CELL-FUNCTION STUDIES IN LONG-TERM BONE-MARROW CULTURE AFTER CELL CONCENTRATION AND CRYOPRESERVATION FOR AUTOLOGOUS TRANSPLANTATION

Autologous bone marrow transplantation (ABMT) is frequently used in th e treatment of neoplastic diseases. It involves several manipulations in vitro that can damage the stem cells responsible for grafting. Long -term bone marrow cultures (LTBMC) reproduce the bone marrow microenvi ronment and support haematopoiesis in vitro for several weeks. This te chnique provides information on the extent of the injury of stem cells during manipulations of bone marrow cells in vitro. We studied the ef fect of the usual process of bone marrow cells in vitro in autologous transplantation: cell concentration and cell cryopreservation. 28 bone marrows from healthy donors and 45 from patients who had undergone AB MT were assayed in LTBMC. The cultures were initiated after manual or automatic buffy coat cell separation or after density gradient or auto matic mononucleated cells suspension. Bone marrows were studied pre- a nd post-cryopreservation. The results show that several manipulations can disturb normal cell behaviour in LTBMC. Following automatic mononu cleated cell separation and after thawing, the development of adherent cell layer in LTBMC is anomalous. These manipulations led to a delay in covering 50% of the flask surface, absence of adypocytes in adheren t cell layer and absence of haemopoietic precursors in the supernatant at the 4(th) week of culture. The results suggest that LTBMC can be u sed for control of the manipulations in vitro related with ABMT.