Development of a bone marrow culture for maintenance and growth of normal human B cell precursors

The absence of long term bone marrow cultures for studying the growth and differentiation of human B cell precursors (BCP) has placed restrictions on the ability to analyze the early stages of human B cell ontogeny. We now describe a bone marrow-derived adherent cell microenvironment that maintains human BCP for several weeks in vitro. The adherent cells are maintained in a serum-free tissue culture medium, and consist of a predominant population of CD10⁺ fibroblast-like cells and a minor population of CD10⁺/nonspecific esterase⁺ macrophages. Adherent cell cultures seeded with fresh or cryopreserved fetal bone marrow, or purified CD10⁺/ surface IgM^- cells, provide a supportive microenvironment for lymphoid cells with a predominant phenotype of CD10⁺/CD19⁺/HLA-DR⁺/surface IgM^-. Supplementation of the adherent cell cultures with human IL-7 induces active growth of BCP during the first 14 to 21 days of culture. However, the expansion of these cells does not continue past 21 days, and the cultures undergo a steady decline in BCP. Analysis of adherent cell conditioned medium revealed the presence of an unidentified soluble factor (or factors) that acts in concert with IL-7 to promote the growth of CD10⁺/surface IgM^- cells. This culture system will be useful in elucidating the patterns of gene expression and growth factor requirements that characterize normal human B cell ontogeny, and perturbations of normal B cell ontogeny that lead to immunodeficiency and leukemia.