Arthritis is the most prevalent cause of chronic morbidity among those over 65 years of age, with over 30 million Americans suffering from the various arthritedes. The actual cost of treating these orthopaedic conditions represents billions of dollars in health care costs. The cost in human suffering is impossible to quantify. Despite tremendous research efforts, our understanding of the etiology and pathogenesis of these diseases is still incomplete. This proposal is designed to utilize gene therapy to systematically study molecular questions about arthritis, as well as test new treatments for this disabling disease. Specifically, this project focuses on a novel anti- TNFalpha gene therapy protocol to treat the polyarthritis of the hTNFalpha-transgenic mouse. A current paradigm is that TNFalpha is at the apex of the pro- inflammatory cytokine cascade which is responsible for joint destruction in arthritis. At the cellular level, the transcription factor NFkappaB is of paramount importance because it is both directly involved in the transcription of many inflammatory cytokines genes like TNFalpha and is activated by cell stimulation with these proteins. In turn NFkappaB amplifies these stimuli by initiating the transcription of a large number of genes, including matrix metaloproteases (MMPs), which cause the pathology. Our hypothesis is that the TNFalpha/NFkappaB signal transduction pathway is central to the genesis of arthritis and by using a specific inhibitor of this pathway we will prevent the disease process. In our studies of NFkappaB signal transduction, we have generated a dominant-negative IkappaBa (mIkappaB) which represents the specific inhibitor we desire. When over-expressed in mammalian cells, the mIkappaB renders them void of NFkappaB activity. Here we propose the following specific aims: i) identify additional cytokines, adhesion molecules and MMPs which promote disease in the hTNFalpha-transgenic mouse, and determine their cellular source, ii) determine the efficiency of our recombinant adenovirus, adenoassociated virus (AAV) and lentivirus (HIV) vector systems to transduce the cells of the joint in vitro and in vivo; and to modify these vectors with specific promoters to achieve the desired tissue expression, iii) test the ability of our mIkappaB to eliminate NFkappaB gene expression and monitor this effect on anabolic and catabolic mechanisms relevant to joint homeostasis, and iv) evaluate the usefulness of our mIkappaB to treat the arthritis of the hTNFalpha-transgenic mouse by in vivo gene therapy.
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