Induction of Stem Cell Expansion by the Vascular Niche

Project Title: Induction of Stem Cell Expansion by the Vascular Niche

Ancillary Site Principal Investigator: Shahin Rafii, M.D.

Hub Site Principal Investigator: Irwin Bernstein, M.D.

Awarded Organization: Fred Hutchinson Cancer Research Center

Abstract:

A major goal of the NHLBI Progenitor Cell Biology Consortium is to derive hematopoietic stem cells (HSC) from pluripotent stem cell sources for therapeutic and scientific purposes. However, although hematopoietic cells have been generated from embryonic stem cells (ESC) and induced pluripotent stem cells (iPSC), it has not been possible to derive true, long-term repopulating HSC. Thus, the overall goal of this ancillary grant to the NHLBI Progenitor Cell Biology Consortium (U01 HL100395) is to examine the potential of vascular niche models in enhancing the generation of functional hematopoietic stem and progenitor cells (HSPCs) from embryonic stem cells (ESCs). Endothelial cells (ECs) serve an integral role in the supportive niche of HSC throughout development, from the first site of HSC generation in the dorsal aorta of the embryonic AGM (aorta-gonad-mesonephros) region [1], to the EC sinusoids of the fetal liver where HSC undergo significant proliferative expansion [2], and finally in bone marrow where sinusoidal ECs play a role in adult HSC homeostasis [3]. Thus, in vitro recapitulation of the critical signal pathways that regulate these developmentally distinct EC hematopoietic niche environments, which are involved in establishing, maintaining, and expanding HSCs throughout development, will be valuablein the pursuit of generating and expanding HSC from pluripotent stem cells. Our collaborative studies between the Rafii laboratory and NHLBI Progenitor Cell Biology Consortium supported Bernstein laboratory have shown that endothelial cells (ECs) derived from human umbilical vein (HUVEC) by production of paracrine factors, defined as angiocrine factors, support the expansion of adult murine and human cord blood HSPC [4]; however, in follow-up studies these ECs failed to support embryonic-stage (AGM-derived) HSC in vitro. Consequently, our laboratories have explored whether developmental-specific vascular niches are capable of supporting embryonic-stage HSC. Recent preliminary experiments have now determined that AGM-derived ECs significantly enhance engraftment from AGM-derived HSC following in vitro co-culture. Furthermore, our studies of organ-specific ECs [5, 6] have revealed unique transcriptional profiles suggesting that each organ specific vascular bed may be endowed with distinctive functional and angiocrine capacity to instruct generation and self-renewal of stem/progenitor cells at specific developmental stages. Based on this data, we hypothesize that ECs derived from the embryonic hematopoietic vascular niches, by supplying unique instructive signals, will significantly increase the specification and expansion of HSPCs, including true repopulating HSCs. The immediate goals of the present application are to: 1. Determine the optimal vascular niches and ex vivo conditions suited for induction and expansion of embryonic-stage HSPCs; and 2. Extend findings utilizing these vascular niches to expand ESC and iPSC-derived HSPC capable of in vivo engraftment. Furthermore, comparison of transcriptional profiles from various sources of embryonic, fetal, and adult ECs will allow us to begin to deconstruct the critical stage-specific signaling pathways differentially required for induction and expansion of HSC from embryonic sources. Therefore, in this proposal we seek to utilize ourunique embryonic and fetal sources of vascular cells to address the hypotheses proposed in the following specific aims.