Murine bone marrow transplantation: A morphologic and kinetic analysis

We studied the damage and regeneration of murine bone marrow after lethal total body irradiation (TBI) and bone marrow transplantation (BMT). Clinical, hematopoietic, and morphologic recovery were contrasted between intact and splenectomized mice, some treated with granulocyte macrophage-colony stimulating factor (GM-CSF). Alterations in the bone marrow stromal cell components were emphasized, including bone lining cells (BLC) and the barrier cell, an activated fibroblastic stromal cell. We tested the hypothesis that the barrier cell was critical in early hematopoietic recovery and found that, although the barrier cell was not routinely identified until 10 days after TBI and BMT, and the barrier cell was prominent in hematopoietic regrowth, particularly granulopoiesis and megakaryopoiesis. We tested the hypothesis that administration of GM-CSF after TBI and BMT would hasten hematopoietic recovery and result in increased numbers of barrier cells. Although hematopoietic recovery was similar, increased numbers of barrier cells were present in mice treated with GM-CSF. We tested the hypothesis that BLC were critical in bone marrow recovery. Autoradiographic data show that DNA-synthesizing BLC are relatively more abundant in the bone lining layer early after BMT, but as hematopoietic recovery progresses, DNA-synthetic cells are found principally in the marrow cavity. In fact, there are no significant differences between absolute numbers of DNA-synthetic bone lining cells at measured intervals after TBI and BMT. We tested the hypothesis that hematologic recovery would be delayed in splenectomized mice. Recovery of hematocrit concentrations and leukocyte numbers were slower but complete in splenectomized mice. We tested the hypothesis that hematopoietic recovery was secondary to reconstitution with cells of donor origin, but that stromal cell populations were solely of recipient origin. We performed sex-mismatched allogeneic BMT and detected repetitive sequences in the Y chromosome with PCR, demonstrating the presence of male DNA with PCR early after BMT. We prepared bone marrow stromal cells cultures, transfected one population of these cells with retrovirus having both the beta-galactosidase and neomycin-resistance markers and a second population of cells with the NTK-BGEO vector. Selected stromal cells were separately administered concurrently with TBI and BMT. No labeled stromal cells were identified in recipient mice