ANIMAL SERUM-FREE CULTURE OF PURIFIED HUMAN CD34- AMPLIFICATION OF PROGENITORS FROM C-CSF AND GM-CSF-MOBILIZED PERIPHERAL-BLOOD( CELLS )

The isolation and culture of human CD34+ cells could have broad clinic al application for hematologic support following high-dose chemotherap y or bone marrow transplantation. The need for reproducible, animal pr oduct-free conditions for the culture of progenitors is crucial to the widespread clinical implementation of ex vivo cell therapies. In thes e studies, we explored the use of animal serum-free (ASF) medium for t he culture of isolated human bone marrow and peripheral blood CD34+ ce lls. In this ASF system, isolated CD34+ cells were cultured using a va riety of different growth factor combinations. Such ASF culture condit ions yielded equivalent to superior cell and progenitor growth when di rectly compared with cultures containing 10% fetal calf serum (FCS). I n cultures containing IL-1, IL-3, and stem cell factor, total cell num bers increased, on average, 33-fold over the first 2 weeks. On phenoty pic analysis, the ASF cultures demonstrated sustained proliferation of CD33+ myeloid cells throughout the culture period. CD34+ cell numbers increased during the first 7-10 days of culture, with a mean 3.4-fold expansion. Concomitant with the CD34+ cell expansion was an average 8 .2-fold expansion of colony-forming unit-granulocyte-macrophage (CFU-G M) and a 102.0-fold increase in burst-forming units-erythrocytes (BFU- E). Likewise, a mean 4929-fold expansion of CD41a+ megakaryocyte proge nitors was observed in these CD34+ cultures. Different combinations of growth factors affected the fold increase in cell and progenitor numb er. When CD34+ cell cultures from normal healthy volunteers mobilized with either G-CSF or GMCSF were compared, similar expansions of total cell and progenitor cells resulted. However, CD41+ cell expansions wer e greater in those samples from G-CSF-mobilized volunteers in every ca se tested. These studies establish the feasibility of this ASF CD34+ c ell culture system to generate a population of maturing progenitors fo r potential use in transfusion support during cytopenic periods follow ing high-dose chemotherapy or bone marrow transplantation.