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Background: Systemic sclerosis (SSc) is a connective tissue disorder characterized by the development of fibrosis in the skin and internal organs. Increasing evidence suggests that mesenchymal stem cells (MSCs) can be used to a treatment for fibrotic diseases. Recent studies have demonstrated that some of the biological effects of MSCs are due to the secretion of exosomes. However, the precise mechanisms underlying MSCs-derived exosomes in skin fibrosis are not well understood., Objective: We aimed to elucidate the effect of MSCs-derived exosomes on skin fibrosis in SSc and the mechanism underlying their inhibitory action on fibrosis., Methods: Exosome was collected from MSCs by ultracentrifugation method. We examined the suppressive effect of MSCs-derived exosome on skin fibrosis in bleomycin-induced SSc mouse model. Skin samples from the injected site were collected for further examination, and micro-RNA analysis of MSCs-derived exosome was performed., Results: Injection of MSCs-derived exosomes significantly inhibited bleomycin-induced dermal fibrosis in mice. MSCs-derived exosomes significantly reduced the amount of collagen and the number of α-SMA + myofibroblasts and CD68 + macrophages in lesional skin. They also reduced the expression of type I collagen and TGF-β receptor 1 in fibroblasts in vitro. Moreover, micro-RNA analysis revealed that several microRNAs in MSCs-derived exosomes have antifibrotic potential. We confirmed that overexpression of miR-196b-5p in fibroblasts significantly suppressed collagen type I alpha 2 expression., Conclusion: This study demonstrated that inhibition of collagen type I expression by miR-196b-5p in exosomes might be one of the mechanisms by which MSCs suppress skin fibrosis in an SSc mouse model., Competing Interests: Declaration of Competing Interest The authors have no conflict of interest to declare., (Copyright © 2021 Japanese Society for Investigative Dermatology. Published by Elsevier B.V. All rights reserved.)

Oxidative stress has been reported to play an important role in the pathogenesis of skin fibrosis in systemic sclerosis (SSc). We previously identified that botulinum toxin (BTX) injection suppresses pressure ulcer formation in a cutaneous ischemia-reperfusion injury mouse model by regulation of oxidative stress. However, the therapeutic possibility of BTX administration for preventing skin fibrosis in SSc is unclear. The objective of this study was to investigate the effect of BTX-B on skin fibrosis in a murine model of SSc and determine the underlying mechanism. We found that BTX-B injection significantly reduced dermal thickness and inflammatory cell infiltration in bleomycin-induced skin fibrosis lesion in mice. We also identified that the oxidative stress signal detected through bioluminescence in OKD48 mice after bleomycin injection in the skin was significantly decreased by BTX-B. Additionally, mRNA levels of oxidative stress associated factors (NOX2, HO-1, Trx2) were significantly decreased by BTX-B. Apoptotic cells in the lesional skin of bleomycin-treated mice were significantly reduced by BTX-B. Oxidant-induced intracellular accumulation of reactive oxygen species in SSc fibroblasts was also inhibited by BTX-B. In conclusion, BTX-B might improve bleomycin-induced skin fibrosis via the suppression of oxidative stress and inflammatory cells in the skin. BTX-B injection may have a therapeutic effect on skin fibrosis in SSc., (© 2021 Japanese Dermatological Association.)

Tissue injury/hypoxia and oxidative stress induced-extracellular adenosine triphosphate (ATP) can act as damage-associated molecular pattern molecules, which initiate inflammatory response. Our objective was to elucidate the role of extracellular ATP in skin fibrosis in systemic sclerosis (SSc). We identified that hypoxia enhanced ATP release and that extracellular ATP enhanced IL-6 production more significantly in SSc fibroblasts than in normal fibroblasts. There were no significant differences of P2X and P2Y receptor expression levels between normal and SSc fibroblasts. Nonselective P2 receptor antagonist and selective P2Y 2 receptor antagonists, kaempferol and AR-C118925XX, significantly inhibited ATP-induced IL-6 production and phosphorylation of p38 in SSc fibroblasts. ATP-induced IL-6 production was significantly inhibited by p38 inhibitors, SB203580, and doramapimod. Collagen type I production in SSc fibroblasts by ATP-induced IL-6/IL-6 receptor trans-signaling was inhibited by kaempferol and SB203580. The amount of ATP in bleomycin-treated skin was increased, and administration of AR-C118925XX significantly inhibited bleomycin-induced dermal fibrosis in mice. These results suggest that vasculopathy-induced hypoxia and oxidative stress might enhance ATP release in the dermis in SSc and that extracellular ATP-induced phosphorylation of p38 via P2Y 2 receptor might enhance IL-6 and collagen type I production in SSc fibroblasts. P2Y 2 receptor antagonist therapy could be a treatment for skin sclerosis in patients with SSc., (Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.)

