草酸钙结晶肾损伤模型小鼠肾小管上皮细胞转分化的实验研究

目的:研究乙醛酸盐诱导所致草酸钙结晶肾损伤模型小鼠中肾小管上皮细胞转分化的发生情况。方法:选择C57BL/6J小鼠连续腹腔注射乙醛酸盐建立草酸钙结晶肾损伤模型,应用HE染色及冯库萨染色分别观察肾组织结构变化及钙盐沉积情况,并应用免疫荧光双染、Western-blot观察肾小管上皮细胞间充质转分化(epithelial mesenchymal transtion,EMT)的情况。结果:随着连续腹腔注射乙醛酸盐时间的延长,小鼠肾组织HE染色结果显示,近端肾小管管腔逐渐扩张,且肾小管上皮细胞逐渐出现肿胀及变形,基底膜逐渐裸露;冯库萨染色结果显示近端小管腔内黑色钙盐沉积逐渐增加;免疫荧光双染、Western blot结果均显示肾小管上皮标志E-cadherin及Pan-ck逐渐丢失,而间质标志α-SMA及Vimentin的表达则逐渐增加,Western blot检测结果显示随着乙醛酸盐干预的增强,Rho相关卷曲螺旋形成蛋白激酶(rho associated coiled coil forming protein kinase,ROCKI)表达也逐渐增加,且在干预第3天即达高峰。结论:乙醛酸盐诱导所致草酸钙结晶肾损伤模型早期即出现EMT,启动了肾间质纤维化的进程。

Experimental Study of Renal Tubular Epithelial Cells Transition in Glyoxylate Induced Crystallization of Calcium Oxalate Kidney Injury Mice Model

Objective：To study the EMT occurrence in the glyoxylate induced crystallization of calcium oxalate kidney injury model. Methods：To select C57BL/6J model. HE staining and VON KOSSA staining were used to observe renal structural changes and calcium deposition separately, and the double staining of immunofluorescence、Western blot was used to monitor the occurrence of the EMT. Results：With the continuous intraperitoneal injection of glyoxylate time prolonged, HE staining displayed that in the lumen of proximal tubular there were refractive irregular crystals adhesion gradually, and the tubular epithelial cells become swelling and deformation ,and also the basement membrane exposed gradually; VON KOSSA staining showed that the black calcium deposition more and more in the proximal lumen; the double staining of immunofluorescence, Western blot showed that the renal tubular epithelial markers E-cadherin and Pan-ck gradually lost, and the expression of interstitial sign α-SMA and Vimentin gradually increase. In addition, the Western blot refected that with the glyoxylate intervention enhanced, the expression of ROCK1 increased and on the third day of intervention that reached a peak. Conclusion：Glyoxylate induced crystallization of calcium oxalate kidney injury model exists EMT occurrence in the early stage, which starts the process of fibrosis.