血管紧张素Ⅱ 1型受体自身抗体通过上调自噬诱导INS-1胰岛β细胞凋亡

目的:本研究旨在探究血管紧张素Ⅱ1型受体自身抗体(AT1-AA)能否引起胰岛β细胞的凋亡,自噬是否参与其中以及其所发挥的作用。方法:10-6mol/L AT1-AA处理INS-1细胞24 h后,用流式细胞术、Western blot及Hoechst 33258染色检测细胞的凋亡水平;Western blot检测自噬相关蛋白LC3和beclin 1的蛋白水平。使用血管紧张素Ⅱ1型受体拮抗剂替米沙坦以及经典的自噬抑制剂3-甲基腺嘌呤(3-MA)预处理细胞1 h之后再加入AT1-AA处理24 h,检测细胞凋亡、自噬及存活率的变化情况。结果:10-6mol/L AT1-AA处理INS-1细胞24 h可明显降低细胞存活率(P0.05)。与阴性Ig G对照组相比,AT1-AA分别处理细胞12 h、24 h和36 h后细胞凋亡水平明显增高(P0.05);此外,LC3及beclin 1的蛋白水平也随处理时间的延长逐渐升高,且凋亡和自噬水平的升高均可被替米沙坦所阻滞。使用自噬抑制剂3-MA预处理之后,细胞的凋亡率较单独给予AT1-AA处理组明显降低(P0.05)。结论:AT1-AA可通过血管紧张素Ⅱ1型受体上调自噬来诱导INS-1胰岛β细胞凋亡。

Angiotensin II type 1 receptor autoantibodies induces INS-1 islet β-cell apoptosis by upregulation of autophagy

AIM: To explore whether the angiotensin Ⅱ type 1 receptor autoantibodies (AT1-AA) induces islet β-cell apoptosis and whether autophagy is involved in the process. METHODS: The INS-1 cells treated with AT1-AA at 10^-6 mol/L for 24 h, and then the apoptosis was analyzed by flow cytometry, Western blot and Hoechst 33258 staining. In addition, the expression of autophagy-related proteins such as LC3 and beclin 1 were determined by Western blot. The effects of AT1-AA on the apoptosis, autophagy and viability of INS-1 cells with or without 3-methyladenine (3-MA; a common autophagy inhibitor) or telmisartan (an angiotensin Ⅱ type 1 receptor blocker) pretreatment, were detected by flow cytometry, Western blot and CCK-8 assay. RESULTS: Treatment with AT1-AA at 10^-6 mol/L for 24 h significantly reduced the cell viability (P<0.05). Compared with the negative IgG control group, the apoptotic cells increased after incubation with AT1-AA for 12 h, 24 h and 36 h, respectively (P<0.05). Moreover, the protein levels of LC3 and beclin 1 also increased gradually with the prolongation of treatment time, and the elevation of apoptosis and autophagy were blocked by telmisartan. After pretreatment with 3-MA, the apoptotic rate of the cells was obviously decreased compared with the cells treated with AT1-AA alone. CONCLUSION: AT1-AA induces the apoptosis of INS-1 islet β cells by upregulating autophagy via the angiotensin Ⅱ type 1 receptor pathway.