Cord blood (CB) is a clinically relevant source of hematopoietic stem/progenitor cells (HSPC) to treat cancer and genetic diseases. The advantages of using CB include its ready availability, the reduced probability of transmitting vira infections, and the lower risk of inducing graft vs. host disease in HLA-mismatched recipients. Still, CB's delayed speed of engraftment, and the relatively low number of hematopoietic stem cells (HSC) per unit, limit its broader use. Despite the advancements made in CB-HSPC expansion, challenges remain regarding the ability to obtain, from a single unit, sufficient numbers of both long-and short-term repopulating cells, for treatment of an adolescent or adult patient. We have previously shown that CB-HSPC can be expanded and differentiated towards both the myeloid and lymphoid lineages, using a feeder layer of adult human bone marrow-derived stromal cells. Using this system, we optimized the initial progenitor content and cytokine concentrations, and showed that expanded cells had the ability to engraft pre-immune fetal sheep. While the absolute number of long-term engrafting HSC increased in this culture system, still, the relative percentage of these most primitive stem cells decreasd with time. Recently, we have developed three- dimensional (3-D), liver extracellular matrix (ECM)-derived scaffolds and seeded them with fetal hepatoblasts and endothelial cells. These cells engrafted in their putative native locations within the liver ECM scaffolds, and subsequently displayed typical endothelial, hepatic, and biliary epithelial markers, thus creating a hepatic-lik tissue in vitro. It is well known that, during development, the fetal liver is the main site of HSC expansion and differentiation. Within the fetal liver, HSC actively cycle and these cells outcompete adult HSC upon transplantation. Thus, within the hepatic tissue, cellular niches exist that promote asymmetric or symmetric self-renewal divisions, leading to maintenance or expansion of primitive HSC. In addition, the initial divisional behavior of CB-HSPC is highly dependent upon the environment. For example, the stromal cell line AFT024 and fetal hepatoblasts, both of murine origin, have been shown, in 2-D cultures, to effectively preserve the self- renewal capacity of human and mouse HSC, respectively. Therefore, we hypothesize that a functional and efficient expansion of CB-HPSC can be achieved under physiological conditions provided by the bioengineered human hepatic constructs. Our ultimate goal is to develop a novel platform for the efficient expansion of CB- HSPC using bioengineered human liver tissue. To this end, we will: 1) Determine the ability of 3-D bioengineered huma liver tissue constructs to support ex-vivo expansion of CB-derived HSPC~ and 2) Examine and define the functional outcome arising from interactions that occur between CB-HSPC and individual cellular and matrix components of the niches of the bioengineered liver tissue. Upon completion, these studies will add to the understanding of how fetal liver niches support HSC expansion, and, more importantly, will allow the development of a novel strategy to functionally expand CB-HPSC.

R21HL117704

2013-07-19

2016-06-30