Mds1 ^CreERT2-Based Lineage-Tracing Reveals Increasing Contributions of HSCs to Fetal Hematopoiesis and to Adult Tissue-Resident Macrophages in the Marrow

The ontogeny of the hematopoietic system consists of two broad programs. The first, an HSC-independent program, consists of overlapping waves of primitive, erythro-myeloid (EMP), and some lymphoid progenitors. HSC-independent hematopoiesis is required for normal fetal development, and provides self-renewing tissue-resident macrophage populations that persist in the adult. This is followed by the emergence of an HSC-dependent program that arises from arterial vessels within the body of the embryo. The overlapping emergence and lineage output of HSC-independent and HSC-derived hematopoiesis raises important questions regarding the identity and potential functional differences of their mature progeny. However, the transition from HSC-independent to HSC-derived hematopoiesis in the murine fetus remains incompletely characterized, particularly since the maturing erythroid, megakaryocytic and myeloid progeny of EMP and HSCs are currently not easily distinguishable. Additionally, lineage-tracing approaches have been challenging because they have relied largely on the temporal induction of promoters that are expressed both in HSC-independent progenitors and in HSCs, which have significant temporal overlap in their developmental emergence and result in incomplete or in mixed labeling. To help resolve this question, we have developed Mds1 ^CreERT2 mice, utilizing the first transcription start site of MECOM gene, which is expressed in HSC and emerging HSC (Yuasa et al., 2005 EMBO; Hou et al. 2020 Cell Research; Zhu et al. 2020, Blood). When mated with Rosa-YFP reporter mice and induced at E9.5 with tamoxifen, this construct lineage-traces pre-HSCs present in the E11.5 AGM region, as well as HSCs in the fetal liver and adult marrow. Importantly, no labeling of primitive erythroid cells, primitive macrophage-derived microglia, EMP, or EMP-derived cells in the E11.5 or E12.5 fetal liver was detected with tamoxifen induction at either E9.5 or E8.5. Analysis of E9.5 tamoxifen-treated Mds1 ^CreERT2Rosa26 ^LSL-YFP embryos indicates that HSCs have begun to generate small numbers of differentiating erythroid, myeloid and lymphoid progeny in the liver between E12.5 and E14.5. By E16.5, a significant proportion of differentiating erythroid, myeloid and B-cell lineage cells in the liver are HSC-derived, and HSCs have now begun to contribute erythroid and myeloid cells to the rapidly expanding pool of circulating blood cells. In the adult, we found increasing contributions of HSCs to macrophages in liver, lung and kidney. Interestingly, the majority of F4/80+ cells in the adult bone marrow and spleen were also lineage-traced in these mice. Thus, HSCs ultimately provide the majority of adult marrow macrophages that go on to self-maintain in the adult marrow (Hashimoto et al., 2013, Immunity). The Mds1 ^CreERT2 mouse model will serve as a useful to deconvolute the complexity of hematopoiesis as it unfolds in the embryo and functions postnatally.