糖尿病基础研究

　　[摘要] 目的 探讨细胞外信号调节激酶1/2(ERK1/2)信号转导通路在人白细胞介素1β(IL-1β)抑制小鼠胰岛素瘤胰岛β细胞(βTC-6)葡萄糖刺激下胰岛素分泌反应(GSIS)中的作用。 方法 (1)βTC-6细胞分别于含不同浓度(0、1.38、5.5 mmol/L)葡萄糖的KRB缓冲液中孵育60 min，放射免疫法检测细胞上清液中胰岛素浓度;(2)βTC-6细胞分别于含不同浓度(0、1.38、5.5 mmol/L)葡萄糖的KRB缓冲液中孵育5 min，蛋白质免疫印迹法检测细胞蛋白中磷酸化ERK1/2及β-Actin的水平;(3)βTC-6细胞加入IL-1β(0.15、1.5、15 ng/ml)培养24 h后于含1.38mmol/L葡萄糖的KRB缓冲液中孵育60 min,检测上清液中胰岛素浓度;(4)βTC-6细胞加入IL-1β(0.15、1.5、15 ng/ml)培养24 h后于含1.38 mmol/L葡萄糖的KRB缓冲液中孵育5 min，检测细胞蛋白中磷酸化ERK1/2及β-Actin的水平。 结果 (1)βTC-6细胞在1.38 mmol/L葡萄糖刺激时胰岛素分泌及ERK1/2磷酸化水平均达到高峰;(2)IL-1β可抑制βTC-6细胞葡萄糖刺激下的ERK1/2磷酸化及胰岛素分泌，作用与剂量呈正相关。 结论 ERK1/2信号转导通路可能在IL-1β抑制βTC-6细胞葡萄糖刺激下的胰岛素分泌反应中发挥重要作用。

　　[关键词]细胞外信号调节激酶1/2;信号转导通路;小鼠胰岛素瘤胰岛β细胞;糖尿病

　　[Abstract] Objective To investigate effects of extracellular signal-regulated kinase1/2 (ERK1/2) signal transduction pathway on glucose stimulated insulin secretion (GSIS) inhibited by IL-1β in βTC-6 cells. Methods(1)βTC-6 cells were incubated in Krebs-Ringer bicarbonate (KRB) buffer containing 0, 1.38 or 5.5 mmol/L glucose for 60 mins. Supernatants were collected for insulin assays by an insulin RIA kit.(2)βTC-6 cells were incubated in KRB buffer containing 0, 1.38 or 5.5 mmol/L glucose for 5 mins. At the end of the incubation, cells were suspended with lysis buffer and cell lysate was used for Western blot to test the levels of phosphorylated ERK1 / 2 and β-Actin's. (3) βTC-6 cells were incubated in DMEM containing IL-1β(0.15,1.5,15 ng/ml)for 24 hours. DMEM was then replaced with fresh KRB buffer containing 1.38 mmol/L glucose for 60 mins. Supernatants were collected for insulin assays.(4)βTC-6 cells were incubated in DMEM containing IL-1β(0.15,1.5,15ng/ml)for 24 hours. DMEM was then replaced with KRB buffer containing 1.38 mmol/L glucose for 5 mins. At the end of the incubation, cells were suspended with lysis buffer and cell lysate was used for Western blotting. Results (1)The insulin level and ERK1/2 phosphorylation reached a peak in response to 1.38 mmol/L glucose stimulation in βTC-6 cells. (2)IL-1β can inhibit ERK1/2 phosphorylation and insulin secretion stimulated by glucose stimulation in a dose-dependent manner in βTC-6 cells. Conclusion ERK1/2 signal transduction pathway activation may have an important effect on the GSIS inhibited by IL-1β in βTC-6 cells.

　　[Key words] Extracellular signal-regulated kinase1/2(ERK1/2); Signal transduction pathway;βTC-6 cell line;IL-1β; Diabetes mellitus