SPACEPROVIDED.The role of the measles virus nucleocapsid gene (MVNP) and p62p392L in PD are just beginning to beclarified. However, how they may interact to cause PD and the cellular and molecular mechanisms involvedare 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 itsinduction of high levels of IL-6 in cells of the OCL lineage. Project 2 will examine the cellular and in vivoeffects 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 involvedn the effects of MVNP and p62PS92L that result in formation of PD OCL and increased IL-6 production. Totest this hypothesis, we will: 1. Determine if co-expression of MVNP and p62P394L induce Paget's disease inmice. We will mate mice in which the MVNP gene is targeted to the OCL lineage (TRAP-MVNP mice) withp62P394L Kl (p62KI) mice. We will determine if OCLs with the characteristics of PD OCLs form in vitro, andassess 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 thechanges in OCL and osteoblast activity that occur upon co-expression of MVNP and p62p394L in mice will becharacterized in Projects 1 and 3 respectively. 2. Determine if increased IL-6 expression induced by MVNPin 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 osteoblastphenotype of these mice, as well as their capacity to develop focal bone lesions characteristic of PD, will becompared to WT, TRAP-MVNP and IL-6J' mice. As part of these experiments, we will determine if blockingIL-6 activity with a neutralizing antibody to IL-6 inhibits the development of pagetic OCLs in marrow culturesfrom patients with PD. 3. Test the hypothesis that increased expression of IL-6 and the presence of thep62P394L mutation are sufficient to induce pagetic bone lesions and OCL in mice. We have previously shownthat 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, featurescharacteristic of PD OCLs. Therefore, we will generate TRAP-IL-6 mice in which expression of IL-6 istargeted to cells of the OCL lineage, and interbreed them to the p62KI mice. The TRAP-IL-6/p62KI mice wilbe characterized with regard to their bone phenotype and OCL and osteoblast activity in vitro and vivo, andthe effects of increased IL-6 expression and p62P392L mutation on NFicB and p38 MAPK signaling in OCLsand stromal cells from these mice. As part of these experiments, we will treat normal human OCL precursorstransfected with p62P392L with varying concentrations of IL-6 to determine if they form OCLs that express acomplete pagetic phenotype.
