【摘要】 目的:研究不同浓度尾加压素Ⅱ(UⅡ)及其受体拮抗剂urantide对培养大鼠肺动脉平滑肌细胞(PASMC)增殖的影响,探讨丝裂原活化蛋白激酶(MAPK)途径及早期生长反应因子1(Egr-1)在UⅡ诱导大鼠PASMCs增殖中的作用。方法:以组织贴块法原代培养大鼠PASMCs:(1)采用Brd U掺入实验测定不同浓度(1μmol/L、0.1μmol/L和0.01μmol/L)UⅡ及其受体拮抗剂对大鼠PASMCs增殖的影响;(2)采用real-time PCR检测UⅡ及其受体拮抗剂对细胞外信号调节激酶1/2(ERK1/2)、应激活化蛋白激酶(SAPK)、p38 MAPK及Egr-1mRNA表达的影响;(3)采用Western blotting检测UⅡ、urantide及ERK1/2抑制剂PD98059对PASMCs磷酸化ERK1/2(p-ERK1/2)、p-SAPK、p-p38 MAPK和Egr-1蛋白表达的影响。结果:(1)UⅡ在一定浓度范围(1μmol/L、0.1μmol/L和0.01μmol/L)呈浓度依赖性地促进肺动脉平滑肌细胞增殖(P<0.01或P<0.05),这种增殖作用可被urantide拮抗(P<0.05);(2)UⅡ上调ERK1/2、SAPK及Egr-1 mRNA表达(P<0.01或P<0.05),PD98059和(或)urantide可拮抗UⅡ诱导的ERK1/2、SAPK和Egr-1 mRNA的表达(P<0.01或P<0.05),但对p-p38 MAPK无影响;(3)UⅡ可增加PASMCs p-ERK1/2、p-SAPK及Egr-1蛋白表达(P<0.01或P<0.05),urantide及PD98059可拮抗UⅡ诱导的p-ERK1/2、p-SAPK和Egr-1蛋白表达(P<0.01,P<0.05)。结论:UⅡ可能通过ERK1/2途径诱导PASMCs增殖和上调Egr-1表达,Egr-1可能通过激活ERK1/2途径参与了PASMCs的增殖。更多还原

【Abstract】 AIM: To investigate the effect of urotensinⅡ( UⅡ) on the proliferation of cultured rat pulmonary arterial smooth muscle cells( PASMCs),and to explore whether mitogen-activated protein kinase( MAPK) signaling pathways and early growth response factor-1( Egr-1) involved in the regulation of the PASMCs proliferation stimulated by UⅡ.METHODS: The rat PASMCs were isolated and cultured in vitro with explant culture technique. The proliferation of cultured PASMCs stimulated by different doses of UⅡ was detected by Brd U incorporation. The mRNA expression of extracellular signal-regulated kinase 1 /2( ERK1 /2),stress-activated protein kinase( SAPK),p38 MAPK and Egr-1 in cultured PASMCs treated with UⅡ,U Ⅱ-specific antagonist urantide,and ERK1 /2 inhibitor PD98059 was detected by real-time PCR. The protein levels of phosphorylated ERK1 /2( p-ERK1 /2),p-SAPK,p-p38 and Egr-1 in cultured PASMCs were determined by Western blotting. RESULTS: UⅡ at concentrations of 1 μmol / L,0. 1 μmol / L and 0. 01 μmol / L increased the proliferation of cultured PASMCs in a dose-dependent manner( P < 0. 01 or P < 0. 05),with the maximal effect at a concentration of 1 μmol / L. However,urantide inhibited the promotion effect of UⅡ on PASMC proliferation( P < 0. 05).UⅡ up-regulated the mRNA expression of ERK1 /2,SAPK and Egr-1( P < 0. 01 or P < 0. 05),but not the p38 MAPK.However,the up-regulatory effect of U Ⅱ on ERK1 /2 and Egr-1 expression was inhibited by PD98059 and / or urantide( P < 0. 01 or P < 0. 05). UⅡ also increased the protein levels of p-ERK1 /2,p-SAPK and Egr-1( P < 0. 01 or P <0. 05),but the promotion effect was also inhibited by PD98059 and / or urantide( P < 0. 01 or P < 0. 05). CONCLUSION: UⅡ increases the proliferation of PASMCs,and U Ⅱand Egr-1 participates in UⅡ-mediated proliferation of cultured PASMCs through activation of ERK1 /2 signal pathway.更多还原