Transplantable marrow osteoprogenitors engraft in discrete saturable sites in the marrow microenvironment

Abstract

Objective-Based on the recognition that marrow contains progenitors for bone as well as blood, we undertook the first trial of bone marrow transplantation (BMT) for a genetic disorder of bone, osteogenesis imperfecta (OI). While we documented striking clinical benefit soon after transplantation, the measured level of osteopoietic engraftment was low. To improve the efficacy of BMT for bone disorders, we sought to gain insight into the cellular mechanism of engraftment of transplantable marrow osteoprogenitors.Methods-We transplanted unfractionated bone marrow harvested from green fluorescent protein (GFP)-transgenic FVB/N mice into lethally irradiated FVB/N recipients. At 3 weeks posttransplantation, we assessed hematopoietic engraftment by flow cytometry and osteopoietic engraftment by immunohistochemical staining for the GFP protein.Results-We show that the engraftment of transplantable marrow osteoprogenitors is saturable with a maximal engraftment of about 15% of all bone cells in the epiphysis and metaphysis of the femur at 3 weeks after transplantation. The number of engrafting sites is not up-or downregulated in response to initial progenitor cell engraftment and there is no evidence for clonal succession of osteopoietic differentiation of engrafted progenitors.Conclusions-Our findings indicate that the capacity for initial osteopoietic engraftment after BMT is limited and "megadose" stem cell transplantation is unlikely to enhance engraftment. Thus, novel strategies to foster osteopoietic chimerism must be developed.

Transplanted bone marrow mononuclear cells and MSCs impart clinical benefit to children with osteogenesis imperfecta through different mechanisms

“…Surprisingly, the only factor that could be related to a positive response to NABMC transplantation was mixed hematopoietic chimerism as the 2 nonresponders were complete (100%) and the 3 responders were mixed (Ͻ 90%) donor chimeras (supplemental Figure 4). Because engraftment of transplantable osteoprogenitors is saturable, 16 we reasoned that this result might reflect the extent of occupancy of stem cell niches by donor osteoprogenitors.…”

“…The osteoblasts were identified as the large, cuboidal cells with eccentrically placed nuclei along the endosteal surface as previously described. 16,17 GFP ϩ osteoblasts were enumerated by visual examination of stained slides by two investigators, who were blinded to the experimental conditions. Osteopoietic cell engraftment (chimerism) is the percentage of observed bone cells that express GFP (the marker of donor origin).…”

“…Surprisingly, the only factor that could be related to a positive response to NABMC transplantation was mixed hematopoietic chimerism as the 2 nonresponders were complete (100%) and the 3 responders were mixed (Ͻ 90%) donor chimeras (supplemental Figure 4). Because engraftment of transplantable osteoprogenitors is saturable, 16 we reasoned that this result might reflect the extent of occupancy of stem cell niches by donor osteoprogenitors.To test this notion, we transplanted 2 groups (n ϭ 10 each) of lethally irradiated FVB/N mice (H-2k q ) with a CD3 ϩ cell-depleted bone marrow graft consisting of 2 ϫ 10 5 cells from a GFPtransgenic C57BL/6 donor (H2-k b ) and 1.8 ϫ 10 6 cells from either an FVB/N donor or a C57BL/6 donor. The resultant 2 cohorts represent either 10% donor chimeras or complete donor chimeras.…”

“…Whole bone marrow, NABMCs, or MSCs were transplanted into irradiated C57BL/6 with several dosages or into oim/oim mice (B6 background) as previously described. 16 All animal protocols were approved by the Institutional Animal Care and Use Committees of either St Jude Children's Research Hospital or The Children's Hospital of Philadelphia. …”

“…16 All animal protocols were approved by the Institutional Animal Care and Use Committees of either St Jude Children's Research Hospital or The Children's Hospital of Philadelphia.…”