供体淋巴细胞在GVHD模型小鼠组织器官中迁移及分布动态观察.docVIP

供体淋巴细胞在GVHD模型小鼠组织器官中迁移及分布动态观察 　 作者：温宏升，王健民，周虹，夏荣，邱慧颖，高磊，胡晓霞 【摘要】 本研究应用小鼠GVHD模型探讨异基因T淋巴细胞在移植受体体内的迁移和分布。将C57BL/6 的骨髓细胞和转基因荧光C57BL/6 小鼠的脾淋巴细胞输入经8 Gy全身照射的BALB/c小鼠，建立eGFP 标记供体淋巴细胞的GVHD小鼠模型；应用荧光显微镜和流式细胞术观测GVHD模型中eGFP+细胞的分布；ELISA法检测GVHD靶组织中趋化因子MIP-1α水平的改变。结果表明：①输入脾细胞和骨髓细胞第8天后出现GVHD临床及病理表现;②GVHD模型中，受体鼠肝、皮肤、肠、脾、肺、舌有eGFP+ 细胞浸润;③GVHD小鼠肝、脾eGFP+ 细胞中CD4+ 、CD8+ 细胞比例均逐渐升高;④GVHD鼠脾、肝组织中MIP-1α水平升高，脾中MIP-1α水平高峰出现于移植后第3天，肝中MIP-1α水平高峰出现于移植后第7天。结论：除肝、肠、皮肤外，肺、舌可能也是GVHD靶器官。肝、脾组织中供体淋巴细胞浸润伴随MIP-1α水平的升高。 【关键词】 小鼠GVHD模型; T淋巴细胞；增强型绿色荧光蛋白；巨噬细胞炎症蛋白 　　Migration and Distribution of Allogeneic T Lymphocytes in Organs of Graft-Versus-Host Disease Mouse Model 　AbstractThis study was aimed to investigate the migration and distribution processes of allogeneic donor T lymphocytes in the organs of recipient mice. GVHD model was established by transfusion of the splenocytes of eGFP transgeneic C57BL/6 mice together with born marrow cells harvested from C57BL/6 mice into BALB/c mice underwent 8.0 Gy total body irradiation. The migration and homing of eGFP+ cells were tracked by stereo-fluorescent microscopy or inverted fluorescent microscopy and flow cytometry. The enzyme linked immunosorbent assay (ELISA) was performed on supernatants from the tissue homogenates to detect the amount of MIP-1α. The results indicated that GVHD clinical manifestation and pathological changes of organs appeared on day 8 post transplantation. eGFP-positive donor T cells in recipient organs were observed by inverted fluorescence microscope in frozen section, or by stereo-fluorescence microscopy in living organs, such as liver, spleen, skin, lungs, bowels, and tongue. The highest expression of MIP-1α was on day 7 post transplantation in the liver (491.3±32.1 pg/ml), and day 3 post transplantation in the spleen (881.5± 45.2 pg/ml)，respectively (Plt;0.05). It is concluded that GVHD was induced by splenocytes of eGFP transgeneic C57BL/6 mice. eGFP+ cells in the organs can be observed by fluorescent microscopy. In this GVHD model, do