Factors controlling haemopoiesis in ovine long term bone marrow cultures

We describe an adaptation of the Dexter technique for obtaining ovine long term bone marrow cultures able to sustain haemopoiesis in vitro for long periods. Two inocula of bone marrow cells collected at three-five weeks interval, in IMDM supplemented with fetal calf serum (10%), horse serum (10%) and hydrocortisone (5 × 10−7 M), gave rise to the growth of an adherent cell layer which supported, in most cases, active haemopoiesis for up to 15 weeks. The cell layer was composed of macrophages, fibroblasts and adipocytes. Haemopoietic cells formed large foci of “cobblestone” areas. Haemopoietic progenitors were also released into the supernatant medium and were detectable by clonogenic assay. Granulocytes and monocyte-macrophages differentiated in the cultures in constant proportion until week five, when the monocytic lineage superseded the myelocytic one. These cultures, between weeks five and twelve, produced colony forming cells in a constant pattern, indicating the presence and self renewing of early haemopoietic progenitor cells.