Macrophage Migration Inhibitory Factor (MIF) Promotes Increased Proportions of the Highly Permissive Th17-like Cell Profile during HIV Infection.

In this study, we evaluate the role of the MIF/CD74 axis in the functionality of CD4+ T lymphocytes (CD4TL) during HIV infection. MDMs from healthy donors were infected with a R5-tropic or Transmitted/Founder (T/F) HIV strain. At day 11 post-MDM infection, allogeneic co-cultures with uninfected CD4TLs plus MIF stimulus were performed. Cytokine production was evaluated by ELISA. MIF plasma levels of people with HIV (PWH) were evaluated by ELISA. The phenotype and infection rate of CD4TLs from PWH were analyzed after MIF stimulus. Intracellular cytokines and transcription factors were evaluated by flow cytometry. Data were analyzed by parametric or non-parametric methods. The MIF stimulation of HIV-infected MDMs induced an increased expression of IL-6, IL-1ß and IL-8. In CD4TL/MDM co-cultures, the MIF treatment increased IL-17A/RORγt-expressing CD4TLs. Higher concentrations of IL-17A in supernatants were also observed. These results were recapitulated using transmitted/founder (T/F) HIV-1 strains. The MIF treatment appeared to affect memory CD4TLs more than naïve CD4TLs. MIF blocking showed a negative impact on IL17A+CD4TL proportions. Higher MIF concentrations in PWH-derived plasma were correlated with higher IL-17A+CD4TL percentages. Finally, MIF stimulation in PWH-derived PBMCs led to an increase in Th17-like population. MIF may contribute to viral pathogenesis by generating a microenvironment enriched in activating mediators and Th17-like CD4TLs, which are known to be highly susceptible to HIV-1 infection and relevant to viral persistence. These observations establish a basis for considering MIF as a possible therapeutic target.

Pre-cART Immune Parameters in People Living With HIV Might Help Predict CD8+ T-Cell Characteristics, Inflammation Levels, and Reservoir Composition After Effective cART.

BACKGROUND: Combined antiretroviral treatment (cART) for HIV infection is highly effective in controlling viral replication. However, it cannot achieve a sterilizing cure. Several strategies have been proposed to achieve a functional cure, some of them based on immune-mediated clearing of persistently infected cells. Here, we aimed at identifying factors related to CD8TC and CD4TC quality before cART initiation that associate with the persistence of CD8TC antiviral response after cART, inflammation levels, and the size of the viral reservoir. METHODS: Samples from 25 persons living with HIV were obtained before and after (15 months) cART initiation. Phenotype and functionality of bulk and HIV-specific T cells were assayed by flow cytometry ex vivo or after expansion in pre-cART or post-cART samples, respectively. Cell-Associated (CA) HIV DNA (total and integrated) and RNA (unspliced [US] and multiple spliced [MS]) were quantitated by real-time PCR on post-cART samples. Post-cART plasma levels of CXCL10 (IP-10), soluble CD14 (sCD14) and soluble CD163 (sCD163) were measured by ELISA. RESULTS: Pre-cART phenotype of CD8TCs and magnitude and phenotype of HIV-specific response correlated with the phenotype and functionality of CD8TCs post-cART. Moreover, the phenotype of the CD8TCs pre-cART correlated with markers of HIV persistence and inflammation post-cART. Finally, exhaustion and differentiation of CD4TCs pre-cART were associated with the composition of the HIV reservoir post-cART and the level of inflammation. CONCLUSIONS: Overall, this work provides data to help understand and identify parameters that could be used as markers in the development of immune-based functional HIV cure strategies.

Hepatitis C Virus (HCV) Clearance After Treatment With Direct-Acting Antivirals in Human Immunodeficiency Virus (HIV)-HCV Coinfection Modulates Systemic Immune Activation and HIV Transcription on Antiretroviral Therapy.

