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Elite suppressors harbor low levels of integrated HIV DNA and high levels of 2-LTR circular HIV DNA compared to HIV+ patients on and off HAART.

Graf EH, Mexas AM, Yu JJ, Shaheen F, Liszewski MK, Di Mascio M, Migueles SA, Connors M, O'Doherty U - PLoS Pathog. (2011)

Bottom Line: To determine the composition of unintegrated HIV DNA in these samples, we measured circular 2-LTR HIV DNA forms and found ES patients frequently have high levels of 2-LTR circles in PBMC.We further show that these high levels of 2-LTR circles are not the result of inefficient integration in ES cells, since HIV integrates with similar efficiency in ES and normal donor cells.Moreover, they add significantly to our understanding of the mechanisms that allow viral control and reservoir maintenance in this unique patient population.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology and Laboratory Medicine, University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania, United States of America.

ABSTRACT
Elite suppressors (ES) are a rare population of HIV-infected individuals that are capable of naturally controlling the infection without the use of highly active anti-retroviral therapy (HAART). Patients on HAART often achieve viral control to similar (undetectable) levels. Accurate and sensitive methods to measure viral burden are needed to elucidate important differences between these two patient populations in order to better understand their mechanisms of control. Viral burden quantification in ES patients has been limited to measurements of total DNA in PBMC, and estimates of Infectious Units per Million cells (IUPM). There appears to be no significant difference in the level of total HIV DNA between cells from ES patients and patients on HAART. However, recovering infectious virus from ES patient samples is much more difficult, suggesting their reservoir size should be much smaller than that in patients on HAART. Here we find that there is a significant difference in the level of integrated HIV DNA in ES patients compared to patients on HAART, providing an explanation for the previous results. When comparing the level of total to integrated HIV DNA in these samples we find ES patients have large excesses of unintegrated HIV DNA. To determine the composition of unintegrated HIV DNA in these samples, we measured circular 2-LTR HIV DNA forms and found ES patients frequently have high levels of 2-LTR circles in PBMC. We further show that these high levels of 2-LTR circles are not the result of inefficient integration in ES cells, since HIV integrates with similar efficiency in ES and normal donor cells. Our findings suggest that measuring integration provides a better surrogate of viral burden than total HIV DNA in ES patients. Moreover, they add significantly to our understanding of the mechanisms that allow viral control and reservoir maintenance in this unique patient population.

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Related in: MedlinePlus

ES CD4+ T cells are not resistant to in vitro infection.CD4+ T cells from 10 ES and 6 uninfected donors were inoculated with NL4-3 (A). Reverse transcripts and integrated HIV DNA were measured at 2 days post-infection. From these values, integration efficiency was calculated (i.e. integrated copies per reverse transcript). ES were compared to normal donors with a Wilcoxon rank-sum test. Neither reverse transcription, integration nor integration efficiency measurements were statistically different between the two populations. Six ES were also inoculated with 3 serial ten-fold dilutions of the same virus (B). The bars represent the averages of multiple experiments (n is indicated in figures) and the error bars represent the standard deviations.
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ppat-1001300-g006: ES CD4+ T cells are not resistant to in vitro infection.CD4+ T cells from 10 ES and 6 uninfected donors were inoculated with NL4-3 (A). Reverse transcripts and integrated HIV DNA were measured at 2 days post-infection. From these values, integration efficiency was calculated (i.e. integrated copies per reverse transcript). ES were compared to normal donors with a Wilcoxon rank-sum test. Neither reverse transcription, integration nor integration efficiency measurements were statistically different between the two populations. Six ES were also inoculated with 3 serial ten-fold dilutions of the same virus (B). The bars represent the averages of multiple experiments (n is indicated in figures) and the error bars represent the standard deviations.

Mentions: DNA samples from ten ES patients were analyzed for 2-LTR circular HIV DNA. 2-LTR circular HIV DNA levels were compared to those in samples from HIV+ patients on and off-HAART. Only ES and on HAART could be compared due to small samples size. The level of 2-LTR circles was higher in ES as was the frequency with which they were detected (pā€Š=ā€Š0.007, chi-square test). The line represents the median value.


