摘要
目的:探讨环磷腺苷效应元件结合蛋白(CREB)辅激活物-活性调节2传感器(TORC2)在脂联素抑制肝糖异生过程中的作用及机制。方法:实验室前期已构建的人脂联素真核表达质粒,并成功转染人肝细胞系L-02细胞。在此基础上采用蛋白免疫印迹试验、免疫组织化学等方法,对比不同葡萄糖浓度处理后的脂联素转染组和转染空质粒对照组L-02细胞中TORC2细胞内定位,以及磷酸化腺苷酸活化蛋白激酶(p-AMPK)、CREB、p-CREB蛋白表达差异,同时监测各组葡萄糖-6-磷酸酶(G-6-Pase)活性。结果:与对照组相比,转染组p-AMPK蛋白表达量显著增高(P<0.05),CREB、p-CREB蛋白表达量差异无统计学意义(P>0.05)。定位分析显示转染组TORC2主要位于细胞质,而对照组TORC2主要位于细胞核内。葡萄糖关键酶检测发现转染组G-6-Pase活性显著低于对照组(P<0.05)。结论:脂联素抑制糖异生的机制可能与活化p-AMPK、抑制TORC2去磷酸化、阻止TORC2进入细胞核以及抑制糖异生关键酶基因表达有关。
Objective：To explore the effect and mechanism of CREB coactivator-TORC2 in adiponectin inhibiting hepatic gluconeogensis.Method：Eukaryotic human adiponectin expression vector was constructed and transfected into human hepatocyte lined 2cells（L-02）in our preliminary work.On this basis,TORC2 subcellular localization,as well as expression levels of phosphorylated-AMPK（p-AMPK）,CREB and phosphorylated-CREB（pCREB）,and glucose-6-phosphatase（G-6-Pase）activities were analyzed comparatively between adiponectin transfected L-02 cells and null plasmid transfected L-02 cells by molecular biology methods.Result：A significant higher expression of p-AMPK and lower level of G-6-Pase were observed in adiponectin transfected cells when comparing with control group（P〈0.05）,while the significance were not observed in CREB and p-CREB.Subcellular localization analysis showed that TORC2 of adiponectin transfected group was mainly located in cytoplasm,while control group was in the nucleus.Conclusion：The mechanism of adiponectin regulating hepatic gluconeogenesis may be closely related to the activation of p-AMPK,preventing TORC2 from entering nucleus and suppressing the activity of G-6-Pase.Further studies are still needed to determine the signal pathway.
出处
《临床血液学杂志（输血与检验）》
CAS
2016年第4期609-612,共4页
Journal of Clinical Hematology(Blood Transfusion & Laboratory Medicine)
