Single-cell immune profiling reveals thymus-seeding populations, T cell commitment, and multi-lineage development in the human thymus

Development of T lymphocytes in the thymus has been widely studied in mice, but our insights into human T cell development are scarce. Using a combination of single-cell techniques and functional assays, we report deep immune profiling of human T cell development, with a focus on the initial development stages of pre-lineage commitment. We identified three thymus-seeding progenitor populations that also have a counterpart in the bone marrow. We found that the human thymus physiologically supports development of monocytes, DCs, NK cells, and limited development of B lymphocytes. These insights are important to monitor and guide regenerative therapies after hematopoietic stem cell transplantation. To get more insight into human T cell development, we performed single-cell RNA-seq on sorted thymocytes from six healthy donors. We used flow cytometry to sort the DN1 (CD34+CD38-CD1a-), DN2 (CD34+CD38+CD1a-), DN3 (CD34-CD38+CD1a+) thymocytes. Maturating CD4 and CD8 thymocytes were also analyzed from the sorted subsets: CD4 immature single-positive (ISP), CD4/CD8 double-positive (DP), CD4 SP and CD8 SP. From the 8 sorted populations we generated 8 scRNA-seq libraries for gene expression profiling using the 10x Genomics platform.