Vascular Niche-Derived Angiocrine Factors Specify and Maintain Hematopoietic Stem Cells

Organ-specific endothelial cells (ECs) are both conduits for delivery of nutrients and also establish an instructive vascular niche. The vascular niche produces paracrine factors, (i.e., angiocrine factors), that balance self-renewal and differentiation of hematopoietic stem/progenitor cells (HSPCs) (1,2). Activation of Akt-mTOR pathway in sinusoidal ECs (SECs) stimulates physiological expression of angiocrine factors, including Kit-ligand, Notch-ligands, Wnts, FGFs, BMPs and TGFb, that expand long-term repopulating HSPCs. Activation of MAPkinase in ECs upregulates expression of GM-CSF, M-CSF, IL6, IL7, SDF-1 and G-CSF (..others) to accelerate HSPC multi-lineage differentiation. We developed an ex vivo vascular niche in which HSPC/EC co-cultures are maintained and expanded in serum-free conditions. This vascular niche platform produces physiologic levels of angiocrine factors that balance expansion/differentiation of human cord blood, mobilized peripheral blood, and steady state bone marrow HSPCs that maintain their ability to reconstitute hematopoiesis in vivo. In contrast to our vascular platform, co-culture with bone marrow-derived mesenchymal does not support long-term expansion of HSPCs. In collaboration with Drs. Kiem and Gori at Hutchinson Cancer Center, we have shown that ECs expand repopulating nonhuman primate marrow-derived HSPCs. Transplantation of the vascular-niche expanded gene-modified HSPCs reconstituted long-term multi-lineage hematopoiesis in autologous transplantation setting in nonhuman primates. Importantly, intravenous co-infusion of the vascular niche with HSPCs did not cause infusional toxicity. Vascular niche-expanded HSPCs supported robust hematopoietic recovery underscoring the essential function of vascular niche-signals in hematopoietic reconstitution without provoking fibrosis (3). The ECs also supplies key signals that induce emergence of HSPCs from hemogenic ECs. To prove this point, we transduced adult human or mouse ECs with Runx1/Spi1/Gfi1/FosB transcription factors along with vascular niche-induction allowing for conversion of these ECs into stable and long-term engraftable HSPCs, including functional immune cells (4). Importantly, transition through a pluripotent state results in poorly engraftable hematopoietic cells that are unstable and upon exposure to pathophysiological stressors differentiate aberrantly into other cell-types. Remarkably, signals from vascular niche support specification of repopulating multipotent-HSPCs from both human and nonhuman primate pluripotent stem cells (5). In summary, we developed and characterized a vascular niche platform that provides physiologically relevant levels of key angiocrine factors that stimulate safe clinical-scale expansion of authentic adult, cord blood, and primitive HSPCs under GMP-grade culture conditions. We are currently translating the vascular niche platform to the clinical setting, to evaluate the potential of co-transplantation of HSPCs with vascular niche cells to reconstruct injured EC niches thereby accelerating short- and long-term hematopoietic recovery. This first-in-man clinical application will set the stage for repopulation with true hematopoietic stem cells, thereby enabling use of a vascular niche for treatment of a wide range of acquired, inherited, and malignant hematopoietic diseases.

1. Butler JM …… Rafii S. Endothelial cells are essential for the self-renewal and repopulation of Notch-dependent hematopoietic stem cells. Cell Stem Cell, 3:251-64, 2010.

2. Nolan D........Rafii S. Molecular and cellular signatures of tissue-specific vascular heterogeneity in organ maintenance and regeneration. Developmental Cell, 26(2):204-19, 2013.

3. Ding BS …..Rafii S. Divergent angiocrine signals from vascular niche balance liver regeneration and fibrosis.Nature 505(7481):97-102, 2014.

4. Sandler VM, Lis R ...... Butler JM, Scandura JM, Rafii S. Reprogramming of Human Endothelium Into Engraftable Hematopoietic Progenitors by Vascular Niche Induction.Nature, 511(7509):312-8, 2014.

5. Gori J., Butler JM, .....Rafii S, Kiem HP. Vascular niche promotes hematopoietic multipotent progenitor formation from pluripotent stem cells. Journal of Clinical Investigation, 125(3): 1243-54, 2015.