活血注射液对ox-LDL诱导人脐静脉内皮细胞活化的影响

Effect of Huoxue injection on oxidized low-density lipoprotein induced activation of human umbilical vein endothelial cells

目的:观察活血注射液对氧化型低密度脂蛋白(ox-LDL)诱导的人脐静脉内皮细胞(HUVEC)的细胞间黏附分子-1(ICAM-1)、血管细胞黏附分子-1(VCAM-1)的表达及与人单核细胞黏附作用的影响,并探讨核转录因子kappa B(NF-κB)在其中的调控作用。方法:HUVEC与单核细胞的黏附率用蛋白定量法检测;ICAM-1,VCAM-1 mRNA和蛋白表达分别用RT-PCR技术和流式细胞仪检测;NF-κB p65的阳性细胞百分率和细胞核/浆染色的灰度比值用免疫细胞化学法测定。结果:ox-LDL作用HUVEC后12,24 h时,人单核细胞与HUVEC的黏附率显著升高,HUVEC中ICAM-1和VCAM-1的mRNA和蛋白表达水平也均明显升高,HUVEC的NF-κB p65的阳性细胞百分率和细胞核/浆染色的灰度比值明显增高,明显高于正常组(P<0.05,P<0.01),而活血注射液可明显降低人单核细胞与HUVEC的黏附率和HUVEC ICAM-1,VCAM-1 mRNA和蛋白表达水平,以及显著降低HUVEC中NF-κB p65的阳性细胞百分率和细胞核/浆染色的灰度比值,与ox-LDL组相比均有显著性差异(P<0.05,P<0.01),这种作用随着剂量的增加而增强。结论:活血注射液能通过下调内皮细胞表面ICAM-1,VCAM-1 mRNA和蛋白表达抑制单核-血管内皮细胞黏附,其可能机制是其通过抑制内皮细胞NF-κB p65的活性,从而发挥对血管内皮细胞的保护作用。

Objective： To observe effect of Huoxue injection on the expression of intercellular cell adhesion molecule-1 （ICAM-1） and vascular cell adhesion molecule-1 （ VCAM-1 ）, the adherence of monocytes to endothelial cells, and the regulation role of nuclear factor kappa B （NF-KB） in cultured human umbilical vein endothelial cells （HUVEC） injury induced by the oxidized low- density lipoprotein （ox-LDL）. Method： The ox-LDL （100 mg·L^-1） was added to the cultured HUVEC to prepare the injury model of HUVEC. The adhesive percentage between HUVEC treated with ox-LDL and monocytes was determined by protein quantification. Expression of mRNA and protein of ICAM-1 and VCAM-1 were determined by RT-PCR and flow cytometry respectively. The percentage of positive cells and the ratio of nuclei and cytoplasm of NF-κB p65 staining in HUVEC the were examined by cell immunochemistry. Result： Treatment of HUVEC with ox-LDL for 12, 24 hours significantly increased adhesion of monocytes to HUVEC and enhanced the expressions of mRNA and protein of ICAM-1 and VCAM-1. The percentage of positive cells and the ratio of nuclei and cytoplasm of NF-κB p65 staining in HUVEC were significantly increased after treatment with ox-LDL for 24 hours. Huo Xue Injection could significantly inhibit the adhesion between monocyte and HUVEC, the expression of mRNA and protein of ICAM-1 and VCAM-1, and declined the percentage of positive cells and the ratio of nuclei and cytoplasm of NF-κB p65 staining in HUVEC. The effects were strengthened with increasing the deal of Huoxue injection. Conclusion. Huoxue injection has an inhibitory effect on the adherence of monocytes to HUVEC, probably by way of down-regulating the expression of mRNA and protein of ICAM-1 and VCAM-1 in HUVEC. The mechanism is probably associated with inhibiting the activation of NF-κB p65 of HUVEC. The effects of Huoxue injection can bring about the protective effect to endothelial cells injury induced by ox-LDL.