Heart, Lung, Blood and Vascular Medicine Institute (J.C.G., B.G.D., M.P.M., S.A.H., S.Y., K.C.W., A.C.S.) and Department of Pharmacology and Chemical Biology (J.C.G., A.C.S.), University of Pittsburgh, Pittsburgh, Pennsylvania.
Heart, Lung, Blood and Vascular Medicine Institute (J.C.G., B.G.D., M.P.M., S.A.H., S.Y., K.C.W., A.C.S.) and Department of Pharmacology and Chemical Biology (J.C.G., A.C.S.), University of Pittsburgh, Pittsburgh, Pennsylvania
Mol Pharmacol. 2019 Jun;95(6):629-637. doi: 10.1124/mol.118.115386. Epub 2019 Apr 15.
Nitric oxide (NO) stimulates soluble guanylyl cyclase (sGC) activity, leading to elevated intracellular cyclic guanosine 3',5'-monophosphate (cGMP) and subsequent vascular smooth muscle relaxation. It is known that downregulation of sGC expression attenuates vascular dilation and contributes to the pathogenesis of cardiovascular disease. However, it is not well understood how sGC transcription is regulated. Here, we demonstrate that pharmacological inhibition of Forkhead box subclass O (FoxO) transcription factors using the small-molecule inhibitor AS1842856 significantly blunts sGC and mRNA expression by more than 90%. These effects are concentration-dependent and concomitant with greater than 90% reduced expression of the known FoxO transcriptional targets, glucose-6-phosphatase and growth arrest and DNA damage protein 45 (Gadd45). Similarly, sGC and sGC protein expression showed a concentration-dependent downregulation. Consistent with the loss of sGC and mRNA and protein expression, pretreatment of vascular smooth muscle cells with the FoxO inhibitor decreased sGC activity measured by cGMP production following stimulation with an NO donor. To determine if FoxO inhibition resulted in a functional impairment in vascular relaxation, we cultured mouse thoracic aortas with the FoxO inhibitor and conducted ex vivo two-pin myography studies. Results showed that aortas have significantly blunted sodium nitroprusside-induced (NO-dependent) vasorelaxation and a 42% decrease in sGC expression after 48-hour FoxO inhibitor treatment. Taken together, these data are the first to identify that FoxO transcription factor activity is necessary for sGC expression and NO-dependent relaxation.
一氧化氮（NO）刺激可溶性鸟苷酸环化酶（sGC）的活性，导致细胞内环鸟苷酸 3'，5'-单磷酸（cGMP）的升高，随后血管平滑肌松弛。已知 sGC 表达的下调会减弱血管扩张，并导致心血管疾病的发病机制。然而，sGC 转录是如何调节的还不是很清楚。在这里，我们证明使用小分子抑制剂 AS1842856 抑制叉头框亚科 O（FoxO）转录因子的药理作用，可使 sGC 和 mRNA 表达分别显著降低超过 90%。这些效应呈浓度依赖性，并伴随着已知 FoxO 转录靶点葡萄糖-6-磷酸酶和生长停滞和 DNA 损伤蛋白 45（Gadd45）的表达降低超过 90%。同样，sGC 和 sGC 蛋白表达也表现出浓度依赖性下调。与 sGC 和 mRNA 和蛋白表达的丧失一致，用 FoxO 抑制剂预处理血管平滑肌细胞后，NO 供体刺激下的 cGMP 产生所测量的 sGC 活性降低。为了确定 FoxO 抑制是否导致血管舒张的功能障碍，我们用 FoxO 抑制剂培养小鼠胸主动脉，并进行离体双针肌描记研究。结果表明，FoxO 抑制剂处理 48 小时后，主动脉的硝普钠诱导的（NO 依赖性）血管舒张明显减弱，sGC 表达降低了 42%。综上所述，这些数据首次表明 FoxO 转录因子活性是 sGC 表达和 NO 依赖性松弛所必需的。