BACKGROUND: Hepatitis C virus (HCV) coinfection among people with human immunodeficiency virus (HIV) might perturb immune function and HIV persistence. We aimed to evaluate the impact of HCV clearance with direct-acting antivirals (DAAs) on immune activation and HIV persistence in HIV/HCV-coinfected individuals on antiretroviral therapy (ART). METHODS: In a prospective observational study, ART-treated participants with HIV/HCV coinfection received sofosbuvir/daclatasvir ±â€…ribavirin (n = 19). Blood samples were collected before DAA therapy, at the end of treatment, and 12 months after DAA termination (12MPT). T- and natural killer (NK)-cell phenotype, soluble plasma factors, cell-associated (CA)-HIV deoxyribonucleic acid (DNA) forms (total, integrated, 2LTR), CA-unspliced (US) and multiple-spliced ribonucleic acid (RNA), and plasma HIV RNA were evaluated. RESULTS: Hepatitis C virus clearance was associated with (1) a downmodulation of activation and exhaustion markers in CD4+, CD8+ T, and NK cells together with (2) decreased plasma levels of Interferon gamma-induced protein 10 (IP10), interleukin-8 (IL-8), soluble (s)CD163 and soluble intercellular adhesion molecule (sICAM). Cell-associated US HIV RNA was significantly higher at 12MPT compared to baseline, with no change in HIV DNA or plasma RNA. CONCLUSIONS: Elimination of HCV in HIV/HCV-coinfected individuals alters immune function and the transcriptional activity of latently infected cells. This report provides insights into the effects of HCV coinfection in HIV persistence and regards coinfected subjects as a population in which HIV remission might prove to be more challenging.

Inflamación sistémica y desregulación inmune: rol en la patogénesis de la coinfección VIH/VHC / Systemic inflammation and immune dysregulation: role in the pathogenesis of HIV / HCV coinfection

La disfunción inmune asociada a la infección por el virus de la inmunodeficiencia humana (VIH) es generada por una estimulación crónica del sistema inmune secundaria a la imposibilidad del organismo de erradicar el virus. La misma se encuentra exacerbada en el contexto de la coinfección por el virus de la hepatitis C (VHC). La inflamación sistémica producto de la coinfección por ambos virus genera un aumento de la morbilidad y mortalidad en los individuos afectados. Son varios los mediadores solubles de activación inmunológica, como IP-10, TNF-α, IL-6, IL-1ß (marcadores de inflamación sistémica); IL-17 (linfocitos T CD4+ Th17); IL-2, IFN-γ (linfocitos T CD4+ Th1); IL-8 (inducción de neutrofilia); CD23s, ICAMs, CD14s, CD163s (marcadores de activación de monocitos/macrófagos), niveles circulantes de lipopolisacárido (LPS) (translocación bacteriana); entre otros. Actualmente se necesitan más estudios para lograr definir cuáles serían los biomarcadores de progresión óptimos para el seguimiento de los individuos coinfectados por VIH/VHC. El objetivo de esta revisión es realizar una reseña sobre los mecanismos inmunopatológicos de la infección por VIH/VHC involucrados en la inflamación, daño hepático y su impacto en la morbimortalidad de los individuos coinfectados

The immune dysfunction associated with Human Immunodeficiency Virus (HIV) infection is generated by a chronic stimulation of the immune system, because of the inability to eradicate the virus from the host. This immune dysfunction is exacerbated in the context of coinfection with Hepatitis C Virus (HCV). Systemic inflammation caused by coinfection with both viruses generates an increase in morbidity and mortality in affected individuals. There are several soluble mediators of immunological activation, such as IP-10, TNF-α, IL-6, IL-1ß (systemic inflammation markers); IL-17 (CD4+ T cells Th17); IL-2, IFN-γ (CD4+ T cells Th1); IL-8 (neutrophilia); CD23s, ICAMs, CD14s, CD163s, lipopolysaccharide (LPS) (monocyte/macrophage activation markers and bacterial translocation); among others. Currently, more studies are needed to define optimal progression biomarkers for the follow-up of HIV/HCV coinfected individuals. In this review, we focus on the immunopathological mechanisms of HIV/HCV infection involved in inflammation, liver damage and its impact on the morbidity and mortality of affected individuals

Aerobic Exercise Training Prevents Perivascular Adipose Tissue-Induced Endothelial Dysfunction in Thoracic Aorta of Obese Mice.

