Comparative analysis of cell-associated HIV DNA levels in cerebrospinal fluid and peripheral blood by droplet digital PCR
Genre
Journal ArticleDate
2015-10-02Author
De Oliveira, MFGianella, S
Letendre, S
Scheffler, K
Pond, SLK
Smith, DM
Strain, M
Ellis, RJ
Subject
Anti-HIV AgentsAntiretroviral Therapy, Highly Active
CD4-Positive T-Lymphocytes
Cerebrospinal Fluid
DNA, Viral
HIV Infections
HIV-1
Humans
Leukocytes, Mononuclear
Organ Specificity
Polymerase Chain Reaction
RNA, Viral
Retrospective Studies
Viral Load
Viremia
Virus Replication
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http://hdl.handle.net/20.500.12613/5190
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10.1371/journal.pone.0139510Abstract
Background Measurement of HIV DNA-bearing cells in cerebrospinal fluid (CSF) is challenging because few cells are present. We present a novel application of the sensitive droplet digital (dd) PCR in this context. Methods We analyzed CSF cell pellets and paired peripheral blood mononuclear cells (PBMC) from 28 subjects, 19 of whom had undetectable HIV RNA (<48 copies/mL) in both compartments. We extracted DNA from PBMC using silica-based columns and used direct lysis on CSF cells. HIV DNA and the host housekeeping gene (RPP30) were measured in CSF and PBMC by (dd)PCR. We compared HIV DNA levels in virally-suppressed and-unsuppressed subgroups and calculated correlations between HIV DNA and RNA levels in both compartments using non-parametric tests. Results HIV DNA was detected in 18/28 (64%) CSF cell pellets, including 10/19 (53%) samples with undetectable HIV RNA. HIV DNA levels in CSF cell pellets were not correlated with RPP30 (p = 0.3), but correlated positively with HIV RNA in CSF (p = 0.04) and HIV DNA in PBMC (p = 0.03). Cellular HIV DNA in CSF was detected in comparable levels in HIV RNA-suppressed and unsuppressed subjects (p = 0.14). In contrast, HIV DNA levels in PBMC were significantly lower in HIV RNA-suppressed than in unsuppressed subjects (p = 0.014). Among subjects with detectable HIV DNA in both compartments, HIV DNA levels in CSF were significantly higher than in PBMC (p<0.001). Conclusions Despite low mononuclear cell numbers in CSF, HIV DNA was detected in most virally suppressed individuals. In contrast to PBMC, suppressive ART was not associated with lower HIV DNA levels in CSF cells, compared to no ART, perhaps due to poorer ART penetration, slower decay of HIV DNA, or enrichment of HIV DNA-bearing mononuclear cells into the CSF, compared to blood. Future studies should determine what fraction of HIV DNA is replication- competent in CSF leukocytes, compared to PBMC.Citation to related work
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