NO-induced vascular smooth muscle cell motility

说明(申请人提供)：一氧化氮(NO)通常被认为在血管中起到保护作用。然而，其他人和我们已经获得的证据表明，情况可能并不总是如此，但对NO的不同反应背后的机制尚不清楚。这项提议的目的是测试一个令人兴奋的新假说，即长期升高的胰岛素水平将一氧化氮在血管中的作用从保护物质转换为有害物质的能力。我们发现，在长期用胰岛素处理的血管平滑肌细胞中，NO对运动和增殖的抑制作用被消除。这些发现支持一种新的假说，即胰岛素作为血管平滑肌细胞对NO的表型反应的切换器。我们的初步结果表明，慢性胰岛素处理大鼠主动脉平滑肌细胞发现的NO的运动刺激作用与PI 3激酶活性的增加有关，并且需要血管紧张素II、适配蛋白GAB1和蛋白酪氨酸磷酸酶SHP2的功能可用性。其他人的研究发现，GAB1可以通过增加PIPs水平而被招募到质膜上。然而，慢性胰岛素治疗与GAB1和SHP2功能之间的机制联系尚未明确。此外，还没有进行实验来确定类似的机制是否适用于通过慢性胰岛素治疗来消除NO的抗增殖作用。最后，我们的结果的病理生理学意义尚不清楚。我们建议实现以下特定目标：目标1：确定增加血管紧张素II功能是否必要和/或足以解释胰岛素对PI3K活性和NO诱导的细胞运动的影响。目的：探讨慢性胰岛素治疗是否将GAB1重新募集到细胞膜上，以及GAB1和SHP2在细胞膜上的非胰岛素依赖募集是否能与慢性胰岛素治疗中未发现的NO的运动刺激效应相类似。目的：揭示高胰岛素血症抑制血小板衍生生长因子(PDGF)诱导的大鼠主动脉平滑肌细胞DNA合成的作用机制。目的：研究血管损伤中诱导型一氧化氮合酶的表达是否促进高胰岛素血症小鼠的新生内膜形成，而对正常胰岛素血症小鼠或AT1受体拮抗剂治疗的高胰岛素血症小鼠有相反的作用。

DESCRIPTION (provided by applicant): Nitric oxide (NO) is generally considered to play a protective role in blood vessels. However, others and we have obtained evidence that this may not always be the case but the mechanisms underlying diverse responses to NO are not known. The purpose of this proposal is to test an exciting new hypothesis on the capacity of chronically elevated insulin levels to switch the role of nitric oxide from protective to deleterious substance in blood vessels. We have found that the inhibitory effects of NO on both motility and proliferation are abrogated in vascular smooth muscle cells chronically treated with insulin. These findings support a new hypothesis on the role of insulin as a switcher of vascular smooth muscle cell phenotypic responses to NO. Our preliminary results indicate that the motility-stimulatory effect of NO, uncovered by chronic insulin treatment of cultured rat aortic smooth muscle cells, is associated with increased PI 3 kinase activity and requires the functional availability of angiotensin II, of the adapter protein Gab1 and the protein tyrosine phosphatase SHP2. Studies by others have found that Gab1 can be recruited to the plasma membrane via increased PIPS levels. However, the mechanistic linkage of chronic insulin treatment to Gab1 and SHP2 function has not been defined. Moreover, experiments to determine whether similar mechanisms may be applicable to abrogation of the antiproliferative effect of NO by chronic insulin treatment have not been performed. Finally, the pathophysiological significance of our results is unknown. We propose to implement the following specific aims: Aim 1: To determine whether increased angiotensin II function is necessary and/or sufficient to account for the effect of insulin on PI3K activity and NO-induced cell motility. Aim 2: To determine whether chronic insulin treatment recruits Gab1 to the cell membrane and whether insulin- independent recruitment of Gab1 or SHP2 to the cell membrane can mimic the motility-stimulatory effect of NO uncovered by chronic insulin treatment. Aim 3: To uncover mechanisms that describe how hyperinsulinemia attenuates the effect of NO as inhibitor of PDGF-induced DNA synthesis, in cultured rat aortic smooth muscle cells. Aim 4: To determine whether expression of inducible nitric oxide synthase in vascular injury enhances neointima formation in hyperinsulinemic mice, but has the opposite effect in normoinsulinemic mice or in hyperinsulinemic mice treated with an AT1 receptor antagonist.