Zhu M, Wei M F, Liu F, Shi H F, Wang G, Chen S
Institute of Organ Transplantation, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China.
Transplant Proc. 2004 Oct;36(8):2436-7. doi: 10.1016/j.transproceed.2004.08.034.
Genetic modification of donor dendritic cells (DC) is a potential therapy for allograft rejection. We hypothesized that in vitro interleukin-10 (IL)-10-transfected DC (DC-IL-10) may induce allogeneic T-cell apoptosis, resulting in prolonged allograft survival rat small intestine.
Myeloid DC from Wistar-Furth rats (RT-1u) were propagated with rrGM-CSFand rrIL-4,then genetically modified to express the hIL-10 gene. Secretion of IL-10 was quantitated by enzyme-linked immunosorbent assay (ELISA). Allogeneic T cells from Lewis (LEW; RT-1(l)) at proliferative responses were determined by MTT assay in primary mixed leukocyte reactions. We then used a combination of DNA agarose gel electrophoresis, acridine orange staining, and Annexin V/propridium iodide assays to examine apoptosis of allogeneic T cells exposed to DC-IL-10. Then 5 x 10(6) donor-derived DC-IL-10 or untransduced DC were injected intravenously 7 days before small intestine transplantation (WF-->LEW).
DC-IL-10 showed pronounced impairment of T-cell allostimulatory activity. Apoptotic T cells were detected in the DC-IL-10 group. Flow cytometry counting at 72 hours showed 45.1% apoptotic T cells in response to DC-IL-10, whereas the untransduced group did not undergo significant apoptosis (P < .01). DC-IL-10 pretreated recipients showed moderate prolongation of allograft survival compared with controls (20.7 +/- 6.0 days vs 7.5 +/- 2.2 days, P < .01).
DC-IL-10 induced allogeneic T-cell hyporesponsiveness in vitro, possibly due to apoptosis. DC-IL-10 pretreated recipients displayed prolonged intestinal allograft survival rates.
对供体树突状细胞（DC）进行基因改造是一种治疗同种异体移植排斥反应的潜在方法。我们推测，体外转染白细胞介素-10（IL）-10的DC（DC-IL-10）可能诱导同种异体T细胞凋亡，从而延长同种异体移植大鼠小肠的存活时间。
用重组大鼠粒细胞巨噬细胞集落刺激因子（rrGM-CSF）和重组大鼠白细胞介素-4（rrIL-4）培养Wistar-Furth大鼠（RT-1u）的髓样DC，然后进行基因改造以表达人IL-10基因。通过酶联免疫吸附测定（ELISA）对IL-10的分泌进行定量。在初次混合淋巴细胞反应中，采用噻唑蓝（MTT）比色法测定来自Lewis（LEW；RT-1(l)）大鼠的同种异体T细胞的增殖反应。然后，我们联合使用DNA琼脂糖凝胶电泳、吖啶橙染色和膜联蛋白V/碘化丙啶检测法，来检测暴露于DC-IL-10的同种异体T细胞的凋亡情况。在小肠移植（WF→LEW）前7天，静脉注射5×10(6)个供体来源的DC-IL-10或未转导的DC。
DC-IL-10表现出明显的T细胞同种异体刺激活性受损。在DC-IL-10组中检测到凋亡的T细胞。72小时的流式细胞术计数显示，响应DC-IL-10的凋亡T细胞占45.1%，而未转导组未发生明显凋亡（P <.01）。与对照组相比，经DC-IL-10预处理的受体的同种异体移植存活时间适度延长（20.7±6.0天对7.5±2.2天，P <.01）。
DC-IL-10在体外诱导同种异体T细胞低反应性，可能是由于凋亡所致。经DC-IL-10预处理的受体的肠道同种异体移植存活率延长。
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