Cytomegalovirus is a significant opportunistic pathogen in AIDS patients as well as the most common infectious cause of congenital brain disorders. We have recently developed a model of murine cytomegalovirus (MCMV) encephalitis in which immunocompetent and immunodeficient mice are infected through the intracerebroventricular (icv) route. Analogous t o t he situation in humans, immunocompetent m ice a re not susceptible t o M CMV encephalitis, w whereas mice with a severe immune deficit develop uncontrolled brain infection similar to that seen in patients with advanced AIDS. Adoptive transfer of splenocytes from MCMV-primed mice protects immunodeficient animals and this protection is associated with elevated brain levels of several lymphocyte chemoattractants. In this proposal, the central hypothesis to be tested is that glial cell-produced CXCR3 ligands recruit lymphocytes that control the intracerebral spread of MCMV via the production of antiviral cytokines. To test this hypothesis, we will individually deplete splenic lymphocyte subpopulations (i.e., T lymphocytes [CD4+ and CD8+], NK cells, and B-cells) prior to adoptive transfer into immunodeficient mice and determine the effect on viral spread following subsequent icv infection. We will then examine how disruption of chemokine networks by administration of neutralizing antibodies to each of the CXCR3 ligands affects viral expression and lymphocyte trafficking. Additionally, we will examine the role of CXCR3 ligands in trafficking of lymphocytes into infected brains following adoptive transfer of splenocytes from CXCR3 knockout mice. The effect of lymphocytes on chemokine production by glial cells will be investigated by determining if adoptive transfer amplifies CXCR3 ligand levels and identifying which glial cell types produce these ligands in response to MCMV infection. Transfer of lymphocytes from interleukin-10 knockout mice will be performed to determine if this cytokine is used to dampen glial cell chemokine production in response to infection. The role of antiviral cytokines in inhibiting viral replication in the brain will be addressed by examining differences in tumor necrosis factor (TNF)-alpha and interferon (IFN)-gamma, levels in the brains of infected mice with and without adoptively transferred splenocytes. Finally, adoptive transfer of splenic lymphocyte subpopulations, from TNF-alpha and IFN-gamma knockout mice will be used to determine if these cytokines alter viral replication and spread within the brain. The in vivo MCMV brain-infection model presented in this grant application provides us with the ability to investigate neuropathogenesis and defense of the brain occurring during viral encephalitis. The studies proposed in this competitive renewal application will provide new insights into the role of CXCR3 ligand production by activated glial cells in the control of viral replication, chemokine-mediated trafficking of lymphocytes into the brain, and the generation of chemokine-induced cytokine networks. ? ?
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