Neural cell targets of HIV-1 infection in human fetal organotypic cultures

Abstract

Children infected by HIV-1 can demonstrate nervous system dysfunction. Because some HIV-1 positive children are believed to be infected during gestation and it is thought that HIV-1 may infect neural cells, the studies presented in this dissertation tested the hypothesis that HIV-1 infects cells of the developing human fetal central nervous system (CNS) and, as a result, this infection contributes to the neuropathology associated with the acquired immunodeficiency syndrome (AIDS).;CNS organotypic tissue cultures (OTC) derived from human fetal brain enable the study of complex interactions between neural cells. Many such studies are not possible using dissociated tissue culture models. Immunocytochemical (ICC) and Western blot analyses have identified neurons, astrocytes, microglia and oligodendrocytes in these OTC. To test the hypothesis stated above, CNS OTC were exposed to either lymphocytotropic (L-tropic) or monocytotropic (M-tropic) HIV-1 isolates and monitored for viral production and pathologic changes.;CNS OTC exposed for up to 10 days to high titers of distinct cell-free HIV-1 L- & M-tropic isolates displayed some cytopathologic changes consistent with CNS AIDS at the light microscopic level. Additionally, HIV-1 gp41 and p24 antigens were detected by ICC in cells in virus-exposed OTC. Additionally, HIV-1 RNA was detected in the cytoplasm of CNS cells by in situ hybridization (ISH) and viral DNA was detected by PCR in HIV-1 exposed cultures. Double-label ICC detected HIV-1 antigens in both microglia and astrocytes. To determine if HIV-1 infection of OTC is productive, culture supernatants were monitored after HIV-1 exposure for reverse transcriptase activity (RT), p24 antigen concentration and the ability to infect susceptible cells. While there were no significant increases in RT activity, p24 antigen levels increased and HIV-1 was cultured from culture supernatants. Viral titration and time course experiments determined the minimal infectious dose to infect OTC with HIV-1.;These results demonstrate that both L- and M-tropic HIV-1 isolates infect microglia and astrocytes in human fetal OTC. This model system will permit further examination of the interaction of HIV-1 with the developing human CNS and may contribute to the understanding of the mechanisms associated with AIDS neuropathology.