非酒精性脂肪变性肝细胞模型中ATF4基因的表达及其意义

Expression and significance of activating transcription factor 4 in oleic acid-induced hepatocyte steatosis in vitro

目的研究内质网应激相关蛋白活化转录因子4(activating transcription factor4,ATF4)基因在油酸诱导脂肪变性的肝细胞模型中的表达变化及其意义。方法以常规培养的L02肝细胞为对照,采用油酸诱导L02肝细胞脂肪变性,分别于0、24、48、72h采用RT-PCR与Western blot方法动态监测ATF4基因的表达变化。结果随着时间的增长,肝细胞脂肪变性程度加剧,肝细胞中ATF4 mRNA表达量0、24、48、72h分别为(1.01±0.12)、(1.87±0.24)、(2.01±0.13)、(1.79±0.19),48h时达到峰值后开始缓慢下降;ATF4蛋白表达量0、24、48、72h分别为(0.31±0.16)、(0.57±0.14)、(0.91±0.20)、(0.89±0.17),直至72h时仍维持在高水平状态,各实验组(24、48、72h组)与空白对照组(0h组)比较差异有统计学意义(P<0.05)。结论脂肪变性肝细胞中ATF4表达增强,提示肝细胞脂肪变性可能引起内质网发生应激,其可能与非酒精性脂肪性肝病关系密切。

Objective To investigate the expression and significance of activating transcription factor 4 （ATF4） in the hepatocyte steatosis models induced by oleic acid,and to explore the relationship between the expression of ATF4 and nonalcoholic fatty liver disease （NAFLD）. Methods L02 cells were treated by 20 mg/ml oleic acid for 0,24,48 and 72 h respectively to induce the hepatocytes steatosis,and then the cells were collected and the total RNA and protein were extracted. Reverse transcriptional polymerase chain reaction （RT-PCR） and Western blot analysis were used to detect the expressions of ATF4 gene at different time points. Results With the development of deterioration of steatosis,the expression levels of ATF4 mRNA were 1.01±0.12,1.87±0.24,2.01±0.13,and 1.79±0.19 respectively correspondingly from 0 h to 72 h. The peak of ATF4 mRNA expression was reached at the 48th hour,and then showed the tendency of down-regulation. At the same time points,the protein expression of ATF4 were 0.31±0.16,0.57±0.14,0.91±0.20,and 0.89±0.17 separately,and the protein expression was maintained at a high level up to the 72nd hour. There were significantly differences compared with the control group（P0.05）. Conclusion In the oleic acid-induced hepatocyte steatosis model,the expression of the ATF4 is enhanced persistently,suggesting where the endoplasmic reticulum stress response may happen. ATF4 may play an important role in the development and progression of NAFLD by up-regulating ATF4 expression.