Combination of Early and Late Growth Factors to Enrich Transplantable Cord Blood CD34+Cells in Short Term Cultures

Abstract

Objective: Although umbilical cord blood (UCB) is a useful source of stem and progenitor cells for bone marrow transplantation, it is not possible to obtain sufficient number of transplantable cells from a single human umbilical cord for transplantation in adult patients. In this study, we combined early and late growth factors in short term cultures to enrich cord blood CD34+ hematopoietic progenitor cell count for adult patients. Material and Methods: Cord blood mononuclear cells (CBMC) harvested from 15 healthy volunteers, delivering in term, incubated in culture medium supplemented with interleukin-3 (IL-3), IL-6, IL-11, stem cell factor, flt3/flk2 ligand and thrombopoietin for 14 days. Starting cells and cultured cells (day 7 and 14 cells) were evaluated for; CD4, CD8, CD33, CD34, CD38 and HLA-DR by flow cytometry and seeded in semisolid methylcellulose culture for determining BFU-E, CFU-GM and CFU-GEMM colony forming abilities. Results: In the short-term cultures, total CBMC count significantly increased (p= 0.0001). Compared to starting cells, the expressions of CD34 (p= 0.002), CD33 (p= 0.0001), HLA-DR (p= 0.0001) and CD8 (p= 0.002) increased, but those of CD38 (p= 0.057) and CD4 (p= 0.0001) were suppressed on day 14 of cultures. There were an increase in generation of CFU-GM (p= 0.131) and CFU-GEMM (p= 0.134) colonies and a significant decrease in generation of BFU-E colonies (p= 0.0001). Conclusion: Ex vivo expansion of CBMC, under combination of the growth factors, contribute to CD34+ progenitor cell count and may help to reach sufficient amount of transplantable cells for adult patients. Suspension cultures may lead to elimination of graft-versus-host disease risk and protection of graft-versus-leukemia effect, through the decrease of CD4+ T cells and maintenance of CD8+ T lymphocytes, respectively.