Sarkar Oli, Li Yuan, Anand-Srivastava Madhu B
Department of Pharmacology and Physiology, Faculty of Medicine, University of Montréal, Montréal, Canada.
PLoS One. 2017 Jul 10;12(7):e0179301. doi: 10.1371/journal.pone.0179301. eCollection 2017.
Vascular smooth muscle cells (VSMC) from spontaneously hypertensive rats (SHR) exhibit decreased levels of nitric oxide (NO) that may be responsible for the overexpression of Giα proteins that has been shown as a contributing factor for the pathogenesis of hypertension in SHR. The present study was undertaken to investigate if increasing the intracellular levels of NO by NO donor S-Nitroso-N-acetyl-DL-penicillamine (SNAP) could attenuate the enhanced expression of Giα proteins in VSMC from SHR and explore the underlying mechanisms responsible for this response. The expression of Giα proteins and phosphorylation of ERK1/2, growth factor receptors and c-Src was determined by Western blotting using specific antibodies. Treatment of VSMC from SHR with SNAP for 24 hrs decreased the enhanced expression of Giα-2 and Giα-3 proteins and hyperproliferation that was not reversed by 1H (1, 2, 4) oxadiazole (4, 3-a) quinoxalin-1-one (ODQ), an inhibitor of soluble guanylyl cyclase, however, PD98059, a MEK inhibitor restored the SNAP-induced decreased expression of Giα proteins towards control levels. In addition, the increased production of superoxide anion, NAD(P)H oxidase activity, overexpression of AT1 receptor, Nox4, p22phox and p47phox proteins, enhanced levels of TBARS and protein carbonyl, increased phosphorylation of PDGF-R, EGF-R, c-Src and ERK1/2 in VSMC from SHR were all decreased to control levels by SNAP treatment. These results suggest that NO decreased the enhanced expression of Giα-2/3 proteins and hyperproliferation of VSMC from SHR by cGMP-independent mechanism and involves ROS and ROS-mediated transactivation of EGF-R/PDGF-R and MAP kinase signaling pathways.
自发性高血压大鼠（SHR）的血管平滑肌细胞（VSMC）中一氧化氮（NO）水平降低，这可能是Giα蛋白过度表达的原因，而Giα蛋白过度表达已被证明是SHR高血压发病机制的一个促成因素。本研究旨在探讨通过NO供体S-亚硝基-N-乙酰-DL-青霉胺（SNAP）增加细胞内NO水平是否能减弱SHR来源的VSMC中Giα蛋白的增强表达，并探索这种反应的潜在机制。使用特异性抗体通过蛋白质印迹法测定Giα蛋白的表达以及ERK1/2、生长因子受体和c-Src的磷酸化。用SNAP处理SHR来源的VSMC 24小时可降低Giα-2和Giα-3蛋白的增强表达以及细胞过度增殖，而可溶性鸟苷酸环化酶抑制剂1H（1,2,4）恶二唑（4,3-a）喹喔啉-1-酮（ODQ）并不能逆转这种情况，然而，MEK抑制剂PD98059可使SNAP诱导的Giα蛋白表达降低恢复至对照水平。此外，SHR来源的VSMC中超氧阴离子产生增加、NAD(P)H氧化酶活性增强、AT1受体、Nox4、p22phox和p47phox蛋白过度表达、丙二醛（TBARS）和蛋白质羰基水平升高、血小板衍生生长因子受体（PDGF-R）、表皮生长因子受体（EGF-R）、c-Src和ERK1/2磷酸化增强，经SNAP处理后均降至对照水平。这些结果表明，NO通过不依赖cGMP的机制降低了SHR来源的VSMC中Giα-2/3蛋白的增强表达和细胞过度增殖，并且涉及活性氧（ROS）以及ROS介导的EGF-R/PDGF-R反式激活和丝裂原活化蛋白激酶信号通路。
