Abstract
ObjectiveTo address the role of translationally active HIV reservoir in chronic inflammation and non-AIDS related disorders, we first need a simple and accurate assay to evaluate viral protein expression in virally suppressed subjects.DesignWe optimized an HIV Nef enzyme-linked immunosorbent assay (ELISA) and used it to quantify plasma Nef levels as an indicator of the leaky HIV reservoir in an HIV-infected cohort.MethodsThis study accessed 134 plasma samples from a well-characterized cohort study of HIV-infected and uninfected adults in San Francisco (the SCOPE cohort). We optimized an ELISA for detection of plasma Nef in HIV-negative subjects and HIV-infected non-controllers, and evaluated its utility to quantify plasma Nef levels in a cross-sectional study of ART-suppressed and elite controller HIV-infected subjects.ResultsHere, we describe the performance of an optimized HIV Nef ELISA. When we applied this assay to the study cohort we found that plasma Nef levels were correlated with plasma HIV RNA levels in untreated disease. However, we were able to detect Nef in plasma of approximately half of subjects on ART or with elite control, despite the lack of detectable plasma HIV RNA levels using standard assays. Plasma Nef levels were not consistently associated with CD4+ T-cell count, CD8+ T-cell count, self-reported nadir CD4+ T-cell count or the CD4+/CD8+ T-cell ratio in HIV-infected subjects.ConclusionSince plasma HIV RNA levels are undetectable in virally suppressed subjects, it is reasonable to assume that viral protein expression in leaky reservoir, and not plasma virions, is the source of Nef accumulating in plasma. To examine this further, improvements of the assay sensitivity, by lowering the background through improvements in the quality of Nef antibodies, and detailed characterization of the HIV reservoirs are needed.
Highlights
Effective antiretroviral therapy (ART) reduces plasma HIV RNA levels to below the limit of quantification (< 20 to 50 RNA copies/ml) in the vast majority of treated HIV-infected individuals [1], still the virus persists indefinitely [2]
We describe the performance of an optimized HIV Nef enzyme-linked immunosorbent assay (ELISA)
When we applied this assay to the study cohort we found that plasma Nef levels were correlated with plasma HIV RNA levels in untreated disease
Summary
Effective antiretroviral therapy (ART) reduces plasma HIV RNA levels to below the limit of quantification (< 20 to 50 RNA copies/ml) in the vast majority of treated HIV-infected individuals [1], still the virus persists indefinitely [2]. The quantitative viral outgrowth assay is laborious, expensive and only provides a minimal estimate of the true reservoir size [5]. Reverse-transcription PCR-detection of intracellular and virion HIV RNA provides an (over)estimate of the more transcriptionally active reservoir [3, 4]. Fluorescent in situ hybridization connected to flow cytometry enables single-cell characterization of HIV RNA and proteins in re-activated HIV reservoirs, but assay sensitivity may be limiting [7, 8]. Viral proteins Nef and Env were detected in plasma of some virally suppressed subjects, indicating a need for evaluating the translationally active reservoir [9, 10]
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
