Comprehensive Transplant Center, Department of Surgery, Northwestern University Feinberg School of Medicine, Chicago, IL, United States.
Department of Microbiology-Immunology, Northwestern University Feinberg School of Medicine, Chicago, IL, United States.
Front Immunol. 2024 Feb 22;15:1366972. doi: 10.3389/fimmu.2024.1366972. eCollection 2024.
Donor hematopoietic stem cell (DHSC) infusions are increasingly being studied in transplant patients for tolerance induction.
To analyze the fate of infused DHSCs in patients, we developed an culture system utilizing CD34DHSCs stimulated with irradiated allogeneic cells in cytokine supplemented medium long-term.
Flow cytometric analyses revealed loss of the CD34 marker and an increase in CD33 myeloid and CD3 T-cell proportion by 10.4% and 72.7%, respectively, after 21 days in culture. T-cells primarily expressed TcR-αβ and were of both CD4 and CD8 subsets. Approximately 80% of CD3 T cells lacked expression of the co-stimulatory receptor CD28. The CD4 compartment was predominated by CD4CD25CD127-FOXP3 Tregs (>50% CD4CD127- compartment) with <1% of all leukocytes exhibiting a CD4CD127 phenotype. Molecular analyses for T-cell receptor excision circles showed recent and increased numbers of TcR rearrangements in generated T cells over time suggesting differentiation from DHSCs. CD33 myeloid cells mostly expressed HLA-DR, but lacked expression of co-stimulatory receptors CD80 and CD83. When studied as modulators in primary mixed lymphocyte reactions where the cells used to stimulate the DHSC were used as responders, the DHSC-lines and their purified CD8, CD4, CD33 and linage negative subsets inhibited the responses in a dose-dependent and non-specific fashion. The CD8 cell-mediated inhibition was due to direct lysis of responder cells.
Extrapolation of these results into the clinical situation would suggest that DHSC infusions into transplant recipients may generate multiple subsets of donor "chimeric" cells and promote recipient Treg development that could regulate the anti-donor immune response in the periphery. These studies have also indicated that T cell maturation can occur in response to allogeneic stimulation without the pre-requisite of a thymic-like environment or NOTCH signaling stimulatory cell line.
供者造血干细胞（DHSC）输注在移植患者中越来越多地被用于诱导耐受。
为了分析输注的 DHSC 在患者中的命运，我们开发了一种培养系统，利用 CD34+DHSC 在细胞因子补充的培养基中长期刺激照射的同种异体细胞。
流式细胞术分析显示，在培养 21 天后，CD34 标记物丢失，CD33 髓系和 CD3 T 细胞的比例分别增加 10.4%和 72.7%。T 细胞主要表达 TCR-αβ，并且是 CD4 和 CD8 亚群。大约 80%的 CD3 T 细胞缺乏共刺激受体 CD28 的表达。CD4 细胞群主要由 CD4+CD25+CD127-FOXP3 Tregs（>50%的 CD4+CD127-细胞群）组成，所有白细胞中只有<1%表现出 CD4+CD127 表型。T 细胞受体切除环的分子分析显示，随着时间的推移，生成的 T 细胞中 TcR 重排的数量增加，表明来自 DHSC 的分化。CD33 髓系细胞大多表达 HLA-DR，但缺乏共刺激受体 CD80 和 CD83 的表达。当作为原发性混合淋巴细胞反应中的调节剂进行研究时，用于刺激 DHSC 的细胞作为反应细胞，DHSC 系及其纯化的 CD8、CD4、CD33 和谱系阴性亚群以剂量依赖和非特异性的方式抑制反应。CD8 细胞介导的抑制是由于反应细胞的直接溶解。
将这些结果外推到临床情况表明，DHSC 输注到移植受者中可能会产生多种供体“嵌合”细胞亚群，并促进受者 Treg 的发展，从而调节外周的抗供体免疫反应。这些研究还表明，T 细胞成熟可以在没有胸腺样环境或 NOTCH 信号刺激细胞系的情况下，对同种异体刺激作出反应。
