敲低Raf激酶抑制蛋白促进LX-2人肝星状细胞增殖

发布时间：2018-02-26 04:23

本文关键词： Raf激酶抑制蛋白(RKIP) 胞外信号调节激酶(ERK) 丝裂原活化蛋白激酶(MAPK) 增殖胶原蛋白肝纤维化　出处：《细胞与分子免疫学杂志》2017年01期 　论文类型：期刊论文

【摘要】：目的研究Raf激酶抑制蛋白(RKIP)在LX-2人肝星状细胞增殖中作用。方法将RKIP小干扰RNA(siRNA-RKIP)重组质粒转染至LX-2细胞,培养5 d,并筛选出稳定转染的细胞。MTT法检测沉默RKIP后LX-2细胞的增殖情况;流式细胞术检测细胞凋亡、细胞周期;实时定量PCR检测α平滑肌肌动蛋白(α-SMA)和1型胶原蛋白(Col1)mRNA的表达。Western blot法检测RKIP、α-SMA、Col1及胞外信号调节激酶/丝裂原活化蛋白激酶(ERK/MAPK)信号通路相关蛋白的表达水平。结果与空白对照组相比,RKIP低表达明显诱导LX-2细胞增殖,抑制细胞凋亡,增加G2期细胞比例,提高Col1、α-SMA mRNA和蛋白表达。此外,RKIP的沉默明显提高ERK/MAPK磷酸化水平。结论敲低RKIP的水平促进LX-2细胞的增殖,其机制与激活ERK/MAPK信号通路有关。
[Abstract]:Objective to study the role of Raf kinase inhibitor protein (RKIP) in the proliferation of LX-2 human hepatic stellate cells. Methods the recombinant plasmid of RKIP small interfering RNA-siRNA-RKIPs was transfected into LX-2 cells for 5 days, and the stable transfected cells were screened out to detect the proliferation of LX-2 cells after RKIP silencing. Apoptosis and cell cycle were detected by flow cytometry. Real-time quantitative PCR was used to detect the expression of 伪 -smooth muscle actin (伪 -SMA-1) and collagen type 1 (Collagen 1) mRNA. Western blot was used to detect the expression of RKIP, 伪 -SMA-Col1 and extracellular signal-regulated kinase / mitogen-activated protein kinase (ERK / MAPK) signal pathway related proteins. Compared with the control group, the low expression of RKIP induced the proliferation of LX-2 cells. Inhibition of apoptosis, increased proportion of G2 phase cells, increased expression of Col1, 伪 -SMA mRNA and protein. In addition, silencing of RKIP significantly increased the level of ERK/MAPK phosphorylation. Conclusion knockdown of RKIP can promote the proliferation of LX-2 cells, and its mechanism is related to the activation of ERK/MAPK signaling pathway.
【作者单位】：广西医科大学药学院;广西中医药大学第一附属医院药学部;
【基金】：国家自然科学基金(81260505,81473431) 广西自然科学基金(2014GXNSFAA118155,2014GXNSFAA118154)
【分类号】：R575.2

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