Use of RT-Defective HIV Virions: New Tool to Evaluate Specific Response in Chronic Asymptomatic HIV-Infected Individuals

Abstract

BackgroundGeneration of new reagents that can be used to screen or monitor HIV-1-specific responses constituted an interesting field in the development of HIV vaccines to improve their efficacy.MethodsWe have evaluated the specific T cell response against different types of NL4-3 virions (including NL4-3 aldrithiol-2 treated, NL4-3/ΔRT and R5 envelopes: NL4-3/ΔRT/ΔEnv[AC10] and NL4-3/ΔRT/ΔEnv[Bal]) and against pools of overlapping peptides (15 mer) encompassing the HIV-1 Gag and Nef regions. Cryopreserved PBMC from a subset of 69 chronic asymptomatic HIV positive individuals have been employed using different techniques including IFN-γ ELISPOT assay, surface activation markers and intracellular cytokine staining (ICS) by flow cytometry.ResultsThe differential response obtained against NL4-3 aldrithiol-2 treated and NL4-3/ΔRT virions (25% vs 55%, respectively) allow us to divide the population in three groups: “full-responders” (positive response against both viral particles), “partial-responders” (positive response only against NL4-3/ΔRT virions) and “non-responders” (negative responses). There was no difference between X4 and R5 envelopes. The magnitude of the total responses was higher against NL4-3/ΔRT and was positively correlated with gender and inverse correlated with viral load. On the contrary CD4+ T cell count was not associated with this response. In any case responses to the viruses tended to be lower in magnitude than those detected by the overlapping peptides tested. Finally we have found an increased frequency of HLA-B27 allele (23% vs 9%) and a significant reduction in some activation markers (CD69 and CD38) on T cells surface in responders vs non-responders individuals.ConclusionsIn summary these virions could be considered as alternative and useful reagents for screening HIV-1-specific T cell responses in HIV exposed uninfected people, HIV infected patients and to assess immunogenicity of new prototypes both in vitro and in vaccine trials, by a feasible, simply, effective and low cost assay.

Prolonged administration of maraviroc reactivates latent HIV in vivo but it does not prevent antiretroviral-free viral rebound

“…ELISpot assay. ELISpot assay was performed to measure the numbers of PBMCs producing interferon (IFN)-γ as a response against HIV sequences 78 . Briefly, experiments were performed with cryopreserved PBMCs using 10 pools of peptides, each consisting of 15-mers overlapping 11 peptides, grouped in pools of 10 to 12 peptides each, and covering the whole HIV gag, nef, pol, and env-gp41 sequences.…”

The HIV-1 integrase-LEDGF allosteric inhibitor MUT-A: resistance profile, impairment of virus maturation and infectivity but without influence on RNA packaging or virus immunoreactivity

“…In fact, previous work has indicated that, for MDDC, small amounts of antigen are sufficient for presentation on MHC molecules and activation of T cells. Some of these approaches have been used with antigens from non-replicating viruses, including aldrithiol-2 (AT-2)-inactivated HIV-1 [ 41 – 43 ] or defective virus [ 44 ]. As we describe in this report, MUT-A-inactivated virus conserves conformational and functionally intact surface envelope proteins that can interact with T cells as well as with surface receptors present on circulating dendritic cells and monocytes.…”

Protease inhibitor monotherapy is associated with a higher level of monocyte activation, bacterial translocation and inflammation

“…EDTA-anticoagulated blood was obtained by venipuncture; PBMCs were immediately isolated by density gradient centrifugation using Ficoll-Hypaque (Sigma Chemical Co., St. Louis, MO). We used a comprehensive approach of simultaneous measurement of different immunological parameters [ 12 ] in CD4+ and CD8+ T lymphocyte: activation (using CD38 and HLADR markers), senescence (using CD28 and CD57 marker), exhaustion (using PD-1 marker), co-receptor expression (CCR5, CXCR4), differentiation stage (using CD45RA and CD45RO markers) and monocyte activation (using CD14, CD16 and CD163 markers). The stained cells were analyzed on a FACSCalibur (Becton Dickinson, San Jose, CA) cytometer.…”