目的: 观察姜黄素(Curcumin,Cur)对高糖环境下人肾小管上皮细胞(Human kidney 2,HK-2)线粒体自噬及氧化应激的影响,探究姜黄素对HK-2细胞的保护作用及机制。方法: 体外培养HK-2细胞,随机分为4组:对照组(5.5 mmol/L葡萄糖)、姜黄素组(5.5 mmol/L葡萄糖+10 μmol/L姜黄素)、高糖组(30 mmol/L葡萄糖)、姜黄素高糖组(30 mmol/L葡萄糖+10 μmol/L姜黄素)。各组干预刺激48 h,收集细胞的总蛋白,采用Western blot检测线粒体自噬相关蛋白Pink1、Parkin、LC3B以及beclin1的表达;JC-1检测各组线粒体膜电位水平;采用活性氧检测试剂盒检测细胞内活性氧(Reactive oxygen species, ROS)生成情况。结果: 与对照组比较,姜黄素组各项指标无明显改变(P＞0.05);与姜黄素组比较,高糖组Pink1、Parkin、LC3B、beclin1蛋白表达降低(P＜0.01),细胞内ROS表达升高(P＜0.01),线粒体膜电位活性下降(P＜0.01);与高糖组比较,姜黄素高糖组Pink1、Parkin、LC3B、beclin1蛋白表达升高(P＜0.01);细胞内ROS表达降低(P＜0.01);线粒体膜电位活性升高(P＜0.01)。结论: 姜黄素可逆转高糖诱导的HK-2细胞线粒体自噬,抑制氧化应激。
Objective: To observe the effects of curcumin on mitochondrial autophagy and oxidative stress in human renal tubular epithelial cells (HK-2) under high glucose environment, and to explore the protective effect and mechanism of curcumin on HK-2 cells. Methods: HK-2 cells were divided into 4 groups: control group(5.5 mmol/L glucose), curcumin group(5.5 mmol/L glucose +10 μmol/L Cur), high glucose group(30 mmol/L glucose) and curcumin high glucose group(30 mmol/L glucose+10 μmol/L Cur). All groups were stimulated for 48h. Total protein was collected, and the expression of mitochondrial autophagy related proteins Pink1, Parkin, LC3B and beclin1 was detected by Western blot. Mitochondrial membrane potential was detected by JC-1. Reactive oxygen species (ROS) detection kit was used to detect intracellular ROS production. Results: Compared with the control group, the indexes in the curcumin group did not change significantly(P＞0.05). Compared with curcumin group, the protein expressions of Pink1, Parkin, LC3B and Beclin1 in high glucose group were significantly decreased(P＜0.01), intracellular ROS expression increased significantly(P＜0.01), and mitochondrial membrane potential activity decreased(P＜0.01). Compared with high glucose group, the protein expression of Pink1, Parkin, LC3B and Beclin1 in curcumin high glucose group was significantly increased(P＜0.01), intracellular ROS expression was significantly decreased(P＜0.01), mitochondrial membrane potential activity increased(P＜0.01). Conclusion: Curcumin can reverse the mitochondrial autophagy induced by high glucose and inhibit oxidative stress in HK-2 cells.
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