The polycomb group protein Bmi1 maintains hematopoietic stem cell (HSC) functions. We previously reported that Bmi1-deficient mice showed a progressive fatty change of bone marrow (BM). A large portion of HSCs reside in perivascular niche created partly by endothelial cells and leptin receptor+ (LepR+) mesenchymal stromal cells. To clarify how Bmi1 regulates HSC niche, we deleted Bmi1 specifically in LepR+ cells in mice. The deletion of Bmi1 promoted the adipogenic differentiation of LepR+ stromal cells and caused a progressive fatty change of BM of the limb bones with age, resulting in reduction in the numbers of HSCs and progenitors. The adipogenic change was also evident during BM regeneration after irradiation. Several genes involved in adipogenesis, such as Nr2f2 (COUP-TFII), appeared to be the direct targets of Bmi1. Our results indicate that Bmi1 keeps adipogenic differentiation program repressed in BM stromal cells to maintain the integrity of HSC niche.