3rd Department of Medicine, University Medical Center of Johannes Gutenberg-University, Mainz, Germany.
Haematologica. 2011 Jul;96(7):1024-32. doi: 10.3324/haematol.2010.037481. Epub 2011 Apr 12.
HLA mismatch antigens are major targets of alloreactive T cells in HLA-incompatible stem-cell transplantation, which can trigger severe graft-versus-host disease and reduce survival in transplant recipients. Our objective was to identify T-cell subsets with reduced in vitro reactivity to allogeneic HLA antigens.
We sorted CD4 and CD8 T-cell subsets from peripheral blood by flow cytometry according to their expression of naive and memory markers CD45RA, CD45RO, CD62L, and CCR7. Subsets were defined by a single marker to facilitate future establishment of a clinical-grade procedure for reducing alloreactive T-cell precursors and graft-versus-host disease. T cells were stimulated in mixed lymphocyte reactions against HLA-deficient K562 cells transfected with single HLA-A/-B/-C/-DR/-DQ mismatch alleles. Alloreactivity was measured by interferon-γ spot production and cell proliferation.
We observed that allogeneic HLA-reactivity was preferentially derived from subsets enriched for naïve T cells rather than memory T cells in healthy donors, irrespective of the HLA mismatch allele. This separation was most efficient if CD45RA (versus other markers) was used for sorting. The numbers of allogeneic HLA-reactive effector cells were in median 7.2-fold and 16.6-fold lower in CD45RA(neg) memory CD8 and CD4 T cells than in entire CD8 and CD4 T cells, respectively. In contrast, proliferation of memory T cells in response to allogeneic HLA was more variably reduced (CD8) or equivalent (CD4) when compared to that of naïve T cells. We also demonstrated in HLA-matched donor-patient pairs that leukemia-reactive CD8 cytotoxic T-lymphocytes were mainly derived from subsets enriched for naïve T cells compared to memory T cells.
Memory T-cell subsets of most healthy individuals showed decreased allogeneic HLA-reactivity, but lacked significant anti-leukemia responses in vitro. The clinical use of memory or naïve-depleted T cells might be beneficial for HLA-mismatched patients at high risk of graft-versus-host disease and low risk of leukemia relapse. Preferred allografts are those which contain leukemia-reactive memory T cells. Alternatively, replenishment with leukemia-reactive T cells isolated from naïve subsets is desirable.
HLA 错配抗原是 HLA 不相容干细胞移植中同种反应性 T 细胞的主要靶标，可引发严重的移植物抗宿主病并降低移植受者的存活率。我们的目的是鉴定对同种异体 HLA 抗原体外反应性降低的 T 细胞亚群。
我们通过流式细胞术根据外周血中 CD4 和 CD8 T 细胞的幼稚和记忆标志物 CD45RA、CD45RO、CD62L 和 CCR7 的表达对其进行分选。通过单一标志物定义亚群，以方便未来建立一种用于减少同种反应性 T 细胞前体和移植物抗宿主病的临床级程序。T 细胞在混合淋巴细胞反应中针对 HLA 缺陷的 K562 细胞进行刺激，这些细胞转染了单个 HLA-A/-B/-C/-DR/-DQ 错配等位基因。通过干扰素-γ斑点产生和细胞增殖来测量同种异体反应性。
我们观察到，无论 HLA 错配等位基因如何，同种异体 HLA 反应性主要来自健康供体中富含幼稚 T 细胞的亚群，而不是记忆 T 细胞。如果使用 CD45RA（与其他标志物相比）进行分选，这种分离效率最高。与整个 CD8 和 CD4 T 细胞相比，CD45RA(neg)记忆 CD8 和 CD4 T 细胞中的同种异体 HLA 反应性效应细胞数量中位数分别低 7.2 倍和 16.6 倍。相比之下，与幼稚 T 细胞相比，记忆 T 细胞对同种异体 HLA 的增殖反应差异更大（CD8）或相当（CD4）。我们还在 HLA 匹配的供体-患者对中证明，白血病反应性 CD8 细胞毒性 T 淋巴细胞主要来自幼稚 T 细胞富集的亚群，而不是记忆 T 细胞。
大多数健康个体的记忆 T 细胞亚群表现出降低的同种异体 HLA 反应性，但在体外缺乏明显的抗白血病反应。在移植物抗宿主病风险高且白血病复发风险低的 HLA mismatched 患者中，使用记忆或幼稚细胞耗尽的 T 细胞可能是有益的。首选的同种异体移植物是那些含有白血病反应性记忆 T 细胞的移植物。或者，从幼稚亚群中分离出白血病反应性 T 细胞进行补充是可取的。
