Mechanism of arsenic induced vascular disease

拟议研究的主要目的是确定暴露于低水平亚砷酸盐后促进闭塞性心血管疾病的血管细胞表型和增殖变化的细胞和分子机制。这些研究的假设是，亚砷酸盐通过刺激内皮细胞和平滑肌细胞中氧化介导的信号传导而引起血管疾病。此外，接触亚砷酸盐引起的氧化剂可能会剥夺血管舒张和抑制平滑肌细胞增殖所需的一氧化氮。第一个资助期的研究区分了对氧化敏感的细胞调节和对亚砷酸盐增加作出反应的氧化应激。与环境相关的低水平的亚砷酸盐和氧化剂被证明具有调节和增殖作用，而高水平的亚砷酸盐和氧化剂则激活应激途径和细胞死亡。拟议的研究将继续使用原代内皮细胞和平滑肌细胞来确定亚砷酸盐刺激活性氧的来源以及促进表型变化和增殖的下游信号。重点将放在启动NAD(P)H氧化酶产生超氧化物的信号级联上。高表达腺病毒载体的显性阴性策略将证明单个GTPase Rac1在启动这种活性和促进NF-B激活方面的作用，NF-B是一种促进细胞保护基因表达和细胞增殖的氧化敏感转录因子。最后，将小鼠长期暴露于低水平的亚砷酸盐中，以验证亚砷酸盐是否会降低血管扩张剂诱导的一氧化氮释放并促进NF-B依赖性脑血管增厚。这些研究将通过体内电子顺磁共振波谱和抑制NF-B激活的腺病毒结构来促进。

The primary objective of the proposed studies is to define the cellular and molecular mechanisms responsible for changes in vascular cell phenotype and proliferation, which promote occlusive cardiovascular disease following exposure to low levels of arsenite. The hypothesis for these studies is that arsenite causes vascular disease by stimulating oxidant- mediated signaling in endothelial and smooth muscle cells. In addition the oxidants caused by arsenite exposure may deprive the vasculature of nitric oxide required for vasodilation and suppression of smooth muscle cell proliferation. Studies in the first funding period made the distinction between oxidant-sensitive cell regulation and oxidant stress in response to increasing amounts of arsenite. Low, environmentally relevant levels of arsenite and oxidants were shown to be regulatory and proliferative, while high levels activate stress pathways and cell death. The proposed studies will continue to use primary endothelial and smooth muscle cells to define the source of arsenite-stimulated reactive oxygen and the downstream signals that promote phenotypic change and proliferation. Focus will be on the signal cascades that initiate superoxide production by NAD(P)H oxidase. Dominant negative strategies, with highly expressed adenoviral vectors, will demonstrate the role of the monomeric GTPase, Rac1, in initiating this activity and in promoting the activation NF-B, an oxidant-sensitive transcription factor that promotes expression of cytoprotective genes and cell proliferation. Finally, mice will be chronically exposed to low levels of arsenite to test the hypothesis that arsenite decreases vasodilator-induced nitric oxide release and promotes NF-B dependent thickening of brain blood vessels. These studies will be facilitated by in vivo electron paramagnetic resonance spectroscopy and an adenoviral construct that suppresses NF-B activation.