骨髓间充质干细胞保护阿霉素诱导的肾病综合征模型鼠肾小球的实验研究

Experimental study of bone marrow mesenchymal stem cells in protecting the glomeruli of model rats with Adriamycin-induced nephrotic syndrome

目的观察骨髓间充质干细胞(BMSCs)移植对肾病综合征模型鼠肾小球的作用,探讨BMSCs保护肾病综合征模型鼠肾小球的可能性及其机制。方法从SD大鼠骨髓中提取、分离、纯化、扩增和鉴定BMSCs。21日龄刚断奶的SD大鼠适应性喂养后,筛选尿蛋白浓度<0.8 g/L作为实验用鼠。随机选取5只实验用鼠作为正常对照称A组。其余实验鼠按照Bertani法尾静脉注射盐酸阿霉素制作肾病综合征模型鼠。成模后随机分成B、C和D组,每组5只,分别给予尾静脉注射生理盐水、泼尼松灌胃和尾静脉移植BMSCs细胞悬液。各组大鼠造模前、后,干预后1、4、8周分别进行24 h尿蛋白定量;干预结束后,处死各组实验鼠进行肾脏病理检查和nephrin、podocin mRNA的测定。结果造模前,4个实验组的尿蛋白定量比较,差异无统计学意义(P>0.05)。造模后,B、C和D组的尿蛋白定量高于A组,差异有统计学意义(P<0.05),但B、C和D组内两两比较,差异无统计学意义(P>0.05)。干预后,B、C和D组的尿蛋白定量均高于同期A组,差异有统计学意义(P<0.05);B、C和D组内两两比较:C、D两组的尿蛋白定量均低于同期B组,差异有统计学意义(P<0.05),但同期C、D两组的尿蛋白定量比较,差异无统计学意义(P>0.05)。干预结束后,在电镜下观察实验各组的肾脏损伤程度为B>C>D>A;B、C和D组的肾小球足细胞相关分子nephrin mRNA的表达均低于A组,差异有统计学意义(P<0.05),B、C和D组内两两比较:C、D组高于B组,差异有统计学意义(P<0.05),但C、D组间比较,差异无统计学意义(P>0.05);相关分子podocin mRNA的表达实验各组比较,差异无统计学意义(P>0.05)。结论BMSCs移植可保护阿霉素诱导的肾病综合征模型鼠的肾小球损伤,其机制可能为通过上调肾小球足细胞的nephrin mRNA的表达减轻肾小球损伤和降低蛋白尿。

Objective To observe the effect of bone marrow mesenchymal stem cells(BMSCs)transplantation on the glomeruli of model rats with nephrotic syndrome(NS),and to explore the possibility and mechanism of BMSCs in protecting the glomeruli of model rats with NS.Methods The BMSCs were extracted,isolated,purified,amplified and identified form bone marrow of SD rats.After adaptive feeding of 21-day-old weaned SD rats,urinary protein concentration<0.8 g/L was selected as experimental rats.Five experimental rats were randomly selected as normal control named group A.The other experimental mice were injected with Adriamycin Hydrochloride into tail vein according to Bertani method to make NS model rats.After molding,the rats were randomly divided into group B,C and D,and each group had 5 rats,and each group was given saline by tail vein injection,Prednisone by stomach and BMSCs cell suspension by tail vein transplantation.Urinary protein was quantified 24 hours before and after the model establishment and 1,4 and 8 weeks after the intervention.After the intervention,rats in each group were sacrificed for renal pathological examination and nephrin and podocin mRNA determination.Results There was no statistically significant difference in urinary protein quantitation among the four experimental groups before model establishment (P>0.05). The urinary protein quantitation of group B, C and D was higher than that of group A after model establishment respectively, and the differences were statistically significant (P<0.05), but there was no significant difference between groups B, C and D (P>0.05). After intervention, the urinary protein quantitation of group B, C and D was higher than that of group A at the same time respectively, and the differences were statistically significant (P<0.05). Two comparisons in group B, C and D: the urinary protein quantitation in group C and D was lower than that in group B at the same time respectively, with significant differences (P<0.05), but there was no significant difference in the urinary protein quantitation between group C and D at the same time (P>0.05). After the intervention, the degree of kidney injury in the experimental groups was B>C>D>A under electron microscope. The expression of nephrin in glomerular podocyte-related molecule in group B, C and D was lower than that in group A respectively, and the differences were statistically significant (P<0.05), two comparisons in group B, C and D: group C and D was higher than that of group B respectively, and the differences were statistically significant (P<0.05), but there was no significant difference between groups C and D (P>0.05). There were no significant differences in the expression of podocin mRNA between experimental groups (P>0.05). Conclusion BMSCs transplantation can protect the glomerular injury of model rats with Adriamycin-induced NS. The mechanism may be to reduce glomerular injury and proteinuria by up-regulating the expression of nephrin mRNA in glomerular podocytes.