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Background:  Investigations characterizing central nervous system cellular reservoirs of HIV have generally been performed on tissues from subjects with AIDS or markedly decreased CD4⁺ cell counts with significant plasma viremia. The locus of HIV reservoirs in less-compromised patients has not been directly examined. In brain tissue from AIDS patients, productively infected CNS cells are predominately macrophages/microglia infected by R5 HIV. Whether other types of CNS cells, such as astrocytes, harbor latent or minimally productive virus in vivo is unclear. In vitro experiments imply that astrocytes host restricted infection by X4 HIV. If this astrocytic X4 infection occurs in humans, recrudescent astrocyte-derived virus may reseed plasma and other compartments during interruptions in antiretroviral therapy. Our preliminary study examined the extent and nature of astrocyte infection in difficult-to-acquire tissues from HIV⁺ individuals with substantially intact or reconstituted CD4⁺ cell counts and controlled peripheral viral loads.

Methods:  We investigated autopsy brain specimens from 4 HIV-infected individuals who died with absolute CD4 counts > 400 cells/mm³ and undetectable plasma viral loads. HIV RNA in situ hybridization was performed on tissues from basal ganglia and other CNS regions. Sequences of HIV DNA gag, env, or nef were PCR amplified from gross tissue extracts or on astrocytes collected with laser capture microdissection. Env V3 DNA PCR amplification products were sequenced and analyzed for co-receptor tropism.

Results:  In situ hybridization showed very sparse productive infection in all 4 brains. By PCR, gag, nef, and env were amplifiable from all cases. In laser-captured pools of 500 to 1000 astrocytes, gag DNA was detected in every brain, and env DNA was amplified from 2 of the 4. Astrocyte-derived V3-loop sequences aligned with X4 strains in the Los Alamos database.

Conclusions:  This preliminary data suggests central nervous system infection with HIV is present before development of AIDS and persists after immune reconstitution is achieved despite suppression of plasma viremia by antiretroviral drugs. X4 viral DNA is present in astrocytes; this may indicate these cells are a discrete central nervous system viral reservoir.

Keywords: reservoir; CNS; astrocyte