title: Preclinical ex vivo expansion of G-CSF-mobilized peripheral blood stem cells: effects of serum-free media, cytokine combinations and chemotherapy.
abstract: OBJECTIVE
  Ex vivo expansion of granulocyte-colony stimulating factor (G-CSF)-mobilized peripheral blood stem cells (PBSC) is a promising approach for overcoming the developmental delay of bone marrow (BM) reconstitution after transplantation. This project investigated the effects of culture duration, serum-free media, cytokine combinations, and chemotherapy on the outcomes of expansion.
  
  
  METHODS
  Enriched CD34+ cells were cultured for 8 or 10 d in serum-free media (QBSF-60 or X-Vivo 10) and four combinations of cytokines consisting of recombinant human pegylated-megakaryocyte growth and development factor, stem cell factor, flt-3 ligand, G-CSF, interleukin (IL)-6, platelet-derived growth factor (PDGF), and IL-1beta.
  
  
  RESULTS
  Eight days of culture in QBSF-60 significantly supported efficient expansions of CD34+ cells, CD34+ CD38- cells, colony-forming units (CFU) of myeloid, erythroid, megakaryocytic, and mixed lineages to 3.76-, 14.4-, 28.3-, 24.0-, 38.1-, and 15.7-fold, respectively. Whilst PDGF or IL-6 enhanced the expansion of early, myeloid, and erythroid progenitors, IL-1beta specifically promoted the megakaryocytic lineage. Engraftment of human CD45+ cells were detectable in all non-obese diabetic/severe-combined immunodeficient mice transplanted with expanded PBSC from donor samples, being 5.80 +/- 3.34% of mouse BM cells. The expansion and engraftment capacity of CD34+ cells from subjects postchemotherapy were significantly compromised across the panel of progenitor cells.
  
  
  CONCLUSIONS
  Our results provided an optimized protocol for PBSC expansion, applicable to ameliorating neutropenia and thrombocytopenia in post-BM transplant patients by the prompt provision of progenitor cells. For postchemotherapy patients, expansion products might provide committed progenitors for improving short-term engraftment, but not self-renewable stem cells.
authors.affiliation: Department of Paediatrics, Prince of Wales Hospital, The Chinese University of Hong Kong, Hong Kong, China. lipang@cuhk.edu.hk
authors.forename: Karen
authors.initials: K
authors.lastname: Li
authors.forename: Chi Kong
authors.initials: CK
authors.lastname: Li
authors.forename: Carmen Ka Yee
authors.initials: CK
authors.lastname: Chuen
authors.forename: Kam Sze
authors.initials: KS
authors.lastname: Tsang
authors.forename: Tai Fai
authors.initials: TF
authors.lastname: Fok
authors.forename: Anthony Edward
authors.initials: AE
authors.lastname: James
authors.forename: Shuk Man
authors.initials: SM
authors.lastname: Lee
authors.forename: Matthew Ming Kong
authors.initials: MM
authors.lastname: Shing
authors.forename: Kai Wai
authors.initials: KW
authors.lastname: Chik
authors.forename: Patrick Man Pan
authors.initials: PM
authors.lastname: Yuen
mesh_terms: D036541:ADP-ribosyl Cyclase / Q000378:metabolism; D051997:ADP-ribosyl Cyclase 1; D000293:Adolescent; D000328:Adult; D015703:Antigens, CD / Q000378:metabolism; D018952:Antigens, CD34 / Q000378:metabolism; D001853:Bone Marrow / Q000473:pathology / Q000502:physiology; D016026:Bone Marrow Transplantation; D049109:Cell Proliferation / Q000187:drug effects*; D002478:Cells, Cultured; D002648:Child; D016895:Culture Media, Serum-Free; D016207:Cytokines / Q000494:pharmacology*; D016179:Granulocyte Colony-Stimulating Factor / Q000494:pharmacology*; D006801:Humans; D008562:Membrane Glycoproteins; D008875:Middle Aged; D023461:Myeloid Progenitor Cells / Q000378:metabolism; D009369:Neoplasms / Q000473:pathology / Q000628:therapy; D013234:Stem Cells / Q000187:drug effects
chemical_list: D015703:Antigens, CD; D018952:Antigens, CD34; D016895:Culture Media, Serum-Free; D016207:Cytokines; D008562:Membrane Glycoproteins; D016179:Granulocyte Colony-Stimulating Factor; D036541:ADP-ribosyl Cyclase; C498154:CD38 protein, human; D051997:ADP-ribosyl Cyclase 1