Systemic sclerosis (SSc) is a multisystem connective tissue disease characterized by immune abnormalities, vasculopathy, and fibrosis of the skin and certain internal organs with unknown etiology. Although the established treatment for SSc does not exist, recent studies have demonstrated that bosentan, a dual endothelin receptor antagonist, prevents the development of new digital ulcers in patients with SSc. Furthermore, there are a couple of clinical reports suggesting the potential disease modifying effect of bosentan on SSc vasculopathy. Therefore, in this study, to elucidate the molecular mechanism underlying the effect of bosentan on SSc vasculopathy, we investigated the impact of bosentan on Fli1^+/- mice, which reproduce the histological and functional abnormalities of SSc vasculopathy. A series of experiments demonstrated the following findings: (i) endothelin-1 (ET-1) induces the phosphorylation of Fli1, which results in the decrease of Fli1 DNA binding ability and the promotion of Fli1 degradation through proteasome pathway, via “c-Abl-PKC-δ” pathway in human dermal microvascular endothelial cells (HDMECs), (ii) bosentan suppresses Fli1 phosphorylation, while increasing Fli1 DNA binding ability and Fli1 expression levels, in HDMECs by inhibiting the autocrine ET-1 stimulation, (iii) bosentan improves vasculopathy of Fli1^+/- mice at least partially by increasing the expression levels of Fli1 in dermal microvascular endothelial cells. Given that Fli1 deficiency due to epigenetic suppression is deeply associated with the developmental process of SSc, these results suggest that bosentan may exert a disease modifying effect on SSc vasculopathy at least partially by reversing the expression levels of Fli1 in dermal microvascular endothelial cells.Skin Research, Suppl. 22: 1-6, 2015

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We analyzed the S100A9 protein expression level during wound healing in mouse excisional wounds applied basic fibroblast growth factor (bFGF). We prepared 6mm, round, full-thickness wounds on the back skin of mice and applied bFGF (1.0μg/cm²) once daily for three days. The mRNA level of TNF-α and IL1β in granulation tissues extracted from the bFGF-applied group was determined higher than from the vehicle group by quantitative RT-PCR. The mRNA level of S100A9 extracted from the bFGF-applied group and vehicle group on day 5, day 7, and day 11 was expressed three times higher than in the vehicle group (p=0.0397). The S100A9 protein in granulation tissues from the bFGF-applied group was increased more than that from the vehicle group. We speculate that S100A9 protein plays an important role in wound healing, particularly in the inflammatory reaction.Skin Research, Suppl. 22: 7-12, 2015

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The secreted glycoprotein MFG-E8 has many functions including promoting phagocytosis of apoptotic cells, regulation of regulatory T-cells and angiogenesis. The possible role of MFG-E8 in cutaneous wound healing has not been studied previously. Herein, we analyzed skin wound healing using MFG-E8 wild-type (WT) and knockout (KO) mice. Wound healing in MFG-E8 KO mice was delayed compared with that in WT mice. In addition, administration of rMFG-E8 into wound corrected the delayed wound healing in MFG-E8 KO mice. At 7 days after wounding, the number of CD31⁺ endothelial cells and NG2⁺ pericytes in wound areas in MFG-E8 KO mice was decreased compared with that in WT mice. Furthermore, addition of rMFG-E8 into wound in db/db mice promoted wound healing and angiogenesis in granulation tissue compared with those in control mice. These findings suggest that MFG-E8 might positively regulate cutaneous wound healing during the tissue formation phase by enhancing angiogenesis. Furthermore, we found that exogenous application of rMFG-E8 enhanced wound healing in diabetic mice with similar extent of the application of bFGF. rMFG-E8 application also enhanced angiogenesis in wound area in diabetic mice, suggesting that exogenous MFG-E8 administration has possible therapeutic potential for diabetic skin ulcers.Skin Research, Suppl. 22: 13-18, 2015

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Ischemia-reperfusion (IR) is considered a key mechanism of pressure ulcer. The purpose of this study is to assess a role of inflammatory cells, cytokines, and nitric oxide synthase (NOS) in the ulcer formation process using a cutaneous IR mouse model. iNOS inhibition resulted in marked reduction of IR injury. Both IFN-γ-deficient and IL-6-deficient mice that received IR cycles showed improvement of tissue damage. In these mutant mice, reduced macrophage infiltration and increased expression of iNOS were observed. In vitro culture of macrophages from wild type mice stimulated with IFN-γ resulted in increased production of NO. The ratio of M1 macrophages were more dominant in early phase of IR cycles at the wound site, whereas M2 macrophages ratio was gradually increased as time passes. Our study indicates that IR injury is mediated by M1 and M2 macrophage infiltration that is induced by inflammatory cytokines including IFN-γ and IL-6, with subsequent increasing expression of iNOS and production of NO.Skin Research, Suppl. 22: 19-24, 2015

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Basic fibroblast growth factor (bFGF) stimulates proliferation of fibroblasts and endothelial cells. The one of main effect of bFGF is acceleration wound healing, forming granulation tissue with angiogenesis. On the other hand, there are some reports about synergistic effect of bFGF with other growth factor on angiogenesis. We summarize the combined effect of bFGF and report the synergistic effect of bFGF and prostaglandin E on mouse embryonic fibroblast.Skin Research, Suppl. 22: 25-27, 2015

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In Japan, treatment with basic fibroblast growth factor (bFGF) for wounds has been utilized to accelerate its healing process. Recently, some reports have revealed anti-scaring effect of bFGF on the cutaneous wounds. We examined the influences of bFGF to extracellular matrix in wound healing. Daily application of 0.01% bFGF to the wound shortened the period until wound closure. Wound treated with bFGF resulted in thicker and more contracted scar formation, which contained more amounts of thin collagen fibers. These results may imply scarless effect of bFGF.Skin Research, Suppl. 22: 28-33, 2015

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Endothelin-1 (ET-1) is known as a most potent vasoconstrictive peptide, which released mostly from endothelial cells. Diligent researches in this field indicate that ET-1 is involved in many important processes such as development, cancer biology, and autoimmune disorders and cardiovascular diseases. The aim of this study was to investigate whether the endothelial cell specific ET-1 would influence the wound healing using endothelial cell specific ET-1 knockout mice. We observed that the skin wound healing in endothelial cell specific ET-1 knockout mice was accelerated than in wild-type (WT) littermates. Investigations of the overall regulatory mechanisms of wound healing by ET-1 may lead to a new therapeutic approach for wound cure.Skin Research, Suppl. 22: 34-38, 2015

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