Immune Activation Is Associated with CD8 T Cell Interleukin-21 Production in HIV-1-Infected Individuals

Interleukin-21 (IL-21) can be produced by CD8 T cells from HIV-1-infected individuals and those with autoimmune disease, but the mechanism remains poorly understood. Here we demonstrate that IL-21-producing CD8 T cells are not associated with CD4 depletion and are absent in patients with idiopathic CD4 lymphocytopenia. Instead, IL-21 production by CD8 T cells was associated with high levels of activation, suggesting that these cells emerge as a consequence of excessive chronic immune activation rather than CD4 lymphopenia.     

Detection of HIV-Gag p24-Specific Antibodies in Sera and Saliva of HIV-1-Infected Adults and in Sera of Infants Born to HIV-1-Infected Mothers

Secretory immunoglobulin A (IgA) is known to play an important role in the mucosal defense against a variety of pathogens. Although the role of IgA antibodies during sexual transmission of HIV is not clear, HIV-specific IgA antibodies have been detected in various mucosal secretions of HIV-infected individuals. Using a monoclonal antibody against human IgA, we established an ELISA system to detect anti-HIV p24 IgA antibodies in sera and saliva. We have analyzed the levels of anti-HIV p24 IgG and IgA antibodies in sera and saliva of 107 and 119 adults, respectively, with HIV infection at different clinical stages, and in the sera of 13 infants born to HIV-infected mothers. The level of anti-HIV p24 IgA antibodies was lower in sera and higher in saliva as compared to that of anti-HIV p24 IgG antibodies. Where the percentage of HIV-specific serum antibody-positive cases decreased with disease progression, that of saliva antibody-positive cases increased in AIDS patients. Among the 13 infants born to HIV-infected mothers, 7 infants were HIV-p24-specific serum IgA positive. These sera were negative for anti-HIV p24 secretory IgA, suggesting that some infants develop their own immune responses against HIV infection. Thus, the detection of HIV-specific IgA antibodies, especially in saliva, could be a simple and reliable test for the diagnosis of HIV infection.     

Elimination of HIV-1-Infected Primary T Cell Reservoirs in an In Vitro Model of Latency

Establishment of long-lived cellular reservoirs of HIV-1 represents a major therapeutic challenge to virus eradication. In this study, we utilized a human primary cell model of HIV-1 latency to evaluate the requirements for efficient virus reactivation from, and the selective elimination of, latently infected human T cells. Ectopic expression of BCL2 supported the replication and spread of R5-tropic HIV-1 in activated CD4⁺ T cells. After IL-2 withdrawal, the HIV-1-infected T cells survived as resting cells for several months. Unexpectedly, these resting T cells continue to produce detectable levels of infectious virus, albeit at a lower frequency than cells maintained in IL-2. In the presence of HIV-1 inhibitors, reactivation of the resting T cells with γc-cytokines and allogeneic dendritic cells completely extinguished HIV-1 infectivity. We also evaluated the ability of the bacterial LukED cytotoxin to target and kill CCR5-expressing cells. After γc-cytokine stimulation, LukED treatment eliminated both HIV-1-infected resting cells and the non-infected CCR5⁺ cells. Importantly, complete clearance of in vitro HIV-1 reservoirs by LukED required a lower threshold of cytokine signals relative to HIV-1 inhibitors. Thus, the primary T cell-based HIV-1 latency model could facilitate the development of novel agents and therapeutic strategies that could effectively eradicate HIV-1.     

Regulatory T Cell Suppression of Gag-Specific CD8+ T Cell Polyfunctional Response After Therapeutic Vaccination of HIV-1-Infected Patients on ART

