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Comparative Analysis of Cell-Associated HIV DNA Levels in Cerebrospinal Fluid and Peripheral Blood by Droplet Digital PCR.

de Oliveira MF, Gianella S, Letendre S, Scheffler K, Kosakovsky Pond SL, Smith DM, Strain M, Ellis RJ - PLoS ONE (2015)

Bottom Line: Among subjects with detectable HIV DNA in both compartments, HIV DNA levels in CSF were significantly higher than in PBMC (p<0.001).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.

View Article: PubMed Central - PubMed

Affiliation: University of California, San Diego, La Jolla, California, United States of America.

ABSTRACT

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.

No MeSH data available.


Related in: MedlinePlus

HIV DNA levels in PBMC and CSF cells between HIV RNA-suppressed and non-suppressed subjects.Comparison of HIV DNA levels in PBMC (A) and CSF pellet (B) between individuals with detectable and undetectable HIV RNA levels in blood plasma or CSF supernatant, respectively. Rank-based Mann-Whitney test p values are indicated.
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pone.0139510.g003: HIV DNA levels in PBMC and CSF cells between HIV RNA-suppressed and non-suppressed subjects.Comparison of HIV DNA levels in PBMC (A) and CSF pellet (B) between individuals with detectable and undetectable HIV RNA levels in blood plasma or CSF supernatant, respectively. Rank-based Mann-Whitney test p values are indicated.

Mentions: To evaluate how viral suppression on ART influenced HIV DNA levels, we compared levels of HIV DNA in virally suppressed and unsuppressed participants across compartments. As might be expected, levels of HIV DNA in PBMC were significantly higher in individuals with detectable levels of HIV RNA in blood plasma when compared to subjects with undetectable HIV RNA levels (p = 0.014; Fig 3A). There was, however, no significant difference in HIV DNA levels of CSF cell pellets between participants with detectable or undetectable HIV RNA viral load in CSF supernatant, (p = 0.97 only among people with detectable HIV DNA and p = 0.14 when including the entire cohort; Fig 3B). The frequency of subjects with detectable HIV DNA in CSF was higher among those with detectable HIV RNA in CSF when compared to subjects with undetectable HIV RNA in CSF, however this was not statistically significant (89% versus 52.6%, p = 0.1).


Comparative Analysis of Cell-Associated HIV DNA Levels in Cerebrospinal Fluid and Peripheral Blood by Droplet Digital PCR.

de Oliveira MF, Gianella S, Letendre S, Scheffler K, Kosakovsky Pond SL, Smith DM, Strain M, Ellis RJ - PLoS ONE (2015)

HIV DNA levels in PBMC and CSF cells between HIV RNA-suppressed and non-suppressed subjects.Comparison of HIV DNA levels in PBMC (A) and CSF pellet (B) between individuals with detectable and undetectable HIV RNA levels in blood plasma or CSF supernatant, respectively. Rank-based Mann-Whitney test p values are indicated.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4592012&req=5

pone.0139510.g003: HIV DNA levels in PBMC and CSF cells between HIV RNA-suppressed and non-suppressed subjects.Comparison of HIV DNA levels in PBMC (A) and CSF pellet (B) between individuals with detectable and undetectable HIV RNA levels in blood plasma or CSF supernatant, respectively. Rank-based Mann-Whitney test p values are indicated.
Mentions: To evaluate how viral suppression on ART influenced HIV DNA levels, we compared levels of HIV DNA in virally suppressed and unsuppressed participants across compartments. As might be expected, levels of HIV DNA in PBMC were significantly higher in individuals with detectable levels of HIV RNA in blood plasma when compared to subjects with undetectable HIV RNA levels (p = 0.014; Fig 3A). There was, however, no significant difference in HIV DNA levels of CSF cell pellets between participants with detectable or undetectable HIV RNA viral load in CSF supernatant, (p = 0.97 only among people with detectable HIV DNA and p = 0.14 when including the entire cohort; Fig 3B). The frequency of subjects with detectable HIV DNA in CSF was higher among those with detectable HIV RNA in CSF when compared to subjects with undetectable HIV RNA in CSF, however this was not statistically significant (89% versus 52.6%, p = 0.1).

Bottom Line: Among subjects with detectable HIV DNA in both compartments, HIV DNA levels in CSF were significantly higher than in PBMC (p<0.001).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.

View Article: PubMed Central - PubMed

Affiliation: University of California, San Diego, La Jolla, California, United States of America.

ABSTRACT

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.

No MeSH data available.


Related in: MedlinePlus