缺氧状态下去乙酰化酶SIRT1对MHCⅡ的反式激活因子的调控机制研究

The regulatory effect of SIRT1 on CⅡTA under hypoxia

目的:探讨缺氧对SIRT1表达和活性的影响,及SIRT1对人体免疫功能的调节机制。方法:将人的原代外周血巨噬细胞在常氧和缺氧(1%O2)条件下培养,用Western blot方法检测细胞内SIRT1的蛋白表达变化,Real-time PCR检测SIRT1、NAMPT、HLA-DRα的mRNA水平,检测SIRT1的酶活性及NAD+/NADH的值。结果:SIRT1蛋白表达随缺氧时间的延长先降低后升高,缺氧12 h蛋白下降最明显,24、36 h后回升,但表达量仍低于常氧;缺氧12 h后,SIRT1、NAMPT的mRNA水平与常氧组比较明显降低(P<0.05);SIRT1的酶活性和NAD+/NADH的值显著低于常氧组(P<0.05);SIRT1激动剂白藜芦醇能够逆转缺氧诱导的HLA-DRα的mRNA表达下调。结论:缺氧通过调节巨噬细胞中SIRT1的mRNA水平、蛋白表达及酶活性,使得依赖Ⅱ类反式激活因子(classⅡtrans-activator,CⅡTA)的HLA-DRα表达下降,提示SIRT1可能是一个新的值得关注的免疫调节相关蛋白。

Objective：To investigate the effects of hypoxia on the expression and activity of SIRT1 and its regulatory mechanisms on human immune function.Methods：Human primary peripheral blood monocytes（HPBMs） were cultured in vitro under normal oxygen and hypoxia（1% O2） conditions.The protein expression of SIRT1 was detected by Western blot.Real-time PCR was performed to examine the mRNA levels of SIRT1,NAMPT and HLA-DRα.Activity assay kits were used to detect the activity of SIRT1 and value of NAD＋ / NADH.Results：SIRT1 protein level was inhibited with peak inhibition occurring at 12 h post exposure to hypoxia.The mRNA levels of SIRT1 and NAMPT in cells cultured in hypoxia were significantly lower than that cultured in normal oxygen（P〈0.05）.In addition,the enzyme activity of SIRT1 and the value of NAD＋ / NADH were also significantly decreased（P〈0.05）.Resveratrol,which is the agonist of SIRT1,rescued the decreased expression of HLA-DRα induced by hypoxia.Conclusion：The CⅡTAdependent HLA-DRα expression was decreased,and accompanied with a decrease in the expression and enzyme activity of SIRT1 in macrophages exposed to hypoxia.These results revealed that SIRT1 may play a critical role in regulating adaptive immunity.