業(yè)已證明,Caveolae及其蛋白caveolin-1參與了細(xì)胞膜的膽固醇轉(zhuǎn)運和細(xì)胞膜的信號轉(zhuǎn)導(dǎo).我們前期工作發(fā)現(xiàn)降鈣素基因相關(guān)肽(CGRP)抑制血管平滑肌細(xì)胞(VSMC)增殖的信號通路與抑制ERK1/2活性和上調(diào)caveolin-1表達(dá)有關(guān).本文研究Caveolae及caveolin-1在CGRP抑制VSMC增殖中的作用,進(jìn)一步研究caveolin-1表達(dá)增加是否有直接抑制ERK1/2信號激酶活性的作用.采用大鼠主動脈貼塊法培養(yǎng)VSMC,取3～10代VSMC用于實驗,10%小牛血清(FBS)用于刺激VSMC增殖,用β-環(huán)糊精(cyclodextrin)或菲律賓菌素(filipin)剝奪膽固醇破壞Caveolae結(jié)構(gòu);MTT法和流式細(xì)胞儀用于檢測細(xì)胞增殖;蛋白質(zhì)印跡和免疫共沉淀法分別用于檢測目的蛋白的表達(dá)或蛋白質(zhì)間相互作用.結(jié)果顯示,CGRP呈時間和濃度依賴性顯著抑制10%FBS誘導(dǎo)的VSMC增殖.細(xì)胞Caveolae結(jié)構(gòu)的破壞能降低CGRP抑制VSMC增殖作用,同時也增加了ERK1/2的磷酸化;β-環(huán)糊精孵育細(xì)胞能降低caveolin-1的表達(dá).免疫共沉淀發(fā)現(xiàn)10%FBS和/或CGR...

【文章頁數(shù)】：9 頁

【文章目錄】：
1 Materials and methods
    1.1 Drugs and reagents
    1.2 Rat vascular smooth muscle cell culture
    1.3 Determination of cell proliferation
    1.4 Western blotting
    1.5 Immunoprecipitation
    1.6 Statistical analysis
2 Results
    2.1 Inhibitory effect of CGRP on VSMC proliferation
    2.2 Effect ofβ-CD on the VSMC proliferation
    2.3 Effect ofβ-CD on phosphorylation of ERK1/2
    2.4 Effect of CGRP on expression of caveolin-1in the presence ofβ-CD
    2.5 CGRP enhanced the interaction between caveolin-1 and p-ERK1/2
3 Discussion



本文編號：4036909