Elite suppressors harbor low levels of integrated HIV DNA and high levels of 2-LTR circular HIV DNA compared to HIV+ patients on and off HAART.

Graf EH, Mexas AM, Yu JJ, Shaheen F, Liszewski MK, Di Mascio M, Migueles SA, Connors M, O'Doherty U - PLoS Pathog. (2011)

ES CD4+ T cells are not resistant to in vitro infection.CD4+ T cells from 10 ES and 6 uninfected donors were inoculated with NL4-3 (A). Reverse transcripts and integrated HIV DNA were measured at 2 days post-infection. From these values, integration efficiency was calculated (i.e. integrated copies per reverse transcript). ES were compared to normal donors with a Wilcoxon rank-sum test. Neither reverse transcription, integration nor integration efficiency measurements were statistically different between the two populations. Six ES were also inoculated with 3 serial ten-fold dilutions of the same virus (B). The bars represent the averages of multiple experiments (n is indicated in figures) and the error bars represent the standard deviations.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3044690&req=5

ppat-1001300-g006: ES CD4+ T cells are not resistant to in vitro infection.CD4+ T cells from 10 ES and 6 uninfected donors were inoculated with NL4-3 (A). Reverse transcripts and integrated HIV DNA were measured at 2 days post-infection. From these values, integration efficiency was calculated (i.e. integrated copies per reverse transcript). ES were compared to normal donors with a Wilcoxon rank-sum test. Neither reverse transcription, integration nor integration efficiency measurements were statistically different between the two populations. Six ES were also inoculated with 3 serial ten-fold dilutions of the same virus (B). The bars represent the averages of multiple experiments (n is indicated in figures) and the error bars represent the standard deviations.
Mentions: DNA samples from ten ES patients were analyzed for 2-LTR circular HIV DNA. 2-LTR circular HIV DNA levels were compared to those in samples from HIV+ patients on and off-HAART. Only ES and on HAART could be compared due to small samples size. The level of 2-LTR circles was higher in ES as was the frequency with which they were detected (pā€Š=ā€Š0.007, chi-square test). The line represents the median value.

Bottom Line: To determine the composition of unintegrated HIV DNA in these samples, we measured circular 2-LTR HIV DNA forms and found ES patients frequently have high levels of 2-LTR circles in PBMC.We further show that these high levels of 2-LTR circles are not the result of inefficient integration in ES cells, since HIV integrates with similar efficiency in ES and normal donor cells.Moreover, they add significantly to our understanding of the mechanisms that allow viral control and reservoir maintenance in this unique patient population.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology and Laboratory Medicine, University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania, United States of America.

ABSTRACT
Elite suppressors (ES) are a rare population of HIV-infected individuals that are capable of naturally controlling the infection without the use of highly active anti-retroviral therapy (HAART). Patients on HAART often achieve viral control to similar (undetectable) levels. Accurate and sensitive methods to measure viral burden are needed to elucidate important differences between these two patient populations in order to better understand their mechanisms of control. Viral burden quantification in ES patients has been limited to measurements of total DNA in PBMC, and estimates of Infectious Units per Million cells (IUPM). There appears to be no significant difference in the level of total HIV DNA between cells from ES patients and patients on HAART. However, recovering infectious virus from ES patient samples is much more difficult, suggesting their reservoir size should be much smaller than that in patients on HAART. Here we find that there is a significant difference in the level of integrated HIV DNA in ES patients compared to patients on HAART, providing an explanation for the previous results. When comparing the level of total to integrated HIV DNA in these samples we find ES patients have large excesses of unintegrated HIV DNA. To determine the composition of unintegrated HIV DNA in these samples, we measured circular 2-LTR HIV DNA forms and found ES patients frequently have high levels of 2-LTR circles in PBMC. We further show that these high levels of 2-LTR circles are not the result of inefficient integration in ES cells, since HIV integrates with similar efficiency in ES and normal donor cells. Our findings suggest that measuring integration provides a better surrogate of viral burden than total HIV DNA in ES patients. Moreover, they add significantly to our understanding of the mechanisms that allow viral control and reservoir maintenance in this unique patient population.

Show MeSH
Related in: MedlinePlus