MULTILINEAL HEMATOPOIESIS IN A 3-DIMENSIONAL MURINE LONG-TERM BONE-MARROW CULTURE

The highly packed cell density and the three-dimensional structure in the hematopoietic compartment of bone marrow facilitate cell-to-cell a nd cell-to-matrix interactions known to be important for hematopoietic activities. To provide a similar;environment in vitro, we developed a long-term bone marrow culture (LTBMC) system, continuously perfused w ith Dexter's medium, employing packed, highly porous bovine collagen m icrospheres as the matrix support for marrow cell growth. Using murine bone marrow as a model, we found that the culture system differed fro m the conventional flask culture in the following ways: 1) as revealed by the electron microscopy, the bone marrow cells in the culture syst em grew in a three-dimensional configuration, similar to that in vivo, 2) the cell output from the culture system at 37 degrees C was virtua lly the same as that at 33 degrees C, and 3) in the absence of exogeno us growth factors, except those in the serum, the culture system produ ced lymphoid cells and all stages of committed cells (i.e., erythrocyt es, granulocytes, macrophages, and megakaryocytes), thus indicating mu ltilineal differentiation of the hematopoietic stem cells. Furthermore , cell clusters resembling erythroblastic islands were observed in the absence of exogenous erythropoietin (Epo). The culture system appears to provide a different microenvironment than that of the flask cultur e and may be used as an alternative model for hematopoiesis.