Wang Xue-Ling, Wu Li-Yan, Zhao Long, Sun Li-Na, Liu Hai-Ying, Liu Gang, Guan Guang-Ju
Nephrology Research Institute of Shandong University, The Second Hospital of Shandong University, Shandong University, Jinan, Shandong, China.
Room for Hemodialysis, Affiliated Hospital of Shandong University of Traditional Chinese Medicine, Jinan, Shandong, China.
Biomed Pharmacother. 2016 Oct;83:41-50. doi: 10.1016/j.biopha.2016.06.009. Epub 2016 Jun 21.
We aimed to explore the role of SIRT1 in apoptosis in human kidney proximal tubule epithelial (HK-2) cells, and to determine whether resveratrol (RSV, a SIRT1 activator) could ameliorate apoptosis in rats with streptozotocin-induced diabetes mellitus (DM) and/or in high glucose (HG, 30mM) - stimulated HK-2 cells. Rats were distributed randomly into three groups: 1) control group, 2) DM group, and 3) DM with RSV group (DM+RSV; rats treated with 30mg/kg/d of RSV for 16 weeks). The physical, biochemical, and morphological parameters were then examined. Additionally, the deacetylase activity of SIRT1, and the expression levels of SIRT1 and of representative apoptosis markers, such as p53, acetylated p53, cleaved caspase-3, caspase-9, and cleaved PARP, were measured. HK-2 cells were stimulated by HG for different lengths of time to study the effect of HG on apoptosis. HK-2 cells were treated with or without RSV (25μM) to investigate if RSV has a protective effect on HG-induced apoptosis. A gene-specific small interfering RNA against SIRT1 was used to study the role of SIRT1 in apoptosis. More apoptosis was found in the DM rats than in the control rats. Similarly, the expression levels of cleaved caspase-3, cleaved PARP, and acetylated p53 were significantly higher, and the level of SIRT1 was significantly lower, in the HK-2 cells that were cultured under HG conditions than those in the HK-2 cells that were cultured under low glucose (5.5mM) conditions. Notably, treatment with RSV lessened the HG-induced changes in the levels of apoptosis indicators, and this inhibition of HG-induced apoptosis in HK-2 cells by RSV treatment was abolished by SIRT1 silencing. Our study showed that hyperglycemia contributes to apoptosis in rat kidney and HK-2 cells. SIRT1 activation by RSV can reduce urinary albumin excretion and proximal tubule epithelial apoptosis both in vitro and in vivo. Based on our study, SIRT1/p53 axis played an important role in the hyperglycemia induced apoptosis. These findings indicated that the increased expression of SIRT1, mediated by RSV, is a possible mechanism by which RSV prevents renal tubular injury in diabetic nephropathy (DN). So RSV has great clinical significance and could provide the basis for the new way to effective treatment to contain the morbidity and mortality associated with DN.
我们旨在探讨沉默调节蛋白1（SIRT1）在人肾近端小管上皮（HK-2）细胞凋亡中的作用，并确定白藜芦醇（RSV，一种SIRT1激活剂）是否能改善链脲佐菌素诱导的糖尿病（DM）大鼠和/或高糖（HG，30mM）刺激的HK-2细胞的凋亡。将大鼠随机分为三组：1）对照组，2）糖尿病组，3）糖尿病+RSV组（DM+RSV；大鼠以30mg/kg/d的RSV治疗16周）。然后检查其生理、生化和形态学参数。此外，还测量了SIRT1的去乙酰化酶活性，以及SIRT1和代表性凋亡标志物如p53、乙酰化p53、裂解的半胱天冬酶-3、半胱天冬酶-9和裂解的聚（ADP-核糖）聚合酶的表达水平。用HG刺激HK-2细胞不同时间，以研究HG对凋亡的影响。用或不用RSV（25μM）处理HK-2细胞，以研究RSV是否对HG诱导的凋亡有保护作用。使用针对SIRT1的基因特异性小干扰RNA来研究SIRT1在凋亡中的作用。发现糖尿病大鼠的凋亡比对照大鼠更多。同样，在HG条件下培养的HK-2细胞中，裂解的半胱天冬酶-3、裂解的聚（ADP-核糖）聚合酶和乙酰化p53的表达水平显著更高，而SIRT1水平显著更低，与在低糖（5.5mM）条件下培养的HK-2细胞相比。值得注意的是，RSV处理减轻了HG诱导的凋亡指标水平的变化，而SIRT1沉默消除了RSV处理对HK-2细胞中HG诱导凋亡的这种抑制作用。我们的研究表明，高血糖会导致大鼠肾脏和HK-2细胞凋亡。RSV激活SIRT1可在体外和体内降低尿白蛋白排泄和近端小管上皮细胞凋亡。基于我们的研究，SIRT1/p53轴在高血糖诱导的凋亡中起重要作用。这些发现表明，RSV介导的SIRT1表达增加是RSV预防糖尿病肾病（DN）肾小管损伤的一种可能机制。因此，RSV具有重要的临床意义，并可为有效治疗以控制与DN相关的发病率和死亡率的新方法提供依据。
