Detection of HIV-1 Gene Sequences in Hippocampal Neurons Isolated from postmortem AIDS Brains by Laser Capture Microdissection

Abstract

We employed laser capture microdissection to remove individual pyramidal neurons from the CA1, CA3, and CA4 regions of formalin-fixed, paraffin-embedded hippocampus from 8 AIDS brains and 2 HIV-1-seronegative normal brains. We amplified HIV-1 gag and nef gene sequences using separate, double round PCR reactions for each of the primer sets. In all 3 hippocampal regions, amplification efficiency was best with sequence length between 284 and 324 bp; HIV-1 nef gene sequences were more common than HIV-1 gag sequences; and rank order for percent positive amplification was CA3 > CA4 > CA1 samples. These results are the first to detect HIV-1 gene sequences in microdissected human tissue. They indicate that brain neurons in vivo contain HIV-1 DNA sequences consistent with latent infection by this virus, and suggest that neurons display a selective vulnerability for HIV infection. Neuronal HIV infection could contribute to neuronal injury and death or act as a potential viral reservoir if reactivated.

Differential long‐term neurotoxicity of HIV‐1 proteins in the rat hippocampal formation: A design‐based stereological study

“…Both animal and tissue culture studies have shown that gp120 produces cell death in rodent hippocampal neurons (Brenneman et al, 1988;Lipton et al, 1995). A human study assessing the CA1, CA3, and CA4 regions of the hippocampus from eight AIDS-infected brains further supports the finding that specific regions of the hippocampus are particularly vulnerable to the virus or its components (Torres-Munoz et al, 2001). The selective vulnerability of hippocampal regions for HIV-1 infection is demonstrated by the expression of HIV-1 chemokine co-receptors on hippocampal pyramidal neurons.…”

“…Results revealed evidence of direct HIV-1 infection of neurons as one potential mechanism for neuronal injury or neuronal death. In contrast to studies demonstrating the presence of HIV-1 in neurons (Bagasra et al, 1996;Torres-Munoz et al, 2001;Trillo-Pazos et al, 2003) other studies report the lack of neurons infected by HIV-1 (Takahashi et al, 1996;Thompson et al, 2004). A study which reported HIV-1 infection of macrophages and microglia in addition to a more latent infection of astrocytes, could not detect HIV-1 infected neurons and/or oligodendrocytes (Takahashi et al, 1996).…”

“…A study assessing the hippocampus in brains from six adult and two pediatric AIDS patients demonstrated the latent infection of neurons by the virus in vivo (Torres-Munoz et al, 2001). Brain neurons containing HIV-1 DNA sequences were found in the CA1, CA3, and CA4 regions.…”

Gene expression for HIV-associated dementia and HIV encephalitis in microdissected neurons 1: preliminary analysis

“…31 Laser capture microdissection has been used successfully in the study of several human brain diseases, including the HIV-1-infected brain, subacute sclerosing panencephalitis, Parkinson's disease, and Huntington's disease. [32][33][34][35][36][37][38][39] In the current study, only cases with HAD and HIVE (as well as HIV-1-positive controls) were used. Thus, work in NeuroAIDS has progressed to the point where cell-specific studies will be able to elucidate additional information using novel approaches.…”

Combining laser capture microdissection with quantitative real-time PCR: Effects of tissue manipulation on RNA quality and gene expression

“…More recent investigations have focused on applying LCM to the study of the central nervous system. These have included genome-wide gene expression analyses using DNA microarrays, as well as expression analyses of individual genes using reverse-transcriptase polymerase chain reaction (RT-PCR) (Jin et al, 1999;Kamme et al, 2003;Lu et al, 2004;Mutsuga et al, 2004;Torres-Munoz et al, 2001;Ye et al, 2003).…”