TRANSIENT AND PERMANENT ENGRAFTMENT POTENTIAL OF MURINE HEMATOPOIETICSTEM-CELL SUBSETS - DIFFERENTIAL-EFFECTS OF HOST CONDITIONING WITH GAMMA-RADIATION AND CYTOTOXIC DRUGS

Transplant of sorted donor (BG-Gpi-1(a)) hematopoietic stem cell subse ts and host (BG-Gpi-1(b)) treatment with total body irradiation (TBI) or cytotoxic drugs were compared for induction of short- and long-term engraftment in a murine chimera model of congenic bone marrow transpl antation (BMT). Parallel studies on donor and host marrow were perform ed in vitro in long-term bone marrow cultures to determine early and l ate cobblestone area forming cell (CAFC) frequencies in the grafts or in the transplant recipients 1 day after conditioning. Bone marrow cel ls (BMC) sorted for high wheat germ agglutinin affinity (WGA(+++)) wer e enriched about 30-fold for early developing CAFC (colony-forming uni t-spleen [CFU-S]) but not for primitive late CAFC (pre-CFU-S). This fr action showed only temporary engraftment when transplanted in irradiat ed recipients. In contrast, the low affinity (WGA(+)) fraction were pr eferentially enriched (200- to 300-fold) for late developing CAFC and were very effective for providing stable long-term engraftment followi ng transplantation. Substituting radiation for chemotherapy in the hos t conditioning treatment also had diverse effects on the development o f bone marrow engraftment. Pretreatment with 5-fluorouracil (5-FU, 200 mg/kg) allowed a discrete wave of donor engraftment that peaked at 2 to 4 weeks and then subsided to leave mostly host cells at 10 weeks an d beyond. Busulfan preparation gave over 50% engraftment at 1 month af ter BMT but this continued to increase to reach stable donor chimerism of 80 to 90% beyond 10 weeks. The level of long-term engraftment give n by 50 mg/kg busulfan appeared similar to that induced by doses of 6 to 8 Gy TBI. The changing patterns of erythroid chimerism for each pre parative agent showed a remarkable correlation with depletion of defin ed hematopoietic stem cell subsets as quantified using the CAFC assay at 1 day after recipient treatment. These findings collectively show t hat the level of depletion of host CFU-S (CAFC-10) determines the exte nt of early and transient repopulation, whereas the degree of pre-CFU- S (CAFC-35) depletion determines the percentage of stable chimerism. P reliminary data on bone marrow CAFC content at 9 months after busulfan and BMT revealed a long-term reduction of the stem cell pool by about 50% but with relatively minor effects on supporting bone marrow strom as. The differential cytotoxic effects on stem cell subsets in relatio n to subsequent donor marrow engraftment offer a systematic and mechan istic approach toward identifying more effective chemotherapeutic comp ounds for use in clinical BMT conditioning regimens.