| Objective To investigate the expression of Pn in injured carotid artery and the relationship between Pn expression and the intimal hyperplasia in the injured carotid artery; to investigate the expression of Pn in vitro cultured vascular smooth muscle cells (VSMCs) induced by TGF-β1and the relationship between Pn expression and the migration and proliferation of the VSMCs; to investigate the effects of atorvastatin on the above-mentioned processes and the molecular mechanisms of atorvastatin inhibiting TGF-β1-induced Pn production. To provide new target point for preventing and curing vascular remodeling and proliferating vascular disease.Methods Animal models were prepared by the following way:the rats right carotid arteries were injured by brief drying with a gentle stream of air along the lumen of the vesse. Forty-five rats were randomly divided into nomal control (no operation,15rats),atorvastatin group(15rats) and model control(15rats).Five rats of each group were killed to get the right carotid artery to study at the end of1week,2weeks and4weeks. The intimal hyperplasia was studied by histological study. The expression of Pn was measured by immunohistochemical method and RT-PCR.The smooth muscle cells of rat aorta were cultivated by the method of tissue explants adherent. Cells of generation3rd to6th were used to test. Primary cultured rat vascular smooth muscle cells were treated by TGF-β1and atorvastatin of different concentration for24hours. The expression of Pn was measured by RT-PCR and western-blot. The Boyden chamber assay was used to measure cell migration and the MTT test was used to measure cell proliferation.Primary cultured rat vascular smooth muscle cells were stimulated by TGF-β1,which were treated by atorvastatin, Y-2763(Rho kinase inhibitor),atorvastatin plus MVA.24hours later, the expression of Pn was measured by RT-PCR and western-blot.Results (1) Intimal area in arteries of control group had no change at any time. Compared with that of the control group, the intimal area and the ratio of intima/media area in injured carotid arteries increased significanyly at the end of1week,2weeks and4weeks.(2) There was no Pn expression in normal carotid arteries at any time point. Pn expression of injured carotid arteries increased signifcantly at the end of1week,2weeks and4weeks.(3) Compared with those of model control, the Pn expression,the intimal area and the ratio of intima/media area in atorvastatin group decreased significantly at the end of2weeks and4weeks.(4) Pn expression in rat VSMCs stimulated by TGF-β1increased significantly, VSMCs migration and proliferation also increased significantly. Pn antibody can inhibit VSMC migration and proliferation induced by TGF-β1.(5) TGF-β1-induced Pn production in rat VSMCs would be inhibited by Atorvastatin significantly and VSMCs migration and proliferation inhibited in a dose-dependent manner significantly(6) TGF-β1-induced Pn production in rat VSMCs would be inhibited by Rho kinase inhibitor Y-27632significantly, as shown by the inhibition of atorvastatin.(7) The inhibitory effect of atorvastatin on Pn upregulation induced by TGF-β1was reversed by mevalonate.Conclusions (1) Pn could promote rat VSMCs migrating and proliferating. Pn acts as an important role in the process of intimal hyperplasia in injured arteries.(2) The promotive effect of TGF-β1on Pn expression in VSMCs was mediated by Rho/Rho kinase signaling.(3) The inhibitory effect of atorvastatin on Pn upregulation induced by TGF-β1in VSMCs might be mediated by inhibiting the production of MVA and other isoprene compounds and blocking the Rho/Rho kinase signaling.(4)That Atorvastatin inhibiting VSMCs migration, proliferation and intimal hyperplasia in rat injured arteries might be by means of inhibiting Pn expression. |
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