肿瘤坏死因子促进内皮细胞微颗粒释放的实验研究

Tumor necrosis factor-α induces release of endothelial microparticles from human endothelial cells

目的研究肿瘤坏死因子损伤人脐静脉内皮细胞促进内皮细胞微颗粒释放,提出内皮细胞微颗粒是反映内皮损伤和功能障碍的新指标。方法应用肿瘤坏死因子-α刺激人脐静脉内皮细胞,扫描电镜观察细胞表面形态及内皮细胞微颗粒的生成,应用流式细胞仪检测细胞培养液中血小板细胞黏附分子(CD31+)和玻连蛋白(CD51+)微颗粒的水平。结果扫描电镜观察到静息状态下内皮细胞生成的微颗粒较少,经肿瘤坏死因子-α刺激24h后内皮细胞表面呈“出疹样”改变,小泡数目明显增多,直径大小约1.0μm。流式细胞仪检测证实细胞培养液中肿瘤坏死因子-α刺激组较正常对照组内皮细胞微颗粒水平明显增高[CD31+内皮细胞微颗粒,(164±63)/1000内皮细胞比(42±10)/1000内皮细胞,P<0.05;CD51+内皮细胞微颗粒,(260±108)/1000内皮细胞比(19±4)/1000内皮细胞,P<0.05]。结论肿瘤坏死因子损伤内皮细胞导致内皮细胞微颗粒释放增多,内皮细胞微颗粒有望成为评估内皮损伤替代指标之一。

Objective The present study was designed to investigate whether Tumor necrosis factor （TNF）-α stimulates release of endothelial microparticles（ EMPs ） by human endothelial cells, and whether EMPs may serve as a promising marker for endothelial injury and dysfunction. Methods Human umbilical venous endothelial cells （HUVEC） were incubated with or without TNF-α for 24 hours at 37℃. EMPs generated on the surface of HUVEC were observed with a scanning electron microscopy. The CD31 and CD51 positive EMPs in culture supernatants were measured by flow cytometer. Results Fewer vesicles were observed on cell surface of control group, in TNF-et-stimulated one, however, cells manifested a blebby surface （eruption phenomenon ） and more vesicles on surface were observed. The levels of EMPs were significantly increased in TNF-et stimulated cells compared with controls [ CD31 ＋ EMP, （164 ± 63 ）/1000 cells vs. （42 ± 10）/1000 cells,P 〈0. 05;CD51 ＋ EMP, （260 ± 108）/1000 cells vs. （ 19±4）/1000 cells, P 〈0. 05]. Conclusion TNF-α can stimulate HUVEC to release EMPs which may serve as a surrogate marker for endothelial injury and dysfunction.