Osteomacs maintain the endosteal hematopoietic stem cell niche and participate in mobilization

OsteoMacs are tissue macrophages that reside within bone lining tissues. OsteoMacs enhance osteoblast function in vitro and maintain osteoblast bone forming surface in vivo. Given the endosteum is a preferred niche for hematopoietic stem cells (HSC), we investigated whether OsteoMacs participate in HSC mobilization induced by granulocyte colony-stimulating factor (G-CSF). Immunohistochemistry (IHC) using the F4/80 antibody demonstrated that mobilization induced by G-CSF resulted in a sharp reduction in the number of F4/80+ OsteoMacs within the endosteal niche from days 2–6 post initiation of G-CSF administration. At day 2, clustering of F4/80+ cells in the central marrow and in particular around endothelial sinuses was evident. In contract, localization and maintenance of OsteoMacs within the periosteum was unaffected by G-CSF administration. Flow cytometry demonstrated increased numbers of F4/80+ macrophages in blood from days 1–6, indicating egress of macrophages from bone marrow to blood. Static and dynamic histomorphemtry and IHC for osteocalcin expression demonstrated a concomitant significant reduction in endosteal osteoblast number and function as well as osteoid. Notably, the kinetics of OsteoMac loss strikingly parallels, if not precedes, that of endosteal osteoblasts loss and mobilization of HSC to blood. In contrast, osteoclast numbers were not significantly altered by G-CSF administration and treatment with the bisphosphonate zoledronate increased HSC mobilization in response to G-CSF. F4/80+ macrophages repopulation of bone marrow and endosteal surfaces initiated at day 8, 2 days after cessation of G-CSF administration, and was quickly followed at day 10 by recovery of OsteoMac canopy covering cubiodal osteoblast-like cells. Prior experimentation has indicated that mobilization induced by G-CSF acts through an intermediary G-CSF receptor expressing myeloid population. Finally, we demonstrate a pivotal role of macrophages in maintaining endosteal HSC niches in transgenic MAFIA mice in which macrophages can be specifically deleted in vivo using AP20187. At day 5 following macrophages deletion, we observed a collapse in the number of osteoblasts lining the endosteum and robust mobilization of HSC in peripheral blood and spleen. Given macrophages can express the G-CSF receptor and the observations reported here, we propose that G-CSF induces HSC mobilization through OsteoMac-directed collapse of the endosteal HSC niches