Godfrey Wayne R, Krampf Mark R, Taylor Patricia A, Blazar Bruce R
Department of Pediatrics, University of Minnesota Cancer Center, Minneapolis, MN 55455, USA.
Blood. 2004 Feb 1;103(3):1158-65. doi: 10.1182/blood-2003-04-1098. Epub 2003 Oct 2.
Eliminating alloreactive cells from T-cell populations would enable the transfer of immune function to patients who receive stem cell transplants. However, high-efficiency depletion has proved difficult to achieve. We sought to develop ex vivo approaches for the maximal depletion of alloreactive CD4(+) T cells. Using a flow cytometric cell sorting approach after mixed lymphocyte reaction (MLR) culture, we have found that sorted CFSE(bright) (5-(and-6)-carboxyfluorescein diacetate succinmidyl ester) (nondivided) and activation antigen-negative cells are markedly depleted of alloreactivity. With HLA-mismatched peripheral blood mononuclear cell (PBMC) stimulators we have consistently attained (90%-95%) depletion of alloreactivity. Importantly, when purified matured monocyte-derived dendritic cells (DCs) are used as stimulators, a 100-fold (99%) reduction in alloreactivity was attained, resulting in abrogation of the secondary MLR. Significantly, the CFSE(bright) CD25(-) cells recovered from these cultures retained general immunoreactivity, including responses to Candida and cytomegalovirus (CMV) antigens. In addition, a CFSE-based approach was tested and found to be sufficient for graft-versus-host disease (GVHD) prevention in vivo, in a major histocompatibility complex (MHC) class II disparate murine model. This efficient approach to selectively deplete mature alloantigen-specific T cells may permit enhanced immune reconstitution without GVHD.
从T细胞群体中清除同种反应性细胞，将使免疫功能能够转移到接受干细胞移植的患者身上。然而，已证明难以实现高效清除。我们试图开发体外方法，以最大限度地清除同种反应性CD4(+) T细胞。在混合淋巴细胞反应（MLR）培养后，使用流式细胞术细胞分选方法，我们发现分选的CFSE(亮)（5-(和-6)-羧基荧光素二乙酸琥珀酰亚胺酯）（未分裂）和活化抗原阴性细胞的同种反应性明显降低。使用HLA不匹配的外周血单核细胞（PBMC）刺激物，我们始终能够实现（90%-95%）的同种反应性清除。重要的是，当使用纯化的成熟单核细胞衍生树突状细胞（DC）作为刺激物时，同种反应性降低了100倍（99%），导致二次MLR消除。值得注意的是，从这些培养物中回收的CFSE(亮) CD25(-)细胞保留了一般免疫反应性，包括对念珠菌和巨细胞病毒（CMV）抗原的反应。此外，在主要组织相容性复合体（MHC）II类不同的小鼠模型中，测试了一种基于CFSE的方法，发现该方法足以在体内预防移植物抗宿主病（GVHD）。这种选择性清除成熟同种抗原特异性T细胞的有效方法，可能允许在无GVHD的情况下增强免疫重建。
