title: The impact of progenitor enrichment, serum, and cytokines on the ex vivo expansion of mobilized peripheral blood stem cells: a controlled trial.
abstract: The aim of this study was to verify, and possibly improve, culture conditions to expand human mobilized peripheral blood stem cells (PBSCs). We investigated the role of three parameters: A) the culture medium (serum-free versus serum-dependent); B) the initial cell population (Ficoll-separated mononucleated cells versus CD34(+)-selected cells), and C) the low concentration of recombinant cytokines, flt3 ligand, and thrombopoietin in association with a basic cocktail of stem cell factor, interleukin (IL)-6, IL-3, GM-CSF, and erythropoietin. Eighteen leukapheresis samples were monitored in static culture for 15 days. The expansion potential was assessed at day 10 and 15 by total nuclear cells, colony-forming-units (CFUs) (burst-forming units-erythroid [BFU-E], colony-forming units-granulocyte-macrophage [CFU-GM], and colony-forming units-granulocyte-erythroid-macrophage-megakaryocyte [CFU-GEMM]), and flow cytometry immunophenotyping (CD34(+)/CD38(-), CD38(+), CD33(+), CD41(+), GlyA(+) progenitor cells). The results, evaluated by multivariate analysis of variance, emphasize that some variables affected the outcome of stem and progenitor cell expansion. CD34(+) enrichment increased expansion of total nuclear cells, number of CD38(+) and CD33(+) late precursors, and number of the CFU-GM compartment. Interestingly, however, quantitative expansion of GlyA(+) and the early progenitor cells (CD34(+)/CD38(-), CFU-GEMM, BFU-E) are favored by the use of unselected mononucleated cells. Regarding the role of serum, no significant difference was observed except for expansion of total nuclear cells, CFU-GM, and BFU-E. Cytokine combinations, in particular the use of flt3 ligand, stimulated expansion of almost all the cellular subsets, reaching a statistical significance for total nuclear cells and CFU-GM. Our study indicates that progenitor and late precursor multilineage cell compartments of mobilized PBSCs may be significantly expanded in short-term cultures by well-defined experimental conditions. Furthermore, these data might be useful when evaluating ex vivo expansion of hematopoietic cells for clinical purposes.
authors.affiliation: Department of Oncology and Hemato-Oncology, PF Calvi Hospital, Noale (VE), Italy.
authors.forename: Elena
authors.initials: E
authors.lastname: Balducci
authors.forename: Giuseppe
authors.initials: G
authors.lastname: Azzarello
authors.forename: Maria Teresa
authors.initials: MT
authors.lastname: Valenti
authors.forename: Gian Maria
authors.initials: GM
authors.lastname: Capuzzo
authors.forename: Giovanni Luigi
authors.initials: GL
authors.lastname: Pappagallo
authors.forename: Irene
authors.initials: I
authors.lastname: Pilotti
authors.forename: Simonetta
authors.initials: S
authors.lastname: Ausoni
authors.forename: Mario
authors.initials: M
authors.lastname: Bari
authors.forename: Francesco
authors.initials: F
authors.lastname: Rosetti
authors.forename: Donata
authors.initials: D
authors.lastname: Sartori
authors.forename: Antonino
authors.initials: A
authors.lastname: Ciappa
authors.forename: Adolfo
authors.initials: A
authors.lastname: Porcellini
authors.forename: Orazio
authors.initials: O
authors.lastname: Vinante
mesh_terms: D018929:Cell Culture Techniques / Q000379:methods; D002455:Cell Division / Q000502:physiology; D002469:Cell Separation / Q000379:methods; D002478:Cells, Cultured; D016207:Cytokines / Q000502:physiology*; D005260:Female; D019650:Hematopoietic Stem Cell Mobilization / Q000379:methods*; D006801:Humans; D016130:Immunophenotyping; D007963:Leukocytes, Mononuclear / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism; D036102:Peripheral Blood Stem Cell Transplantation / Q000379:methods; D013234:Stem Cells / Q000737:chemistry / Q000187:drug effects / Q000502:physiology*
chemical_list: D016207:Cytokines