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目的观察止消通脉宁药物血清对转化生长因子-β1(TGF-β1)诱导人肾小管上皮细胞(HK-2)表型转化的影响,并探讨其机制。方法将HK-2细胞用含10%胎牛血清的DMEM/F12(1∶1)培养基培养并分为空白对照组、TGF-β1诱导组(TGF-β110 ng/mL)、空白血清对照组(TGF-β110 ng/mL+10%空白血清)、干预1组(TGF-β110 ng/mL+10%低剂量止消通脉宁药物血清)、干预2组(TGF-β110 ng/mL+10%中剂量止消通脉宁药物血清)、干预3组(TGF-β110 ng/mL+10%高剂量止消通脉宁药物血清)。药物干预24 h后,免疫组化检测α-平滑肌肌动蛋白(α-SMA)和E-钙黏蛋白(E-cadherin)的表达。在24、48、72 h应用酶联免疫吸附法检测细胞培养上清中Ⅰ型胶原(ColⅠ)、Ⅲ型胶原(ColⅢ)和纤连蛋白(FN)的含量。结果正常HK-2细胞表达E-cadherin,不表达α-SMA,经TGF-β1刺激后细胞表型发生转变,E-cadherin的表达明显减弱,α-SMA表达明显增强,且ColⅠ、ColⅢ和FN的分泌明显增加,与空白对照组比较差异有统计学意义(P<0.05)。止消通脉宁药物血清干预后α-SMA表达明显减弱,E-cadherin表达明显增强;同时抑制了ColⅠ、ColⅢ和FN的分泌,与TGF-β1诱导组比较差异有统计学意义(P<0.05)。结论止消通脉宁在一定程度上能够抑制人肾小管上皮细胞表型转化,减少细胞外基质成分的分泌,具有防治肾间质纤维化的作用。
Objective To observe the effect of Zhixiaotongmaining serum on phenotypic transformation of human renal tubular epithelial cells (HK-2) induced by transforming growth factor-β1 (TGF-β1) and to explore its mechanism. Methods HK-2 cells were cultured in DMEM / F12 (1: 1) medium containing 10% fetal bovine serum and divided into blank control group, TGF-β1 induction group (TGF-β110 ng / mL), blank serum control group (TGF-β110 ng / mL + 10% blank serum), intervention group 1 (TGF-β110 ng / mL + 10% low dose Zhixiaotongmaining serum) % Of the dose of anti-Tongmai Ning serum), intervention group 3 (TGF-β110 ng / mL + 10% high dose Zhi Tong Tong Mai Ning serum). After 24 h of drug intervention, the expressions of α-SMA and E-cadherin were detected by immunohistochemistry. The levels of ColⅠ, Col Ⅲ and fibronectin (FN) in the cell culture supernatants were detected by enzyme-linked immunosorbent assay 24, 48 and 72 h after transfection. Results The expression of E-cadherin and the expression of α-SMA in normal HK-2 cells were normal while the expressions of E-cadherin and α-SMA were not significantly increased in HK-2 cells. The secretion increased significantly compared with the blank control group, the difference was statistically significant (P <0.05). The effect of Zhuxiaotongmaining serum on α-SMA expression was significantly weakened and the expression of E-cadherin was significantly increased. At the same time, the secretion of ColⅠ, Col Ⅲ and FN was inhibited, and the difference was statistically significant (P <0.05) ). Conclusion Zhuxiaotongmaining can inhibit the phenotypic transformation of human renal tubular epithelial cells to a certain extent and reduce the secretion of extracellular matrix components, and has the effect of preventing and treating renal interstitial fibrosis.