title: Clinical-scale cultures of cord blood CD34(+) cells to amplify committed progenitors and maintain stem cell activity.
abstract: We developed a clinical-scale cord blood (CB) cell ex vivo procedure to enable an extensive expansion of committed progenitors--colony-forming cells (CFCs) without impairing very primitive hematopoietic stem cells (HSCs). CD34(++) cells, selected from previously cryopreserved and thawed CB units, were cultured in two steps (diluted 1:4 after 6 days) in the presence of stem cell factor (SCF), fms-related tyrosine kinase 3 ligand (Flt-3L), megakaryocyte growth and development factor (MGDF) (100 ng/ml each), granulocyte-colony stimulating factor (G-CSF) (10 ng/ml) in HP01 serum-free medium. HSC activity was evaluated in a serial transplantation assay, by detection of human cells (CD45, CD33, CD19 and CFC of human origin) in bone marrow (BM) of primary and secondary recipient NOD/SCID mice 6-8 weeks after transplantation. A wide amplification of total cells (∼350-fold), CD34(+) cells (∼100-fold), and CFC (∼130-fold) without impairing the HSC activity was obtained. The activity of a particular HSC subpopulation (SRC(CFC)) was even enhanced.Thus, an extensive ex vivo expansion of CFCs is feasible without impairing the activity of HSCs. This result was enabled by associating antioxidant power of medium with an appropriate cytokine cocktail (i.e., mimicking physiologic effects of a weak oxygenation in hematopoietic environment).
authors.affiliation: Etablissement Français du Sang Aquitaine-Limousin, Bordeaux, France. zoran.ivanovic@efs.sante.fr
authors.forename: Zoran
authors.initials: Z
authors.lastname: Ivanovic
authors.forename: Pascale
authors.initials: P
authors.lastname: Duchez
authors.forename: Jean
authors.initials: J
authors.lastname: Chevaleyre
authors.forename: Marija
authors.initials: M
authors.lastname: Vlaski
authors.forename: Xavier
authors.initials: X
authors.lastname: Lafarge
authors.forename: Bernard
authors.initials: B
authors.lastname: Dazey
authors.forename: Elodie
authors.initials: E
authors.lastname: Robert-Richard
authors.forename: Frédéric
authors.initials: F
authors.lastname: Mazurier
authors.forename: Jean-Michel
authors.initials: JM
authors.lastname: Boiron
mesh_terms: D000818:Animals; D018952:Antigens, CD34 / Q000378:metabolism*; D002452:Cell Count; D018929:Cell Culture Techniques / Q000379:methods*; D019070:Cell Lineage; D002465:Cell Movement; D049109:Cell Proliferation; D002478:Cells, Cultured; D003114:Colony-Forming Units Assay; D005312:Fetal Blood / Q000166:cytology*; D006801:Humans; D016130:Immunophenotyping; D051379:Mice; D016688:Mice, Inbred NOD; D016513:Mice, SCID; D033581:Stem Cell Transplantation; D013234:Stem Cells / Q000166:cytology* / Q000378:metabolism
chemical_list: D018952:Antigens, CD34