The mechanisms through which the bone marrow (BM) microenvironment regulates hematopoietic stem cell (HSC) fate remain incompletely understood. We examined the role of the heparin-binding growth factor pleiotrophin (PTN) in regulating HSC function in the niche. PTN−/− mice displayed significantly decreased BM HSC content and impaired hematopoietic regeneration following myelosuppression. Conversely, mice lacking protein tyrosine phosphatase receptor zeta, which is inactivated by PTN, displayed significantly increased BM HSC content. Transplant studies revealed that PTN action was not HSC autonomous, but rather was mediated by the BM microenvironment. Interestingly, PTN was differentially expressed and secreted by BM sinusoidal endothelial cells within the vascular niche. Furthermore, systemic administration of anti-PTN antibody in mice substantially impaired both the homing of hematopoietic progenitor cells to the niche and the retention of BM HSCs in the niche. PTN is a secreted component of the BM vascular niche that regulates HSC self-renewal and retention in vivo. Display omitted ► Pleiotrophin (PTN) regulates hematopoietic stem cell self-renewal in vivo ► Deletion of protein tyrosine phosphatase receptor zeta amplifies the hematopoietic stem cell pool ► PTN is expressed and secreted by bone marrow endothelial cells ► PTN regulates hematopoietic progenitor cell homing and retention Much remains unknown regarding how the bone marrow (BM) microenvironment regulates hematopoietic stem cell (HSC) fate. Chute and colleagues demonstrate that BM endothelial cells secrete pleiotrophin (PTN), a heparin-binding growth factor, in the HSC niche. Mice lacking PTN contained decreased BM HSCs and a severe defect in hematopoietic regenerative capacity. Antibody blockade of PTN inhibited HSC homing to the niche and promoted HSC mobilization to the blood. Thus, PTN is a secreted component of the vascular niche that regulates HSC retention and self-renewal.