Tumor necrosis factor-alpha (TNF) is a major contributor to skeletal pathophysiology in osteoporosis and inflammatory disorders. Although a great deal of research has been focused on TNF stimulation of bone resorption, there is compelling evidence that TNF also inhibits bone formation in osteoporosis and other TNF excess states (arthritis). Identification of the mechanism through which TNF blocks the differentiation of osteoblasts will provide targets for treatments that stimulate the formation of bone in addition to the current approach of inhibiting resorption. Osteoblast differentiation is under the regulation of many factors that influence the proliferation of pluripotent precursor cells, their phenotype selection, and finally, acquisition of the functional osteoblast phenotype. The death of osteoblasts by apoptosis is also under regulation. Sound preliminary data will be presented to support the hypothesis that TNF inhibits osteoblastogenesis at the stage of phenotype selection through inhibition of the transcription factor Cbfa-l. The experimental aims will seek to prove this hypothesis and to identify the cellular pathway of TNF action. To achieve these aims, the Principal Investigator will use novel methods of recombinant retroviral expression and antisense technology to introduce a blockade of several TNF pathways including NF-kB, AP-1 nuclear protein activation, and the apoptotic death pathway. The use of available mice harboring null mutations for elements of these TNF signal pathways will also aid in testing our hypothesis. Similarly, the Principal Investigator expects retro virally forced expression of Cbfa-1 to overcome the inhibitory effect of TNF. The viral vectors constructed for this project could have future utility as gene therapy agents in vivo.
