Ex vivo expansion of human cord blood Lin-CD34-cells using indian hedgehog stromal cells

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Transplantation of hematopoietic stem cells (HSCs) is usually based on intravenous injection, a complex process that requires circulation through blood, recognition and extravasation through bone marrow vasculature, and migration to a supportive microenvironment. However, some of HSCs including CB Lin-CD34- stem cells rarely engraft in bone marrow. We cocultured CB Lin-CD34- cells with human stromal cells in the presence of TPO, FL and SCF. As a result, cocultured CB Lin-CD34- cells remarkably expressed chemokine receptor CXCR4 with remarkable proliferative potential although CB Lin-CD34- cells undergo apoptosis without stromal free culture (Total cells, 92±10 vs. 1542±161(x10^3) ; CD34+ cells, 0±0 vs. 156±16 (x10^3) ; CFU-C, 192±46 vs. 120320±34762 ; CFU-Mix, 8±5 vs. 6513±3791 from five hundred thousand CB Lin-CD34- cells). Moreover, these cells acquired homing activity to BM and engrafted in NOD/SCID/IL2R _chain null mice for 24 weeks although Lin-CD34- cells cultured without human stromal cells hardly engrafted. Therefore, stromal interaction with CB Lin-CD34- cells confers the ability of engraftment to BM niche and responsiveness to hematopoietic growth factors. These findings may provide novel strategy of BM transplantation and utilization of CB Lin-CD34- fraction for ex vivo production of blood cells.