Aida Sivro

Epigenetic control of IRF1 responses in HIV-exposed seronegative versus HIV-susceptible individuals

Not all individuals exposed to HIV become infected. Understanding why these HIV-exposed seronegative individuals remain uninfected will help inform the development of preventative measures against HIV infection. Interferon regulatory factor-1 (IRF1) plays a critical role both in host antiviral immunity and in HIV-1 replication. This study examined IRF1 expression regulation in the ex vivo peripheral blood mononuclear cells of HIV-exposed seronegative commercial sex workers who can be epidemiologically defined as relatively resistant to HIV infection (HIV-R), versus HIV-uninfected, susceptible controls (HIV-S). Whereas HIV-susceptible individuals demonstrated a biphasic, prolonged increase in IRF1 expression after interferon-γ stimulation, HIV-R individuals demonstrated a robust, but transient response. We also found that the IRF1 promoter in HIV-R was primed by increased basal histone deacetylase-2 binding, independently of transcription regulators, STAT1 and nuclear factor-κB/p65, implicating an epigenetic silencing mechanism. Interestingly, the transitory IRF1 response in HIV-R was sufficient in comparable regulation of interleukin-12 and interleukin-4 expression compared with the HIV-susceptible controls. This is the first study characterizing IRF1 responsiveness in individuals who demonstrate altered susceptibility to HIV infection. These data suggest that transitory IRF1 responsiveness in HIV-R may be one of the key contributors to the altered susceptibility to HIV infection during the early stages of primary HIV infection.

Age and menopause affect the expression of specific cytokines/chemokines in plasma and cervical lavage samples from female sex workers in Nairobi, Kenya

BackgroundAging of the immune system, known as immunosenescence, is associated with profound changes in both innate and adaptive immune responses, resulting in increased susceptibility to infection and a decreased ability to respond to vaccination. The purpose of this study was to investigate the effect of age and menopause on the expression of 22 different cytokines/chemokines in both plasma and cervical lavage samples from female sex-worker cohort from Nairobi, Kenya (age range 20–65).ResultsCytokine/chemokine levels were measured using a Miliplex multiplex assay (Millipore). We found that age positively correlated with MCP-1 (p = 0.0002) and IP-10 (p = 0.03) systemic cytokine expression, and that women over 50 expressed the highest levels of these cytokines, but also had elevated expression of MIG (ANOVA p = 0.0096) and MIP-3β(ANOVA p = 0.0434). We also found that IL-8 (p = 0.047) and sCD40L (p = 0.01) systemic expression negatively correlated with age. Further, MIG (p = 0.0081) and MCP-1 (p = 0.0157) were present at higher levels in post-menopausal women suggesting a potential estrogen dependant systemic regulation of these cytokines. In cervical lavage samples, age did not directly correlate with the expression of any of the tested cytokines/chemokines, however sIL-2Rα (ANOVA p = 0.0170) and IL-15 (ANOVA p = 0.0251)were significantly higher in women over 50. Menopause was shown to have a more profound effect on cytokine expression in the cervical mucosa with MIG (p = 0.0256), MIP-3α (p = 0.0245), IL-1β (p = 0.0261), IL-6 (p = 0.0462), IL-8 (p = 0.007), IP-10 (p = 0.0357) and MCP-1 (p = 0.0427) all significantly under-expressed in post-menopausal women.ConclusionsThis study demonstrates that aging and menopause-associated hormonal changes are associated with significant changes in systemic and mucosal cytokine/chemokine expression, which may have implications for the age-related decline in the ability to fight against infections.

Integrin α4β7 expression on peripheral blood CD4+ T cells predicts HIV acquisition and disease progression outcomes.

CD4+ T cells expressing α4β7 play an important role in HIV pathogenesis. Indicative integrins in HIV The gut is thought to be a major viral reservoir in HIV infection, and studies in nonhuman primates suggest that targeting the α4β7 integrin on T cells may be a viable therapy. Sivro et al. now extend these findings to humans by examining HIV acquisition in multiple African cohorts. Higher frequencies of α4β7+ circulating CD4+ T cells before infection were associated with increased HIV acquisition, viral load at set point, and more rapid CD4+ T cell decline. These exciting data confirm that integrin targeting could help reduce the spread of HIV. The gastrointestinal (GI) mucosa is central to HIV pathogenesis, and the integrin α4β7 promotes the homing of immune cells to this site, including those that serve as viral targets. Data from simian immunodeficiency virus (SIV) animal models suggest that α4β7 blockade provides prophylactic and therapeutic benefits. We show that pre-HIV infection frequencies of α4β7+ peripheral blood CD4+ T cells, independent of other T cell phenotypes and genital inflammation, were associated with increased rates of HIV acquisition in South African women. A similar acquisition effect was observed in a Kenyan cohort and in nonhuman primates (NHPs) after intravaginal SIV challenge. This association was stronger when infection was caused by HIV strains containing V2 envelope motifs with a preference for α4β7 binding. In addition, pre-HIV α4β7+ CD4+ T cells predicted a higher set-point viral load and a greater than twofold increased rate of CD4+ T cell decline. These results were confirmed in SIV-infected NHPs. Increased frequencies of pre-HIV α4β7+ CD4+ T cells were also associated with higher postinfection expression of lipopolysaccharide binding protein, a microbial translocation marker, suggestive of more extensive gut damage. CD4+ T cells expressing α4β7 were rapidly depleted very early in HIV infection, particularly from the GI mucosa, and were not restored by early antiretroviral therapy. This study provides a link between α4β7 expression and HIV clinical outcomes in humans, in line with observations made in NHPs. Given the availability of a clinically approved anti-α4β7 monoclonal antibody for treatment of inflammatory bowel disease, these data support further evaluation of targeting α4β7 integrin as a clinical intervention during HIV infection.

