CD4(+) T follicular helper and IgA(+) B cell numbers in gut biopsies from HIV-infected subjects on antiretroviral therapy are similar to HIV-uninfected individuals

BACKGROUND: Disruption of gastrointestinal tract epithelial and immune barriers contribute to microbial translocation, systemic inflammation, and progression of HIV-1 infection. Antiretroviral therapy (ART) may lead to reconstitution of CD4(+) T cells in gut-associated lymphoid tissue (GALT), but its impact on humoral immunity within GALT is unclear. Therefore, we studied CD4(+) subsets, including T follicular helper cells (Tfh), as well as resident B cells that have switched to IgA production, in gut biopsies, from HIV(+) subjects on suppressive ART compared to HIV-negative controls (HNC). METHODS: Twenty-three HIV(+) subjects on ART and 22 HNC undergoing colonoscopy were recruited to the study. Single-cell suspensions were prepared from biopsies from left colon (LC), right colon (RC), and terminal ileum (TI). T and B lymphocyte subsets, as well as EpCAM(+) epithelial cells, were accurately enumerated by flow cytometry, using counting beads. RESULTS: No significant differences in the number of recovered epithelial cells were observed between the two subject groups. However, the median TI CD4(+) T cell count/10(6) epithelial cells was 2.4-fold lower in HIV(+) subjects versus HNC (19,679 versus 47,504 cells; p = 0.02). Similarly, median LC CD4(+) T cell counts were reduced in HIV(+) subjects (8,358 versus 18,577; p = 0.03) but were not reduced in RC. Importantly, we found no significant differences in Tfh or IgA(+) B cell counts at either site between HIV(+) subjects and HNC. Further analysis showed no difference in CD4(+), Tfh, or IgA(+) B cell counts between subjects who commenced ART in primary compared to chronic HIV-1 infection. Despite the decrease in total CD4 T cells, we could not identify a selective decrease of other key subsets of CD4(+) T cells, including CCR5(+) cells, CD127(+) long-term memory cells, CD103(+) tissue-resident cells, or CD161(+) cells (surrogate marker for Th17), but there was a slight increase in the proportion of T regulatory cells. CONCLUSION: While there were lower absolute CD4(+) counts in the TI and LC in HIV(+) subjects on ART, they were not associated with significantly reduced Tfh cell counts or IgA(+) B cells, suggesting that this important vanguard of adaptive immune defense against luminal microbial products is normalized following ART.John Zaunders, Mark Danta, Michelle Bailey, Gerald Mak, Katherine Marks, Nabila Seddiki, Yin Xu, David J. Templeton, David A. Cooper, Mark A. Boyd, Anthony D. Kelleher and Kersten K. Koelsc

HIV DNA subspecies persist in both activated and resting memory CD4+ T cells during antiretroviral therapy

The latent HIV reservoir is a major impediment to curing HIV infection. The contribution of CD4 T cell activation status to the establishment and maintenance of the latent reservoir was investigated by enumerating viral DNA components in a cohort of 12 individuals commencing antiretroviral therapy (ART) containing raltegravir, an integrase inhibitor. Prior to ART, the levels of total HIV DNA were similar across HLA-DR and HLA-DR (HLA-DR) CD38 memory CD4 T cell phenotypes; episomal two-long terminal repeat (2-LTR) HIV DNA levels were higher in resting (HLA-DR CD38) cells, and this phenotype exhibited a significantly higher ratio of 2-LTR to integrated HIV DNA (P = 0.002). After 1 year of ART, there were no significant differences across each of the memory phenotypes of any HIV DNA component. The decay dynamics of integrated HIV DNA were slow within each subset, and integrated HIV DNA in the resting HLA-DR CD38 subset per mm of peripheral blood exhibited no significant decay (half-life of 25 years). Episomal 2-LTR HIV DNA decayed relative to integrated HIV DNA in resting cells with a half-life of 134 days. Surprisingly, from week 12 on, the decay rates of both total and episomal HIV DNA were lower in activated CD38 cells. By weeks 24 and 52, HIV RNA levels in plasma were most significantly correlated with the numbers of resting cells containing integrated HIV DNA. On the other hand, total HIV DNA levels in all subsets were significantly correlated with the numbers of HLA-DR CD38 cells containing integrated HIV DNA. These results provide insights into the interrelatedness of cell activation and reservoir maintenance, with implications for the design of therapeutic strategies targeting HIV persistence

