| Summary: | 碩士 === 臺北醫學大學 === 醫學研究所 === 97 === Atherosclerosis and its complications remain the principal causes of death in developed countries. Although the pathogenesis of atherosclerosis is not fully elucidated, one theory holds that atherosclerosis is an immune response of the vascular wall to injury. In response to injury and to various stimuli, the activated vascular endothelium produces cytokines and growth factors to promote the growth and the migration of vascular smooth muscle cells, key events in the formation of atherosclerotic lesions in humans. Thus, one of the most important goals in the study of prevention of atherosclerosis is to identify factors that inhibit vascular smooth muscle cell proliferation and migration. Previously, we have demonstrated that progesterone (P4) inhibits the growth of vascular smooth muscle cells in vitro. P4 inhibits rat aortic smooth muscle cells (RASMC) proliferation by increasing the levels of p21 and p27 protein, which in turn inhibit CDK2 kinase activity, and finally interrupt the cell cycle. In this thesis research, we examined the effect of P4 on vascular smooth muscle cell migration and its molecular mechanisms underlying. We demonstrated that progesterone at physiologic levels (5-500 nM) concentration-dependently inhibited migration of cultured rat aortic smooth muscle cells (RASMC). The effect is blocked by pretreatment with the progesterone receptor (PR) antagonist RU486, indicating that migration inhibition is mediated through the PR. Western blot analyses demonstrated that the protein levels of RhoA, but not RhoB, RhoC, Ras and Rac1, were reduced in the progesterone-treated RASMC. Progesterone also inhibited the membrane translocation of RhoA protein. Moreover, the progesterone-induced migration inhibition in RASMC was prevented by over-expression of constitutively active RhoA construct (RhoA V14). However, pretreatment of RASMC with a ROCK (a kinase associated with RhoA for transducing RhoA signaling) inhibitor, Y27632, abolished the over-expression of RhoA-induced prevention effect on the progesterone-induced migration inhibition. These data suggest that the inhibition of Rho GTPases might account for the progesterone-induced migration inhibition of RASMC. Surprisingly, we found that the levels of phosphorylated Src (p-c-Src) and PR-p-c-Src complex in RASMC were increased at 10 sec after progesterone treatment. Moreover, treatment of RASMC with a Src inhibitor, PP2, blocked the progesterone-induced RhoA down-regulation and migration inhibition, suggesting that activation of Src might contribute to the RhoA inactivation leading to the migration inhibition caused by progesterone in RASMC. These findings expend our knowledge of the basis of progesterone’s effect on vascular smooth muscle cell migration and highlight novel pathways of signaling transduction of progesterone through PR-mediated nongenomic mechanisms.
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