高平进 朱鼎良 陈玉英 易静 200025 上海第二医科大学附属瑞金医院上海市高血压研究所(高平进、朱鼎良)；上海第二医科大学生物物理教研室(陈玉英、易静) 中华心血管病杂志 1999 0 27 6


    关键词：细胞；表型；蛋白激酶类；聚合酶链反应 期刊 zhxxgbzz 0 基础研究 fur -->


    

【摘要】 目的 建立血管平滑肌细胞(VSMC)的收缩及合成表型,探讨蛋白激酶Cα(PKCα)亚型对其转化的影响。方法 用酶消化法及传代培养分别获2种VSMC表型细胞,细胞特性通过免疫细胞化学及透射电镜鉴定。用竞争性逆转录聚合酶链反应(竞争RT-PCR),观察2种表型细胞的PKCα表达。结果 酶消化法分离的 VSMC较传代培养的VSMC显示缬蛋白(desmin)表达呈强阳性。2种表型细胞经超微结构观察分别显示典型的收缩表型及合成表型变化。PKCα表达在酶消化法分离的VSMC中明显高于传代培养的VSMC(t=3.568;P<0.05)。结论 PKCα亚型在收缩表型的表达量明显高于合成表型,提示该亚型对维持 VSMC的收缩表型起了一定作用。研究中选用竞争RT-PCR方法克服了以往使用β-actin的局限性。

Phenotypic transition of vascularsmooth muscle cells and protein kinase Cα

GAO Pingjin, ZHU Dingliang, CHEN Yuying, et al. Shanghai Instituteof Hypertention, Rui-jin Hospital, Shanghai Second Medical University, Shanghai, 200025

【 Abstract】 Objective The purpose of this study was to investigate theexpression of protein kinase Cα (PKCα ) in two phenotypic states of vascular smooth muscle cell (VSMC) byestablishing contractile and synthetic phenotype.Methods VSMC enzymatically isolated from rat aorta were ofcontractile phenotype while the passaged cells were of synthetic phenotype. The cellphenotypes were identified by transmission electron microscopy and immunocytochemicalstaining. The gene expressions of PKCα were determined by competitive reverse transcriptase-PCR(RT-PCR).Results Thefreshly dispersed VSMC were positively stained by desmin antibodies compared with passagedVSMC. The two type of cells showed typical morphology of contractile and syntheticphenotypes, respectively. The expression of PKCα in freshly dispersed VSMC was significantly higher thanthat of VSMC of synthetic phenotype (t=3.568;P<0.05). Conclusion The present data demonstrate an association between PKCα expression and phenotypictransition. It is proposed that PKCα plays a distinct role in sustaining contractile phenotype of VSMC.The competitive RT-PCR used in this study overcomes the limitations of β -actin method.

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