TNF-α对人脐静脉内皮细胞活性及PD-L1蛋白表达的影响

The effects of recombinant human TNF-α on the expression of programmed death-1 ligand-1 and proliferation of human umbilical vein endothelial cells

目的研究外源性肿瘤坏死因子α(TNF-α)对原代人脐静脉内皮细胞(HUVECs)活性及程序性死亡因子配体PD-L1蛋白表达的影响,探讨动脉粥样硬化的免疫机制。方法体外培养HUVECs,加入不同浓度TNF-α持续刺激48h,CCK-8法检测450nm波长处吸光度值(OD值),计算细胞生长存活率。选同代细胞重复培养并干预,Westernblot方法检测细胞PD-L1蛋白表达。使用SB203580阻断p38MAPK通路后再用TNF-α干预HUVECs,比较各组细胞PD-L1蛋白表达。结果实验组HUVECs在450nm波长处的吸光度值较空白对照组明显降低(P<0.05),细胞存活率依次下降。实验组细胞PD-L1蛋白表达随TNF-α浓度增高逐渐降低(P<0.05)。TNF-α单独刺激组PD-L1蛋白表达较TNF-α与SB203580共刺激组明显降低(P<0.05),SB203580阻断效应明显。结论 TNF-α可明显抑制HUVECs细胞活性,并降低PD-L1蛋白表达,p38MAPK通路可能参与了这一过程。

Objective This study aims to （i） investigate the effect of TNF-α on the expression of PD-L1 and on the proliferation of human umbilical vein endothelial cells （HUVECs）, and （ii） study the immunopathogenesis of atherosclerosis. Methods HUVECs were incubated with various concentrations of TNF-α for 48 hours. Cell viability and proliferation were determined by CCK-8 assay. The protein of PD-L1 was analyzed by Western blotting. p38MAPK inhibitor SB203580 was added before TNF-α treatment. Results As the concentration of TNF-α increased from 5 ng / ml to 80 ng / ml, the OD value at 450 nm in the CCK-8 assay decreased significantly （P〈0.05）. The expression of PD-L1 protein was suppressed by TNF-α significantly when compared with the control group （P〈0.05）. The level of PD-L1 protein in TNF-α-treated group was lower than the normal group and TNF-α ＋ SB203580 group （P〈0.05）. Conclusion TNF-α was found to suppress the expression of PD-L1 and inhibit the proliferation of HUVECs. p38MAPK pathway may play a role in this process.