Characterizing human ILC development from early hematopoietic progenitors: regulation by intrinsic and extrinsic signals

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Innate lymphoid cells (ILC) are early acting lymphocytes that lack antigen-specific receptors and either exert T helper-like (ILC1-3) or cytotoxic effector functions (NK cells). In humans, ILC progenitors and the molecular events leading to the specification of group 1, 2, and 3 ILCs are not well understood. Except the ILC3p, progenitor populations for other ILC subsets were so far only described in the mouse. One major challenge for the faithful identification of human ILC progenitors is to provide suitable developmental niches, which mimic the in vivo environment and provide the necessary cell-contact dependent and independent extrinsic signals. In this regard, studies of human ILC differentiation are so far based on murine stem cell niches or stroma-free culture systems, which might not provide the adequate species-specific environmental signals that are present in the respective locations such as human bone marrow (BM). In this project, as part of the DFG Priority Program Innate Lymphoid Cells, we would like to identify early human ILC progenitors in BM and cord blood and explore their differentiation potential using novel human co-culture models. In this regard, we have recently developed an in vitro differentiation protocol employing human mesenchymal stem cells (MSC) to support NK cell development in a fully human system. We would thus like to employ MSC from bone marrow, cord blood, and tonsils to compare how ILC development from early HPC is influenced by different niche conditions. By differentiating early BM progenitors on tonsil MSC we hope to mimic the conditions found in vivo assuming that BM progenitors are trafficking to peripheral lymphoid organs where they would get the necessary signals for differentiation towards ILC effector subsets. We would like to monitor early transcriptional changes in these cultures by RNAseq and identify regulatory modules that are induced by variation in niche conditions. Furthermore, we would like to address the role of the two transcription factors E4BP4/NFIL3 and ID2, which play key roles in ILC development by modulating their expression in this system. Besides increasing basic knowledge about the regulation of human ILC development, the study will provide defined conditions for in vitro differentiation and expansion of human ILC from hematopoietic progenitors without xenogeneic feeder cells. In this way, the project will help to develop suitable protocols for production of clinical-grade products, for example from cord blood, for the future use of ILC as cellular therapeutics.