Single cell RNA sequencing of mutant bone marrow stromal cells lacking HIF2 and controls.

Osteoblast lineage cells within adult bone marrow are exposed to varying oxygen levels. Hypoxia-Inducible Factor-1α (HIF1α) and HIF2α are pivotal in the cellular response to hypoxia. Our study explores the effects of targeted HIF2α deletion in mesenchymal progenitors and their descendants. We demonstrate that HIF2α acts as a negative regulator of osteoblastogenesis and bone mass accrual. Therapeutically targeting HIF2α could be beneficial in treating low bone mass conditions, such as those observed in chronic diseases, osteoporosis, or during aging. To elucidate the mechanisms underlying the increased bone mass resulting from HIF2α loss, we performed single-cell RNA sequencing (scRNA-seq) on bone marrow stromal cells from both mutant mice lacking HIF2α in PRX1 lineage cells and control mice. To investigate how the deletion of HIF2 affects the transcriptional profile of bone marrow stromal cells, we conditionally knocked out HIF2 in uncommitted mesenchymal progenitors and their descendants using PRX1-Cre driver, and a mouse line homozygous for a floxed Hif2 allele (HIF2f/f). PRX1-Cre mice express Cre recombinase in mesenchymal cells that give rise to chondrocytes and osteoblasts in long bones and calvaria. The Ai14 reporter was used to recognize Ai14-positive bone marrow stromal cells. PRx1-Cre+/Ai14+/-/HIF2f/+ mice were crossed with HIF2f/f or C57 mice to generate PRx1-Cre+/Ai14+/-/HIF2f/f mutants and PRx1-Cre+/Ai14+/-/HIF-/- control littermates. scRNA-seq was performed on bone marrow stromal cells isolated from the long bones hindlimbs of 12-week-old mice.