白细胞介素22抗体抑制Snail1高表达对糖尿病肾病的影响

Effect of interlukin-22 antibody on diabetic nephropathy via inhibition on Snail1 expression

目的:探讨白细胞介素22(IL-22)在糖尿病肾病(DN)发生发展中的作用及机制。方法:将C57 BL/6小鼠随机分成正常对照(NC)组、DN组、重组IL-22(r IL-22)干预组(DN+r IL-22组)和IL-22中和抗体(anti-IL-22)干预组(DN+anti-IL-22组)。糖尿病造模成功8周后,DN+rIL-22组和DN+anti-IL-22组分别给予rIL-22(200μg/kg)和anti-IL-22(200μg/kg)腹腔注射,NC组和DN组分别给予等量的0.1%牛血清白蛋白腹腔注射,每周2次,连续4周。干预结束后,检测各组小鼠血糖、肾功能、24 h尿微量白蛋白(m-Alb)和24 h尿肌酐(UCr)水平,光镜下观察肾脏组织的病理结构改变,qPCR法检测肾脏组织Snail1的mRNA表达,Western blot法检测肾脏组织纤连蛋白(FN)和E-钙黏素(E-cadherin)的表达水平。结果:干预结束后,与NC组小鼠比较,各模型组24 hm-Alb/UCr比值均显著升高(P<0.05);相较于DN组,DN+rIL-22组24 hm-Alb和24 hUCr均显著升高(P<0.05);而DN+anti-IL-22组24 hm-Alb、24 h UCr和24 hm-Alb/UCr比值较DN组和DN+rIL-22组均得到改善(P<0.05)。光镜下可见糖尿病小鼠肾小管上皮细胞空泡变性、蛋白管型形成和肾小球系膜扩张,DN+rIL-22组上述病变更广泛,而DN+anti-IL-22组病变程度得以改善。qPCR发现糖尿病小鼠Snail1的mRNA表达显著升高(P<0.05),anti-IL-22阻断4周后Snail1的mRNA显著下降(P<0.05)。Western blot发现DN+rIL-22组细胞外基质蛋白FN较NC组和DN组显著升高(P<0.05),且肾小管上皮-间充质转化标志物E-cadherin显著下降(P<0.05)。结论:IL-22中和抗体可能通过抑制肾小管上皮细胞Snail1信号分子高表达,减轻DN小鼠微量白蛋白尿,延缓DN进程。

AIM： To investigate the effect of interlukin-22（ IL-22） on diabetic nephropathy（ DN） and its possible mechanism. METHODS： C57 BL/6 mice were randomized to normal control（ NC） group,DN group,DN ＋ recombinant IL-22（ rIL-22） group and DN ＋ IL-22 antibody（ anti-IL-22） group. After successful establishment of diabetes model for 8 weeks,the mice in DN ＋ rIL-22 group and DN ＋ anti-IL-22 group were intraperitoneally injected with rIL-22（ 200 μg/kg） and anti-IL-22（ 200 μg/kg）,respectively,and the mice in NC group and DN group were intraperitoneally injected with 0. 1 % bovine serum albumin,twice a week for 4 weeks. After the intervention,blood glucose,kidney function,24 h urine microalbumin（ m-Alb） and 24 h urine creatinine（ UCr） were measured. The pathological changes of renal tissues were observed under light microscope. The mRNA expression of Snail1 was detected by q PCR. The protein levels of fibronetin（ FN） and E-cadherin were determined by Western blot. RESULTS： After the intervention,the ratio of24 h m-Alb/UCr increased significantly in other model groups compared with NC group（ P〈0. 05）. The levels of 24 h mAlb and 24 hUCr increased significantly in DN ＋ rIL-22 group compared with DN group（ P〈0. 05）. However,in DN ＋anti-IL-22 group,the levels of 24 hm-Alb,24 hUCr and 24 hm-Alb/UCr ratio were significantly lower than those in DN group and DN ＋ rIL-22 group（ P〈0. 05）. The tubular epithelial cell vacuolar degeneration,protein cast formation and glomerular mesangial expansion in the renal tissues from diabetic mice were observed under light microscope. The lesions were more severe in DN ＋ rIL-22 group,but attenuated in DN ＋ anti-IL-22 group. The mRNA expression of Snail1 increased significantly in diabetic mice（ P〈0. 05）,but decreased significantly after a 4-week intervention by anti-IL-22（ P〈0. 05）.The expression of FN,an extracellular matrix protein,increased significantly in DN ＋ rIL-22 group（ P〈0. 05）. The expression of E-cadherin,an epithelial-mesenchymal transition marker,decreased significantly in DN ＋ rIL-22 group as well（ P〈0. 05）. CONCLUSION： IL-22 neutralizing antibody may attenuate microalbuminuria and delay the progression of DN via inhibition of Snail1 expression in the renal tubular epithelial cells.