Interferon Regulatory Factor 1 Polymorphisms Previously Associated with Reduced HIV Susceptibility Have No Effect on HIV Disease Progression

Introduction Interferon regulatory factor 1 (IRF1) is induced by HIV early in the infection process and serves two functions: transactivation of the HIV-1 genome and thus replication, and eliciting antiviral innate immune responses. We previously described three IRF1 polymorphisms that correlate with reduced IRF1 expression and reduced HIV susceptibility. Objective To determine whether IRF1 polymorphisms previously associated with reduced HIV susceptibility play a role in HIV pathogenesis and disease progression in HIV-infected ART-naïve individuals. Methods IRF1 genotyping for polymorphisms (619, MS and 6516) was performed by PCR in 847 HIV positive participants from a sex worker cohort in Nairobi, Kenya. Rates of CD4+ T cell decline and viral loads (VL) were analyzed using linear mixed models. Results Three polymorphisms in the IRF1, located at 619, microsatellite region and 6516 of the gene, previously associated with decreased susceptibility to HIV infection show no effect on disease progression, either measured by HIV-1 RNA levels or the slopes of CD4 decline before treatment initiation. Conclusion Whereas these three polymorphisms in the IRF1 gene protect against HIV-1 acquisition, they appear to exert no discernable effects once infection is established.

Genital inflammation undermines the effectiveness of tenofovir gel in preventing HIV acquisition in women

Several clinical trials have demonstrated that antiretroviral (ARV) drugs taken as pre-exposure prophylaxis (PrEP) can prevent HIV infection, with the magnitude of protection ranging from −49 to 86% (refs. ). Although these divergent outcomes are thought to be due primarily to differences in product adherence, biological factors likely contribute. Despite selective recruitment of higher-risk participants for prevention trials, HIV risk is heterogeneous even within higher-risk groups. To determine whether this heterogeneity could influence patient outcomes following PrEP, we undertook a post hoc prospective analysis of results from the CAPRISA 004 trial for 1% tenofovir gel (n = 774 patients), one of the first trials to demonstrate protection against HIV infection. Concentrations of nine proinflammatory cytokines were measured in cervicovaginal lavages at >2,000 visits, and a graduated cytokine score was used to define genital inflammation. In women without genital inflammation, tenofovir was 57% protective against HIV (95% confidence interval (CI): 7–80%) but was 3% protective (95% CI: −104–54%) if genital inflammation was present. Among women who highly adhered to the gel, tenofovir protection was 75% (95% CI: 25–92%) in women without inflammation compared to −10% (95% CI: −184–57%) in women with inflammation. Immunological predictors of HIV risk may modify the effectiveness of tools for HIV prevention; reducing genital inflammation in women may augment HIV prevention efforts.

Plasma Cytokine Predictors of Tuberculosis Recurrence in Antiretroviral-Treated Human Immunodeficiency Virus-infected Individuals from Durban, South Africa

