白细胞介素-22在多囊卵巢综合征大鼠胰岛素抵抗中的作用机制

Mechanism of interleukin-22 regulating insulin resistance in polycystic ovary syndrome rats

目的研究胰腺白细胞介素-22(interleukin-22,IL-22)信号通路在多囊卵巢综合征(polycystic ovary syndrome,PCOS)大鼠胰岛素抵抗(insulin resistance,IR)中的作用机制。方法选取21 d的SD大鼠按照计算机随机生成数字的方式分为对照组和PCOS组,每组各10只。PCOS组大鼠皮下埋置双氢睾酮(dihydrotestosterone,DHT)硅胶缓释管90 d。测定大鼠体质量,检测腹腔葡萄糖耐量试验(intraperitoneal glucose tolerance test,IPGTT)、胰岛素耐量试验(intraperitoneal insulin tolerance test,IPITT),酶联免疫吸附法(enzyme-linked immunosorbent assay,ELISA)检测血清IL-22和空腹胰岛素(fasting insulin,FINS),计算胰岛素稳态指数(homeostasis model assessment of insulin resistance,HOMA-IR),免疫荧光染色观察胰岛β细胞中IL-22受体1(interleukin-22 receptor 1,IL-22R1)的定位与差异性表达,蛋白印迹法检测大鼠胰腺组织IL-22、酪氨酸蛋白激酶1(Janus kinase 1,JAK1)、信号转导与转录激活因子3(signal transducer and activator of transcription 3,STAT3)蛋白表达水平。结果PCOS组体质量[(420.50+10.26)g]明显高于对照组[(314.40+6.54)g,P=0.003],血清IL-22浓度[(9.86±1.41)ng/L]低于对照组[(16.07±4.68)ng/L,P=0.038],PCOS组FINS[(17.97±2.86)mU/L]、HOMA-IR(4.46±0.80)明显高于对照组[(13.05±2.12)mU/L,P=0.001;2.99±0.61,P=0.001]。PCOS组IPGTT 30 min、60 min、90 min血糖[(15.58±1.86)mmol/L、(10.71±2.07)mmol/L、(7.47±1.36)mmol/L]均明显高于对照组[(12.54±1.60)mmol/L,P=0.010;(8.33±1.02)mmol/L,P=0.026;(6.28±0.72)mmol/L,P=0.013];PCOS组的糖耐量曲线下面积[area under curve,AUC;(1248.71±164.23)mmol/(L·min)]明显大于对照组[(1007.29±102.29)mmol/(L·min),P=0.010]。PCOS组IPITT 60 min、90 min、120 min血糖[(3.12±0.28)mmol/L、(2.62±0.17)mmol/L、(2.28±0.25)mmol/L]均明显高于对照组[(2.60±0.28)mmol/L,P=0.031;(2.10±0.33)mmol/L,P=0.026;(1.90±0.24)mmol/L,P=0.033];PCOS组胰岛素耐量AUC[(414.60±23.06)mmol/(L·min)]明显大于对照组[(359.40±39.95)mmol/(L·min),P=0.044]。免疫荧光染色显示,IL-22R1与INS在胰岛β细胞内共表达,PCOS组大鼠IL-22R1表达(48.26±3.83)较对照组(63.44±2.66)显著降低(P=0.031)。PCOS组大鼠胰腺内IL-22、p-STAT3蛋白表达水平(0.82±0.09、0.77±0.09)较对照组明显下降(1.01±0.06,P=0.025;1.01±0.07,P=0.015)。结论在PCOS大鼠胰腺中,IL-22通过影响STAT3信号通路活化,降低p-STAT3水平,参与IR的发生。

Objective To evaluate the mechanism of pancreatic interleukin-22(IL-22)signaling pathway regulating insulin resistance(IR)in polycystic ovary syndrome(PCOS)rats.Methods Postnatal 21 d SD rats were divided into control group and PCOS group with 10 rats in each group based on numbers randomly generated by computer.PCOS rats implanted with a silastic tube filled with dihydrotestosterone(DHT)at dosage of 7.5 mg.Their body weight was monitored weekly.At the end of the experiment,the intraperitoneal glucose tolerance test(IPGTT)and intraperitoneal insulin test(IPITT)were conducted,the serum IL-22 and fasting insulin(FINS)were monitored through the enzyme-linked immunosorbent assay(ELISA),and homeostasis model assessment of insulin resistance(HOMA-IR)was calculated.The location and differential expression of the interleukin-22 receptor 1(IL-22R1)in pancreatic β cells were observed by immunofluorescence staining,and the expressions of the IL-22,Janus kinase 1(JAK1),signal transducer and activator of transcription 3(STAT3)proteins were detected by Western blotting.Results Compared with control group,the body weight was increased[(420.50±10.26)g vs.(314.40±6.54)g,P=0.003]and the serum level of IL-22 in PCOS group was decreased[(9.86±1.41)ng/L vs.(16.07±4.68)ng/L,P=0.038].The levels of FINS and HOMA-IR were higher in PCOS group than in control group[(17.97±2.86)mU/L vs.(13.05±2.12)mU/L,P=0.001;4.46±0.80 vs.2.99±0.61,P=0.001].The glucose levels in IPGTT test were significantly elevated in PCOS group in 30 min,60 min and 90 min compared with control group[(15.58±1.86)mmol/L vs.(12.54±1.60)mmol/L,P=0.010;(10.71±2.07)mmol/L vs.(8.33±1.02)mmol/L,P=0.026;(7.47±1.36)mmol/L vs.(6.28±0.72)mmol/L,P=0.013],as well as the area under curve(AUC)[(1248.71±164.23)mmol/(L·min)vs.(1007.29±102.29)mmol/(L·min),P=0.010].Following IPITT,compared with control group,PCOS group showed significantly higher 60 min[(3.12±0.28)mmol/L vs.(2.60±0.28)mmol/L,P=0.031],90 min[(2.62±0.17)mmol/L vs.(2.10±0.33)mmol/L,P=0.026]and 120 min glucose level[(2.28±0.25)mmol/L vs.(1.90±0.24)mmol/L,P=0.033],as well as the AUC[(414.60±23.06)mmol/(L·min)vs.(359.40±39.95)mmol/(L·min),P=0.044].The immunofluorescence results suggested that IL-22R1 and INS were colocalized in pancreatic β cells and the expression of IL-22R1 was lower in PCOS group than in control group(48.26±3.83 vs.63.44±2.66,P=0.031).Further,the levels of pancreatic IL-22 and p-STAT3 protein were significantly lower in PCOS group than in control group(0.82±0.09 vs.1.01±0.06,P=0.025;0.77±0.09 vs.1.01±0.07,P=0.015).Conclusion Our results demonstrated that the abnormality of IL-22 may result in IR through attenuating the activation of STAT3 signaling pathway and decreasing the level of p-STAT3 in the pancreatic tissue of PCOS rats.