Circulating hematopoietic stem/progenitor cell subsets contribute to human hematopoietic homeostasis

In physiological conditions, few circulating hematopoietic stem/progenitor cells (cHSPC) are present in the peripheral blood but their contribution to hematopoietic homeostasis in humans remain unsolved. By integrating advanced immunophenotyping, cellular indexing of transcriptomes and epitopes by sequencing (CITE-seq), functional single-cell assays and integration site (IS) clonal tracking, we unveiled the phenotypic composition, the transcriptional features and the biological role of human cHSPC subpopulations in relationship to their bone marrow (BM) counterpart. We found that cHSPC progressively reduced in cell count over aging and are enriched for primitive, lymphoid and erythroid subpopulations, showing pre-activated transcriptional and functional state. Moreover, cHSPC have low expression of multiple BM-retention molecules, but maintain their homing potential after xenotransplantation. By generating a comprehensive Human Organ-Resident HSPC (HuOR) dataset based on scRNAseq data, we detected organ-specific seeding properties of the distinct trafficking HSPC subpopulations. Of note, circulating multi-lymphoid progenitors (MLP) are primed for seeding the thymus and actively contribute to T-cell production at steady state in patients treated with HSPC-gene therapy (GT). Human clonal tracking data from GT patients also showed that cHSPC connect distant BM niches and participate to steady-state hematopoietic production, with primitive cHSPC having the highest re-circulation capability to travel in and out the BM. Finally, in case of hematopoietic impairment, cHSPC composition reflects the BM-HSPC content and might represent a biomarker of the BM state for clinical and research purposes. Overall, our comprehensive work unveiled fundamental insights into the in vivo dynamics of human HSPC trafficking and its role in sustaining hematopoietic homeostasis.

生理状态下，外周血中仅存在极少量循环造血干/祖细胞（circulating hematopoietic stem/progenitor cells, cHSPC），但其对人类造血稳态的贡献仍未阐明。本研究整合了先进的免疫分型技术、转录组与表位细胞索引测序（cellular indexing of transcriptomes and epitopes by sequencing, CITE-seq）、功能性单细胞检测以及整合位点（integration site, IS）克隆追踪技术，阐明了人类循环造血干/祖细胞亚群与其骨髓（bone marrow, BM）对应群体的表型组成、转录特征及生物学功能。研究发现，循环造血干/祖细胞的数量随衰老逐渐减少，且富集于原始、淋巴系及红系亚群，呈现预激活的转录与功能状态。此外，循环造血干/祖细胞低表达多种骨髓驻留分子，但在异种移植后仍保留归巢潜能。本研究基于单细胞RNA测序（single-cell RNA sequencing, scRNAseq）数据构建了一套完整的人类器官驻留造血干/祖细胞（Human Organ-Resident HSPC, HuOR）数据集，揭示了不同循环造血干/祖细胞亚群的器官特异性定植特性。值得注意的是，循环多淋巴系祖细胞（circulating multi-lymphoid progenitors, MLP）具备定植胸腺的预编程特性，在接受造血干/祖细胞基因治疗（HSPC-gene therapy, GT）的患者稳态条件下，可主动参与T细胞生成。来自基因治疗患者的人类克隆追踪数据同样表明，循环造血干/祖细胞可连接远端骨髓微环境，并参与稳态造血生成，其中原始循环造血干/祖细胞具备最强的往返骨髓的再循环能力。最后，当造血功能受损时，循环造血干/祖细胞的组成可反映骨髓造血干/祖细胞的含量，有望成为临床与科研中评估骨髓状态的生物标志物。综上，本研究通过系统性工作阐明了人类造血干/祖细胞循环的体内动态变化及其在维持造血稳态中的核心作用。