Abstract

Previously, we demonstrated that activation of Notch receptors by culture of murine lin Sca-1+c-kit+ (LSK) hematopoietic progenitors with the Notch ligand Delta1ext-IgG, consisting of the extracellular domain of Delta1 fused to the Fc domain of human IgG, promoted early T cell differentiation and increased the number of progenitors capable of short-term lymphoid and myeloid reconstitution. Here we show accelerated thymic engraftment and T cell recovery after hematopoietic cell transplantation (HCT) in recipients of LSK cells cultured with Delta1ext-IgG compared to recipients of non-cultured LSK cells or control human IgG-cultured LSK cells. Furthermore, data suggest that addition of Delta1ext-IgG cultured LSK cells to non-cultured LSK cells have the potential to accelerated T cell recovery by the non-cultured LSK cells by facilitating thymic engraftment by the non-cultured LSK cells. 103 Ly5a LSK cells cultured with Delta1ext-IgG for 21 days generated a 108-fold increase in total number of cells compared to culture with control IgG. Lethally irradiated C57Bl/6 mice received 105 C57Bl/6 GFP+ LSK cells alone or along with 103 Ly5a LSK cells or 106 Lya5a Delta1ext-IgG-cultured cells or 106 Ly5a IgG cultured cells. We monitored egraftment by difference in CD45 as well as GFP. Mice were sacraficed and blood, bone marrow and thymus were harvested at 1,2,3 and 4 wks after HCT. At 3 wks after HCT, blood samples from recipients of Delta1ext-IgG-cultured cells contained at least 4-fold higher total number of CD3+ cells compared to the other control groups (p<0.05). Furthermore, recipients of Delta1ext-IgG-cultured cells contained at least 3-fold higher numbers of CD3+ cells derived from the GFP+ LSK cells compared to the other control groups (p<0.05). 1 wk after HCT, thymuses and BM from recipients of Delta1ext-IgG-cultured cells showed at lest a 3- and 7-fold increase, respectively, in the number of GFP+ cells compared to the other control groups (p<0.05and p<0.007). Previously, we have shown our system generates committed and non-committed T cell progenitors with enhanced T cell reconstitution potential. Here, we demonstrate that the addition of Delta1ext-IgG-cultured LSK cells to a non-cultured LSK graft has the potential to facilitate BM and thymic engraftment of the non-cultured LSK cells. By using the GFP+ LSK cells, we were able to demonstrate that the facilitation of early BM and thymic recovery was due to the increased engraftment/homing of the GFP+ cells rather than autologous recovery of the host. Thus this report describes a novel and clinically feasible ex-vivo culture system using Delta1 for the generation of T cell progenitors as a means to accelerate initial T-cell recovery as well as facilitating BM and thymic engraftment after HCT.

Disclosures: No relevant conflicts of interest to declare.

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