目的:研究游离脂肪酸(FFA)诱导的非酒精性脂肪性肝炎(NASH)细胞模型,动态观测细胞脂肪变性、炎症、过氧化损伤及糖转运缺陷发生发展的特点。方法:以HepG2细胞为实验对象,采用不同浓度游离脂肪酸(0.25mM,0.5mM,1mM)对HepG2细胞进行不同时间(12小时,18小时,24小时)的干预刺激,并通过油红O染色,酶联免疫的方法分别对细胞内脂肪变及细胞培养上清液中白介素-8(IL-8)、肿瘤坏死因子-α(TNF-α)、超氧化物歧化酶(SOD)、丙二醛(MDA)、葡萄糖转运蛋白4(GLUT4)进行测定评估。结果:FFA作用的实验各组,随着时间的延长及浓度的增加,细胞脂肪样变、IL-8、TNF-α、SOD、MDA及GLUT4均有不同程度的改变,以24小时0.5mM FFA、24小时1mM FFA最为明显,与同期空白对照组相比,其在IL-8、TNF-α、SOD、MDA、GLUT4的表达方面均有统计学意义(P〈0.05),油红O染色可见典型的"印戒"状。结论:应用游离脂肪酸诱导建立体外NASH模型成模效果好,重复率高,以0.5mM FFA 24小时为佳。
Objective: To establish a cell model of nonalcoholic steatohepatitis (NASH) by free fatty acid (FFA) and to dynamically monitor the pathological features of steatosis, oxidative damage and inflammation in these cells. Methods: Liver cells HepG2 were exposed to increasing concentrations of a fresh mixture of exogenous FFA (0. 25mM, O. 5mM, lmM) in molar ratio 1:2 palmitic: oleic at different time points (12h, 18h, 24h) respectively. Oil red O staining and the IL-8, TNF-ot, SOD, MDA, GLUT4 in supernatant were evaluated. Results: Steatosis could be seen in FFA groups to different extents. IL-8, TNF- ct, SOD, MDA and GLUT4 gradually differed according to the change of stimulation time by different FFA concentrations, espe- cially in the groups with 0. 5mM and lmM FFA concentrations for 24h, of which groups there could be seen a typical steatosis. Conclusion: A cell model of NASH has been created successfully by FFA with 0. 5mM for 24h.