GRP78-c-Src信号通路对周期性机械拉伸诱导的牙周韧带成纤维细胞成骨细胞分化的作用

Jing Hu, Zhihua Cui, Keqiang Huang, Rongjian Su, Song Zhao
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引用次数: 0

摘要

研究目的本研究旨在探讨葡萄糖调节蛋白(GRP)78对周期性机械拉伸下牙周韧带成纤维细胞(PDLFs)成骨细胞分化的影响,并确定其潜在机制:方法:应用 FlexCell 5000 细胞机械装置模拟正畸牙齿的应力环境。方法:应用 FlexCell 5000 细胞机械装置模拟正畸牙齿的应力环境,通过流式细胞分选获得 GRP78High 和 GRP78Low 亚群。进行基因转染以敲除 GRP78 和 c-Src 的表达,并过表达 c-Src。采用 Western 印迹分析检测 Runt 相关基因 2(RUNX2)、Osterix、骨钙蛋白(OCN)和骨营养素(OPN)的蛋白表达。免疫沉淀试验用于确定 GRP78 与 c-Src 的相互作用。茜素红染色法测定细胞矿化结节的形成:结果:GRP78在PDLFs中异质性表达,通过流式细胞分拣获得了GRP78高亚群和GRP78低亚群。GRP78High亚群的成骨分化能力和c-Src激酶的磷酸化水平在循环机械拉伸(PPP)后均较GRP78Low亚群显著提高:GRP78通过与c-Src激酶相互作用上调c-Src表达,促进PDLFs在循环机械拉伸条件下的成骨分化。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
Role of the GRP78-c-Src signaling pathway on osteoblast differentiation of periodontal ligament fibroblasts induced by cyclic mechanical stretch.

Objectives: This study aims to investigate the influence of glucose regulated protein (GRP) 78 on osteoblast differentiation in periodontal ligament fibroblasts (PDLFs) under cyclic mechanical stretch and determine the underlying mechanism.

Methods: FlexCell 5000 cell mechanical device was applied to simulate the stress environment of orthodontic teeth. GRP78High and GRP78Low subpopulation were obtained by flow sorting. Gene transfection was performed to knockdown GRP78 and c-Src expression and overexpress c-Src. Western blot analysis was used to detect the protein expression of Runt-related gene 2 (RUNX2), Osterix, osteocalcin (OCN), and osteopontin (OPN). Immunoprecipitation assay was used to determine the interaction of GRP78 with c-Src. The formation of cellular mineralized nodules was determined by alizarin red staining.

Results: GRP78 was heterogeneously expressed in PDLFs, and GRP78High and GRP78Low subpopulations were obtained by flow sorting. The osteogenic differentiation ability and phosphorylation level of c-Src kinase in the GRP78High subpopulation were significantly increased compared with those in GRP78Low subpopulation after cyclic mechanical stretch (P<0.05). GRP78 interacted with c-Src in PDLFs. The overexpression c-Src group showed significantly increased osteogenic differentiation ability than the vector group (P<0.05), and the sic-Src group showed significantly decreased osteogenic differentiation ability (P<0.05) after cyclic mechanical stretch.

Conclusions: GRP78 upregulates c-Src expression by interacting with c-Src kinase and promotes osteogenic differentiation under cyclic mechanical stretch in PDLFs.

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