摘要
目的研究探讨骨髓间充质干细胞(MSC)是否能够诱导肾脏移植物免疫耐受的产生以及吲哚胺2,3-双加氧酶(IDO)在MSC介导免疫调节反应中的作用。方法在BALB/c小鼠接受C57BL/6小鼠移植肾脏后24小时,C57BL/6小鼠来源的正常(wt)或IDO基因敲除(IDO-/-)的MSC(1×106)经静脉注入到移植受体中,分别作为wt-MSC治疗组和IDO-/--MSC治疗组,每组6只。以6只未注射的BALB/c移植小鼠受体为未治疗组。以移植物排斥反应所致小鼠死亡或术后100天设定为研究终点。长期存活的BALB/c肾移植受体在移植术后100天,接受来自C57BL/6供体或第三方移植物供体C3H(H-2k)小鼠的皮肤移植,监测移植皮肤情况。对3组小鼠进行移植物组织病理学观察,免疫组化评估肾脏组织Foxp3+细胞水平。用流式细胞技术进行抗原特异性抗体和细胞表型检测,用混合淋巴细胞反应(MLR)评估树突细胞(DC)和T细胞功能。结果本研究发现wt-MSC治疗可诱导受体产生同种异体移植物免疫耐受,表现为移植物病理检查结果正常、未发现抗原特异性抗体水平升高、免疫耐受受体中耐受性树突细胞(Tol-DC)数量显著增多等。同时,在免疫耐受的受体脾脏和肾移植物中均可发现大量CD4+CD25+Foxp3+调节性T细胞(Treg)。这些结果均提示Treg在MSC诱导免疫耐受中的重要作用。值得关注的是,经IDO-/--MSC处理的移植受体中,MSC丧失了其诱导同种异体移植物的免疫耐受的能力,肾移植物很快被排斥,同时机体移植物的排斥反应变化与未治疗组相同。结论由MSC分泌的IDO通过促进Treg的生成,在诱导肾脏移植免疫耐受中起着关键性的作用。本研究将为MSC在器官移植中的临床应用提供理论及临床转化依据。
Objective To determine whether mesenchymal stem cells(MSCs)can induce the establishment of kidney allograft tolerance,as well as indoleamine 2,3-dioxygenase(IDO)contributes to the immunoregulatory functions of MSCs in that process. Methods MSCs(1×106)from wild-type(wt-MSCs)or IDO-knockout (IDO-/--MSCs)C57BL/6 mice were injected intravenously into BALB/c recipients 24 hours after receiving a life-supporting orthotopic C57BL/6 renal graft. Recipients treated with either wt-MSCs or IDO-/--MSCs were used as two study groups(n=6/group). In addition,6 native BALB/c mice were used as controls. Samples were collected at endpoint of graft rejection or on postoperative day(POD)100. Long-term surviving BALB/c kidney allograft recipients received full-thickness skin grafts(1 cm×1 cm)from C57BL/6 donor and C3H mouse strains on POD100 and were monitored daily. The graft pathology,immunohistochemistry,flow cytometry,mixed lymphocyte reaction were used for detecting graft rejection,intragraft Foxp3+ cell infiltration,donor-reactive antibody and cellular phenotypic expression,dendritic cell(DC)and T cell function,respectively. Results wt-MSC-treated recipients achieved allograft tolerance with normal histology and undetectable antidonor antibody levels. Tolerant recipients demonstrated significantly high frequencies of tolerogenic dendritic cells(Tol-DCs). In addition,high frequencies of CD4+CD25+Foxp3+regulatory T-cells(Tregs)were found in recipient spleens and donor grafts,implying the important role of Tregs in the MSC-induced tolerance. Interestingly,renal allograft recipients treated with IDO-/--MSCs, were unable to achieve allograft tolerance,suggesting that functional IDO was necessary for the immunosuppression observed with wt-MSC treatment. Conclusion IDO secreted by MSCs was responsible for induction of kidney allograft tolerance through generation of Tregs. This study supports the clinical application of MSCs in transplantation.
出处
《实用器官移植电子杂志》
2013年第3期138-146,共9页
Practical Journal of Organ Transplantation(Electronic Version)
基金
国家自然科学基金面上项目(81273257)
天津市外国专家局引进国外技术
管理人才项目(Y2012087)
天津医科大学总医院科研启动基金