We tested the hypothesis that therapeutic vaccination against HIV-1 can increase the frequency and suppressive function of regulatory, CD4⁺ T cells (Treg), thereby masking enhancement of HIV-1-specific CD8⁺ T cell response. HIV-1-infected subjects on antiretroviral therapy (N = 17) enrolled in a phase I therapeutic vaccine trial received 2 doses of autologous dendritic cells (DC) loaded with HIV-1 peptides. The frequency of CD4⁺CD25^hiFOXP3⁺ Treg in blood was determined prior to and after vaccination in subjects and normal controls. Polyfunctional CD8⁺ T cell responses were determined pre- and post-vaccine (N = 7) for 5 immune mediators after in vitro stimulation with Gag peptide, staphylococcal enterotoxin B (SEB), or medium alone. Total vaccine response (post-vaccine–pre-vaccine) was compared in the Treg(+) and Treg-depleted (Treg-) sets. After vaccination, 12/17 subjects showed a trend of increased Treg frequency (P = 0.06) from 0.74% to 1.2%. The increased frequency did not correlate with CD8⁺ T cell vaccine response by enzyme linked immunosorbent assay for interferon γ production. Although there was no significant change in CD8⁺ T cell polyfunctional response after vaccination, Treg depletion increased the polyfunctionality of the total vaccine response (P = 0.029), with a >2-fold increase in the percentage of CD8⁺ T cells producing multiple immune mediators. In contrast, depletion of Treg did not enhance polyfunctional T cell response to SEB, implying specificity of suppression to HIV-1 Gag. Therapeutic immunization with a DC-based vaccine against HIV-1 caused a modest increase in Treg frequency and a significant increase in HIV-1-specific, Treg suppressive function. The Treg suppressive effect masked an increase in the vaccine-induced anti-HIV-1-specific polyfunctional response. The role of Treg should be considered in immunotherapeutic trials of HIV-1 infection.     

Analysis of Memory B Cell Responses and Isolation of Novel Monoclonal Antibodies with Neutralizing Breadth from HIV-1-Infected Individuals

Background

The isolation of human monoclonal antibodies (mAbs) that neutralize a broad spectrum of primary HIV-1 isolates and the characterization of the human neutralizing antibody B cell response to HIV-1 infection are important goals that are central to the design of an effective antibody-based vaccine.

Methods and Findings

We immortalized IgG⁺ memory B cells from individuals infected with diverse clades of HIV-1 and selected on the basis of plasma neutralization profiles that were cross-clade and relatively potent. Culture supernatants were screened using various recombinant forms of the envelope glycoproteins (Env) in multiple parallel assays. We isolated 58 mAbs that were mapped to different Env surfaces, most of which showed neutralizing activity. One mAb in particular (HJ16) specific for a novel epitope proximal to the CD4 binding site on gp120 selectively neutralized a multi-clade panel of Tier-2 HIV-1 pseudoviruses, and demonstrated reactivity that was comparable in breadth, but distinct in neutralization specificity, to that of the other CD4 binding site-specific neutralizing mAb b12. A second mAb (HGN194) bound a conserved epitope in the V3 crown and neutralized all Tier-1 and a proportion of Tier-2 pseudoviruses tested, irrespective of clade. A third mAb (HK20) with broad neutralizing activity, particularly as a Fab fragment, recognized a highly conserved epitope in the HR-1 region of gp41, but showed striking assay-dependent selectivity in its activity.

Conclusions

This study reveals that by using appropriate screening methods, a large proportion of memory B cells can be isolated that produce mAbs with HIV-1 neutralizing activity. Three of these mAbs show unusual breadth of neutralization and therefore add to the current panel of HIV-1 neutralizing antibodies with potential for passive protection and template-based vaccine design.     

Expansion of Monocytic Myeloid-Derived Suppressor Cells Dampens T Cell Function in HIV-1-Seropositive Individuals

