Osteoblast progenitors engraft from whole bone marrow transplants in nonablated mice

Allogeneic, and even more often autologous bone marrow transplants are routinely being done to correct a wide variety of diseases. In addition, autologous marrow transplants poientially provide an opportune means of delivering genes in transfected, engrafting cells. However, despite the wide spread clinical use and promising gene therapy applications of bone marrow transplants, relatively little is known about the mechanisms of engraftmenl in marrow transplant recipients. This is especially so in the nonublated recipient setting, and with regards to cells not of hemopoielic progenitor origin, in particular this includes stromal cells, and cells of the osteoblastic lineage. We have demonstrated for the first time, that a whole bone marrow transplantation contains bone progenitor cells which engraft, and proliferate and differentiate to form mature cells capable of forming bone. Using a male into non-ablated female transplant model, cells of donor origin were evident ex vivo by fluorescence in situ hybridization, as osteocytes encapsulated within the bone lacuna. In addition, donor cells were evident as flattened lining cells on the periostea! bone surface and as cells within the endosteal growth region. Very few transplanted bone cells were detected 3 weeks post-transplant, but these cells were easily detected both 6 weeks and 6 months post transplant. A technical limitation of the in situ hybridization procedure is the removal of the actual bone matrix during the denaturation process. To confirm that the cells analyzed were both of donor and bone origin, we cut serial, longitudinal femoral sections and stained every second section by in situ hybridization, and every alternate section with haemotoxylin and eosin. Individual cells could then be identified in the bone lacuna or periosteum prior to the procedure, and where the bone matrix used to he following the in situ hybridization. Our data allows circumventing of osteoblast stem and progenitor cell isolation and purification, as well as irradiation of recipients, demonstrating that a normal bone marrow transplant into a nonablated recipient contains bone cell progenitors which are capable of bone formation. The data points the way to the treatment of many bone disorders by bone marrow transplantation, and to bone directed gene therapy.