TY - JOUR
T1 - Successful application of hyperbranched multidisplacement genomic amplification to detect HIV-1 sequences in single neurons removed from autopsy brain sections by laser capture microdissection
AU - Torres-Muñoz, Jorge E.
AU - Núñez, Mariana
AU - Petito, Carol K.
N1 - Copyright:
Copyright 2017 Elsevier B.V., All rights reserved.
PY - 2008/7
Y1 - 2008/7
N2 - To confirm studies suggesting that HIV-1 infects neurons and to determine whether CD8+ T lymphocytes traffic to HIV-1-infected neurons, we used laser capture microdissection to remove hippocampal neurons with and without perineuronal CD8+ T cells from AIDS patients with HIV-1 encephalitis (HIVE) or without HIVE and from normal controls. We used hyperbranched multidisplacement amplification for whole gene amplification (MDA-WGA) plus two rounds of PCR to amplify housekeeping sequences (HK+) and, in HK + samples, to amplify HIV-1 gag, nef, and pol sequences. Sample size and, in single neurons, MDA-WGA correlated with housekeeping gene amplification (P < 0.05), whereas patient group and postmortem interval did not (P > 0.05). Neuronal viral sequences correlated with HIVE (43% vs. 13% and 0 in non-HIVE and controls, respectively) and, in HIVE cases, with perlneuronal CD8+ T lymphocytes (70% in CD8+ samples vs. 37% of CD8- samples). Our results suggest that MDA-WGA is a useful technique when analyzing DNA from single cells from autopsy brains, supporting prior studies that show that neurons may contain HIV-1 neuronal sequences in vivo. The association between neuronal infection and perineuronal CD8+ T cells supports our hypothesis that these cells specifically traffic to infected neurons but raises the possibility that CD8+ T cells, if infected, could transmit virus to neurons.
AB - To confirm studies suggesting that HIV-1 infects neurons and to determine whether CD8+ T lymphocytes traffic to HIV-1-infected neurons, we used laser capture microdissection to remove hippocampal neurons with and without perineuronal CD8+ T cells from AIDS patients with HIV-1 encephalitis (HIVE) or without HIVE and from normal controls. We used hyperbranched multidisplacement amplification for whole gene amplification (MDA-WGA) plus two rounds of PCR to amplify housekeeping sequences (HK+) and, in HK + samples, to amplify HIV-1 gag, nef, and pol sequences. Sample size and, in single neurons, MDA-WGA correlated with housekeeping gene amplification (P < 0.05), whereas patient group and postmortem interval did not (P > 0.05). Neuronal viral sequences correlated with HIVE (43% vs. 13% and 0 in non-HIVE and controls, respectively) and, in HIVE cases, with perlneuronal CD8+ T lymphocytes (70% in CD8+ samples vs. 37% of CD8- samples). Our results suggest that MDA-WGA is a useful technique when analyzing DNA from single cells from autopsy brains, supporting prior studies that show that neurons may contain HIV-1 neuronal sequences in vivo. The association between neuronal infection and perineuronal CD8+ T cells supports our hypothesis that these cells specifically traffic to infected neurons but raises the possibility that CD8+ T cells, if infected, could transmit virus to neurons.
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U2 - 10.2353/jmoldx.2008.070074
DO - 10.2353/jmoldx.2008.070074
M3 - Article
C2 - 18556769
AN - SCOPUS:47649130437
VL - 10
SP - 317
EP - 324
JO - Journal of Molecular Diagnostics
JF - Journal of Molecular Diagnostics
SN - 1525-1578
IS - 4
ER -