T lymphocyte dysfunction contributes to human immunodeficiency virus type 1 (HIV-1) disease progression by impairing antivirus cellular immunity. However, the mechanisms of HIV-1 infection-mediated T cell dysfunction are not completely understood. Here, we provide evidence that expansion of monocytic myeloid-derived suppressor cells (M-MDSCs) suppressed T cell function in HIV-1-infected individuals. We observed a dramatic elevation of M-MDSCs (HLA-DR^−/low CD11b⁺ CD33^+/high CD14⁺ CD15⁻ cells) in the peripheral blood of HIV-1-seropositive subjects (n = 61) compared with healthy controls (n = 51), despite efficacious antiretroviral therapy for nearly 2 years. The elevated M-MDSC frequency in HIV-1⁺ subjects correlated with prognostic HIV-1 disease markers, including the HIV-1 load (r = 0.5957; P < 0.0001), CD4⁺ T cell loss (r = −0.5312; P < 0.0001), and activated T cells (r = 0.4421; P = 0.0004). Functional studies showed that M-MDSCs from HIV-1⁺ subjects suppressed T cell responses in both HIV-1-specific and antigen-nonspecific manners; this effect was dependent on the induction of arginase 1 and required direct cell-cell contact. Further investigations revealed that direct HIV-1 infection or culture with HIV-1-derived Tat protein significantly enhanced human MDSC generation in vitro, and MDSCs from healthy donors could be directly infected by HIV-1 to facilitate HIV-1 replication and transmission, indicating that a positive-feedback loop between HIV-1 infection and MDSC expansion existed. In summary, our studies revealed a novel mechanism of T cell dysfunction in HIV-1-infected individuals and suggested that targeting MDSCs may be a promising strategy for HIV-1 immunotherapy.     

Emerging Variability in HIV-1 Genetics among Recently Infected Individuals in Yunnan,China

Background

Yunnan has the longest endured Human Immunodeficiency Virus-1 (HIV-1) epidemic in China, and the genetic diversity of HIV-1 constitutes an essential characteristic of molecular epidemiology in this region. To obtain a more comprehensive picture of the dynamic changes in Yunnan’s HIV-1 epidemic, a cross-sectional molecular epidemiological investigation was carried out among recently infected individuals.

Methodology/Principal Findings

We sequenced partial gag (HXB2∶781–1861) and env (HXB2∶7002–7541) genes from 308 plasma samples of recently infected patients. With phylogenetic analysis, 130 specimens generated interpretable genotyping data. We found that the circulating genotypes included: CRF08_BC (40.8%), unique recombinant forms (URFs, 27.7%), CRF01_AE (18.5%), CRF07_BC (9.2%), subtype B (2.3%) and C (1.5%). CRF08_BC was the most common genotype, and was predominant in both intravenous drug users (IDUs) and heterosexually transmitted populations. CRF08_BC and CRF07_BC still predominated in eastern Yunnan, but CRF08_BC showed increasing prevalence in western Yunnan. Strikingly, the URFs raised dramatically in most regions of Yunnan. Seven different types of URFs were detected from 12 prefectures, suggesting that complicated and frequent recombination is a salient feature of Yunnan’s HIV-1 epidemic. Among URFs, two BC clusters with distinctive recombination patterns might be potential new CRF_BCs. CRF01_AE was no longer confined to the prefectures bordering Myanmar, and had spread to the eastern part of Yunnan, especially the capital city of Kunming, with a large number of infections in the transient population. The ratios of the main genotypes showed no statistical differences between infected IDUs and heterosexually transmitted infections.

Conclusions/Significance

The changing patterns of the dominant HIV-1 genotypes in Yunnan indicate the complex evolving dynamic nature of the epidemic. Understanding new trends in molecular epidemiology of HIV-1 infection is critical for adjusting current prevention strategies and vaccine development in Yunnan.     

Durability and Effectiveness of Maraviroc-Containing Regimens in HIV-1-Infected Individuals with Virological Failure in Routine Clinical Practice

Introduction

Limited data are available on the durability and effectiveness of maraviroc in routine clinical practice. We assessed the durability of maraviroc-containing regimens during a 30-month period, as well as their immunovirological and clinical efficacy, according to viral tropism in treatment-experienced individuals with viral load (VL) >50 copies/ml in the French Hospital Database on HIV.

Methods

Virological success was defined as VL<50 copies/ml, immunological success as a confirmed increase of at least 100 CD4 cells/mm³ measured twice at least one month apart, and clinical failure as hospitalization for a non-AIDS event, an AIDS event, or death. Multivariable Cox regression models adjusted for potential confounders were used to assess the influence of viral tropism on durability, the immunovirological responses, and clinical outcome.

