糖尿病诱导硫氧还蛋白相互作用蛋白(TXNIP)对小鼠胰岛β细胞衰老的影响*

目的: 本研究旨在观察糖尿病中硫氧还蛋白相互作用蛋白(TXNIP)的表达是否影响胰岛β细胞衰老。方法: 正常小鼠(db/m)、糖尿病小鼠(db/db)随机各取6只,血糖仪检测其空腹血糖值,Western blot检测胰腺组织TXNIP蛋白表达、免疫化学染色检测胰腺组织中衰老相关β-半乳糖苷酶活性,Western blot检测胰腺组织衰老相关指标p16、p21、Rb表达的变化。INS-1胰岛β细胞随机分7组(n=6),用各组慢病毒(30 μl)转染4~6 h后,嘌呤霉素(PM, 3 μg/m)筛选7 d,构建Normal组(正常组)、Scramble ShRNA组(干扰空病毒组)、TXNIP-ShRNA-1(TXNIP沉默一组)组、TXNIP-ShRNA-2(TXNIP沉默二组)组、TXNIP-ShRNA-3组(TXNIP沉默三组)、Ad-GFP组(过表达空病毒组)、Ad-TXNIP-GFP组(TXNIP过表达组)稳转INS-1胰岛β细胞株,检测其TXNIP蛋白表达、衰老相关β -半乳糖苷酶活性、衰老相关指标。结果: 与正常小鼠相比,db/db组小鼠空腹血糖显著上升(P＜0.01),胰腺组织TXNIP蛋白表达显著升高(P＜0.05),胰腺组织β -半乳糖苷酶阳性染色率增加,p16、p21、Rb蛋白表达显著升高(P＜ 0.05)。与Ad-GFP组相比,Ad-TXNIP-GFP组β -半乳糖苷酶阳性染色率增加,p16、p21、Rb蛋白表达均显著增加(P＜0.01)。与Scramble ShRNA组相比,TXNIP-ShRNA组β -半乳糖苷酶阳性染色率降低,p16、p21以及Rb蛋白表达均降低(P＜0.05)。结论: 糖尿病可通过上调TXNIP表达,诱导胰岛β细胞衰老。

Effect of diabetic induced thioredoxin interacting protein (TXNIP) on islet cell senescence

Objective: To investigate whether the increased expression of thioredoxin interacting protein (TXNIP) in diabetes affects the senescence of islet β cells. Methods: Six normal mice (db/m) and six diabetic mice (db/db) were randomly selected. Fasting blood glucose was measured by blood sugar meter, the expression levels of TXNIP protein, p16, p21 and Rb in pancreatic tissues were detected by Western blot, senescence-associated beta-galactosidase activity in pancreatic tissue was determined by immunochemical staining. INS-1 islet beta cells were randomly divided into 7 groups (n=6), and transfected with lentiviruses (30 μl) for 4 to 6 hours, then was screened with puromycin (PM, 3 μg/m) for 7 days to construct normal group, scramble ShRNA group (interference with airborne poison group), TXNIP-ShRNA-1 group (TXNIP silence group-1), TXNIP-ShRNA-2 group (TXNIP silence group 2), TXNIP-ShRNA-3 group (TXNIP silence group 3), Ad-GFP group (overexpression of the air virus group), Ad-TXNIP-GFP group (TXNIP overexpression group) stably transferred INS-1 islet beta cell line. TXNIP protein expression was detected by Western blot, aging-related beta-galactosidase activity was detected by immunochemical staining, the changes of expression of p16, p21 and Rb was determined by Western blot. Results: Compared with normal mice, the fasting blood glucose of db/db group was increased significantly (P＜0. 01), the expression of TXNIP protein was increased significantly in pancreatic tissues(P＜0. 05), positive staining rate of β- galactosidase was increased significantly in pancreatic tissues, p16/p21/Rb protein expression levels were increased significantly (P＜0. 05). Compared with Ad-GFP group, the positive staining rate of β- galactosidase in Ad-TXNIP-GFP group was increased significantly, p16/p21/Rb protein expression levels were increased significantly (P＜0. 01). Compared to the scramble ShRNA group, the positive staining rate of β- galactosidase in TXNIP-ShRNA group was decreased, p16/p21/Rb protein expression levels were decreased significantly (P＜0. 05). Conclusion: Diabetes can induce islet β-cell senescence by up-regulating TXNIP expression.