Department of Pharmaceutical and Pharmacological Sciences, University of Padova, Padova, Italy.
PLoS One. 2013 May 29;8(5):e65666. doi: 10.1371/journal.pone.0065666. Print 2013.
Sirtuins enzymes are a conserved family of nicotinamide adenine dinucleotide (NAD)-dependent deacetylases and ADP-ribosyltransferases that mediate responses to oxidative stress, fasting and dietary restriction in mammals. Vascular smooth muscle cells (VSMCs) are involved in many mechanisms that regulate vascular biology in vivo but the role of SIRT1 has not been explored in much detail. Therefore, we investigated the regulation of SIRT1 in cultured VSMCs under various stress conditions including diabetes. Sprague-Dawley rats were made diabetic by injecting a single dose of streptozotocin (65 mg/Kg), and aortic VSMCs were isolated after 4 weeks. Immunocytochemistry showed that SIRT1 was localized predominantly in the nucleus, with lower staining in VSMCs from STZ-diabetic as compared with normoglycemic rats. Previous diabetes induction in vivo and high glucose concentrations in vitro significantly downregulated SIRT1 amounts as detected in Western blot assays, whereas TNF-α (30 ng/ml) stimulation failed to induce significant changes. Because estrogen signaling affects several pathways of oxidative stress control, we also investigated SIRT1 modulation by 17β-estradiol. Treatment with the hormone (10 nM) or a selective estrogen receptor-α agonist decreased SIRT1 levels in VSMCs from normoglycemic but not in those from STZ-diabetic animals. 17β-estradiol treatment also enhanced activation of AMP-dependent kinase, which partners with SIRT1 in a signaling axis. SIRT1 downregulation by 17β-estradiol could be observed as well in human peripheral blood mononuclear cells, a cell type in which SIRT1 downregulation is associated with insulin resistance and subclinical atherosclerosis. These data suggest that SIRT1 protein levels are regulated by diverse cellular stressors to a variable extent in VSMCs from diabetic and normoglycemic rats, warranting further investigation on SIRT1 as a modulator of VSMC activity in settings of vascular inflammation.
Sirtuins 酶是一类保守的烟酰胺腺嘌呤二核苷酸 (NAD) 依赖性去乙酰化酶和 ADP-核糖基转移酶家族，介导哺乳动物对氧化应激、禁食和饮食限制的反应。血管平滑肌细胞 (VSMCs) 参与许多调节体内血管生物学的机制，但 SIRT1 的作用尚未得到充分探索。因此，我们研究了在各种应激条件下，包括糖尿病，培养的 VSMCs 中 SIRT1 的调节。通过单次注射链脲佐菌素 (65mg/Kg) 将 Sprague-Dawley 大鼠制成糖尿病模型，4 周后分离主动脉 VSMCs。免疫细胞化学显示，SIRT1 主要定位于细胞核，而与正常血糖大鼠相比，STZ 糖尿病大鼠的 VSMCs 中染色较少。体内先前的糖尿病诱导和体外高葡萄糖浓度显著下调了 Western blot 检测到的 SIRT1 含量，而 TNF-α (30ng/ml) 刺激未能诱导显著变化。由于雌激素信号影响氧化应激控制的几个途径，我们还研究了 17β-雌二醇对 SIRT1 的调节。用激素 (10nM) 或选择性雌激素受体-α激动剂处理可降低正常血糖 VSMCs 中的 SIRT1 水平，但不能降低 STZ 糖尿病动物中的 SIRT1 水平。17β-雌二醇处理还增强了 AMP 依赖性激酶的激活，SIRT1 与 AMP 依赖性激酶在信号轴中是伙伴关系。在人外周血单核细胞中也可以观察到 17β-雌二醇对 SIRT1 的下调，在人外周血单核细胞中，SIRT1 的下调与胰岛素抵抗和亚临床动脉粥样硬化有关。这些数据表明，SIRT1 蛋白水平在糖尿病和正常血糖大鼠的 VSMCs 中受到不同程度的多种细胞应激的调节，需要进一步研究 SIRT1 作为血管炎症环境中 VSMC 活性调节剂的作用。
