目的 研究万古霉素诱导人肾小管上皮细胞（HK-2）损伤及其Smad同源物4（Smad4）表达，并探讨其潜在的作用机制。方法 使用不同终浓度（0、1、2、3、4、5 mmol/L）的万古霉素溶液作用于HK-2细胞24 h，显微镜下观察万古霉素对细胞形态的影响；蛋白质免疫印迹法检测Smad4、B淋巴细胞瘤-2（Bcl-2）相关X蛋白（Bax）、Bcl-2的蛋白水平；TUNEL法检测HK-2细胞凋亡情况；CCK8法检测HK-2细胞活力来研究万古霉素对肾脏的毒性作用。构建Smad4过表达质粒，通过CCK8法和TUNEL实验检测Smad4过表达对万古霉素诱导的HK-2细胞损伤的影响。结果 与对照组比较，随着万古霉素浓度升高，HK-2细胞逐渐变形，数量逐渐减少；细胞活力显著降低，凋亡率显著增加（P＜0.05、0.01）。与对照组比较，促凋亡蛋白Bax表达显著增高，而抗凋亡蛋白Smad4、Bcl-2表达则显著降低（P＜0.05、0.01、0.001）。过表达Smad4后，使万古霉素诱导的HK-2的细胞活力显著增加，凋亡率显著降低（P＜0.05、0.001）。结论 万古霉素能够诱导人肾小管上皮细胞损伤，其作用机制可能与调控Smad4的蛋白降解密切相关。

Objective To study the injury of human renal tubular epithelial cells (HK-2) induced by vancomycin and the expression of Smad4, and to explore the potential mechanism of action. Methods HK-2 cells were treated with vancomycin solution at different final concentrations (0, 1, 2, 3, 4, and 5 mmol/L) for 24 h, and the effect of vancomycin on cell morphology was observed under microscope. The protein levels of Smad4, Bax, and Bcl-2 were detected by Western blotting. The apoptosis of HK-2 cells was detected by TUNEL method. The activity of HK-2 cells was detected by CCK8 method to study the toxic effect of vancomycin on kidney. Smad4 overexpression plasmid was constructed, and the effect of Smad4 overexpression on vancomycin-induced HK-2 cell damage was detected by CCK8 assay and TUNEL assay. Results Compared with the control group, with the increase of vancomycin concentration, the number of HK-2 cells was gradually deformed and decreased. The cell viability was significantly decreased and the apoptosis rate was significantly increased (P＜ 0.05, 0.01). Compared with the control group, the expressions of pro-apoptotic protein Bax were significantly increased, while the expressions of anti-apoptotic proteins Smad4 and Bcl-2 were significantly decreased (P＜ 0.05, 0.01, 0.001). After overexpression of Smad4, vancomycin-induced HK-2 cell viability was significantly increased, and apoptosis rate was significantly decreased (P＜0.05, 0.001). Conclusion Vancomycin can induce the injury of human renal tubular epithelial cells, and its mechanism may be closely related to the regulation of Smad4 protein degradation.

R965

天津市滨海新区卫健委科技项目（2019BWKQ029）