关键词: HUVEC, 肿瘤坏死因子-α, 内皮型一氧化氮合酶, 蛋白质降解途径, MG-132

Abstract: Objective To investigate whether intracellular protein degradation pathway play a important role in the decrease of endothelial nitric oxide synthase (eNOS) in human umbilical vein endothelial cells (HUVECs), Which is induced by tumor necrosis factor-alpha (TNF-α). Methods Establish a primary HUVEC culture methods, the HUVECs were incubated with concentration gradient group of TNF-α (0.01, 0.1, 1 and 10 ng/mL) under different time periods (24, 48 and 72 h). The HUVECs were pretreated with NH4Cl or treated with caspase inhibitor or MG-132 1.5 h prior to incubation for an additional 24 h with TNF-α. The expression of eNOS was detected via Western blot assay. Results Compared with the control group, treatment of the HUVECs with TNF-α (0.01-10 ng/ml) led to a dose-dependent reduction in the expression of eNOS. And treatment with TNF-α (1 ng/ml) reduced the eNOS expression in a time-depended manner. Compared with the TNF-α group, the protein expression level of eNOS was obviously increased in the co-working group of MG-123 and TNF-α. Conclusions TNF-α induced the degradation of eNOS through the ubiquitin-proteasome pathway.

Key words: HUVEC, TNF-α, eNOS, protein degradation pathway, MG-132