Higher Protein Kinase C ζ in Fatty Rat Liver and Its Effect on Insulin Actions in Primary Hepatocytes

We previously showed the impairment of insulin-regulated gene expression in the primary hepatocytes from Zucker fatty (ZF) rats, and its association with alterations of hepatic glucose and lipid metabolism. However, the molecular mechanism is unknown. A preliminary experiment shows that the expression level of protein kinase C ζ (PKCζ), a member of atypical PKC family, is higher in the liver and hepatocytes of ZF rats than that of Zucker lean (ZL) rats. Herein, we intend to investigate the roles of atypical protein kinase C in the regulation of hepatic gene expression. The insulin-regulated hepatic gene expression was evaluated in ZL primary hepatocytes treated with atypical PKC recombinant adenoviruses. Recombinant adenovirus-mediated overexpression of PKCζ, or the other atypical PKC member PKCι/λ, alters the basal and impairs the insulin-regulated expressions of glucokinase, sterol regulatory element-binding protein 1c, the cytosolic form of phosphoenolpyruvate carboxykinase, the catalytic subunit of glucose 6-phosphatase, and insulin like growth factor-binding protein 1 in ZL primary hepatocytes. PKCζ or PKCι/λ overexpression also reduces the protein level of insulin receptor substrate 1, and the insulin-induced phosphorylation of AKT at Ser473 and Thr308. Additionally, PKCι/λ overexpression impairs the insulin-induced Prckz expression, indicating the crosstalk between PKCζ and PKCι/λ. We conclude that the PKCζ expression is elevated in hepatocytes of insulin resistant ZF rats. Overexpressions of aPKCs in primary hepatocytes impair insulin signal transduction, and in turn, the down-stream insulin-regulated gene expression. These data suggest that elevation of aPKC expression may contribute to the hepatic insulin resistance at gene expression level.我们此前已证实，Zucker肥胖（Zucker fatty, ZF）大鼠的原代肝细胞（primary hepatocytes）中，胰岛素调控基因表达存在损伤，且该现象与肝脏糖脂代谢（hepatic glucose and lipid metabolism）紊乱相关。然而，其具体分子机制尚未阐明。 前期实验发现，非典型PKC家族（atypical PKC family）成员蛋白激酶Cζ（protein kinase C ζ, PKCζ）在ZF大鼠肝脏及原代肝细胞中的表达水平，高于Zucker瘦型（Zucker lean, ZL）大鼠。 本研究拟探讨非典型蛋白激酶C在肝脏基因表达调控中的作用。我们对经编码非典型PKC的重组腺病毒（recombinant adenoviruses）处理的ZL原代肝细胞的胰岛素调控肝脏基因表达进行了评估。 结果显示，通过重组腺病毒介导的PKCζ或另一非典型PKC家族成员PKCι/λ过表达，可改变ZL原代肝细胞的基础基因表达水平，并损伤胰岛素对葡萄糖激酶（glucokinase）、固醇调节元件结合蛋白1c（sterol regulatory element-binding protein 1c）、胞质型磷酸烯醇式丙酮酸羧激酶（the cytosolic form of phosphoenolpyruvate carboxykinase）、葡萄糖-6-磷酸酶催化亚基（catalytic subunit of glucose 6-phosphatase）以及胰岛素样生长因子结合蛋白1（insulin like growth factor-binding protein 1）的表达调控。 PKCζ或PKCι/λ过表达还可降低胰岛素受体底物1（insulin receptor substrate 1）的蛋白水平，并抑制胰岛素诱导的AKT在Ser473与Thr308位点的磷酸化。 此外，PKCι/λ过表达可损伤胰岛素诱导的Prckz基因表达，提示PKCζ与PKCι/λ之间存在信号串扰。 综上，胰岛素抵抗ZF大鼠的肝细胞中PKCζ表达水平升高。 在原代肝细胞中过表达非典型PKCs（aPKCs）可损伤胰岛素信号转导，进而影响下游胰岛素调控的基因表达。上述数据表明，非典型PKC表达升高可能在基因表达层面参与肝脏胰岛素抵抗的发生。