982. Adaptive Immune System Down-Regulates Transgene Expression from Adenovirally Infected Neurons in the Brain In Vivo
Authors: Jeffrey M. Zirger, Carlos Barcia, Josee Bergeron, Chunyan Lui, Maria G. Castro, Pedro R. Lowenstein
Journal: Molecular Therapy (2006)

Studies on mechanisms by which the immune system eliminates virally infected cells from the brain have been limited by the difficulty in differentiating damage induced by a replicating virus from that induced by the immune system. To understand the cellular and molecular mechanisms by which the immune system can control virus gene expression in neurons we utilized a replication incompetent virus and a marker transgene expressed exclusively in neurons as a surrogate for expression of genes from the viral genome. Upon systemic immunization against the virus, we demonstrate that CD4+ and CD8+ T cells are necessary to eliminate expression of the marker gene expression from the CNS through a mechanism dependent upon IFNγ, perforin and tumor necrosis factor γ; almost complete loss of expression ensues at 30 days post-immunization and does not recover up to a year later. To differentiate whether the immune system eliminates transgene expression through killing of transduced cells, or functional inhibition of expression from the viral genome, we performed detailed anatomical analysis, studies in mice deficient in important immunological mediators, studied simultaneously the expression of marker genes from the viral genome and the cellular genome of infected cells, and performed adoptive transfer and immune-suppression experiments. Detailed confocal microscopical analysis discovered that CD8 and CD4 T cells infiltrated specifically infected regions of the brain, established close anatomical contacts with infected neurons, and could be detected within the infected brains for up to one year after the systemic immunization. In addition, we detected a significant infiltration of macrophages, which at later stages could be seen phagocytosing transduced cells, illustrating terminal stages of cell death of adenovirally transduced brain cells. Using ROSA26 mice and viruses expressing Cre, we demonstrated that the immune system abolished expression from both the viral genome and the cellular genome, supporting the idea that at least some loss of marker gene expression is due to target cell death. Surprisingly however, while adoptive transfer of CD4+ and CD8+ T cells induced a transient loss of viral genome expression, immune suppression of animals more than two months post-immunization only induced a partial recovery. Thus, we demonstrate that CD4+ and CD8+ T cells eliminate viral gene expression from the brain through mechanisms that require constant surveillance of the CNS, and are mediated in part through the elimination of infected cells and partly, through the functional inhibition of gene expression. These data have important implications for our understanding of mechanisms by which the immune system clears virally infected cells from the brain, and consequently the safety of long term gene therapy strategies for the treatment of brain diseases. Finally, our data also appear to throw light on mechanisms by which the immune system controls 'slow replicating' viruses such as the JC virus, or HSV-1, which remain latent in the human nervous system for long periods of time, but reactivate upon immune-suppression.