Bian Che, Bai Bowen, Gao Qian, Li Siyi, Zhao Yuyan
Department of Endocrinology, The First Affiliated Hospital of China Medical University, Shenyang, China.
Department of Endocrinology and Metabolism, The Fourth Affiliated Hospital of China Medical University, Shenyang, China.
Front Endocrinol (Lausanne). 2019 Aug 6;10:531. doi: 10.3389/fendo.2019.00531. eCollection 2019.
To explore the molecular mechanism by which 17β-estradiol (estrogen 2, E2) regulates glucose transporter 2 (GLUT2) and insulin secretion in islet β cells through G protein-coupled estrogen receptor (GPER) via Akt/mTOR pathway. SPF-grade SD male rats were used to establish an type 2 diabetes model treated with E2. Rat insulinoma cells (INS-1) were cultured in normal or high glucose media with or without E2. Immunofluorescence double staining was used to detect GPER, GLUT2, insulin, and glucagon immunolocalization in rat islet tissues. Western blot was used to detect GPER, Akt, mTOR, and GLUT2 protein immunocontent. Real-time PCR detected Slc2a2 and glucose kinase (GK) content, and ELISA was used to detect insulin levels. Glucose uptake, GK activity and pyruvate dehydrogenase (PDH) activity were analyzed with glucose detection, GK activity and PDH activity assay kit. Immunofluorescence double staining confocal indicated that E2 treatment up-regulated expression levels of GPER, GLUT2, and insulin, while down-regulated glucagon. Western blot results revealed E2 increased GPER, Akt/mTOR pathway, and GLUT2 protein immunocontent. Real-time PCR showed E2 elevated Slc2a2, GK content. Moreover, E2 improved insulin secretion, glucose uptake, GK activity, and PDH activity. Our findings indicated that exogenous E2 up-regulated GPER via the Akt/mTOR pathway to increase GLUT2 protein content and insulin secretion in islet β cells.
探讨17β-雌二醇（雌激素2，E2）通过G蛋白偶联雌激素受体（GPER）经Akt/mTOR途径调节胰岛β细胞中葡萄糖转运蛋白2（GLUT2）和胰岛素分泌的分子机制。使用SPF级SD雄性大鼠建立E2治疗的2型糖尿病模型。将大鼠胰岛素瘤细胞（INS-1）在含或不含E2的正常或高糖培养基中培养。采用免疫荧光双染色法检测大鼠胰岛组织中GPER、GLUT2、胰岛素和胰高血糖素的免疫定位。采用蛋白质免疫印迹法检测GPER、Akt、mTOR和GLUT2蛋白免疫含量。实时荧光定量PCR检测Slc2a2和葡萄糖激酶（GK）含量，酶联免疫吸附测定法检测胰岛素水平。使用葡萄糖检测试剂盒、GK活性和丙酮酸脱氢酶（PDH）活性测定试剂盒分析葡萄糖摄取、GK活性和PDH活性。免疫荧光双染色共聚焦显示，E2处理上调了GPER、GLUT2和胰岛素的表达水平，同时下调了胰高血糖素。蛋白质免疫印迹结果显示，E2增加了GPER、Akt/mTOR途径和GLUT2蛋白免疫含量。实时荧光定量PCR显示，E2提高了Slc2a2、GK含量。此外，E2改善了胰岛素分泌、葡萄糖摄取、GK活性和PDH活性。我们的研究结果表明，外源性E2通过Akt/mTOR途径上调GPER，以增加胰岛β细胞中GLUT2蛋白含量和胰岛素分泌。
