CGRP与VEGF对内皮细胞成血管的作用比较研究

[目的]比较降钙素基因相关肽(calcitonin gene-related peptide,CGRP)与血管内皮细胞生长因子(vascular endothelial growth factor,VEGF)对血管内皮细胞增殖、迁移、体外成血管作用的大小.[方法]体外分离培养获取脐静脉血管内皮细胞(human umbilical vein endothelial cells,HUVECs).并通过血管性血友病因子(vonWillebrand factor,Vwf)与CD31抗原鉴定.实验分9组,(10-12～10-8)CGRP,(5 ～20 ng/ml) VEGF165,另设置一个空白对照组.采用细胞免疫荧光脱察HUVECs的CGRP受体(calcitonin gene-related peptide receptors,CGRPR)表达情况,alarmar Blue法动态检测各组HUVECs的增殖率,Transwell小室检测各组HUVECs的迁移能力,采用体外成管实验检测各组促血管生成作用大小.Q-PCR检测CGRP和VEGF165刺激细胞时,CGRP受体的mRNA表达情况.[结果]CGRP及VEGF165时间-浓度依赖性促进HUVECs增殖,VEGF165促内皮细胞增殖的能力强于CGRP,差异有显著性.CGRP及VEGF165都能明显促进HUVECs迁移及促进HUVECs在基质胶中形成毛细血管样结构.并且两者的最大作用基本相当.CGRP与VEGF165都能促进内皮细胞中CGRP受体表达水平增加.但是CGRP促CGRP受体的生成作用明显强于VEGF165.[结论]CGRP是强的促血管生成因子,其促血管生成的作用与低浓度(5 ～20ng/ml) VEGF165相当.

The effect on angiogenesis of human umbilical vein endothelial cells(HUVECs):CGRP VS VEGF

[Objective]To compare the effects on proliferation,migration and tube formation of human umbilical vein endothelial cells(HUVECs) between calcitonin gene-related peptide(CGRP) and vascular endothelial growth factor(VEGF).[Method]HUVECs were collected from human umbilical core,and identified through von Willebrand factor(VWF) and CD31 immunofluorescence.Experiments were divided in totally 9 groups:CGRP(10-12-10-8M) group,VEGF165(5-20 ng/ml) and control group.Immunoflurescence observed CGRPR on HUVECs.The growth rate of HUVECs was detected through alarmarflue at 1,2,3,4 and 5 day.Transwell chamber detected the ability of cell migration and tube formation experiment detected the role of angiogenesis.The mRNA expression levels of CGRP receptors were detected through real-time flurescence quantitative PCR.[Result]HUVECs were identified through morphology,VWF and CD31 immunofluorescence.HUVECs expressed CGRP receptors.GRPM and VEGF165 stimulated the proliferation of HUVECs in a concentration and time dependent manner and the effect of VEGF 165 was stronger than CGRP.CGRP and VEGF165 can also significantly promoted HUVECs migration and tube formation.The maximum effect of CGRP(10-12-10-10M) on migration and tube formation were equivalent with that of VEGF165(5-20 ng/ml).The Q-PCR results showed that CGRP and VEGF165 can significantly promoted the mRNA expression level of CGRP receptors.But,the level of CGRP receptors mRNA were significantly higher in CGRP-treated group.[Conclusion] CGRP is a strong proangiogenic growth factor and the effect of CGRP on angiogenesis is equivalent or stronger than low dose VEGF165(5-20 ng/ml).