HIV-1 DNA predicts disease progression and post-treatment virological control

In HIV-1 infection, a population of latently infected cells facilitates viral persistence despite antiretroviral therapy (ART). With the aim of identifying individuals in whom ART might induce a period of viraemic control on stopping therapy, we hypothesised that quantification of the pool of latently infected cells in primary HIV-1 infection (PHI) would predict clinical progression and viral replication following ART. We measured HIV-1 DNA in a highly characterised randomised population of individuals with PHI. We explored associations between HIV-1 DNA and immunological and virological markers of clinical progression, including viral rebound in those interrupting therapy. In multivariable analyses, HIV-1 DNA was more predictive of disease progression than plasma viral load and, at treatment interruption, predicted time to plasma virus rebound. HIV-1 DNA may help identify individuals who could safely interrupt ART in future HIV-1 eradication trials

Switching Virally Suppressed, Treatment-Experienced Patients to a Raltegravir-Containing Regimen Does Not Alter Levels of HIV-1 DNA

Background: Current HIV-1 antiretroviral therapy (ART) greatly reduces virus replication but does not significantly affect the viral reservoir. Raltegravir, a recently introduced integrase inhibitor, could, at least theoretically, reduce residual viremia in patients on ART and affect the viral reservoir size. The aim of this study was to assess whether switching therapy in treatment-experienced patients that were virally suppressed to a raltegravir-containing regimen reduces the size of the viral reservoir, and if such treatment leads to a change in levels of HIV 2-LTR circles in this patient group. Methods: 14 ART experienced individuals with a suppressed viral load (,50 HIV-1 RNA copies/mL plasma) at baseline (for at least 2 months) were switched to a raltegravir-containing regimen. Blood samples were taken at baseline and at $2 timepoints up to 4866 weeks. Levels of total HIV-1 DNA and 2-LTR circles in peripheral blood mononuclear cells (PBMCs) were measured using real-time PCR assays. Results: There was no significant change in HIV-1 total DNA levels over the study duration (p = 0.808), median slope 0.24 (conservative nonparametric 95 % CI: 211.78, 26.23). Low levels of 2-LTR circles were detected in 2 patients. One had 16 copies/10 6 PBMCs at baseline and the other had 34 copies/10 6 PBMCs at week 51. Conclusions: The switch to a raltegravir containing regimen was not associated with a significant change in HIV-1 total DNA levels in this cohort. There were no observed changes in the levels of HIV-1 2-LTR circles associated with raltegravi

Progressive Activation of CD127+132− Recent Thymic Emigrants into Terminally Differentiated CD127−132+ T-Cells in HIV-1 Infection

AIM: HIV infection is associated with distortion of T-cell homeostasis and the IL-7/IL7R axis. Progressive infection results in loss of CD127+132- and gains in CD127-132+ CD4+ and CD8+ T-cells. We investigated the correlates of loss of CD127 from the T-cell surface to understand mechanisms underlying this homeostatic dysregulation. METHODS: Peripheral and cord blood mononuclear cells (PBMCs; CBMC) from healthy volunteers and PBMC from patients with HIV infection were studied. CD127+132-, CD127+132+ and CD127-132+ T-cells were phenotyped by activation, differentiation, proliferation and survival markers. Cellular HIV-DNA content and signal-joint T-cell receptor excision circles (sjTRECs) were measured. RESULTS: CD127+132- T-cells were enriched for naïve cells while CD127-132+ T-cells were enriched for activated/terminally differentiated T-cells in CD4+ and CD8+ subsets in health and HIV infection. HIV was associated with increased proportions of activated/terminally differentiated CD127-132+ T-cells. In contrast to CD127+132- T-cells, CD127-132+ T-cells were Ki-67+Bcl-2(low) and contained increased levels of HIV-DNA. Naïve CD127+132- T-cells contained a higher proportion of sjTRECs. CONCLUSION: The loss of CD127 from the T-cell surface in HIV infection is driven by activation of CD127+132- recent thymic emigrants into CD127-132+ activated/terminally differentiated cells. This process likely results in an irreversible loss of CD127 and permanent distortion of T-cell homeostasis

Divergent Expression of CXCR5 and CCR5 on CD4+ T Cells and the Paradoxical Accumulation of T Follicular Helper Cells during HIV Infection

