Ex vivo depletion of alloreactive cells based on CFSE dye dilution, activation antigen selection, and dendfitic cell stimulation
Ex vivo depletion of alloreactive cells based on CFSE dye dilution, activation antigen selection, and dendfitic cell stimulation
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DOI：
10.1182/blood-2003-04-1098
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发表时间：
2004-02-01
期刊：
影响因子：
20.3
通讯作者：
Blazar, BR
中科院分区：
文献类型：
--
作者：
Godfrey, WR;Krampf, MR;Blazar, BR
从T细胞群中消除同种异体反应性细胞将使接受干细胞移植的患者能够转移免疫功能。然而，高效率的耗尽已被证明难以实现。我们试图开发出最大限度地清除同种异体反应性CD 4（+）T细胞的体外方法。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CIFSEbright（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明显缺乏同种异体反应性。使用HILA不匹配的外周血单核细胞（PBMC）刺激剂，我们始终达到（90%-95%）同种异体反应性的耗竭。重要的是，当纯化的成熟单核细胞来源的树突状细胞（DC）被用作刺激，同种异体反应性降低100-fofd（99%），导致废除的第二MLR。值得注意的是，从这些培养物中回收的CIFSEbright CD 25（-）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，CFSE为基础的方法进行了测试，发现是足够的移植物抗宿主病（GVHD）预防体内，在主要组织相容性复合体（MHC）II类不同的小鼠模型。这种选择性耗尽成熟同种异体抗原特异性T细胞的有效方法可能会增强免疫重建而不会发生GVHD。
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 CIFSEbright (5-(and-6)-carboxyfluorescein diacetate succinmidyl ester) (nondivided) and activation antigen-negative cells are markedly depleted of alloreactivity. With HILA-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-fofd (99%) reduction in alloreactivity was attained, resulting in abrogation of the secondary MLR. Significantly, the CIFSEbright 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.