Shaohai Qi

Wnt and Notch signaling pathway involved in wound healing by targeting c-Myc and Hes1 separately

Wnt and Notch signaling pathways are critically involved in relative cell fate decisions within the development of cutaneous tissues. Moreover, several studies identified the above two pathways as having a significant role during wound healing. However, their biological effects during cutaneous tissues repair are unclear. We employed a self-controlled model (Sprague–Dawley rats with full-thickness skin wounds) to observe the action and effect of Wnt/β-catenin and Notch signalings in vivo. The quality of wound repair relevant to the gain/loss-of-function Wnt/β-catenin and Notch activation was estimated by hematoxylin-and-eosin and Masson staining. Immunofluorescence analysis and Western blot analysis were used to elucidate the underlying mechanism of the regulation of Wnt and Notch signaling pathways in wound healing. Meanwhile, epidermal stem cells (ESCs) were cultured in keratinocyte serum-free medium with Jaggedl or in DAPT (N-[(3,5-difluorophenyl)acetyl]-L-alanyl-2-phenyl]glycine-1,1-dimethylethyl) to investigate whether the interruption of Notch signaling contributes to the expression of Wnt/β-catenin signaling. The results showed that in vivo the gain-of-function Wnt/β-catenin and Notch activation extended the ability to promote wound closure. We further determined that activation or inhibition of Wnt signaling and Notch signaling can affect the proliferation of ESCs, the differentiation and migration of keratinocytes, and follicle regeneration by targeting c-Myc and Hes1, which ultimately lead to enhanced or delayed wound healing. Furthermore, Western blot analysis suggested that the two pathways might interact in vivo and in vitro. These results suggest that Wnt and Notch signalings play important roles in cutaneous repair by targeting c-Myc and Hes1 separately. What’s more, interaction between the above two pathways might act as a vital role in regulation of wound healing.

Granulocyte/macrophage colony-stimulating factor influences angiogenesis by regulating the coordinated expression of VEGF and the Ang/Tie system.

Granulocyte/macrophage colony-stimulating factor (GM-CSF) can accelerate wound healing by promoting angiogenesis. The biological effects of GM-CSF in angiogenesis and the corresponding underlying molecular mechanisms, including in the early stages of primitive endothelial tubule formation and the later stages of new vessel maturation, have only been partially clarified. This study aimed to investigate the effects of GM-CSF on angiogenesis and its regulatory mechanisms. Employing a self-controlled model (Sprague-Dawley rats with deep partial-thickness burn wounds), we determined that GM-CSF can increase VEGF expression and decrease the expression ratio of Ang-1/Ang-2 and the phosphorylation of Tie-2 in the early stages of the wound healing process, which promotes the degradation of the basement membrane and the proliferation of endothelial cells. At later stages of wound healing, GM-CSF can increase the expression ratio of Ang-1/Ang-2 and the phosphorylation of Tie-2 and maintain a high VEGF expression level. Consequently, pericyte coverages were higher, and the basement membrane became more integrated in new blood vessels, which enhanced the barrier function of blood vessels. In summary, we report here that increased angiogenesis is associated with GM-CSF treatment, and we indicate that VEGF and the Ang/Tie system may act as angiogenic mediators of the healing effect of GM-CSF on burn wounds.

Effects of asiaticoside on the expression of Smad protein by normal skin fibroblasts and hypertrophic scar fibroblasts

Background.  The precise mechanism of asiaticoside in molecular and gene expression levels of Smad protein and mRNA still remains unknown. We hypothesised that asiaticoside might inhibit the formation of hypertrophic scarring by affecting the expression of Smad protein and interfering with the Smad signalling pathway.

Expression of Smad Protein by Normal Skin Fibroblasts and Hypertrophic Scar Fibroblasts in Response to Transforming Growth Factor β1

BACKGROUND Smad proteins are important intracellular mediators of transforming growth factor (TGF)-β signaling. Little has been known about the specific relationship between TGF-β and TGF-β/Smad signaling in hypertrophic scars. OBJECTIVE The objective was to investigate the expression of Smads and the specific relationship between TGF-β and TGF-β/Smad signaling in hypertrophic scars. METHODS In this study, we initially determined the endogenous protein levels of Smad2 and Smad7 in hypertrophic scar fibroblast (HSFs) and normal skin fibroblast (NSFs). Second, we stimulated HSFs and NSFs with recombinant human TGF-β1 for 24 hours to determine whether the TGF-β1 could potentiate its effect by further stimulating the production of Smad by reverse transcription–polymerase chain reaction and Western blot analysis. RESULTS When compared with NSFs, the endogenous expression of Smad2 in HSFs was up-regulated and TGF-β1 could further stimulate the production of Smad2. Although the levels of Smad7 were similar between HSFs and NSFs, TGF-β1 up-regulated the expression of Smad7 for NSFs only, with no discernible effect on HSFs. These changes were paralleled by a significant increase in cytoplasm-to-nuclear translocation of Smad2. CONCLUSION These data substantiated the model of an autocrine positive loop in hypertrophic scars pathogenesis.

Epidermal stem cells (ESCs) accelerate diabetic wound healing via the Notch signalling pathway.

Epidermal stem cells (ESCs) accelerate diabetic wound healing via the Notch signalling pathway.

