SIRT1抗视网膜色素上皮细胞氧化应激作用的实验研究

目的 观察去乙酰化酶-1(SIRT1)对视网膜色素上皮细胞(RPE)氧化应激的防护作用.方法 应用50 μmol/L H₂O₂与ARPE-19细胞孵育24 h诱导RPE产生氧化应激病理生理过程,观察细胞增殖及凋亡率、细胞活性氧水平(ROS), 氧化剂诱导RPE细胞衰老状况,了解氧化应激对RPE的损伤;进一步采用SIRT1激动剂白藜芦醇(RSV)及抑制剂烟酰胺(NA)分别处理经H₂O₂干预的RPE细胞,观察SIRT1抗RPE氧化应激效果.结果 ①与对照组相比,H₂O₂干预RPE后细胞增殖显著下降(P=0.020),凋亡率及内生ROS水平均显著增高(P=0.013, P=0.040),细胞衰老水平显著升高(P=0.045);②NA可显著提高H₂O₂的毒性作用,降低RPE细胞增殖率(P=0.049),刺激凋亡(P=0.028),使ROS累积和衰老速度增加;③RSV具有显著的抗H₂O₂毒性作用;④RT-qPCR分析显示,SIRT1mRNA水平显著下调.结论 氧化应激状况下SIRT1表达下调,表明SIRT1对氧化应激诱发的视网膜色素上皮细胞损伤有显著的防护作用.适当提高SIRT1活性也许是延缓AMD的一种治疗策略.

The experiment of SIRT1 on against oxidative stress to retinal pigmented epithelium cells

Objective To observe the protection role of the sirtuin 1 (SIRT1) towards the retinal pigmented epithelium(RPE) cells following exposure to oxidative stress. Methods The rates of proliferation and apoptosis, levels of intracellular reactive oxygen species(ROS) and cell senescence of RPEs, induced by oxidants (H₂O₂), were evaluated. Results The results revealed a down-regulation of SIRT1 expression during oxidative stress. Furthermore, SIRT1 activator resveratrol (RSV) and inhibitors nicotinamide (NA) were respectively treated on RPE cells following exposure by H₂O₂ intervention; the expression of SIRT1 gene were detected by RT-PCR technology after RNA interfering on SIRT1. The experiments indicated that cell proliferation, apoptosis and ROS levels were significantly different compared with the control group after intervention by H₂O₂, and cell senescence were significantly increased; NA was significantly increased the toxicity of H₂O₂, however RSV had the opposite anti-cytotoxicity of H₂O₂; RT-PCR analysis showed that SIRT1 levels were significantly reduced. Conclusion These results therefore suggested that down-regulation of SirT1 expression during oxidative stress, further investigation indicated that SIRT1 protected RPEs from oxidative stressinduced damage. Appropriate increase of the activity of SIRT1 may be a therapeutic strategy to delay AMD.