Background Immune correlates of tuberculosis (TB) risk in populations infected with human immunodeficiency virus (HIV) remain understudied, despite HIV being associated with a high burden of TB disease. Here we describe plasma cytokine correlates of TB recurrence in a well-characterized cohort of HIV-infected individuals on antiretroviral therapy (ART) with a history of prior TB cure. Methods Study participants were drawn from a prospective cohort study initiated at the conclusion of a randomized clinical trial in which individuals presented with untreated HIV infection and active pulmonary TB. At baseline, ART was initiated, and TB successfully cured. Participants were screened for TB recurrence quarterly for up to 4 years. TB recurrent cases (n = 63) were matched to controls (n = 123) on sex, study arm assignment in the original trial, and month of enrollment with a subset of cases sampled longitudinally at several time-points. Results Three cytokines were associated with increased rates of TB recurrence in univariate models: interleukin 6 (IL6) (odds ratio [OR] 2.66, 95% confidence interval [CI] 1.34-5.28, P = .005), IP10 (OR 4.62, 95% CI 1.69-12.65, P = .003), monokine induced by IFN-γ (MIG) (OR 3.11, 95% CI 1.10-8.82, P = .034). Conversely, interferon β (IFNβ) was associated with decreased TB risk (OR 0.34, 95% CI 0.13-0.87, P = .025). Following multivariate analyses adjusting for covariates IL6, interleukin 1β (IL1β), and interleukin 1Rα (IL1Rα) were associated with increased risk and IFNβ with decreased TB risk. Longitudinal analysis showed that levels of many TB-associated markers, including IL6, IP10, sCD14, and interferon γ (IFNγ) are reduced following TB treatment. Conclusion These data show that TB recurrence, in HIV-infected individuals on ART is predicted by biomarkers of systemic inflammation, many of which are implicated in more rapid HIV disease progression. Clinical Trials Registration NCT 01539005.

Ex vivo HIV entry into blood CD4+ T cells does not predict heterosexual HIV acquisition in women

Background A blood-based assay that could quantify HIV susceptibility would be very valuable for HIV prevention research. Previously, we developed and validated an ex vivo, flow-based, HIV entry assay to assess genital HIV susceptibility in endocervical CD4+ T cells. Methods Here we assessed whether this tool could be used to predict HIV risk using blood-derived CD4+ T cells in a rigorously-blinded, nested case-control study using blood samples collected from high-risk, HIV-uninfected South African women enrolled in the CAPRISA 004 clinical trial. Cases, subsequently acquiring HIV were sampled prior to HIV infection and compared with controls, who remained HIV-uninfected. The primary endpoint was ex vivo entry of a CCR5-tropic HIV founder virus into blood CD4+ T cells. Secondary endpoints included HIV entry into CD4+ central (TCM) and effector (TEM) memory T cells, and into CD4+ T cell subsets expressing CCR5, CD69, CCR6, α4β1 or α4β7. Results Compared to bulk CD4+ T cells (4.9% virus entry), CD4+ T cells expressing CCR5, CCR6 or α4β1 and TEM were highly susceptible (15.5%, 8.8%, 8.2% and 10.8% entry, respectively, all p<0.0001), while TCM, CD69+ or α4β7+ CD4+ cells were moderately susceptible (6.4%, 6.0% and 5.8% respectively, p ≤ 0.003). While the proportion of the aforementioned highly susceptible cells correlated with overall virus entry into CD4+ T cells within an individual (r = 0.68, 0.47, 0.67, and 0.60 respectively, p<0.0001), blood virus entry did not predict subsequent mucosal HIV acquisition after controlling for sexual behaviour and condom use (OR 0.92, 95% CI 0.77–1.11, p = 0.40). Conclusions Although virus entry identified several previously known highly susceptible cellular HIV targets, blood HIV entry did not predict subsequent heterosexual HIV acquisition. Assessment of mucosal HIV susceptibility may require sampling at the site of HIV exposure.

Reducing IRF-1 to Levels Observed in HESN Subjects Limits HIV Replication, But Not the Extent of Host Immune Activation.

Cells from women who are epidemiologically deemed resistant to HIV infection exhibit a 40–60% reduction in endogenous IRF-1 (interferon regulatory factor-1), an essential regulator of host antiviral immunity and the early HIV replication. This study examined the functional consequences of reducing endogenous IRF-1 on HIV-1 replication and immune response to HIV in natural HIV target cells. IRF-1 knockdown was achieved in ex vivo CD4+ T cells and monocytes with siRNA. IRF-1 level was assessed using flow cytometry, prior to infection with HIV-Bal, HIV-IIIB, or HIV-VSV-G. Transactivation of HIV long terminal repeats was assessed by p24 secretion (ELISA) and Gag expression (reverse transcription–polymerase chain reaction (RT–PCR)). The expression of IRF-1–regulated antiviral genes was quantitated with RT–PCR. A modest 20–40% reduction in endogenous IRF-1 was achieved in >87% of ex vivo–derived peripheral CD4+ T cells and monocytes, resulted in >90% reduction in the transactivation of the HIV-1 genes (Gag, p24) and, hence, HIV replication. Curiously, these HIV-resistant women demonstrated normal immune responses, nor an increased susceptibility to other infection. Similarly, modest IRF-1 knockdown had limited impact on the magnitude of HIV-1–elicited activation of IRF-1–regulated host immunologic genes but resulted in lessened duration of these responses. These data suggest that early expression of HIV-1 genes requires a higher IRF-1 level, compared to the host antiviral genes. Together, these provide one key mechanism underlying the natural resistance against HIV infection and further suggest that modest IRF-1 reduction could effectively limit productive HIV infection yet remain sufficient to activate a robust but transient immune response.