) and macrophages (p<0.01) in tissue analyses, down regulation of the inflammatory transcription factor pNF-κB protein expression, and reduced production of inflammatory cytokine, such as TGF-β, TNF-α (p<0.0001), and IL-6 (p<0.001). These findings suggest a positive role of BRPE oral administration in the wound healing process via suppressing the inflammatory response during tissue repair.
Models for the study of skin wound healing. The role of Nrf2 and NF-κB.
Ambrozova Nikola,Ulrichova Jitka,Galandakova Adela
Biomedical papers of the Medical Faculty of the University Palacky, Olomouc, Czechoslovakia
Nrf2 and NF-κB transcription factors act in wound healing via their anti-inflammatory and anti-oxidant effects or through the immune response. Studying this process is a matter of some importance given the high cost of wound treatment. A major contribution in this regard is being made by models that enable investigation of the involvement of multiple factors in wound healing and testing new curative substances. This literature review was carried out via searches in the PubMed and Web of Science databases up to 2016. It covers skin wound healing, available models for its study (part I), the role of Nrf2 and NF-κB, substances that influence them and whether they can be used as markers (part II). Was found that in vitro assays are used for their availability but a holistic view must be established in vivo. In silico approaches are facilitating assessment of a vast amount of research data. Nfr2 and NF-κB play a crucial and reciprocal role in wound healing. Nrf2 controls repair-associated inflammation and protects against excessive accumulation of ROS while Nf-κB activates the innate immune reaction, proliferation and migration of cells, modulates expression of matrix metalloproteinases, secretion and stability of cytokines and growth factors for wound healing.
DPSC Products Accelerate Wound Healing in Diabetic Mice through Induction of SMAD Molecules.
Despite advances in diabetic wound care, many amputations are still needed each year due to their diabetic wounds, so a more effective therapy is warranted. Herein, we show that the dental pulp-derived stem cell (DPSC) products are effective in wound healing in diabetic NOD/SCID mice. Our results showed that the topical application of DPSC secretory products accelerated wound closure by inducing faster re-epithelialization, angiogenesis, and recellularization. In addition, the number of neutrophils producing myeloperoxidase, which mediates persisting inflammation, was also reduced. NFκB and its downstream effector molecules like IL-6 cause sustained pro-inflammatory activity and were reduced after the application of DPSC products in the experimental wounds. Moreover, the DPSC products also inhibited the activation of NFκB, and its translocation to the nucleus, by which it initiates the inflammation. Furthermore, the levels of TGF-β, and IL-10, potent anti-inflammatory molecules, were also increased after the addition of DPSC products. Mechanistically, we showed that this wound-healing process was mediated by the upregulation and activation of Smad 1 and 2 molecules. In sum, we have defined the cellular and molecular mechanisms by which DPSC products accelerated diabetic wound closure, which can be used to treat diabetic wounds in the near future.
Silk protein/polyvinylpyrrolidone nanofiber membranes loaded with puerarin accelerate wound healing in mice by reducing the inflammatory response.
In modern clinical applications, wound healing remains a considerable challenge. Excessive inflammatory response is associated with delayed wound healing. In this study, we prepared composite nanofibrous membranes by mixing the Chinese herbal extract puerarin (PUE) with natural silk protein (SF) and synthetic polymer polyvinylpyrrolidone (PVP) using electrostatic spinning technique, and conducted a series of studies on the structural and biological properties of the fibrous membranes. The results showed that the loading of PUE increased the diameter, porosity and hydrophilicity of nanofibers, which were more favorable for cell adhesion and proliferation. ABTS radical scavenging assay also showed that the loading of PUE enhanced the antioxidant properties of the fibrous membranes. In addition, SF/PVP/PUE nanofibers are non-toxic and can be used as wound dressings. In vitro experiments showed that SF/PVP/PUE nanofibers could effectively alleviate lipopolysaccharide (LPS)-induced inflammation in Immortalized human keratinocytes (HaCaT) cells and down-regulate pro-inflammatory cytokine expression in cells. In vivo studies further showed that the SF/PVP/PUE nanofibers could effectively accelerate wound repair. The mechanism is that SF/PVP/PUE nanofibers can inhibit the activation and transduction of toll-like receptor 4/myeloid differentiation factor88/nuclear factor kappa B (TLR4/MyD88/NF-κB) and phosphatidylinositol 3-kinase/protein kinase B (PI3K/AKT) signaling pathways, thereby reducing the inflammatory response and achieving wound healing.
Epigenetic regulation of the PGE2 pathway modulates macrophage phenotype in normal and pathologic wound repair.
