大鼠骨髓间充质干细胞移植治疗肾小管间质损伤的实验研究

Study on the Treatment of Bone Marrow Mesenchymal Stem Cells Transplanting to Renal Tubulointerstitial Damage in Rat

目的:探讨骨髓间充质干细胞(MSCs)移植对大鼠单侧输尿管梗阻(UUO)后肾小管间质损伤的治疗效果。方法:密度梯度离心法和贴壁细胞培养法结合分离纯化MSCs。取成年雄性Wistar大鼠40只,随机分为移植组(组A,n=15)、对照组(组B,n=15)、假手术组(组C,n=10)。移植组和对照组分别结扎左侧输尿管,制作肾小管间质纤维化模型。移植组于结扎当日肾脏多点注射Brd U标记的含MSCs 1×10~6的DMEM培养基50μl,对照组注射等量DMEM培养基。假手术组开腹但不结扎输尿管。术后第14天处死各组大鼠,行HE染色,观察肾脏病理变化;免疫组化方法测定BrdU阳性细胞在肾脏的分布以及TGF-β_1,Ki-67的表达情况。结果:HE染色显示移植组与对照组肾脏间质损伤严重,两组相比无明显差异,假手术组正常。移植组肾实质内可见Brd U标记的MSCs,向周围组织分散,形态上与肾间质成纤维细胞类似,肾小管上皮细胞中未发现MSCs掺入。移植组较对照组TGF-β_1表达差异无统计学意义。移植组Ki-67表达较对照组有明显增强。结论:UUO大鼠骨髓间充质干细胞移植后,肾小管及间质细胞增生明显增高,提示骨髓间充质干细胞同种异体移植可能通过促进肾脏细胞的增生对肾间质损伤起到一定治疗作用。具体机制尚待进一步研究。

Objective:To investigate the effects of transplanting bone marrow mesenchymal stem cells(MSCs)in treatment of renal tubulointerstitial damage after unilateral ureteral obstruction(UUO)in rats.Methods:Bone marrow MSCs were separated and purified by density gradient centrifugation and by adhering to the culture plastic.Forty adult male wistar rats were divided into transplantation group(group A,n=15)、control group(group B,n=15)and sham-operated group(group C,n=10)on random.Rat kidneys underwent UUO in transplantation group were injected 1×106 MSCs which were labeled with BrdU and were resolved in 50μl DMEM.Control group were injected 50μl DMEM.Sham-operated rats underwent the surgery but not transplantation.All rats were killed at 14 days and the kidney underwent UUO were got.Histological changes were observed by HE.Immunohistochemistry study was performed on renal tissue for BrdU、TGF-β1 and Ki-67.Results:There were great damages in both transplantation group and control group in kidneys by HE,with sham-operated group no changes.There were no significant differences between transplantation group and control group.There were some BrdU-positive cells in kidneys,part of which deversed to surrounding.The cell shape was like fibroblast Expression of TGF-β1 in transplantation group was less heavy than control group,but no significant differences.There was a significant increase in immunohistochemistry of Ki-67 in transplantation group than in control group,with sham-operated group no or little expression.Conclusion:There was a tendency of lessen in renal tubulointerstitial damage after MSCs transplantation,with a higher proliferation of renal cells.It suggested that MSCs transplantation maybe had effects to the renal tubulointerstitial damage.It needed more work to make out the mechanism.