George James F, Gooden Christie W, Guo Lingling, Kirklin James K
Department of Surgery, University of Alabama at Birmingham, AL 35294, USA.
J Heart Lung Transplant. 2008 Jun;27(6):616-22. doi: 10.1016/j.healun.2008.02.015.
Extracorporeal photopheresis (ECP) is used to treat recurrent severe rejection in clinical heart and lung recipients. The mechanisms of the salutary effects of ECP are poorly understood, but appear to involve regulation of T-cell-mediated alloreactive responses, possibly by induction of regulatory T cells. We created a mouse model of ECP to determine the effects of ECP on T-cell responses in vivo and the contribution of CD4(+)CD25(+) T cells.
In this study, 1 x 10(7) splenocytes were treated with 8-methoxypsoralen (8-MOP, 200 ng/ml), followed by ultraviolet A (UVA) irradiation (2 J/cm(2), 350 nm), and then injected intravenously into syngeneic mice. Thirty minutes later, the treated animals received heterotopic cardiac allografts with no immunosuppression. Treated graft recipients were analyzed to determine the effect of ECP on graft survival, deletion of allospecific T cells, and the frequency and in vivo suppressive activity of CD4(+)CD25(+) T cells.
ECP extends cardiac allograft survival in at least two different strain combinations. For CBA/Ca recipients of C57BL/6 allografts, median survival time (MST) in ECP-treated animals was 16 days vs 10 days in graft recipients treated with cells exposed only to 8-MOP (p = 0.04). The frequency of splenic CD4(+)CD25(+) cells expressing FoxP3(+) increased 2-fold in ECP-treated CBA/Ca mice (82.6 +/- 5.2%, n = 4) relative to untreated mice (44.9 +/- 4.5%, n = 4, p < 0.001). Adoptive transfer of 3 x 10(5) sorted CD4(+)CD25(+) splenocytes from ECP-treated graft recipients to untreated cardiac allograft recipients 30 minutes after transplantation resulted in extended graft survival compared with animals that received the same number of CD4(+)CD25(+) splenocytes from cardiac allograft recipients not treated with ECP (MST: 24 days vs 13 days, respectively, p = 0.001). Analyses of 5,6-carboxy-fluorescein-succinimidyl-ester (CFSE)-labeled H-2K(b)-specific T cells in the spleen and lymph node showed no evidence of peripheral deletion after ECP treatment.
ECP extends graft survival even in fully histoincompatible strain combinations with no immunosuppression. It increases the frequency of FoxP3(+)CD4(+)CD25(+) splenic T cells, and its effects can be transferred to untreated recipients using minimal numbers of CD4(+)CD25(+) T cells, indicating that CD4(+)CD25(+) T cells may play a key role in the immunomodulatory effects of ECP.
体外光化学疗法（ECP）用于治疗临床心脏和肺移植受者的复发性严重排斥反应。ECP有益作用的机制尚不清楚，但似乎涉及调节T细胞介导的同种异体反应，可能是通过诱导调节性T细胞实现的。我们建立了一个ECP小鼠模型，以确定ECP对体内T细胞反应的影响以及CD4(+)CD25(+) T细胞的作用。
在本研究中，用8-甲氧基补骨脂素（8-MOP，200 ng/ml）处理1×10(7) 脾细胞，然后进行紫外线A（UVA）照射（2 J/cm(2)，350 nm），随后静脉注射到同基因小鼠体内。30分钟后，给处理过的动物进行异位心脏同种异体移植，不进行免疫抑制。对处理过的移植受者进行分析，以确定ECP对移植物存活、同种异体特异性T细胞缺失以及CD4(+)CD25(+) T细胞频率和体内抑制活性的影响。
ECP可延长至少两种不同品系组合的心脏同种异体移植存活时间。对于C57BL/6同种异体移植的CBA/Ca受者，ECP处理的动物中位存活时间（MST）为16天，而仅用暴露于8-MOP的细胞处理的移植受者为10天（p = 0.04）。相对于未处理的小鼠（44.9 +/- 4.5%，n = 4），ECP处理的CBA/Ca小鼠中表达FoxP3(+) 的脾CD4(+)CD25(+) 细胞频率增加了2倍（82.6 +/- 5.2%，n = 4，p < 0.001）。在移植后30分钟，将3×10(5) 分选的来自ECP处理的移植受者的CD4(+)CD25(+) 脾细胞过继转移到未处理的心脏同种异体移植受者中，与接受相同数量来自未用ECP处理的心脏同种异体移植受者的CD4(+)CD25(+) 脾细胞的动物相比，移植物存活时间延长（MST分别为24天和13天，p = 0.001）。对脾和淋巴结中5,6-羧基荧光素-琥珀酰亚胺酯（CFSE）标记的H-2K(b) 特异性T细胞的分析显示，ECP处理后没有外周缺失的证据。
即使在完全组织不相容的品系组合且不进行免疫抑制的情况下，ECP也能延长移植物存活时间。它增加了FoxP3(+)CD4(+)CD25(+) 脾T细胞的频率，并且使用最少数量的CD4(+)CD25(+) T细胞就能将其作用转移到未处理的受者身上，这表明CD4(+)CD25(+) T细胞可能在ECP的免疫调节作用中起关键作用。