Davis Frank M,Tsoi Lam C,Wasikowski Rachael,denDekker Aaron,Joshi Amrita,Wilke Carol,Deng Hongping,Wolf Sonya,Obi Andrea,Huang Steven,Billi Allison C,Robinson Scott,Lipinski Jay,Melvin William J,Audu Christopher O,Weidinger Stephan,Kunkel Steven L,Smith Andrew,Gudjonsson Johann E,Moore Bethany B,Gallagher Katherine A
Macrophages are a primary immune cell involved in inflammation, and their cell plasticity allows for transition from an inflammatory to a reparative phenotype and is critical for normal tissue repair following injury. Evidence suggests that epigenetic alterations play a critical role in establishing macrophage phenotype and function during normal and pathologic wound repair. Here, we find in human and murine wound macrophages that cyclooxygenase 2/prostaglandin E2 (COX-2/PGE2) is elevated in diabetes and regulates downstream macrophage-mediated inflammation and host defense. Using single-cell RNA sequencing of human wound tissue, we identify increased NF-κB-mediated inflammation in diabetic wounds and show increased COX-2/PGE2 in diabetic macrophages. Further, we identify that COX-2/PGE2 production in wound macrophages requires epigenetic regulation of 2 key enzymes in the cytosolic phospholipase A2/COX-2/PGE2 (cPLA2/COX-2/PGE2) pathway. We demonstrate that TGF-β-induced miRNA29b increases COX-2/PGE2 production via inhibition of DNA methyltransferase 3b-mediated hypermethylation of the Cox-2 promoter. Further, we find mixed-lineage leukemia 1 (MLL1) upregulates cPLA2 expression and drives COX-2/PGE2. Inhibition of the COX-2/PGE2 pathway genetically (Cox2fl/fl Lyz2Cre+) or with a macrophage-specific nanotherapy targeting COX-2 in tissue macrophages reverses the inflammatory macrophage phenotype and improves diabetic tissue repair. Our results indicate the epigenetically regulated PGE2 pathway controls wound macrophage function, and cell-targeted manipulation of this pathway is feasible to improve diabetic wound repair.
MDL-800, the SIRT6 Activator, Suppresses Inflammation via the NF-B Pathway and Promotes Angiogenesis to Accelerate Cutaneous Wound Healing in Mice.
Oxidative medicine and cellular longevity
Sirtuin 6 (SIRT6) is an NAD-dependent deacetylase belonging to the sirtuin family. It has been shown to participate in wound healing and some inflammation-related disorders. However, the effect of MDL-800, a highly efficient and selective SIRT6 activator, on wound healing and inflammation has not been reported. Therefore, this study investigated whether MDL-800 confers anti-inflammatory effects and promotes wound healing and uncovered the molecular mechanisms involved. This was achieved using mouse models of full-thickness wounds. Results showed that MDL-800 significantly downregulated inflammation by attenuating the release of inflammatory mediators and improved collagen deposition and neovascularization of wounds, thereby accelerating cutaneous wound healing. Furthermore, MDL-800 significantly downregulated expression levels of TNF- and IL-6 in the dorsal skin tissue of mice via the NF-B pathway. These results demonstrated that MDL-800 exerted anti-inflammatory and prohealing effects, indicating that the SIRT6/NF-B/IB signaling pathway may play an important role in wound healing.
From Inflammation to Cutaneous Repair: Topical Application of Lupeol Improves Skin Wound Healing in Rats by Modulating the Cytokine Levels, NF-κB, Ki-67, Growth Factor Expression, and Distribution of Collagen Fibers.
Pereira Beserra Fernando,Sérgio Gushiken Lucas Fernando,Vieira Ana Júlia,Augusto Bérgamo Danilo,Luísa Bérgamo Patrícia,Oliveira de Souza Mariana,Alberto Hussni Carlos,Kiomi Takahira Regina,Henrique Nóbrega Rafael,Monteiro Martinez Emanuel Ricardo,John Jackson Christopher,Lemos de Azevedo Maia Gabriela,Leite Rozza Ariane,Helena Pellizzon Cláudia
International journal of molecular sciences
Skin wound healing is a highly complex event that involves different mediators at the cellular and molecular level. Lupeol has been reported to possess different biological activities, such as anti-inflammatory, antioxidant, antidiabetic, and in vitro wound healing properties, which motivated us to proceed with in vivo studies. We aimed to investigate the wound healing effect of lupeol-based cream for 3, 7, and 14 days. Wound excisions were induced on the thoraco-lumbar region of rats and topically treated immediately after injury induction. Macroscopic, histopathological, and immunohistochemical analyses were performed. Cytokine levels were measured by ELISA and gene expression was evaluated by real-time RT-qPCR. Our results showed a strong wound-healing effect of lupeol-based cream after 7 and 14 days. Lupeol treatment caused a reduction in proinflammatory cytokines (TNF-a, IL-1β, and IL-6) and gene and protein NF-κB expression, and positively altered IL-10 levels, showing anti-inflammatory effects in the three treatment periods. Lupeol treatment showed involvement in the proliferative phase by stimulating the formation of new blood vessels, increasing the immunostaining of Ki-67 and gene expression, and immunolabeling of vascular endothelial growth factor (VEGF) and epidermal growth factor (EGF), and increasing gene expression of transforming growth factor beta-1 (TGF-β1) after seven days of treatment. Lupeol was also involved in the tissue regeneration phase by increasing the synthesis of collagen fibers noted in the three treatment periods analyzed. Our findings suggest that lupeol may serve as a novel therapeutic option to treat cutaneous wounds by regulating mechanisms involved in the inflammatory, proliferative, and tissue-remodeling phases.