Objective: Several studies have demonstrated that the secreted glycoprotein and integrin ligand milk fat globule-associated protein with epidermal growth factor- and factor VIII-like domains (MFG-E8) negatively regulates fibrosis in the liver, lungs, and respiratory tract. However, the mechanisms and roles of MFG-E8 in skin fibrosis in systemic sclerosis (SSc) have not been characterized. We undertook this study to elucidate the role of MFG-E8 in skin fibrosis in SSc., Methods: We assessed expression of MFG-E8 in the skin and serum in SSc patients. We examined the effect of recombinant MFG-E8 (rMFG-E8) on latent transforming growth factor β (TGFβ)-induced gene/protein expression in SSc fibroblasts. We examined the effects of deficiency or administration of MFG-E8 on fibrosis mouse models., Results: We demonstrated that MFG-E8 expression around dermal blood vessels and the serum MFG-E8 level in SSc patients (n = 7 and n = 44, respectively) were lower than those in healthy individuals (n = 6 and n = 28, respectively). Treatment with rMFG-E8 significantly inhibited latent TGFβ-induced expression of type I collagen, α-smooth muscle actin, and CCN2 in SSc fibroblasts (n = 3-8), which suggested that MFG-E8 inhibited activation of latent TGFβ as well as TGFβ signaling via binding to αv integrin. In a mouse model of bleomycin-induced fibrosis (n = 5-8) and in a TSK mouse model (a genetic model of SSc) (n = 5-10), deficient expression of MFG-E8 significantly enhanced both pulmonary and skin fibrosis, and administration of rMFG-E8 significantly inhibited bleomycin-induced dermal fibrosis., Conclusion: These results suggest that vasculopathy-induced dysfunction of pericytes and endothelial cells, the main cells secreting MFG-E8, may be associated with the decreased expression of MFG-E8 in SSc and that the deficient inhibitory regulation of latent TGFβ-induced skin fibrosis by MFG-E8 may be involved in the pathogenesis of SSc and may be a therapeutic target for fibrosis in SSc patients., (© 2018, American College of Rheumatology.)

Objective: Apelin/APJ signaling has been determined to regulate cardiac and arterial fibrosis and to be involved in the pathogenesis of pulmonary arterial hypertension. Our objective was to elucidate the role of apelin in skin fibrosis in systemic sclerosis (SSc)., Methods: Expression of apelin/APJ in normal and SSc fibroblasts was compared. Effects of small interfering RNA depletion and the addition of apelin in fibroblasts were analyzed. The effect of apelin injections on bleomycin-induced dermal fibrosis in mice was investigated. We analyzed the effects of the biased agonist of APJ, MM07, on skin fibrosis in vitro and in vivo., Results: The expression of apelin in SSc fibroblasts was significantly lower than that in normal fibroblasts. Serum apelin levels were negatively correlated with the modified Rodnan skin thickness score in SSc patients. Stimulation with transforming growth factor β1 (TGFβ1) inhibited apelin expression in fibroblasts, suggesting that activation of TGFβ1 signaling in SSc might be responsible for reduced apelin expression in SSc fibroblasts. Small interfering RNA depletion of apelin from fibroblasts significantly enhanced fibrosis-related gene expression, and treatment with apelin protein significantly inhibited TGFβ1 signaling in fibroblasts. Administration of apelin significantly inhibited bleomycin-induced dermal fibrosis in mice. We demonstrated that MM07 had greater potential than apelin to inhibit fibrosis in vivo and in vitro., Conclusion: Collectively, TGFβ1 signaling and apelin signaling may counteract each other in the fibrotic process of SSc. Inhibitory regulation of TGFβ1-induced skin fibrosis by apelin/APJ signaling may be involved in the pathogenesis of SSc and could be a therapeutic target for fibrosis in SSc patients., (© 2018, American College of Rheumatology.)