Results

356 individuals started maraviroc with VL>50 copies/ml of whom 223 harbored R5 viruses, 44 non-R5 viruses and 89 viruses of unknown tropism. Individuals with non-R5 viruses were more likely than individuals with R5 viruses to discontinue maraviroc (75% vs 34%, p<0.0001). At 30 months, the estimated rates of virological and immunological success were respectively 89% and 51% in individuals with R5 viruses and 48% and 23% in individuals with non-R5 viruses. In multivariable analysis, non-R5 viruses were associated with a lower likelihood of both virological success (hazard ratio (HR): 0.42; 95% confidence interval (CI), 0.25–0.70) and immunological success (HR: 0.37; 95% CI, 0.18–0.77). No difference in clinical outcome was found between individuals with R5 and non-R5 viruses. The effectiveness of maraviroc-containing regimens in individuals with unknown viral tropism was not significantly different from that in individuals with R5 viruses. A limitation of the study is the absence of genotypic susceptibility score.

Conclusion

In this observational study, maraviroc-containing regimens yielded high rates of viral suppression and immunological responses in individuals with R5 viruses in whom prior regimens had failed.     

Tumor Necrosis Factor Receptor Expression in HIV-1-Infected CD4+ T Cells

We investigated whether HIV-1 can regulate tumor necrosis factor receptor (TNFR) expression in SupT-1, a CD4 + T-cell line. The cells were infected with HIV-1 containing 1,000 cpm RT activity, as early as day 3 after infection and all along the culture the supernatant level of core protein p24 was >250 pg/ml, and on days 6 and 9 after infection, p24 was found in 10 % of the cells as determined by indirect immunofluorescence assay. The cells were growing without loss of viability. The study of TNFR expression was based on a microassay for measurement of binding of ¹²⁵I-TNFα to cells, in which free and cell-bound ligand separation was performed by centrifugation through oil. Scatchard analysis of TNFα binding on days 6 and 9 after infection revealed a 90 % increase in the expression of high-affinity membrane receptors in HIV + SupT-1 culture compared with uninfected cells (mean +/-S.D. = 501 +/-148.5 vs. 263 +/-77.8 receptors/cell, n = 9, P< 0.001) with no change in dissociation constants (mean +/? S.D. = 4.36 +/?1.06 vs. 4.00 +/?1.12 × 10^?10 m ).     

Analysis of Suppressor and Non-Suppressor FOXP3+ T Cells in HIV-1-Infected Patients

Recently, it was shown that peripheral blood FOXP3+CD4+ T cells are composed of three phenotypic and functionally distinct subpopulations. Two of them having in vitro suppressive effects were characterized as resting Treg cells (rTregs) and activated Treg cells (aTregs). A third subset, identified as FOXP3+ non-Tregs, does not display any suppressor activity and produce high levels of Th1 and Th17 cytokines upon stimulation. In the present study we focus on the characteristics of these three subsets of FOXP3+CD4+ T cells in untreated HIV-1-infected patients. We found that the absolute counts of rTregs, aTregs and FOXP3+ non-Tregs were reduced in HIV-1 patients compared with healthy donors. The relative frequency of rTregs and aTregs was similar in HIV-1 patients and healthy donors, while the frequency of FOXP3+ non-Tregs was significantly higher in HIV-1 patients, reaching a maximum in those patients with the lower values of CD4 counts. Contrasting with the observations made in FOXP3- CD4+ T cells, we did not find a negative correlation between the number of rTregs, aTregs or FOXP3+ non-Tregs and virus load. Studies performed with either whole PBMCs or sorted aTregs and FOXP3+ non-Tregs cells showed that these two populations of FOXP3+ T cells were highly permissive to HIV-1 infection. Upon infection, FOXP3+ non-Tregs markedly down-regulates its capacity to produce Th1 and Th17 cytokines, however, they retain the ability to produce substantial amounts of Th2 cytokines. This suggests that FOXP3+ non-Tregs might contribute to the polarization of CD4+ T cells into a Th2 profile, predictive of a poor outcome of HIV-1-infected patients.     