Overexpression of S100A9 in obesity impairs macrophage differentiation via TLR4-NFkB-signaling worsening inflammation and wound healing.
Franz Sandra,Ertel Anastasia,Engel Kathrin M,Simon Jan C,Saalbach Anja
: In obesity the fine-tuned balance of macrophage phenotypes is disturbed towards a dominance of pro-inflammatory macrophages resulting in exacerbation and persistence of inflammation and impaired tissue repair. However, the underlying mechanisms are still poorly understood. : Impact of obesity on macrophage differentiation was studied in high fat diet induced obese and db/db mice during skin inflammation and wound repair, respectively. Mechanisms of S100A9-mediated effects on macrophage differentiation was studied on generated macrophages by genomic and proteomic approaches. The role of S100A9 on macrophage differentiation was investigated by pharmacological inhibition of S100A9 during skin inflammation and wound repair in obese and db/db mice. : We demonstrate an overexpression of S100A9 in conditions of obesity-associated disturbed macrophage differentiation in the skin. We show that saturated free fatty acids (SFA), which are increased in obesity, together with S100A9 induce TLR4 and inflammasome-dependent IL-1β release in macrophages which in turn amplifies S100A9 expression initiating a vicious cycle of sustained S100A9 overexpression in skin inflammation in obesity. We reveal a yet unrecognized impact of obesity-associated S100A9 overexpression on macrophage differentiation. S100A9 binding to TLR4 and activation of NFkB attenuates development of M2-like macrophages and induces pro-inflammatory functions in these cells. Consequently, inhibition of S100A9 restores disturbed M2-like macrophage differentiation in mouse models of obesity-associated skin inflammation and wound repair. Similarly, breaking the vicious cycle of S100A9 overexpression by dietary reduction of SFA restored M2-like macrophage activation. Improvement of skin inflammation and wound repair upon reduction of S100A9 by pharmacological inhibition or by reduction of SFA uncovers the pathogenic role of S100A9 overexpression in obesity. : This study identifies S100A9 as a previously unrecognized vital component in obesity-associated disturbed macrophage differentiation and subsequent impaired regulation of inflammation and wound repair. The findings open new opportunities for therapeutic implications for inflammatory diseases and wound repair in obesity.
Whey Derivatives and Galactooligosaccharides Stimulate the Wound Healing and the Function of Human Keratinocytes through the NF-kB and FOXO-1 Signaling Pathways.
Skin repair requires the activation of keratinocytes and is mediated by controlled inflammation and cell migration and proliferation, ending with the regeneration of well-differentiated cell layers. Whey derivatives contain galactooligosaccharides (GOS), which have potential beneficial effects on wound healing due to their activity as toll-like receptor ligands, although their direct nonprebiotic effects in the skin have not yet been described. In this study, we investigated the effects of different whey-derived products and purified GOS on a human keratinocyte cell line. We found that the inflammatory cytokine interleukin-8 (IL-8) was upregulated by nuclear factor kappa B (NF-kB) signaling triggered by whey derivatives and GOS and that wound healing was accelerated by promoting cell migration and the loss of E-cadherin in the absence of epithelial-mesenchymal transition. Interestingly, the treatments enhanced the mitochondrial function in association with the translocation of the Forkhead Box O1 (FOXO-1) transcription factor. Finally, we detected the increased expression of the differentiation markers induced by GOS and whey derivatives. All together, our results show that GOS-containing products can promote wound closure and skin health by direct activity on keratinocyte functions. Among the preparations tested, the fermented compound produced by autochthonous microorganisms was the most active in modulating keratinocyte activity, supporting the biological value of whey derivatives for health.