miR-590-3p调控人肝星状细胞LX-2TGF-β/SMAD通路抗肝纤维化的研究Effect of miR-590-3p on Anti-Liver Fibrosis by Regulating ECM and TGF-β/SMAD Pathway in Human LX-2 Cell Line
曹聪,许晨舒,唐欲博,陈杰,陈孝
CAO Cong,XU Chenshu,TANG Yubo,CHEN Jie,CHEN Xiao
摘要(Abstract):
目的建立转化生长因子-β1(TGF-β1)诱导人肝星状细胞LX-2体外纤维化模型,观察miR-590-3p对肝纤维化影响。方法设计并合成miR-590-3p模拟物和miR-590-3p抑制剂,瞬时转染至LX-2细胞,再经TGF-β1诱导24 h后,荧光定量PCR检测胞内miR-590-3p表达,Western Blot检测胞内纤维化相关蛋白Smad 4及CollagenⅠ表达变化,并分析胞内miR-590-3p表达改变对细胞纤维化指标的影响。结果 miR-590-3p过表达下调Smad 4蛋白水平表达50%(P<0.05),抑制miR-590-3p功能上调Smad 4蛋白表达50%(P<0.05)。结论 miR-590-3p可能通过下调LX-2 Smad4的表达,影响LX-2活化,在肝纤维化进程中发挥重要作用。
OBJECTIVE To explore the establishment of a fibrosis model in vitro by inducing human LX-2 with transforming growth factor β1( TGF-β1),and to observe the effects of miR-590-3p on the expression of liver fibrosis markers. METHODS The miR-590-3p mimics and miR-590-3p inhibitor were designed,synthesized and transfacted into LX-2 cells. Transfacted cells were then induced by TGF-β1of 10 ng / m L for 24 h. The expression of miR-590-3p was detected by real-time PCR. The variations of expressions of fibrosis related protein Smad4 and Collagen Ⅰ were detected by the Western blotting to analyze the effects of the miR-590-3p expression on cell fibrosis indexes. RESULTS Compared to the control group,the expression of Smad4 protein significantly decreased 50% after LX-2 cells were transfacted with miR-590-3p mimics and increased 50% after LX-2 cells were transfacted with miR-590-3p inhibitor respectively( P < 0. 05). CONCLUSION The miR-590-3p may affect the activation of LX-2 cells through the depression of Smad 4 and then modulate liver fibrosis.
关键词(KeyWords):
miR-590-3p;肝星状细胞;肝纤维化
miR-590-3p;HSCs;liver fibrosis
基金项目(Foundation): 国家自然科学基金(30972917);; 广东省医学科研基金(A2013193);; 广东省药学会肝炎用药研究基金(2011G25)
作者(Author):
曹聪,许晨舒,唐欲博,陈杰,陈孝
CAO Cong,XU Chenshu,TANG Yubo,CHEN Jie,CHEN Xiao
参考文献(References):
- [1]夏云红.肝星状细胞的基因差异表达以及抑制T细胞功能与肝细胞癌转移的实验研究[D].上海:复旦大学中山医院,2009.
- [2]Jiang X,Xiang G,Wang Y,et al.Micro RNA-590-5p regulates proliferation and invasion in human hepatocellular carcinoma cells by targeting TGF-beta RII[J].Mol Cells,2012,33(6):545-551.
- [3]杨红飞,郑文宏,赵文淘,等.mi R-590-5p和mi R-590-3p参与肝细胞肝癌发展的机制[J].南方医科大学学报,2013,33(6):804-811.
- [4]肖琳,成军,郭江,等.转化生长因子β1刺激肝星状细胞差异表达下调基因筛选[J].中华实验和临床感染病杂志,2010,4(03):236-243.
- [5]Tu X,Zhang H,Zhang J,et al.Micro RNA-101 suppresses liver fibrosis by targeting the TGF-beta signalling pathway[J].J Pathol,2014,234(1):46-59.
- [6]Li J,Ghazwani M,Zhang YF,et al.mi R-122 regulates collagen production via targeting hepatic stellate cells and suppressing P4HA1 expression[J].Journal of Hepatology,2013,58(3):522-528.
- [7]Murphy FR,Issa R,Zhou XY,et al.Inhibition of apoptosis of activated hepatic stellate cells by tissue inhibitor of metalloproteinase-1 is mediated via effects on matrix metalloproteinase inhibition:implications for reversibility of liver fibrosis[J].J Biol Chem,2002,277(13):11069-11076.
- [8]He Y,Huang C,Sun X,et al.Micro RNA-146a modulates TGF-beta1 induced hepatic stellate cell proliferation by targeting SMAD4[J].Cellular Signalling,2012,29(24):1923-1930.