Identification and Isolation of the Hematopoietic Stem Cell Niche Initiating Cell Population

Introduction: Identification and understanding of the cells and processes that can generate, sustain and influence the HSC niche and hematopoiesis are critical for the development of a more comprehensive knowledge of normal hematopoiesis, stem cell homing, trafficking, differentiation and hematopoietic pathology. Growth and renewal in many tissues are initiated by stem cells, supported by the microenvironment (niche) in which they reside. While recent work has begun to describe functional interactions between stem cells and their niches, little is known about the formation of stem cell niches.

Methods & Results: We established a functional, in vivo assay (via implantation of cells under the renal capsule) to isolate the determinants of hematopoietic stem cell (HSC) niche formation and activity. Using this novel assay, we show that a population of progenitor cells (CD45−Tie2-aV+CD105+Thy1.1−; CD105+Thy1−) sorted from 15.5 dpc fetal limbs and transplanted under the adult mouse renal capsule recruit host-derived vasculatures in a VEGF dependent manner, produce donor-derived ectopic bones through endochondral ossification, and generate a marrow cavity populated by host-derived long term reconstituting HSC (LT-HSC). In contrast, CD45−Tie2-aV+CD105+Thy1a+ (CD105+Thy1+) progenitors form bone that does not contain a marrow cavity. While analyzing these and other sorted populations, we did not observe any instances where niche was present without bone, suggesting that skeletal progenitors are necessary for initiating an HSC niche but osteoblasts alone cannot initiate and support niche activity. Suppression of factors important for HSC maintenance, such as steel factor (SLF), in progenitor populations prior to transplant did not alter their ability to initiate and support an HSC niche. On the other hand, suppression of factors involved in endochondral ossification, such as osterix and VEGF, inhibited niche generation. Furthermore, CD105+Thy1− progenitor populations derived from regions of the fetal mandible or calvaria that do not undergo endochondral ossification form only bone without marrow in our assay.

Conclusions: In addition to identifying the limb-derived skeletal progenitor capable of endochondral ossification involved and the basic mechanisms of HSC niche initiation, our study provides a functional framework by which future studies on HSC-niche interactions at the cellular level can be carried out.