淫羊藿苷对TGF-β1诱导的肾小管上皮细胞上皮间质转分化的作用及机制

The effect and mechanism of icariin on the epithelial-mesenchymal transdifferentiation of renal tubular epithelial cells induced by TGF-β1

关键词:  转化生长因子-β1  肾小管上皮细胞  淫羊藿苷  上皮-间质转化  Smad信号通路

Key words:  transforming growth factor-β1  renal tubular epithelial cell  icariin  epithelial-mesenchymal transition  Smad signaling pathway

摘要:

[目的] 观察淫羊藿苷对转化生长因子-β1(TGF-β1)诱导的肾小管上皮-间质转化的作用以及对Smad信号通路的影响。[方法] 将肾小管上皮细胞(HK-2)分为空白对照组、TGF-β1组、TGF-β1+淫羊藿苷(10、20、40 μmol/L)组。采用CCK-8法检测10、20、40 μmol/L的淫羊藿苷干预10 μg/L TGF-β1诱导的肾小管上皮细胞的增殖影响; 使用实时定量聚合酶链式反应法检测检测上皮间质转化分管关键因子α-平滑肌肌动蛋白(α-SMA)、E-钙黏蛋白(E-cadherin)、波形蛋白(vimentin)mRNA的表达量; 采用Western blot检测α-SMA、E-cadherin、Smad2/3和p-Smad2/3蛋白表达量; 划痕实验和侵袭迁移小室法(Transwell小室)分别检测HK-2细胞的迁移和侵袭能力。[结果] 与空白对照组比较, TGF-β1组的HK-2细胞增殖、迁移和侵袭能力显著增高(P<0.05), α-SMA、vimentin蛋白和mRNA以及p-Smad2/3蛋白表达量显著增加(P<0.05), 而E-cadherin、Smad2/3蛋白和mRNA表达量显著降低(P<0.05)。与TGF-β1组比较, TGF-β1+淫羊藿苷(10、20、40 μmol/L)组的HK-2细胞增殖、迁移和侵袭能力显著降低(P<0.05), α-SMA、vimentin蛋白和mRNA以及p-Smad2/3蛋白表达量显著降低(P<0.05), 而E-cadherin、Smad2/3蛋白和mRNA表达量显著增加(P<0.05)。淫羊藿苷对TGF-β1诱导HK-2细胞抑制增殖、迁移和侵袭能力以及上皮间质转化无剂量依赖性。[结论] 淫羊藿苷可以抑制TGF-β1诱导的HK-2细胞的上皮-间质转化的分化, 可能与抑制Smad信号通路有关。

Abstract:

[Objective] To observe the effect of icariin on the renal tubular epithelial-mesenchymal transition induced by TGF-β1 and its influence on the Smad signaling pathway.[Methods] Renal tubular epithelial cells(HK-2) were divided into blank control group, TGF-β1 group, and TGF-β1+icariin(10, 20, 40 μmol/L) group. We used CCK-8 method to detect the effect of 10, 20, 40 μmol/L icariin on the proliferation of renal tubular epithelial cells induced by 10 μg/L TGF-β1;RT-qPCR method to detect epithelial-mesenchymal transition in charge of the key factors α-smooth muscle actin(α-SMA), E-cadherin, vimentin mRNA expression; Western blot to detect α-SMA, E-cadherin, Smad2/3 and p-Smad2/3 protein expression; and scratch test and invasion and migration chamber method(Transwell chamber) to detect the migration and invasion ability of HK-2 cells.[Results] Compared with the blank control group, the HK-2 cell proliferation, migration and invasion ability of the TGF-β1 group were significantly increased(P<0.05), and the expression of α-SMA, vimentin protein and mRNA, and p-Smad2/3 protein were significantly increased(P<0.05), while the expression of E-cadherin, Smad2/3 protein and mRNA were significantly reduced(P<0.05). Compared with the TGF-β1 group, the HK-2 cell proliferation, migration and invasion ability of the TGF-β1+ icariin(10, 20, 40 μmol/L) group were significantly reduced(P<0.05), α-SMA, vimentin protein, mRNA and p-Smad2/3 protein expression were decreased significantly(P<0.05), while E-cadherin, Smad2/3 protein and mRNA expression were increased significantly(P<0.05). Icariin had not a dose-dependent effect on the ability of TGF-β1 to induce HK-2 cells to inhibit proliferation, migration and invasion, and epithelial-mesenchymal transition.[Conclusion] Icariin could inhibit TGF-β1 induced differentiation of epithelial-mesenchymal transition of HK-2 cells, which may be related to the inhibition of Smad signaling pathway.