LONG-TERM RECONSTITUTION OF MICE AFTER EX-VIVO EXPANSION OF BONE-MARROW CELLS - DIFFERENTIAL ACTIVITY OF CULTURED BONE-MARROW AND ENRICHED STEM-CELL POPULATIONS

In this report, we evaluated the short-term expansion of murine bone m arrow mononuclear cells (BMMNC) and enriched stem cell populations to determine the capacity of these cells for long-term rescue and engraft ment to lethally irradiated recipients. In our study, nonadherent bone marrow mononuclear cell (NBM-MNC) and Thy1(+)Lin(-) stem cell populat ions were cultured with interleukin-3 (IL-3) or IL-3 plus stem cell fa ctor (SCF) for periods up to 6 days. By day 6 of culture, the mononucl ear cells (MNC) decreased to 6% of input cell number, whereas Thy1(+)L in(-) cells increased by 2310%. Doses of 95,000; 100,000; 50,000; and 250,000 NBM-MNCs at O, 1, 2, and 6 days of culture, respectively, resc ued 50% of lethally irradiated mice. When 250,000 MNCs were cultured f or 0, 1, 2, and 6 days, 71, 61, 100, and 50% of the animals survived l ethal irradiation for greater than 24 weeks. In contrast, doses of 8,0 00 and 21,000 Thy1(+)Lin(-) cells cultured 0 and 1 day, respectively, yielded 50% survival rates. These same cells cultured for 6 days faile d to rescue recipients even at high doses. Twenty thousand Thy1(+)Lin( -) cells cultured for 0, 1, 2, and 6 days, even in the presence of SCF , produced decreasing survival rates of 86, 43, 26, and 0%, respective ly. The proliferative responses of these different populations in comb ination with their longterm rescue abilities indicated that the absolu te number of long-term rescue units (LDS,, 24 weeks) in the cultured T hy1(+)Lin(-) population decreased faster than in similarly cultured NB M-MNCs. Studies evaluating donor cell engraftment demonstrated that an imals rescued with cultured Thy1(+)Lin(-) and NBM-MNCs maintained high levels of donor reconstitution [7]. The percent donor T cell engraftm ent did not significantly change between 2 and 17 months post-bone mar row transplantation (post-BMT). Therefore, those animals who received sufficient cells to survive lethal irradiation generally established a nd maintained high levels of donor engraftment. The data suggest a rol e for accessory cells and/or factors in the preservation of stem cell activity.