L-NAME对人脐静脉内皮细胞VE-cadherin表达的影响

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。

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

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.