摘要
目的初步探讨Mibefradil对高糖作用下胰岛细胞胰岛素分泌的作用及其机制。方法体外培养大鼠胰岛瘤β细胞株(INS-1),将INS-1细胞分为对照组(11.1 mmol/L葡萄糖)、高糖组(33.3 mmol/L葡萄糖)、药物组(11.1 mmol/L葡萄糖+1μmol/L Mibefradil、1μmol/L NNC 55-0396、10μmol/L尼卡地平)、高糖+药物组(33.3 mmol/L葡萄糖+1μmol/L Mibefradil、1μmol/L NNC 55-0396、10μmol/L尼卡地平),先用不同浓度葡萄糖孵育细胞48 h,再按分组加入药物继续孵育24 h。采用放射免疫法检测细胞培养上清胰岛素含量,RT-PCR及Western blot检测T型钙通道cav3.1、cav3.2亚基表达。结果与对照组相比,高糖组能够明显增加细胞胰岛素分泌(P<0.05)和胰岛细胞T型钙通道cav3.1、cav3.2基因及蛋白表达(P<0.05),而药物组胰岛细胞胰岛素分泌水平和T型钙通道cav3.1、cav3.2亚基基因及蛋白表达则与对照组无显著差异(P>0.05);与高糖组相比,高糖+药物组可不同程度降低INS-1细胞胰岛素分泌,其中Mibefradil组显著降低胰岛素分泌(P<0.05)和T型钙通道cav3.1、cav3.2亚基基因及蛋白表达(P<0.05)。结论 Mibefradil可能通过下调胰岛细胞T型钙通道cav3.1、cav3.2亚基表达抑制高糖作用下INS-1细胞胰岛素分泌。
Objective To determine the role of mibefradil on the insulin secretion of the islet cells under high glucose environment. Methods Rat insulinoma INS-1 cells were cultured with RPMI 1640 including 10% FBS under 37℃ and 5% CO2 in vitro. The cells were randomly divided into 4 groups, that is, control group (11. 1 mmol/L glucose ), high glucose group (33. 3 mmol/L glucose ), drug group (11.1 mmol/L glucose + 1 μmol/L mibefradil, 1 μmoL/L NNC 55-0396, 10 μmol/L nicardipine), and high glucose+drug group (33.3 mmol/L glucose +1 μmol/L mibefradil, 1 μmol/L NNC 55-0396, 10 μmol/L nicardipine). The cells in each group had been treated with different concentrations of glucose for 48 h, and then incubated with corresponding drugs for another 24 h. The insulin secretion of INS-1 cells in the cell superuatant was measured by radioimmunoassay. The expression of T-type calcium channel subunits cav3.1 and cav3.2 at mRNA and protein levels was detected by RT-PCR and Western blotting respectively. Results Compared with the control group, high glucose inducement resulted in obviously increased insulin release in the cell supernatant ( P 〈 0.05 ), and markedly increased expression levels of cav3. 1 and cav3.2 ( P 〈 0. 05 ), In addition, there was no significant difference in the insulin release and the expression levels of the subunits between the control group and the drug group ( P 〉 0.05 ). Compared with the high glucose group, high glucose + drug treatment decreased the insulin release in the cell supematant. Among them, high glucose + mibefradil showed stronger inhibition on insulin secretion ( P 〈 0.05 ) and the expression of the subunits at mRNA and protein levels (P 〈 0.05 ). Conclusion Mibefradil may inhibits the insulin release of INS-1 cells under high glucose circumstance through down-regulating the expression levels of T-type calcium channels subunits cav3.1 and cav3.2.
出处
《第三军医大学学报》
CAS
CSCD
北大核心
2015年第7期660-665,共6页
Journal of Third Military Medical University