miR-543靶向TCF7L2调控BMSCs成脂分化的实验研究
Experimental study on miR-543 targeting TCF7L2 to regulate the adipogenic differentiation of BMSCs
  
DOI:10.3969/j.issn.1006-7108.2022.12.011
中文关键词:  微小RNA-543  转录因子7类似物2  骨髓间充质干细胞  成脂分化  大鼠
英文关键词:microRNA-543  transcription factor 7-like 2  bone marrow mesenchymal stem cells  adipogenic differentiation  rat
基金项目:武汉市医学科研项目(WX20B22)
作者单位
封宝红 喻业安* 晏莉 伍军 张艳霞 朱戈丽 武汉市第三医院光谷院区肾内科湖北 武汉 430014 
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中文摘要:
      目的 探讨微小RNA-543(miR-543)靶向调控转录因子7类似物2(TCF7L2)对大鼠骨髓间充质干细胞(bone marrow mesenchymal stem cells,BMSCs)成脂分化的影响。方法 分离并鉴定大鼠BMSCs,实时荧光定量PCR(RT-qPCR)检测BMSCs成脂分化诱导过程中miR-543的表达变化;体外培养BMSCs细胞,对BMSCs细胞进行干预,过表达miR-543、抑制TCF7L2表达或同时过表达miR-543与TCF7L2,使用成脂分化诱导培养基培养,油红O染色检测BMSCs成脂分化;免疫印迹法检测BMSCs细胞TCF7L2、脂肪细胞蛋白2(aP2)、CCAAT/增强子结合蛋白α(C/EBPα)、过氧化物酶体增殖物激活受体γ(PPAR-γ)蛋白表达。双荧光素酶报告基因实验验证miR-543与TCF7L2的靶向关系。结果 成功分离大鼠BMSCs;BMSCs成脂分化诱导处理1、2、3、4、5 d后miR-543表达水平降低;双荧光素酶报告基因实验证实miR-543与TCF7L2存在靶向关系;过表达miR-543或抑制TCF7L2表达,BMSCs油红O染色后OD值、aP2、PPAR-γ、C/EBPα蛋白表达水平显著降低(P<0.05);过表达TCF7L2可部分逆转过表达miR-543对BMSCs细胞成脂分化的抑制作用(P<0.05)。结论 miR-543在大鼠BMSCs细胞成脂分化过程中下调表达,过表达miR-543可通过靶向抑制TCF7L2表达而抑制BMSCs细胞成脂分化。
英文摘要:
      Objective To investigate the effect of microRNA-543 (miR-543) on the adipogenic differentiation of rat bone marrow mesenchymal stem cells (BMSCs) by targeting and regulating transcription factor 7-like 2 (TCF7L2). Methods The rat BMSCs were isolated and identified. Real-time fluorescent quantitative PCR (RT-qPCR) was used to detect the expression changes of miR-543 during the induction of adipogenic differentiation of BMSCs. BMSCs cells were cultured in vitro, and were intervened with overexpression of miR-543, inhibition of TCF7L2, or overexpression of both miR-543 and TCF7L2. Cells were cultured in the adipogenic differentiation induction medium. Oil red O staining was used to detect the adipogenic differentiation of BMSCs. Western blotting was used to detect the protein expression of TCF7L2, adipocyte protein 2 (aP2), peroxisome proliferator-activated receptor γ (PPAR-γ), and CCAAT/enhancer binding protein α (C/EBPa) in BMSCs cells. Dual luciferase reporter gene experiment was used to verify the targeting relationship between miR-543 and TCF7L2. Results The rat BMSCs were successfully isolated. The expression of miR-543 decreased after 1, 2, 3, 4, and 5-d induction of adipogenic differentiation of BMSCs. The dual luciferase reporter gene experiment confirmed that miR-543 had a targeting relationship with TCF7L2. Overexpressing miR-543 or inhibiting the expression of TCF7L2 led to significantly reduction of the OD value, aP2, PPAR-γ, and C/EBPa protein expression levels of BMSCs (P<0.05). Overexpression of TCF7L2 partially reversed the inhibitory effect of overexpression of miR-543 on the adipogenic differentiation of BMSCs cells (P<0.05). Conclusion The expression of miR-543 is down-regulated during the adipogenic differentiation of rat BMSCs. Overexpression of miR-543 inhibits the adipogenic differentiation of BMSCs by targeting the inhibition of TCF7L2 expression.
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