■肥厚性瘢痕(HS)是一种常见的纤维化皮肤病,继发于烧伤或受伤。TGF-β信号通路的激活极大地促进了HS的形成。异鼠李素(ISO)是一种类黄酮化合物,其通过TGF-β信号传导抑制发挥抗纤维化作用。然而,ISO是否可以通过TGF-β信号抑制HS形成尚待阐明。本研究旨在探讨ISO对HS发病机制和TGF-β信号通路的影响,特别是促进HS形成的TGF-β信号的下游分子和网络。
■从人HS组织中分离出肥厚性瘢痕成纤维细胞(HSFB)。体外增殖,迁移,收缩能力,细胞周期,ISO处理后HSFB的凋亡采用细胞活力测定法进行测定,EdU染色,伤口愈合试验,胶原蛋白凝胶收缩试验,和流式细胞术。使用定量PCR(qPCR)确定ISO处理的HSFB中TGF-β信号传导相关基因和蛋白及其下游分子的表达,免疫荧光,和西方印迹。在体内,建立了兔HS模型,使用组织学分析研究了ISO对兔HS形成的影响,免疫组织化学染色,和qPCR。
■体外研究表明,ISO处理抑制了增殖,迁移,和HSFB的收缩能力;减弱COLⅠ的表达,COLⅢ,和α-SMA;并通过以剂量依赖性方式降低磷酸化Smad2/3和裂解的CREB3L1的水平来抑制TGF-β1信号诱导的HSFB活化。此外,ISO通过上调促凋亡蛋白Bax和裂解的caspase-3的表达并下调抗凋亡蛋白Bcl-2的表达来增强HSFB的凋亡和G2期细胞周期阻滞。体内研究表明,ISO通过降低瘢痕升高指数改善了兔耳中HS的形成,减弱胶原蛋白密度,促进胶原纤维的规则排列,下调TGF-β1、CREB3L1、COLⅠ、COLⅢ,和α-SMA。
■ISO通过抑制TGF-β1/Smad和TGF-β1/CREB3L1信号通路抑制HS发病机制,这表明它可以作为TGF-β1信号传导的候选抑制剂和具有高治疗潜力的有前途的抗HS药物。
UNASSIGNED: Hypertrophic scar (HS) is a common fibrotic skin disease that occurs secondary to burns or injuries. The activation of the TGF-β signaling pathway contributes immensely to HS formation.
Isorhamnetin (ISO) is a type of flavonoid compound that exerts an antifibrotic effect via TGF-β signaling suppression. However, whether ISO can inhibit HS formation via TGF-β signaling is yet to be elucidated. This study aimed to examine the influence of ISO on HS pathogenesis and TGF-β signaling, especially the downstream molecules and networks of TGF-β signaling that facilitate HS formation.
UNASSIGNED: Hypertrophic scar fibroblasts (HSFBs) were isolated from human HS tissues. The in vitro proliferation, migration, contractile ability, cell cycle, and apoptosis of HSFBs after ISO treatment were determined using cell viability assay, EdU staining, wound healing assay, collagen gel contraction assay, and flow cytometry. The expressions of genes and proteins involved in TGF-β signaling and its downstream molecules in ISO-treated HSFBs were determined using quantitative PCR (qPCR), immunofluorescence, and western blotting. In vivo, a rabbit HS model was established, and the effects of ISO on rabbit HS formation were investigated using histological analysis, immunohistochemical staining, and qPCR.
UNASSIGNED: In vitro studies indicated that ISO treatment suppressed the proliferation, migration, and contractile ability of HSFBs; attenuated the expressions of COL Ⅰ, COL Ⅲ, and α-SMA; and inhibited TGF-β1 signaling-induced activation of HSFBs by decreasing the levels of phosphorylated Smad2/3 and cleaved CREB3L1 in a dose-dependent manner. Furthermore, ISO augmented apoptosis and G2 phase cell cycle arrest of HSFBs by upregulating the expressions of the proapoptotic proteins Bax and cleaved caspase-3 and downregulating the expression of the antiapoptotic protein Bcl-2. In vivo studies revealed that ISO ameliorated HS formation in the rabbit ear by lowering the scar elevation index, attenuating the collagen density, facilitating the regular arrangement of collagen fibers, and downregulating the expressions of TGF-β1, CREB3L1, COL Ⅰ, COL Ⅲ, and α-SMA.
UNASSIGNED: ISO suppressed HS pathogenesis by dampening TGF-β1/Smad and TGF-β1/CREB3L1 signaling pathways, which suggests that it may serve as a candidate inhibitor of TGF-β1 signaling and a promising anti-HS drug with a high therapeutic potential.