炎症状态下西罗莫司对小鼠胰岛β细胞功能的影响

Effect of sirolimus on pancreatic β cell function under inflammation condition

目的研究炎症状态下西罗莫司对胰岛β细胞功能的影响。方法采用8周龄C57BL/6J雄性小鼠,随机分为A,B,C,D组:A组为对照组,小鼠隔天皮下注射生理盐水;B组小鼠隔天皮下注射酪蛋白;C组小鼠隔天皮下注射酪蛋白和溶媒介质;D组小鼠隔天皮下注射酪蛋白和西罗莫司。所有小鼠均给予高脂饮食喂养14周。通过葡萄糖耐量和胰岛素耐量了解小鼠体内糖代谢情况。酶联免疫吸附法(ELISA)检测小鼠血清中肿瘤坏死因子(TNF-α)、血清淀粉样蛋白(SAA)、胰岛素含量。用聚合酶链式反应(PCR)分析小鼠体内凋亡基因(BAX,BCL2)和胰岛素合成分泌相关基因的表达。苏木精-伊红(HE)染色观察小鼠胰岛形态,TUNEL染色观察胰岛细胞的凋亡情况。结果与A组相比,B组小鼠血清中炎症因子(TNF-α,SAA)显著增加(P<0.05),糖耐量及胰岛素耐量受损。葡萄糖耐量实验中,在30,60,120 min时D组小鼠血糖水平明显高于C组。胰岛素耐量实验中,D组小鼠血糖水平在15,30,60,120 min时明显高于C组。与C组相比,D组小鼠血清胰岛素含量明显下降(P<0.05),胰岛体积变小,胰岛β细胞凋亡增加,胰岛组织中BAX/BCL2增加,胰岛素合成分泌相关基因表达明显下降(P<0.05)。结论炎症状态下,西罗莫司使胰岛β细胞凋亡增加,胰岛素的产生和分泌下降,胰岛β细胞的功能降低,最终导致小鼠糖代谢紊乱。

Objective To investigate the mechanism of sirolimus induced insulin resistance under inflammation stress.Methods Eight-week-old male C57 BL /6J mice were randomly assigned to A,B,C,D groups.Mice in group A received normal saline injection;group B received casein injection;group C received casein and vehicle injection;group D received casein and sirolimus injection.All injections were performed every other day.All mice were fed with a high-fat diet for 14 weeks.Then glucose tolerance test（ GTT） and insulin tolerance test（ ITT） were performed to test the glucose and insulin tolerances.The concentration of tumor necrosis factor（ TNF-α）,serum amyloid A（ SAA） and insulin in serum were measured by enzyme-linked immunosorbent assay（ ELISA）.Real-time PCR was used to determine the expression of apoptosis related gene（ BAX,BCL2） and key participants in β-cell functional integrity,such as pancreatic duodenal homeobox-1（ PDX1）,glucokinase（ GK）,glucose transporter 2（ GLUT2）.HE staining was used to observe the morphology of pancreatic islets.The apoptosis of pancreas was stained by TUNEL method.Results The contents of serum TNF-α and SAA in group B were much higher thanthose in group A.Compared with group C,group D showed higher glucose concentration in GTT（ 30,60,120 min）and ITT（ 15,30,60,120 min）.In group D,TUNEL staining showed more pancreatic β-cell apoptosis,BAX /BCL2increased（ P〈0.05）,the key genes participated in pancreatic insulin secretion were significantly reduced（ P〈0.05）.Compared with group C,the shape of islets was smaller to group D and serum insulin content much decreased（ P〈0.05）.Conclusions In inflammation condition,sirolimus treatment induced pancreatic β-cell apoptosis,reduced pancreatic β-cell function and decrease insulin secretion,thereby resulting in glucose metabolism disorders.