Intracerebral xenotransplantation of GFP mouse bone marrow stromal cells in intact and stroke rat brain: Graft survival and immunologic response
Intracerebral xenotransplantation of GFP mouse bone marrow stromal cells in intact and stroke rat brain: Graft survival and immunologic response
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DOI：
10.3727/000000004783983990
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发表时间：
2004-01-01
影响因子：
3.3
通讯作者：
Borlongan, CV
中科院分区：
文献类型：
--
作者：
Irons, H;Lind, JG;Borlongan, CV
本研究的特点是骨髓基质细胞（BMSCs）移植到完整和中风脑后的生存和免疫反应。在第一项研究中，在正常成年Sprague-Dawley雄性大鼠中进行BMSC（60.000/3穆尔）或载体的纹状体内移植，随后每天接受环孢菌素A（CsA，10 mg/kg，IP/3 ml）或载体（橄榄油）。相似体积），从手术当天开始直至移植后3天。在移植后3或30天对动物实施安乐死，并处理脑用于绿色荧光蛋白（GFP）显微镜或流式细胞术（FACS）。GFP荧光和流式细胞仪扫描显示，GFP+ BMSCs在两组移植大鼠与或没有CsA，虽然显着增加（1.6- 3倍以上）的GFP+ BMSCs的生存观察到免疫抑制动物。进一步的组织学检查显示，在非CsA移植的动物中，BMSCs广泛分散远离移植物核心，并伴有许多长的生长过程。而一个非常致密的移植物核心，细胞只表达零星的短生长过程。在CsA移植动物中观察到。在接受CsA或载体的非移植大鼠中没有检测到GFP+ BMSC。通过流式细胞仪分析的免疫应答显示细胞毒性细胞的存在减少，其特征在于几乎完全不存在CD 8+细胞，并且缺乏由CsA处理的移植动物中的低CD 69表达所描述的活化。相比之下，在仅接受溶媒的移植动物中观察到CD 8+细胞水平升高和CD 69表达活化增加。在接受CsA的移植大鼠中几乎检测不到CD 4+辅助细胞，但在接受载体的移植大鼠中也仅轻微升高。接受CsA或溶剂的非移植大鼠显示所有三种淋巴细胞标志物的可检测水平非常低。在第二项研究中，一组新的雄性Sprague-Dawley大鼠首先接受BMSCs的双侧立体定位纹状体内移植，并在3天后进行单侧大脑中动脉短暂闭塞。动物在中风后不使用CsA免疫抑制剂存活3天。荧光显微镜显示，与完整纹状体相比，移植GFP+ BMSC在中风纹状体中的存活率显著更高（5倍以上）。大部分g-筏保留在原背侧纹状体移植部位，在完整纹状体内无明显迁移，但在卒中纹状体内可沿缺血半暗带沿着长距离迁移。此外，流式细胞仪扫描分析显示，移植的BMSCs在中风和完整的纹状体的免疫反应水平低。这些结果。总之，表明小鼠BMSCs异种移植到成年大鼠体内是可行的。免疫抑制治疗可以提高异种移植物的存活率，降低移植物诱导的免疫反应;然而，在移植后的急性期，BMSCs可以在完整脑和中风脑中存活。甚至可以在没有免疫抑制的情况下表现出长距离迁移和增加的向外生长过程。
The present study characterized survival and immunologic response of bone marrow stromal cells (BMSCs) following transplantation into intact and stroke brains. In the first study, intrastriatal transplantation of BMSC (60.000 in 3 mul) or vehicle was performed in normal adult Sprague-Dawley male rats that subsequently received daily cyclosporin A (CsA, 10 mg/kg, IP in 3 ml) or vehicle (olive oil. similar volume) starting on day of surgery up to 3 days posttransplantation. Animals were euthanized at 3 or 30 days posttransplantation and brains were processed either for green fluorescent protein (GFP) microscopy or flow cytometry (FACS). Both GFP epifluorescence and FACS scanning revealed GFP+ BMSCs in both groups of transplanted rats with or without CsA, although significantly increased (1.6- to 3-fold more) survival of GFP+ BMSCs was observed in the immunosuppressed animals. Further histologic examination revealed widespread dispersal of BMSCs away from the graft core accompanied by many long outgrowth processes in non-CsA-transplanted animals. whereas a very dense graft core, with cells expressing only sporadic short outgrowth processes. was observed in CsA-transplanted animals. There were no detectable GFP+ BMSCs in nontransplanted rats that received CsA or vehicle. Immunologic response via FACS analysis revealed a decreased presence of cytotoxic cells, characterized by near complete absence of CD8+ cells, and lack of activation depicted by low CD69 expression in CsA-treated transplanted animals. In contrast, elevated levels of CD8+ cells and increased activation of CD69 expression were observed in transplanted animals that received vehicle alone. CD4+ helper cells were almost nondetectable in transplanted rats that received CsA, but also only minimally elevated in transplanted rats that received vehicle. Nontransplanted rats that received either CsA or vehicle displayed very minimal detectable levels of all three lymphocyte markers. In the second study, a new set of male Sprague-Dawley rats initially received bilateral stereotaxic intrastriatal transplantation of BMSCs and 3, days after were subjected to unilateral transient occlusion of middle cerebral artery. The animals were allowed to survive for 3 days after stroke without CsA immunosuppression. Epifluorescence microscopy revealed significantly higher (5-Fold more) survival of transplanted GFP+ BMSCs in the stroke striatum compared with the intact striatum. The majority of the g-rafts remained within the original dorsal striatal transplant site, characterized by no obvious migration in intact striatum, but with long-distance migration along the ischemic penumbra in the stroke striatum. Moreover, FACS scanning analyses revealed low levels of immunologic response of grafted BMSCs in both stroke and intact striata. These results. taken together, suggest that xenotransplantation of mouse BMSCs into adult rats is feasible. Immunosuppression therapy can enhance xenograft survival and reduce graft-induced immunologic response; however, in the acute phase posttransplantation, BMSCs can survive in intact and stroke brain. and may even exhibit long-distance migration and increased out growth processes without immunosuppression.