糖尿病肾脏病大鼠造模方法的改进及肾组织IL-10表达水平

Improvement of modeling methods in rats with diabetic nephropathy and the expression level of IL-10 in kidney tissue

目的对糖尿病肾脏病(diabetic kidney disease,DKD)大鼠造模方法进行改进并检测IL-10在肾组织的表达水平。方法48只雄性SD大鼠被随机分为正常对照组(NC组,n=24)、糖尿病(diabetes mellitus,DM)组(DM组,n=24),DM组以链脲佐菌素(streptozotocin,STZ)40 mg/kg一次性腹腔注射诱导DM模型。造模成功后分别于0周、4周、8周、12周检测血糖、尿糖、尿微量白蛋白、尿肌酐水平,观察大鼠肾脏病理在不同时间点的差别,应用免疫组化检测IL-10水平的表达。结果DM组大鼠造模成功率为75%,出现明显多饮、多食、多尿症状,至12周时,DM组大鼠平均体质量为(478.0±79.1)g,明显低于NC组(650.0±26.9)g;DM组大鼠血糖在造模成功后0周血糖(25.6±5.3)mmol/L显著高于NC组(5.2±0.2)mmol/L,直至实验结束发现血糖仍维持在(23.0±5.5)mmol/L;DM组大鼠尿微量白蛋白/尿肌酐比在8周、12周分别为(39.0±18.6)mg/g、(77.0±12.3)mg/g,均显著高于同期对照组(15.1±5.4)mg/g、(15.8±7.0)mg/g;光镜观察DM组大鼠0周即出现肾间质水肿、炎细胞浸润,第12周出现肾小球基底膜轻度增厚、肾小管上皮细胞空泡变性;免疫组化结果显示,NC组各期IL-10表达量均较DM组高。结论高糖高脂联合小剂量STZ建立DKD模型,在12周时肾脏才出现肾小球基底膜轻度增厚、肾小管上皮细胞空泡变性等病理改变,而不能单纯应用尾静脉血糖≥16.7 mmol/L判断DKD造模成功,肾组织IL-10表达降低进一步论证IL-10参与DKD发病并有可能作为DKD治疗的靶点。

Objective To improve the modeling method in diabetic nephropathy rats,and to detect the expression of IL-10 in kidney tissue.Methods Forty-eight male SD rats were randomly classified as normal control group(NC group,n=24)and diabetes group(DM group,n=24).In the DM group intraperitoneal injection of STZ at a dose of 40 mg/kg was performed one time to induce a diabetes model.Blood sugar,urine sugar,urinary microalbumin,urinary creatinine levels were detected at 0,4,8,12 weeks after successfully modeling,respectively.At the same time,the kidney pathology of rats at different time points was observed,and IL-10 expression levels was detected with immunohistochemical method.Results Compared with NC group,the modeling success rate of rats in the DM group was 75%,and the modeled rats showed significantly increased drink,food uptake and urine.By 12 weeks the average weight of rats in the DM group was(478.0±79.1)g,significantly lower than that in NC group(650.0±26.9)g.Blood glucose in the DM group(25.6±5.3)mmol/L was significantly higher than that in the NC group(5.2±0.2)mmol/L.Until the end of the experiment,it was found that blood sugar was still maintained at(23.0±5.5)mmol/L.In the DM group rats,urine microalbumin creatinine ratios at 8 weeks and 12 weeks were(39.0±18.6)mg/g and(77.0±12.3)mg/g,respectively,significantly higher than those in the control group(15.1±5.4)mg/g,(15.8±7.0)mg/g).Optical microscope observation indicated that DM group rat experienced renal interstitium edema and inflammatory cell infiltration at 0 weeks,and mild thickening of the glomerular base membrane and renal tubular epithelial vacuolar degeneration at 12 weeks.The results of immunohistochemistry showed that the expression of IL-10 in NC group was higher than that in DM group.Conclusions When high sugar and high-fat combined with a small dose of STZ are used to establish a DKD model,at 12 weeks the kidney has mild thickening of renal glomerular base membrane,vacuolar degeneration of renal tubular epithelial cells and other pathological changes.However,Blood glucose≥16.7 mmol/L in the tail vein≥16.7mmol/L cannot be used alone to judge success of the modeling of DKD.The decreased expression of IL-10 in renal tissues further demonstrates that IL-10 is involved in the pathogenesis of DKD and may be the target of DKD treatment.