β1-AA下调脂联素水平诱发糖尿病的机制研究

Mechanism of diabetes induced by β1-AA down-regulating adiponectin levels

目的探讨脂联素水平在β1-肾上腺素受体自身抗体(autoantibodies against the second extra-cellular loop of theβ1-adrenoceptor,β1-AA)诱导糖尿病发生发展中的作用。方法收集45例诊断糖尿病患者和42例健康体检者的血清,ELISA法检测血清中β1-AA及脂联素水平;建立β1-AA主动免疫小鼠模型,动态检测小鼠血糖变化;标记法芯片技术筛选主动免疫模型小鼠外周血清中发生改变的细胞因子,进一步从大体及细胞水平进行验证。结果与健康体检者相比,糖尿病患者血清β1-AA效价及阳性率均增高[(0.82±0.31)vs.(0.12±0.10),57.7%vs.0.1%,P<0.001],脂联素水平降低[(0.19±0.09)vs.(1.08±0.62),P<0.001];主动免疫8周时,免疫组小鼠空腹血糖明显高于对照组[(14.0±0.81)mmol/L vs.(5.03±0.21)mmol/L,P<0.001)],且空腹葡萄糖耐量试验呈阳性;芯片结果显示,主动免疫8周时,小鼠外周血清中脂联素水平显著低于对照组(变化比值>100);蛋白免疫印迹法和实时荧光定量聚合酶链反应显示,主动免疫8周脂肪组织脂联素蛋白及mRNA水平均低于对照组;细胞实验发现,β1-AA刺激后,3T3-L1细胞脂联素分泌减少。结论脂联素在β1-AA诱导的糖尿病发生发挥重要作用。

Objective The role of adiponectin in the development of diabetes induced by autoantibodies against the second extracellular loop of theβ1-adrenoceptor(β1-AA).Methods Serum samples were collected from 45 diabetic patients and 42 healthy subjects.The levels of serumβ1-AA and adiponectin were detected by ELISA,A passive immunization model ofβ1-AA was established to dynamically monitor blood glucose in mice.The marker microarray technique was used to screen the altered cytokines in the peripheral serum of passive immune mice and further validate them at the gross and cellular levels.Results The serumβ1-AA titer and positive rate of diabetic patients were higher than those of healthy subjects[(0.82±0.31)vs.(0.12±0.10),57.7%vs.0.1%,P<0.001].Adiponectin levels were reduced[(0.19±0.09)vs.(1.08±0.62),P<0.001].After 8 weeks of passive immunization,fasting blood glucose in the immunized mice was significantly higher than that in the control group[(14.0±0.81)mmol/L vs.(5.03±0.21)mmol/L,P<0.001)],and fasting glucose tolerance test was positive.Chip results showed that compared with the control group,the passive immunization lasted for 8 weeks.The level of adiponectin in peripheral serum of mice was decreased significantly(Fold change>100).Western blot and real-time PCR showed that compared with the control group,the levels of adiponectin and mRNA in adipose tissue were decreased after active immunization for 8 weeks.Meanwhile,β1-AA treatment decreased secretion of adiponectin in 3T3-L1 cells.Conclusions These results suggest thatβ1-AA can induce diabetes through down regulating adi-ponectin level.