HIV-1 infection of the brain can result in HIV-1 associated dementia (HAD) and HIV encephalitis (HIVE). HIV-infected cells release viral proteins such as envelope and tat that have direct toxic effects in the brain. Indirect effects include gene expression induced in infected and uninfected cells such as production and release of toxic molecules. The host genes that produce these toxic molecules may be placed in functional groups that include NMDA receptors, chemokines, cytokines, growth factors, their receptors, apoptosis genes, nitric oxide synthases, cell surface markers, adhesion molecules, and toxic glutamic, arachidonic, and quinolinic acid pathways. We will focus on the basal ganglia since this is a region where effects of infection have been demonstrated using brain imaging, clinical, and molecular analyses. AIDS brain tissue diagnosed with and without HAD and HIVE will be provided by the National NeuroAlDS Tissue Consortium. We will micro-dissect individual macrophage-lineage cells, astrocytes, and neurons from tissue sections using LCM to attain specificity. We will use Affymetrix High Density Gene Chip HU 95A Arrays with 12,000 human genes and ESTs to identify changed expression profile of these genes related to HIVE and HAD. Prior techniques such as Northern blots or gel shift assays were more limited in their approach. Affymetrix GeneChip Analysis Suite and GeneCluster will be used for data analysis and we will assess the statistical significance of changes in gene expression observed in each cell type by comparing the results to those obtained from random permutations of the data. We will also confirm gene expression changes using Real Time RT-PCR. In situ hybridization and immunohistochemistry will be used to determine cell specificity of target gene detection to determine the pathological significance of these genes. This exploratory R21 proposal will obtain the preliminary information needed for a larger-scale R0l study to increase the number of cases, refine and increase the number of genes analyzed, and more specifically study those genes whose expression significantly changes as a result of HAD and HIVE. Gene targeting for therapeutic intervention of HAD and HIVE will be feasible.
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