miR-21基因缺失对糖尿病肾病小鼠肾损伤的影响及机制

Effects of miR-21 gene deletion on renal injury in diabetic nephropathy mice and its mechanism

目的观察miR-21基因缺失对糖尿病肾病(DN)小鼠肾损伤的影响,并探讨其作用机制。方法选取野生型(WT)雄性C57BL/6小鼠10只,分成WT-Con组、WT-DN组各5只;另取miR-21基因敲除型(miR-21 KO)C57BL/6小鼠10只,分成KO-Con组、KO-DN组各5只。WT-DN组、KO-DN组腹腔注射STZ制作DN模型,WT-Con组、KO-Con组腹腔注射等量生理盐水作为对照。造模成功12周后,麻醉小鼠,剥离双肾,采用HE染色观察肾组织病理学改变,Masson染色观察肾脏胶原纤维染色聚集程度,透射电镜观察肾脏超微结构变化,免疫组化法检测肾组织转化生长因子β1(TGF-β1)、纤维连接蛋白(FN)、Ⅰ型胶原(ColⅠ)蛋白表达。结果HE染色结果显示,与WT-Con组相比,WT-DN组、KO-DN组有明显的肾小球及肾小管损伤;与WT-DN组相比,KO-DN组肾脏病变程度较轻微。Masson染色结果显示,与WT-Con组相比,WT-DN组、KO-DN组肾脏胶原纤维增多;与WT-DN组比较,KO-DN组肾脏胶原纤维较少。透射电镜结果显示,与WT-Con组相比,WT-DN组、KO-DN组有明显的肾脏超微结构损伤,足突融合,基底膜改变,肾小管堵塞;与WT-DN组相比,KO-DN组肾脏超微结构损伤减轻。与WT-Con组相比,WT-DN组、KO-DN组肾脏TGF-β1、FN、ColⅠ表达水平升高;与WT-DN组比较,KO-DN组TGF-β1、FN、ColⅠ表达水平降低(P均<0.05)。结论miR-21基因缺失能够抑制DN小鼠的肾脏纤维化改变以保护肾脏,其机制可能与调控肾脏纤维化因子TGF-β1及相关基质蛋白FN、ColⅠ表达有关。

Objective To observe the effect of miR-21 gene deletion on renal injury in diabetic nephropathy(DN)mice and to explore its mechanism.Methods Ten male C57BL/6 mice of wild type(WT)were divided into the WT-Con group and WT-DN group,with 5 in each;another 10 C57BL/6 mice of miR-21 knockout type(miR-21 KO)were divided into the KO-Con group and KO-DN group,with 10 in each.The DN model was made by intraperitoneal injection of streptozotocin(STZ)in the WT-DN group and KO-DN group,and the same amount of normal saline was injected in the WT-Con group and KO-Con group as controls.After 12 weeks of successful modeling,the anesthetized mice were deprived of both kidneys;the changes of renal histopathology were observed by HE staining,the degree of collagen fiber aggregation was observed by Masson staining,the ultrastructural changes of kidney were observed by transmission electron microscope,and the expression levels of transforming growth factor 1(TGF-1),fibronectin(FN)and Collagen typeⅠ(ColⅠ)were detected by immunohistochemistry.Results Compared with the WT-Con Group,WT-DN group and KO-DN group had significant glomerular and tubular injury,and the degree of renal lesions in the KO-DN group was milder than that in the WT-DN group.Compared with the WT-Con Group,Masson staining showed that the expression of collagen fibers in kidney of the WT-DN group and KO-DN group was higher than that of the WT-Con group.Compared with WT-Con group,WT-DN group and KO-DN group had significant renal ultrastructural damage,foot process fusion,basement membrane change and renal tubule obstruction;compared with the WT-DN group,KO-DN group had less renal ultrastructural damage.Compared with the WT-Con group,TGF-1,FN and Col I expression increased in kidney of WT-DN group and KO-DN group,and TGF-β1,FN and Col I expression decreased in KO-DN group(all P<0.05).Conclusion The miR-21 gene deletion can inhibit renal fibrosis in DN mice and protect the kidney,which may be related to the regulation of renal fibrosis factor TGF-1 and its related matrix protein FN,ColⅠexpression.