RhoA/ROCK Signaling Regulates TGF-beta 1-Induced Fibrotic Effects in Human Pterygium Fibroblasts through MRTF-A
RhoA/ROCK Signaling Regulates TGF-beta 1-Induced Fibrotic Effects in Human Pterygium Fibroblasts through MRTF-A
复制标题
RhoA/ROCK 信号通过 MRTF-A 调节 TGF-β 1 诱导的人翼状胬肉成纤维细胞纤维化效应
DOI：
10.1080/02713683.2021.1962363
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发表时间：
2021
影响因子：
2
通讯作者：
Ye Juan
中科院分区：
文献类型：
--
作者：
Xie Jiajun;Ning Qingyao;Zhang Huina;Ni Shuang;Ye Juan
目的：翼状胬肉手术切除后TGF-β1的过度表达常导致纤维化，提示翼状胬肉复发。本体外研究的目的是探讨RhoA/ROCK信号在调节原代人翼状胬肉成纤维细胞（HPFs）纤维化效应中的作用，以及探讨这些效应的可能机制。CCK-8法检测细胞增殖情况;伤口愈合法检测细胞迁移情况;实时定量PCR和Western blot检测TGF-β1对HPFs α-平滑肌肌动蛋白（a-SMA）、Ⅰ型和Ⅲ型胶原（COL 1和COL 3）、基质金属蛋白酶9（MMP 9）合成的影响。Western blot检测细胞内信号通路的变化;结果ROCK抑制剂Y-27632可抑制TGF-β1诱导的细胞增殖和迁移，降低TGF-β 1诱导的HPFs促纤维化标志物的表达，并抑制TGF-β1诱导的肌红蛋白相关转录因子A（MRTF-A）的核内积聚以及伴随的F/G-actin比值升高。结论RhoA/ROCK信号通路在TGF-β 1诱导的HPFs纤维化和肌成纤维细胞活化中起关键作用，其机制至少部分是通过抑制下游MRTF-A/SRF转录通路而实现的。
PurposeThe overexpression of transforming growth factor-beta1 (TGF-β1) after surgical excision often leads to excessive fibrosis, indicating the recurrence of pterygium. The aims of the present in vitro study were to investigate the role of RhoA/ROCK signaling in regulating fibrotic effects of primary human pterygium fibroblasts (HPFs), as well as to explore the possible mechanisms of these effects.MethodsPterygium samples were obtained from surgery, and profibrotic activation was induced by TGF-β1. Cell proliferation was detected by CCK-8 assay; cell migration was detected by wound healing assay; quantitative real-time PCR and Western blot were used to detect the effects of TGF-β1 and the role of RhoA/ROCK signaling in the synthesis of alpha-smooth muscle actin (a-SMA), type I and III collagen (COL1 and COL3), and matrix metalloproteinase-9 (MMP9) in HPFs. The changes of signaling pathways were detected by Western blot; and pharmaceutical inhibition of RhoA/ROCK signaling and its downstream MRFT-A/SRF transcription pathway were used to assess their possible mechanism in HPFs fibrosis.ResultsROCK inhibitor Y-27632 decreased TGF-β1-induced cell proliferation and migration, reduced the TGF-β1-induced expression of profibrotic markers in HPFs, and suppressed TGF-β1-induced nuclear accumulation of Myocardin-related transcription factor A (MRTF-A) as well as accompanied elevation of F/G-actin ratio in HPFs. MRTF-A/Serum response factor (SRF) inhibitor CCG-100602 attenuated the TGF-β1-induced α-SMA expression and reduced myofibroblast activation in HPFs.ConclusionsRhoA/ROCK signaling played a pivotal role in TGF-β1-induced fibrosis and myofibroblast activation in HPFs at least in part by inactivating the downstream MRTF-A/SRF transcriptional pathway.