One hypothesis is that human immunodeficiency virus (HIV, HTLV-III/LAV, ARV) acts persistently in brain. There is immunologic neuropathologic, epidemiologic, and molecular evidence supporting this, especially that HIV nucleic acids are present in AIDS CNS tissue, in neurons, endothelial cells, and macrophages. In addition, HIV antibodies are found in CSF in AIDS patients. The purpose of this grant application is to characterize the replication of HIV in brain and compare it to lymphocyte HIV isolated from the same and other patients. Fresh brain tissue will be obtained from patients who died from AIDS and will be subject to two modes of analysis. One part of the tissue will be placed into explant cell culture using techniques developed in this laboratory and another part will be fixed for histology, electron microscopy, and in situ hybridization. HIV riboprobes will be used for in situ hybridization and hybridized cells will be identified using histologic and immunocytochemical techniques. Virus isolation will be attempted from the brain explant cultures up to 30 days. In addition, cells from the brain explants will also be subject to in situ hybridization as above and the cells will be identified using histology, immunocytochemistry, and confirmed by electron microscopy. Recently, this laboratory obtained evidence that HIV can infect and replicate in human neuroblastoma cells, but not in human astrocytoma and Hela cells nor in monkey Vero cells. Comparisons of radioactively labeled viral proteins in brain, lymphocytes, and neuroblastoma cells will be made using immune precipitation followed by one-dimensional (1D) and two dimensional SDS-PAGE, 1D oligopeptide mapping and fluorography. The brain explant culture experiments will provide information as to whether HIV in brain acts persistently or acutely (lytically) compared to culture in lymphocyte and neuroblastoma cells. Furthermore, it will be possible to determine whether neurotropic strains or mutants of HIV exist.
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