小檗碱对TGF-β1诱导的肾小管上皮细胞转分化及TGF-β/Smad通路的影响  

The effect of Berberine on TGF-β1-induced epithelial-to-mesenchymal transition in rat kidney tubular epithelial cells

作　　者：姚璐[1] 张秀丽[2,3] 赵越[2] 张萍[4] 王雪梅[5] 池志宏[6] 

机构地区：[1]中国医科大学科学实验中心,沈阳110122 [2]中国医科大学细胞生物国家重点实验室,沈阳110122 [3]本溪市中心医院肾内科,本溪117000 [4]辽宁医学院组胚教研室,锦州121000 [5]中国医科大学附属口腔医院修复科,沈阳110001 [6]中国医科大学病理生理教研室,沈阳110122

出　　处：《解剖科学进展》2016年第4期361-365,共5页Progress of Anatomical Sciences

基　　金：国家自然科学基金(81170561);辽宁省自然科技计划基金(2012408002;2013225090);中国博士后科学基金(2014MM551144)

摘　　要：目的研究小檗碱(BBr)对转化生长因子-β1(TGF-β1)诱导的肾小管上皮细胞(NRK-52E)转分化(EMT)及TGF-β/Smad通路的影响。方法常规培养肾小管上皮细胞、传代、分组:1正常对照组;2TGF-β1组(10ng/ml TGF-β1作用48h);3小檗碱作用组(30μM小檗碱作用24h后加10ng/ml TGF-β1作用48h)。采用相差显微镜观察各组细胞表型改变,免疫荧光及Western Blot方法检测α平滑肌肌动蛋白(α-SMA)、上皮钙黏素(E-cadherin)、Smad2、Smad3、磷酸化的Smad2、3(P-Smad2、3)蛋白表达;应用流式细胞术,采用活性氧自由基(ROS)检测试剂盒检测各组别的ROS蛋白表达情况。结果 TGF-β1处理可导致NRK-52E细胞的α-SMA蛋白表达明显升高,E-cadherin蛋白表达降低。而小檗碱处理能够减少肾小管上皮细胞α-SMA的高表达,提高E-cadherin蛋白的表达;同时,小檗碱可有效降低由TGF-β1引起的NRK-52E细胞中ROS的过多产生。此外,小檗碱可有效抑制Smad2/3的活化。结论小檗碱可逆转TGF-β1所致的肾小管上皮细胞的EMT,其作用机制可能是通过抑制氧化应激及TGF-β/Smad信号通路而发挥作用。Objective To investigate the effects of berherine （BBr） on transforming growth factor - β 1 （TGF- β1）-induced epithelial-to-mesenchymal transition （EMT） in rat kidney tubular epithelial cell （NRK-52E cells） and examine the underlying molecular mechanisms. Methods NRK-52E ceils were incubated withl0ng/ml TGF-β1 for 48 hours, or pre-incubated with 30 μ M BBr for 24 hours with stimulated by 10ng/ml TGF-β1 for 48 hours. NRK-52E cells in the control group were just incubated with medium. The expression of α -SMA, E-cadherin, Smad2, Smad3, P-Smad2, P- Smad3 was detected by Western Blot or immunofluorescence staining. In addition, reactive oxygen species （ROS） assay experiments were performed using the reactive oxygen species assay kit. Results We found that BBr tratment inhibited TGF- β1-induced NRK-52E cells EMT significantly, by attenuating ROS production. Whereas this TGF-β 1-induced EMT can be attenuated by pre-treating the cells with 30μM BBr which were evidenced by the reduced up-regulation of α-SMA and the ameliorated expression of epithelial protein E-cadherin, further analysis revealed that BBr treatment inhibited TGF- β l-induced NRK-52E cells EMT likely through inhibition of TGF- β/Smad pathways. Conclusion These results indicate that BBr can inhibit EMT in TGF- β1-induced NRK-52E cells may be by inhibition of ROS production as well as TGF- β/Smad pathways activation.

关 键 词：TGF-Β1 小檗碱 转分化 糖尿病肾病 肾小管上皮细胞 

分 类 号：Q291[生物学—细胞生物学]