BMP‐7 attenuates TGF‐β1–induced fibroblast‐like differentiation of rat dermal papilla cells

Dermal papilla cells (DPCs) show phenotypic plasticity during wound healing. The multipotency of DPCs is well recognized, but the signaling pathways that regulate the differentiation of these cells into fibroblasts are poorly understood. A preliminary experiment showed that transforming growth factor beta1 (TGF‐β1) can induce DPCs to differentiate into fibroblast‐like cells, which suggests that DPCs may be a source of wound‐healing fibroblasts. Bone morphogenetic protein‐7 (BMP‐7), a member of the TGF‐β superfamily, can prevent and reverse fibrosis by counteracting the TGF‐β1–mediated profibrotic effect. To determine whether BMP‐7 attenuates the TGF‐β1–induced differentiation of DPCs into fibroblasts, we established an in vitro system for DPC differentiation and recorded the gene expression patterns that distinguished DPCs from fibroblasts. The proportion of fibroblast‐like cells was significantly enhanced in DPCs treated with TGF‐β1, as evidenced by immunocytochemistry, flow cytometry, quantitative real‐time reverse transcriptase polymerase chain reaction, and Western blot analysis. BMP‐7 and TGF‐β1 administration substantially decreased fibroblast‐like differentiation, indicating inhibition of TGF‐β1–induced differentiation. The antagonistic BMP‐7– and TGF‐β1–activated signaling pathways can be used to promote wound healing or suppress hypertrophic scarring.

Effects of Basic Fibroblast Growth Factors on Hypertrophic Scarring in a Rabbit Ear Model

Background: Basic fibroblast growth factor (bFGF) was clinically proven to accelerate acute and chronic wound healing. Accelerated wound healing may lead to improved scarring. These studies suggested a possible antiscarring effect of bFGF during wound healing. Little was known about the precise pathologic mechanisms of bFGF on scarring formation. Aims: The aim of this study was to investigate the effect of bFGF on hypertrophic scarring in a rabbit ear model and clarify the mechanism of bFGF on scar treatment. Methods: The rabbit model of hypertrophic scarring was created and received of a low- or high-dose topical treatment three times daily for 1, 2, or 3 months. Then we examined the changes in the macroscopic and histopathologic characteristics of the scars. The expression of collagen, α1β2 integrin, and matrix metalloproteinase 1 (MMP-1) was studied by applying reverse transcriptase-polymerase chain reaction, enzyme-linked immunosorbent assay, and Western blotting. Result: High-dose bFGF remarkably alleviated the scar in the rabbit ear model and decreased collagen type I expression. Further study revealed that bFGF remarkably enhanced MMP-1 and decreased α1β2 integrin expression. Conclusion: This study supports the hypothesis that bFGF exerted a net negative effect on collagen remodeling, therefore suggesting a potential antiscarring role.

Angiopoietin-1 protects the endothelial cells against advanced glycation end product injury by strengthening cell junctions and inhibiting cell apoptosis.

Endothelial dysfunction is a major characteristic of diabetic vasculopathy. Protection of the vascular endothelium is an essential aspect of preventing and treating diabetic vascular complications. Although Angiopoietin‐1 (Ang‐1) is an important endothelial‐specific protective factor, whether Ang‐1 protects vascular cells undergoing advanced glycation end product (AGE) injury has not been investigated. The aim of the present study was to determine the potential effects of Ang‐1 on endothelial cells after exposure to AGE. We show here that Ang‐1 prevented AGE‐induced vascular leakage by enhancing the adherens junctions between endothelial cells, and this process was mediated by the phosphorylation and membrane localization of VE‐cadherin. Furthermore, Ang‐1 also protected endothelial cells from AGE‐induced death by regulating phosphatidylinositol 3‐kinase (PI3K)/Akt‐dependent Bad phosphorylation. Our findings suggest that the novel protective mechanisms of Ang‐1 on endothelium are achieved by strengthening endothelial cell junctions and reducing endothelial cell death after AGE injury. J. Cell. Physiol. 230: 1895–1905, 2015.

Prostaglandin E2 inhibits collagen synthesis in dermal fibroblasts and prevents hypertrophic scar formation in vivo

Hypertrophic scarring is a common dermal fibroproliferative disorder characterized by excessive collagen deposition. Prostaglandin E2 (PGE2), an important inflammatory product synthesized via the arachidonic acid cascade, has been shown to act as a fibroblast modulator and to possess antifibroblastic activity. However, the mechanism underlying the antifibrotic effect of PGE2 remains unclear. In this study, we explored the effects of PGE2 on TGF‐β1‐treated dermal fibroblasts in terms of collagen production and to determine the regulatory pathways involved, as well as understand the antiscarring function of PGE2 in vivo. We found that PGE2 inhibited TGF‐β1‐induced collagen synthesis by regulating the balance of matrix metalloproteinases (MMPs) and tissue inhibitor of metalloproteinase (TIMP). It did so by upregulating cAMP through the E prostanoid (EP)2 receptor. We determined that inhibition of the TGF‐β1/Smad pathway by PGE2 is associated with its ability to inhibit collagen synthesis. An in vivo study further confirmed that PGE2 inhibits hypertrophic scar formation by decreasing collagen production. Our results demonstrate that the novel anti‐scarring function of PGE2 is achieved by balancing MMPs/TIMP expression and decreasing collagen production.

Notch1 Signaling Regulates Wound Healing via Changing theCharacteristics of Epidermal Stem Cells

During wound healing and reconstruction, epidermal stem cells (ESCs) migrate to the wound site and activate to repair the damaged epithelium. Moreover, there exist complicate signaling pathways to regulate wound regeneration including Notch signaling. The Notch signaling pathway is a regulator of epidermal differentiation, which may be an important mediator of wound regeneration that participates in various processes, from the development of the dermis to the formation of skin appendages. Here, we show that Notch signaling pathways are upregulated by Jagged1 in ESCs and stem cell characteristics of ESCs change when Notch1 signaling varies. By administration of siRNAJagged1 knockdown ESCs in wounds, we observe that the suppression of Jagged1 down regulate expression of Notch signaling and resulted in poor-quality wound healing. Connecting Notch1 pathways activity to ESCs response to wound repairing may develop a new therapeutic strategy for delayed healing.