Description (Taken from the application): Somatic gene therapy may have the potential for treating diseases of bone that are resistant to current therapies. However, the cellular and molecular steps required to develop an integrated effective strategy are not fully developed. During the previous grant cycle, some of the problems were successfully resolved, including the ability to deliver a cell-specific expressed promoter-reporter construct within the context of a retrovector and the production of a pseudotyped retrovector capable of infecting a high proportion of primary marrow stromal cells without antibiotic selection. New models for assessing the effectiveness of bone marrow transplantation were developed using Collal-GFP transgenes whose expression is dependent on the stage of osteoblast differentiation. This proposal utilizes these tools to develop engineered marrow stromal cells that are enabled to engraft to bone, participate in bone turnover and produce a growth factor that will influence endogenous bone cells. We will confirm that viral transduced stromal cells can produce bone in vivo and we will develop a cell-specific promoter that can be activated in a controlled manner. The differentiation of stromal cells will be blocked with growth factors that expand the proportion of immature cells within the culture. The ability of these cells to participate in bone formation will be assessed by direct injection into the diaphyseal space of the femur and by tail vein injection into a murine model undergoing recovery from conditional ablation of the osteoblast lineage. The degree of engraftment, the ability to participate in new bone formation and the contribution to the self-renewing pool of osteoprogenitor cells will be determined based on the expression of GFP lineage markers of osteoblast differentiation of the donor cells. An additional aim will develop the steps required to make the host more receptive to engraftment. By analyzing each component of the strategy, it should be possible to develop a method for somatic gene therapy of bone in the mouse that may be applicable to man.