五味子乙素对缺氧/复氧肾小管上皮细胞凋亡与自噬的影响

Effects of schisandrin B on apoptosis and autophagy in hypoxia/reoxygenation HK-2 cells

目的:体外观察五味子乙素(SchB)对缺氧/复氧肾小管上皮细胞(HK-2)凋亡与自噬的影响,并探讨其可能的作用机制。方法:将体外培养的HK-2细胞随机分为对照组、缺氧/复氧组(H12/R6、H12/R12、H12/R24)、SchB+H/R组(SchB+H12/R6、SchB+H12/R12、SchB+H12/R24),SchB+H/R组首先用SchB预孵育6h后再行缺氧/复氧处理。CCK-8试剂盒检测各组HK-2细胞的活性,Annexin-V/PI染色检测HK-2细胞的凋亡率,Western Blot检测HK-2细胞中凋亡相关蛋白Bcl-2、Bax及自噬相关蛋白LC3的表达量。结果:H/R组与SchB+H/R组的细胞活性均出现明显的下降(P<0.01),但SchB+H/R组较H/R组的细胞活性是增长的,且随复氧时间的延长,各组细胞活性均逐渐增多;H/R组与SchB+H/R组细胞凋亡率均上升(P<0.01),但H/R组上升更为显著(P<0.01);随着复氧时间的延长,两组细胞凋亡率均减少(P<0.01)。与H/R组比较,SchB+H/R组Bcl-2/Bax比值与LC3水平均上升(P<0.01),且随复氧时间的延长,两组Bcl-2/Bax值上升、LC3水平下降。结论:SchB对缺氧/复氧造成HK-2细胞的凋亡有一定的保护作用,其机制可能与调节凋亡相关蛋白Bcl-2、Bax与自噬相关蛋白LC3的表达有关。

Objective: To observe the effects of schisandrin B(SchB) on apoptosis and autophagy of hypoxia/reoxygenation renal tubular epithelial cells(HK-2) in vitro and to explore its possible mechanism. Methods: HK-2 cells cultured in vitro were randomly divided into control group, hypoxia/reoxygenation group(H12/R6, H12/R12, H12/R24) and SchB+H/R group(SchB+H12/R6, SchB+H12/R12, SchB+H12/R24), SchB+H/R group was first incubated with SchB for 6 h before hypoxia/reoxygenation. The activity of HK-2 cells in each group was detected by CCK-8 kit. Annexin-V/PI staining was used to detect the rate of apoptosis in HK-2 cells. The expression of apoptosis-related proteins Bcl-2, Bax and autophagy-related protein LC3 in HK-2 cells were detected by Western Blot. Results: The cell viability of H/R group and SchB+H/R group decreased significantly(P<0.01), but the cell activity of SchB+H/R group was higher than that of H/R group, and with reoxygenation time, the cell viability of each group increased gradually. The apoptosis rate of H/R group and SchB+H/R group increased(P<0.01), but the increase in H/R group was more significant(P<0.01). With the prolongation of reoxygenation time, the apoptosis rate of both groups decreased(P<0.01). Compared with H/R group, the ratio of Bcl-2/Bax and LC3 in SchB+H/R group increased(P<0.01), and with the prolongation of reoxygenation time, the value of Bcl-2/Bax up-regulated, and the levels of LC3 down-regulated. Conclusion: SchB has a protective effect on apoptosis of HK-2 cells induced by hypoxia/reoxygenation. The mechanism may be related to the regulation of apoptosis-related proteins Bcl-2, Bax and autophagy-related protein LC3.