Kuo Yi-Chen, Chen I-Yin, Chang Shin C, Wu Shun-Chi, Hung Tzu-Min, Lee Po-Huang, Shimotohno Kunitada, Chang Ming-Fu
Liver Int. 2014 Oct;34(9):1358-68. doi: 10.1111/liv.12389.
BACKGROUND & AIMS: Hepatitis C virus (HCV) infection is highly associated with the type 2 diabetes mellitus, but the detailed mechanisms by which the viral proteins are involved in the clinical outcome remain unclear.
A cDNA microarray analysis was performed following introducing an NS5A-encoding plasmid or a control vector into a mouse system by hydrodynamics- based transfection. Differentially expressed genes that are associated with gluconeogenesis were selected and their expression levels in HCV patients, in NS5A-expressing systems, and in the viral subgenomic replicon system were further examined by real-time quantitative polymerase chain reaction and Western blot analysis.
Differential gene expression including an upregulation of the gluconeogenic rate-limiting enzyme phosphoenolpyruvate carboxykinase (PEPCK) compared with controls was detected in mouse hepatocytes expressing HCV NS5A and in HCV patients with diabetes. In addition, an NS5A-dependent increase in glucose production was demonstrated in human primary hepatocytes. The upregulation of PEPCK and peroxisome proliferator-activated receptor-c coactivator-1a (PGC-1a) were also detected in NS5A-expressing cells and in the viral genotype 1b subgenomic replicon system. Further studies demonstrated that the NS5A-mediated upregulation of PEPCK and PGC-1a genes were resulted from the activation of PI3K-Akt and JNK signalling pathways. In addition, the expression levels of the forkhead transcription factor FoxO1 and the liver-enriched transcription factor HNF-4a were increased in HCV NS5A expressing cells.
By upregulating the expression of PEPCK gene via its transactivators FoxO1 and HNF-4a, and the coactivator PGC-1a, the NS5A promotes the production of hepatic glucose which may contribute to the development of HCV-associated type 2 diabetes mellitus.
丙型肝炎病毒（HCV）感染与2型糖尿病高度相关，但病毒蛋白参与临床结局的详细机制仍不清楚。
通过基于流体动力学的转染将编码NS5A的质粒或对照载体引入小鼠系统后，进行cDNA微阵列分析。选择与糖异生相关的差异表达基因，并通过实时定量聚合酶链反应和蛋白质印迹分析进一步检测其在HCV患者、NS5A表达系统和病毒亚基因组复制子系统中的表达水平。
在表达HCV NS5A的小鼠肝细胞和糖尿病HCV患者中检测到差异基因表达，包括与对照相比糖异生限速酶磷酸烯醇丙酮酸羧激酶（PEPCK）上调。此外，在人原代肝细胞中证实了NS5A依赖性葡萄糖生成增加。在表达NS5A的细胞和病毒基因型1b亚基因组复制子系统中也检测到PEPCK和过氧化物酶体增殖物激活受体-γ共激活因子-1α（PGC-1α）上调。进一步研究表明，NS5A介导的PEPCK和PGC-1α基因上调是由PI3K-Akt和JNK信号通路激活所致。此外，在表达HCV NS5A的细胞中，叉头转录因子FoxO1和肝脏富集转录因子HNF-4α的表达水平增加。
NS5A通过其反式激活因子FoxO1和HNF-4α以及共激活因子PGC-1α上调PEPCK基因的表达，促进肝葡萄糖生成，这可能有助于HCV相关2型糖尿病的发生发展。