The Effect of Leflunomide on Cycling and Activation of T-Cells in HIV-1-Infected Participants

Background

The pathogenesis of immunodeficiency due to human immunodeficiency virus (HIV)-1 is incompletely understood, but immune activation is believed to play a central role. Immunomodulatory agents that decrease immune activation may be useful in the treatment of HIV-1 infection.

Methodology

A randomized, double blind, placebo-controlled pilot study of leflunomide for 28 days was performed in participants with HIV-1 infection who were not receiving antiretroviral therapy. Participants randomized to leflunomide were subsequently treated with cholestyramine until leflunomide levels were below detection limit.

Findings

Treatment with leflunomide was well tolerated with mostly low-grade adverse events. Leflunomide administration reduced cycling of CD4 T cells (by ex vivo bromodeoxyuridine uptake and Ki67 expression) and decreased expression of activation markers (HLA-DR/CD38 co-expression) on CD8 T cells in peripheral blood. In addition, decreased expression of HIV-1 co-receptors was observed in both CD4 and CD8 T cells in the leflunomide group. There were no significant changes in naïve and memory T cell subsets, apoptosis of T cells or markers of microbial translocation.

Conclusions

Leflunomide was effective in reducing immune activation in the setting of chronic HIV-1 infection suggesting that targeting immune activation with immunomodulatory agents may be a feasible strategy.

Trial Registration

ClinicalTrials.gov {"type":"clinical-trial","attrs":{"text":"NCT00101374","term_id":"NCT00101374"}}NCT00101374     

Psychiatric Disorders in HTLV-1-Infected Individuals with Bladder Symptoms

BackgroundPrevious studies have reported high rates of depression and anxiety in HTLV-1 infected individuals with the neurological disease and in the asymptomatic phase. No study has investigated the rates in individuals that already show bladder symptoms without severe neurological changes; that is, during the oligosymptomatic phase. The present study investigated patients in this intermediate form on the spectrum of the infection.Conclusion/SignificanceThe results suggest that individuals with overactive bladders need a more thorough assessment from the mental health perspective. These patients remain an understudied group regarding psychiatric comorbidities.     

Association of Neisseria gonorrhoeae OpaCEA with Dendritic Cells Suppresses Their Ability to Elicit an HIV-1-Specific T Cell Memory Response

Infection with Neisseria gonorrhoeae (N. gonorrhoeae) can trigger an intense local inflammatory response at the site of infection, yet there is little specific immune response or development of immune memory. Gonococcal surface epitopes are known to undergo antigenic variation; however, this is unlikely to explain the weak immune response to infection since individuals can be re-infected by the same serotype. Previous studies have demonstrated that the colony opacity-associated (Opa) proteins on the N. gonorrhoeae surface can bind human carcinoembryonic antigen-related cellular adhesion molecule 1 (CEACAM1) on CD4⁺ T cells to suppress T cell activation and proliferation. Interesting in this regard, N. gonorrhoeae infection is associated with impaired HIV-1 (human immunodeficiency virus type 1)-specific cytotoxic T-lymphocyte (CTL) responses and with transient increases in plasma viremia in HIV-1-infected patients, suggesting that N. gonorrhoeae may also subvert immune responses to co-pathogens. Since dendritic cells (DCs) are professional antigen presenting cells (APCs) that play a key role in the induction of an adaptive immune response, we investigated the effects of N. gonorrhoeae Opa proteins on human DC activation and function. While morphological changes reminiscent of DC maturation were evident upon N. gonorrhoeae infection, we observed a marked downregulation of DC maturation marker CD83 when the gonococci expressing CEACAM1-specific Opa_CEA, but not other Opa variants. Consistent with a gonococcal-induced defect in maturation, Opa_CEA binding to CEACAM1 reduced the DCs’ capacity to stimulate an allogeneic T cell proliferative response. Moreover, Opa_CEA-expressing N. gonorrhoeae showed the potential to impair DC-dependent development of specific adaptive immunity, since infection with Opa_CEA-positive gonococci suppressed the ability of DCs to stimulate HIV-1-specific memory CTL responses. These results reveal a novel mechanism to explain why infection of N. gonorrhoeae fails to trigger an effective specific immune response or develop immune memory, and may affect the potent synergy between gonorrhea and HIV-1 infection.