A novel stromal cell-dependent B lymphoid stem-like cell line that induces immunoglobulin gene rearrangement

A stroma-dependent B lymphoid cell line (B31-1) has been established by coculturing sorted stem cells on a novel bone marrow stromal cell line (TBR31-1). B31-1 cells express B220, but do not express other B lymphoid differentiation markers including CD43, heat stable antigen (HSA), or surface immunoglobulin (Ig)M (sIgM), and their Ig heavy chain (IgH) gene loci are germ-line in configuration. The addition of interleukin (IL)-7 or coculture with another stromal cell line, ST2, induces D-J rearrangement of the IgH gene and B lymphocyte differentiation markers. B31-1 cells restore an in vivo repopulation activity to lethally irradiated mice, and the repopulated cells differentiate to HSA⁺ pre-B cells. Continuous coculture results in two distinct populations, B220^- c-Kit⁺ cells and B220⁺ c-Kit⁺ cells; B220^- c-Kit⁺ cells are self-renewed and differentiate to B220⁺ c-Kit⁺ cells, while B220⁺ c-Kit⁺ cells produce only B220⁺ c-Kit⁺ cells. Both B220^- and B220⁺ cells similarly express the IgH germ-line transcript (Iμ), mRNAs for recombinase (TdT, Rag-1, and Rag-2), and lymphoid-specific transcription factors (Pax-5, EBF, E12/E47, Oct-2, and Ikaros), but the DNA binding activity of Pax-5, EBF, Oct-2, and E2A are low in B220^- cells and while high in B220⁺ cells. These results suggest the existence of at least two active states in the IgH locus before the induction of IgH gene rearrangement during B lymphopoietic development.