Various techniques are available for distinguishing donor from host cells evaluating the efficacy of conditioning regimen for experimental bone marrow transplantation (BMT). Techniques include the use of extracellular immunological markers, such as Ly5 (CD45), and intracellular biochemical markers, such as glucose-phosphate-isomerase (Gpi). Because Ly5 is an extracellular protein, the disparity between donor (Ly5.1) and host (Ly5.2) antigens may induce a weak immune response whereas with Gpi, no immune response is expected. This difference may be of particular concern in experimental transplantation approaches that use minimal conditioning such as low-dose total body irradiation (TBI). Such mild conditioning may not induce the immunosuppression required to overcome host rejection of Ly5 disparate cells.To ). However, when higher TBI or BMC doses were used, similar engraftment levels were found, suggesting sufficient immune suppression to allow equal engraftment of both sources of BM.These data suggest that even a minor phenotypic disparity between donor and host, such as Ly5, may necessitate high-dose TBI to prevent rejection. The combination of low-dose TBI or other nonmyeloablative conditioning strategies with small numbers of BMC may lead to reduced engraftment when extracellular immunological markers such as Ly5 are used for transplantation studies. Therefore, small immunological differences must be considered when using the Ly5 marker for engraftment.

Despite the absence of differences in peripheral blood cell counts or bone marrow cellularity 14 days after each dose, progenitor cell levels (HPP-CFC, GM-CFC, and CAFC-7) were increased up to 2.5-fold with cytotoxic agent and G-CSF administration compared with cytotoxic agent administration alone. Mice given G-CSF for eight days had the greatest number of progenitors suggesting a dose-response relationship for G-CSF administration. G-CSF resulted in a decrease in hematopoietic stem cell (CAFC-28) content when measured two weeks after each cycle of saline, CY, and BCNU. Twenty weeks after six cycles of BCNU, the reduction in stem cell levels persisted and was further decreased when G-CSF was added to BCNU for four or eight days.Data from this study suggest that the most likely explanation for the damaging effects of G-CSF is that G-CSF directly or indirectly induces stem cells to differentiate into more committed hematopoietic cells resulting in a loss of marrow reserve. This effect is enhanced in animals with an already compromised hematopoietic stem cell compartment as seen with repeated doses of BCNU.