Ex-vivo expansion and maturation of CD34-positive hematopoietic progenitors optimization of culture conditions.
The aim of this study was to look for an ex vivo culture system for clinical application. We evaluated the ex vivo expansion of peripheral blood CD34+ cells in gas-permeable bags and whether or not an exogenous protein source would be required in these kind of cultures. We also evaluated maturation of the cells during culture. Cells were cultured for 15 days in medium supplemented with SCF, G-CSF, IL3 and IL6. The bags supported the expansion of hematopoietic cells in a similar manner to small scale flasks system: (a) the expansion means of total nucleated cells on day +5 were 12.5-fold for bag versus 5-fold for flask, on day +10 were 44.12-fold for bag versus 41-fold for flask and on day +15 were 67.7-fold for bag versus 84.2-fold for flask, (b) the peak values of CFU-GM were reached on day +10 (9.2-fold for bag vs. 12-fold for flask), and (c) maximal expansion of CD15+/CD11b- population occurred on day +10 (517.5-fold for bag vs. 2959.2-fold for flask). So, we did not find any advantages by culturing further than day +10. We subsequently investigated the use of serum-free medium. The study showed better results when we used medium supplemented with autologous plasma versus serum-free system. In summary, these data described a strategy of culture clinically feasible and safe, using gas-permeable bags, and the kinetics and differentiation of neutrophils and neutrophil precursors from selected CD34+ cells in liquid cultures. Ex vivo expansion of this population might result in earlier engraftment as compared with that for selected stem cells alone.
Mellado-Damas
N
N
Servicio de Hematología y Hemoterapia, Hospital Universitario Virgen del Rocio, Sevilla, Spain.
Rodríguez
J M
JM
Carmona
M
M
González
J
J
Prieto
J
J
D000328:Adult; D018952:Antigens, CD34 / Q000032:analysis*; D018929:Cell Culture Techniques; D002455:Cell Division; D002675:Child, Preschool; D005434:Flow Cytometry; D006412:Hematopoietic Stem Cells / Q000166:cytology* / Q000276:immunology; D006801:Humans; D008875:Middle Aged; D009369:Neoplasms / Q000473:pathology; D010641:Phenotype
D018952:Antigens, CD34