SPACE PROVIDED. The role of the measles virus nucleocapsid gene (MVNP) and p62p392L in PD are just beginning to be clarified. However, how they may interact to cause PD and the cellular and molecular mechanisms involved are still unclear. It is our hypothesis that both an environmental factor (MVNP) and a genetic factor (p62P392L) contribute to the development of PD, and that many of the effects of MVNP are mediated through its induction of high levels of IL-6 in cells of the OCL lineage. Project 2 will examine the cellular and in vivo effects of co-expression of MVNP and p62p392L, and the mechanisms involved in the development of PD. Studies in Project 1, which are closely linked to Project 2, will examine the molecular mechanisms involved n the effects of MVNP and p62PS92L that result in formation of PD OCL and increased IL-6 production. To test this hypothesis, we will: 1. Determine if co-expression of MVNP and p62P394L induce Paget's disease in mice. We will mate mice in which the MVNP gene is targeted to the OCL lineage (TRAP-MVNP mice) with p62P394L Kl (p62KI) mice. We will determine if OCLs with the characteristics of PD OCLs form in vitro, and assess the bone phenotype of these mice in terms of OCL and osteoblast activity, OCL morphology, histomorphometry and the development and extent of pagetic bone lesions. The molecular basis for the changes in OCL and osteoblast activity that occur upon co-expression of MVNP and p62p394L in mice will be characterized in Projects 1 and 3 respectively. 2. Determine if increased IL-6 expression induced by MVNP in OCL is responsible for the development of pagetic OCLs and bone lesions in TRAP-MVNP mice. TRAP- MVNP will be mated with IL-6'A mice to generate TRAP-MVNP/IL-B""""""""''mice. The OCL and osteoblast phenotype of these mice, as well as their capacity to develop focal bone lesions characteristic of PD, will be compared to WT, TRAP-MVNP and IL-6J"""""""" mice. As part of these experiments, we will determine if blocking IL-6 activity with a neutralizing antibody to IL-6 inhibits the development of pagetic OCLs in marrow cultures from patients with PD. 3. Test the hypothesis that increased expression of IL-6 and the presence of the p62P394L mutation are sufficient to induce pagetic bone lesions and OCL in mice. We have previously shown that the p62KI mice do not develop pagetic bone lesions, and that OCL from these mice are not hyper- multinucleated or hyper-responsive to 1,25-(OH)2D3, and do not produce elevated levels of IL-6, features characteristic of PD OCLs. Therefore, we will generate TRAP-IL-6 mice in which expression of IL-6 is targeted to cells of the OCL lineage, and interbreed them to the p62KI mice. The TRAP-IL-6/p62KI mice wil be characterized with regard to their bone phenotype and OCL and osteoblast activity in vitro and vivo, and the effects of increased IL-6 expression and p62P392L mutation on NFicB and p38 MAPK signaling in OCLs and stromal cells from these mice. As part of these experiments, we will treat normal human OCL precursors transfected with p62P392L with varying concentrations of IL-6 to determine if they form OCLs that express a complete pagetic phenotype.
