L-NAME对人脐静脉内皮细胞VE-cadherin表达的影响

中国临床解剖学杂志 ›› 2012, Vol. 30 ›› Issue (4): 412-416.

L-NAME对人脐静脉内皮细胞VE-cadherin表达的影响

杨宝林^1，3，　伍校琼²,　罗明英¹, 　朱武¹，　叶峰¹，　刘丽华¹，　关莹露¹，　蔡维君¹

中南大学基础医学院 1.组织与胚胎学系，　2.人体解剖学与神经生物学系，湖南长沙 410013；
3.南昌大学基础医学院人体解剖学教研室，江西南昌 330006

收稿日期:2011-10-26 出版日期:2012-07-25 发布日期:2012-07-27
通讯作者: 蔡维君,教授，E-mail:wjcai@yahoo.com E-mail:ybl0027@sina.com
作者简介:杨宝林（1977-），男，江西瑞金市人，在读博士，讲师，主要从事侧支血管生长机制及神经损伤与修复研究，Tel：（0731）82650436
基金资助:
国家自然科学基金（30971532，30771134）；教育部博士点基金（20090162110063）；湖南省自然科学基金（09JJ5015）

Effects of L-NAME on the expression of VE-cadherin in human umbilical vein endothelial cell

YANG Bao-lin ^1,3, WU Xiao-qiong², LUO Ming-ying¹, ZHU Wu¹, YE Feng¹, LIU Li-hua¹, GUAN Ying-lu¹, CAI Wei-jun¹

1. Department of Histology & Embryology, 2. Department of Anatomy & Neurobiology, School of Basic Medicine, Central South University, Changsha 410013,China; 3. Department of Anatomy, School of Basic Medicine, Nanchang University, Nanchang 330006, China

Received:2011-10-26 Online:2012-07-25 Published:2012-07-27

摘要/Abstract

摘要：

目的　探讨一氧化氮合酶（NOS）抑制剂N-硝基-L-精氨酸甲酯（L-NAME）对人脐静脉内皮细胞（HUVEC）的eNOS、VE-cadherin蛋白表达的影响。方法 HUVEC传代培养并用CD31特异性抗体进行鉴定，实验分为对照组、0.1 mmol/L L-NAME组、1.0 mmol/L L-NAME组，运用MTT比色法检测L-NAME对HUVEC细胞活性的影响，并采用免疫荧光细胞化学法或Western blot检测实验各组HUVEC细胞的eNOS、VE-cadherin等蛋白的表达，以观察L-NAME对HUVEC的eNOS蛋白、VE-cadherin蛋白的影响。结果 95%以上传代后的细胞为CD31免疫阳性细胞。MTT比色法结果显示，随着L-NAME的浓度增加，HUVEC的细胞活力值逐渐减弱。免疫荧光细胞化学或Western blot结果显示，与对照组比较，0.1 mmol/L L-NAME组和1.0 mmol/L L-NAME组 HUVEC中eNOS的表达减弱（P<0.01），并呈剂量依赖性降低；与对照组比较，0.1 mmol/L L-NAME组和1.0 mmol/L L-NAME组 HUVEC中VE-cadherin的表达增强（P<0.01），呈剂量依赖性增加。结论 L-NAME能抑制HUVEC 的eNOS表达和细胞活性，并影响细胞粘附连接的主要蛋白—VE-cadherin的表达。

关键词: N-硝基-L-精氨酸甲酯, 人脐静脉内皮细胞, 血管内皮钙粘蛋白

Abstract:

Objective To investigate the effects of N-nitro-L-arginine methyl ester （L-NAME）as a nitric oxide synthase inhibitor on the expression of eNOS and VE-cadherin in human umbilical vein endothelial cell (HUVEC). Methods Subculturing HUVECs were identified with specific antibody to CD31. 3 groups (control group, 0.1mmol/L L-NAME group, 1.0 mmol/L L-NAME group) were designed to carry out in this study. The effects of L-NAME on HUVEC activity was assessed with MTT assay. Expression of eNOS and Ve-cadherin protein in HUVEV of all group were detected with immunocytochemistry staining or Western blot. Results More than 95% subculturing HUVECs were CD31+ cell. MTT assay showed a L-NAME dose-dependent reduction in HUVEC activity when L-NAME was added into medium. The immunofluorescence intensity of eNOS positive staining in 0.1 mmol/L L-NAME group and 1.0mmol/L L-NAME group were obviously decreased as compared with control group（P<0.01）. The immunofluorescence intensity and the relative expression of VE-cadherin positive staining in 0.1mmol/L L-NAME group and 1.0 mmol/L L-NAME group were obviously increased as compared with control group（P<0.01）. Conclusions L-NAME suppresses the expression of eNOS and cell activity, but improves the expression of VE-cadherin in HUVEC.

Key words: N-nitro-L-arginine methyl ester, Human umbilical vein endothelial cell, VE-cadherin

中图分类号:

R331.31

杨宝林, 伍校琼, 罗明英, 朱武, 叶峰, 刘丽华, 关莹露, 蔡维君. L-NAME对人脐静脉内皮细胞VE-cadherin表达的影响[J]. 中国临床解剖学杂志, 2012, 30(4): 412-416.

YANG Bao-Lin, WU Jiao-Qiong, LUO Meng-Yang, SHU Wu, XIE Feng, LIU Li-Hua, GUAN Ying-Lou, CA Wei-Jun. Effects of L-NAME on the expression of VE-cadherin in human umbilical vein endothelial cell[J]. Chinese Journal Of Clinical Anatomy, 2012, 30(4): 412-416.