RESUMEN
The CD34 antigen is present at all differentiation stages of hematopoietic cells, from immature progenitor cells to committed precursor cells. In vivo, transplantation of CD34+ cells is sufficient to allow hematopoietic recovery after myeloablative chemotherapy, but a neutropenic period of 9-12 days still exists, even when hematopoietic growth factors are given posttransplantation. After ex vivo expansion cultures in the presence of cytokines, CD34+ cells can generate mature precursor cells in a stroma-free liquid culture system. This could lead to a shortening of the aplasia duration, but the persistence of primitive progenitor cells in the expanded CD34+ compartment remains to be demonstrated. In this study, CD34+ cells were isolated from eight peripheral blood (PB) and eight cord blood (CB) samples using either Isolex 50 (n = 6), Ceprate LC CD34 kit (n = 6), or Microcellector T-25 Stem Cell kit (n = 4). We have evaluated the functional potential of CD34+ cells after 7 days of ex vivo expansion culture in the presence of 500 UI/ml of interleukin-1 (IL-1), 10 ng/ml of IL-3, and 10 ng/ml of stem cell factor (SCF). The expansions of nucleated cells, granulocyte-macrophage colony-stimulating factor (GM-CSF)-responsive committed precursors, IL-1 + IL-3 + SCF + erythropoietin (EPO)-responsive multilineage progenitors, and 5-fluorouracil (5-FU)-resistant quiescent progenitor were 8-fold, 59-fold, 4.4-fold, and 2.2-fold, respectively. There was no significant difference in the amplification/expansion parameters between cultures initiated with CD34+ cells from PBSC or CB. Our data confirm that cytokine-mediated ex vivo expansion of blood CD34+ cells can produce large numbers of committed precursors and does not significantly affect the compartment containing more immature progenitors. Cytokine-mediated expansion could be of great interest in autologous transplantation to decrease the duration of marrow aplasia.
Asunto(s)
Antígenos CD34/análisis , Células Madre Hematopoyéticas/citología , Diferenciación Celular/efectos de los fármacos , Diferenciación Celular/inmunología , División Celular/efectos de los fármacos , Células Cultivadas , Resistencia a Medicamentos , Sangre Fetal/citología , Sangre Fetal/efectos de los fármacos , Fluorouracilo/farmacología , Trasplante de Células Madre Hematopoyéticas/métodos , Células Madre Hematopoyéticas/inmunología , Humanos , LeucaféresisRESUMEN
CD34 positive (CD34+) cells contain all hematopoietic progenitors from stem cells to committed precursors. Therefore the transplantation of purified bone marrow or blood CD34+ cells is sufficient for hematopoietic recovery after a myeloablative radiochemotherapy. Using different techniques, CD34+ progenitors can be induced to undergo terminal differentiation in a stroma-free liquid culture system in the presence of cytokines. In the present study, we have evaluated the functional potential of CD34+ blood progenitors after ex-vivo expansion cultures. CD34+ cells were isolated from 16 samples (PBSC n = 8 and Cord Blood (CB) n = 8) using either ISOLEX 50 (n=6), CEPRATE LC CD34 kit (n = 6) or MICROCELLECTOR T-25 Stem Cell kit (n = 4). CD34+ cells were cultured for seven days in the presence of 500 UI/ML of IL-1, 10 ng/ml of IL-3 and 10 ng/ml of SCF. We obtained an 8-fold expansion of nucleated cells. We observed a 59-fold expansion of GM-CSF responsive committed precursors, a 4.4-fold expansion of IL-1+IL-3+SCF+Epo responsive multilineage progenitors and a 2.2-fold expansion of the 5-FU resistant quiescent progenitors. We did not observe any significant difference in the amplification/expansion parameters between cultures initiated with CD34+ cells from PBSC or CB. Our data show that cytokine mediated ex-vivo expansion of blood CD34+ cells can produce a large number of committed precursors without affecting the compartment of the most immature progenitors. These results suggest that cytokine-mediated amplification technology could be of great interest in the autologous transplantation setting.