Viral infection sets in motion a cascade of immune responses, including both CXCR5+CD4+ T follicular helper (Tfh) cells that regulate humoral immunity and CCR5+CD4+ T cells that mediate cell-mediated immunity. In peripheral blood mononuclear cells, the majority of memory CD4+ T cells appear to fall into either of these two lineages, CCR5-CXCR5+ or CCR5+CXCR5-. Very high titers of anti-HIV IgG antibodies are a hallmark of infection, strongly suggesting that there is significant HIV-specific CD4+ T cell help to HIV-specific B cells. We now know that characteristic increases in germinal centers (GC) in lymphoid tissue (LT) during SIV and HIV-1 infections are associated with an increase in CXCR5+PD-1high Tfh, which expand to a large proportion of memory CD4+ T cells in LT, and are presumably specific for SIV or HIV epitopes. Macaque Tfh normally express very little CCR5, yet are infected by CCR5-using SIV, which may occur mainly through infection of a subset of PD-1intermediateCCR5+Bcl-6+ pre-Tfh cells. In contrast, in human LT, a subset of PD-1high Tfh appears to express low levels of CCR5, as measured by flow cytometry, and this may also contribute to the high rate of infection of Tfh. Also, we have found, by assessing fine-needle biopsies of LT, that increases in Tfh and GC B cells in HIV infection are not completely normalized by antiretroviral therapy (ART), suggesting a possible long-lasting reservoir of infected Tfh. In contrast to the increase of CXCR5+ Tfh, there is no accumulation of proliferating CCR5+ CD4 T HIV Gag-specific cells in peripheral blood that make IFN-γ. Altogether, CXCR5+CCR5- CD4 T cells that regulate humoral immunity are allowed greater freedom to operate and expand during HIV-1 infection, but at the same time can contain HIV DNA at levels at least as high as in other CD4 subsets. We argue that early ART including a CCR5 blocker may directly reduce the infected Tfh reservoir in LT and also interrupt cycles of antibody pressure driving virus mutation and additional GC responses to resulting neoantigens

Peripheral blood mononuclear cells HIV DNA levels impact intermittently on neurocognition.

To determine the contribution of peripheral blood mononuclear cells' (PBMCs) HIV DNA levels to HIV-associated dementia (HAD) and non-demented HIV-associated neurocognitive disorders (HAND) in chronically HIV-infected adults with long-term viral suppression on combined antiretroviral treatment (cART).Eighty adults with chronic HIV infection on cART (>97% with plasma and CSF HIV RNA <50 copies/mL) were enrolled into a prospective observational cohort and underwent assessments of neurocognition and pre-morbid cognitive ability at two visits 18 months apart. HIV DNA in PBMCs was measured by real-time PCR at the same time-points.At baseline, 46% had non-demented HAND; 7.5% had HAD. Neurocognitive decline occurred in 14% and was more likely in those with HAD (p<.03). Low pre-morbid cognitive ability was uniquely associated with HAD (p<.05). Log10 HIV DNA copies were stable between study visits (2.26 vs. 2.22 per 106 PBMC). Baseline HIV DNA levels were higher in those with lower pre-morbid cognitive ability (p<.04), and higher in those with no ART treatment during HIV infection 1st year (p = .03). Baseline HIV DNA was not associated with overall neurocognition. However, % ln HIV DNA change was associated with decline in semantic fluency in unadjusted and adjusted analyses (p = .01-.03), and motor-coordination (p = .02-.12) to a lesser extent.PBMC HIV DNA plays a role in HAD pathogenesis, and this is moderated by pre-morbid cognitive ability in the context of long-term viral suppression. While the HIV DNA levels in PBMC are not associated with current non-demented HAND, increasing HIV DNA levels were associated with a decline in neurocognitive functions associated with HAND progression

Data from: HIV dynamics linked to memory CD4+ T cell homeostasis

The dynamics of latent HIV is linked to infection and clearance of resting memory CD4+ T cells. Infection also resides within activated, non-dividing memory cells and can be impacted by antigen-driven and homeostatic proliferation despite suppressive antiretroviral therapy (ART). We investigated whether plasma viral level (pVL) and HIV DNA dynamics could be explained by HIV’s impact on memory CD4+ T cell homeostasis. Median total, 2-LTR and integrated HIV DNA levels per μL of peripheral blood, for 8 primary (PHI) and 8 chronic HIV infected (CHI) individuals enrolled on a raltegravir (RAL) based regimen, exhibited greatest changes over the 1st year of ART. Dynamics slowed over the following 2 years so that total HIV DNA levels were equivalent to reported values for individuals after 10 years of ART. The mathematical model reproduced the multiphasic dynamics of pVL, and levels of total, 2-LTR and integrated HIV DNA in both PHI and CHI over 3 years of ART. Under these simulations, residual viremia originated from reactivated latently infected cells where most of these cells arose from clonal expansion within the resting phenotype. Since virion production from clonally expanded cells will not be affected by antiretroviral drugs, simulations of ART intensification had little impact on pVL. HIV DNA decay over the first year of ART followed the loss of activated memory cells (120 day half-life) while the 5.9 year half-life of total HIV DNA after this point mirrored the slower decay of resting memory cells. Simulations had difficulty reproducing the fast early HIV DNA dynamics, including 2-LTR levels peaking at week 12, and the later slow loss of total and 2-LTR HIV DNA, suggesting some ongoing infection. In summary, our modelling indicates that much of the dynamical behavior of HIV can be explained by its impact on memory CD4+ T cell homeostasis

Baseline and follow-up log₁₀ HIV DNA per 10⁶ PBMC.

<p>Baseline and follow-up log₁₀ HIV DNA per 10⁶ PBMC.</p