Enterovirus infection has been linked to the expression of several human autoimmune diseases including polymyositis, juvenile-onset diabetes, and cardiomyopathy. Evidence of prior enterovirus infection includes elevated antibody titers against virus and the detection of persistent viral RNA in the afflicted tissue. However, results from some studies are equivocal, and hence the role of enterovirus in precipitating autoimmune diseases is still unclear. Techniques for the detection of persistent enteroviruses are hampered by a lack of knowledge about the mechanism which underlies persistence. Similarly, the mariner in which persistent enterovirus provokes the induction or maintenance of immunologically-mediated tissue damage is not understood. This proposal employs a mouse model of coxsackievirus B 1-induced polymyositis to study mechanisms of virus persistence and evaluate the effect of virus persistence on immunopathic muscle disease. Two key attributes of this model are that viral RNA persists for long periods of time in muscle, and immunopathic disease is mediated by T cells. The first objective is to explore the nature of persistent virus and determine whether it is comprised of altered forms of viral RNA and/or infectious virus. Nuclease protection assays will be used to identify genomic deletions, sequence changes, and altered ratios of positive and negative RNA strands. Reactivation of infectious virus will be attempted with several different strategies including immunosuppression and muscle denervation. A somewhat unique strategy for virus reactivation will be to culture muscle in vitro where it is removed from potential anti-viral effects of the immune system. Tissues other than muscle that have previously been found to harbor persistent viral RNA will also be tested as potential reservoirs of virus. If infectious virus is recovered, it will be phenotypically and genotypically characterized and compared with the original infecting strain. The second objective seeks to better understand factors that influence the coevolution of host and virus, and focuses on selective pressures exerted by the host on development of virus persistence. Nude mice and inbred strains of mice with varying susceptibilities to myositis will be used to examine the influence of T cells and host background genes on the establishment of persistent infection. The third objective is to identify muscle genes that are differentially regulated as a result of virus persistence. The differential display technique will be used to analyze muscle for RNA transcripts that are upregulated or downregulated at different stages of the disease. Taken together, these experiments will clarify important interactions between persistent virus and the host that culminate in the development of immunopathic muscle disease. They will also lead to more effective approaches for the epidemiologic study of persistent virus- induced autoimmune disease.