Background: The mechanisms underlying the perivascular adipose tissue (PVAT) dysfunction in obesity are closely related to inflammation and oxidative stress. The present study aimed to investigate the effects of aerobic exercise training on PVAT-induced endothelial dysfunction of thoracic aorta of obese mice. Methods: Male mice C57BL6/JUnib (6-7 weeks) were divided into: sedentary (c-SD), trained (c-TR), obese sedentary (o-SD), and obese trained (o-TR). Obesity was induced by 16 weeks of high-fat diet and exercise training of moderate intensity started after 8 weeks of protocol and was performed on a treadmill, 5 days/week, for more 8 weeks, 60 min per session. The vascular responsiveness was performed in thoracic aorta in the absence (PVAT-) or in the presence (PVAT+) of PVAT. We analyzed circulatory parameters, protein expression, vascular nitric oxide (NO) production, and reactive oxygen species (ROS) in PVAT. Results: The maximal responses to acetylcholine (ACh) were reduced in PVAT+ compared with PVAT- rings in the o-SD group, accompanied by an increase in circulating glucose, insulin, resistin, leptin, and TNF-α. Additionally, the protein expression of iNOS and generation of ROS were increased in PVAT and production of vascular NO was reduced in the o-SD group compared with c-SD. In the o-TR group, the relaxation response to ACh was completely restored and the circulatory TNF-α, iNOS protein expression, and ROS were normalized with increased expression of Mn-SOD in PVAT, resulting in enhanced vascular NO production. Conclusion: The PVAT-induced endothelial dysfunction in thoracic aorta of obese mice, associated with circulatory inflammation and oxidative stress. Aerobic exercise training upregulated the anti-oxidant expression and decreased PVAT oxidative stress with beneficial impact on endothelium-dependent relaxation.

PD-1/PD-L1 Pathway Modulates Macrophage Susceptibility to Mycobacterium tuberculosis Specific CD8+ T cell Induced Death.

CD8+T cells contribute to tuberculosis (TB) infection control by inducing death of infected macrophages. Mycobacterium tuberculosis (Mtb) infection is associated with increased PD-1/PD-L1 expression and alternative activation of macrophages. We aimed to study the role of PD-1 pathway and macrophage polarization on Mtb-specific CD8+T cell-induced macrophage death. We observed that both PD-L1 on CD14+ cells and PD-1 on CD8+T cells were highly expressed at the site of infection in pleurisy TB patients' effusion samples (PEMC). Moreover, a significant increase in CD8+T cells' Mtb-specific degranulation from TB-PEMC vs. TB-PBMC was observed, which correlated with PD-1 and PDL-1 expression. In an in vitro model, M1 macrophages were more susceptible to Mtb-specific CD8+T cells' cytotoxicity compared to M2a macrophages and involved the transfer of cytolytic effector molecules from CD8+T lymphocytes to target cells. Additionally, PD-L1 blocking significantly increased the in vitro Ag-specific CD8+T cell cytotoxicity against IFN-γ-activated macrophages but had no effect over cytotoxicity on IL-4 or IL-10-activated macrophages. Interestingly, PD-L1 blocking enhanced Mtb-specific CD8+ T cell killing of CD14+ cells from human tuberculous pleural effusion samples. Our data indicate that PD-1/PD-L1 pathway modulates antigen-specific cytotoxicity against M1 targets in-vitro and encourage the exploration of checkpoint blockade as new adjuvant for TB therapies.

PD-1 Expression in HIV-Specific CD8+ T cells Before Antiretroviral Therapy Is Associated With HIV Persistence.

BACKGROUND: The persistence of latently infected T cells remains the principal barrier to HIV cure. Understanding how the early immune responses shape persistence of HIV on antiretroviral therapy (ART) will be fundamental for potential eradication. Here, we aimed to determine the relationship between CD8 T-cell function and phenotype before therapy and HIV persistence on ART. METHODS: Blood samples from 29 individuals enrolled during primary HIV infection (at baseline and every 3 months up to 2 years post-ART initiation) were obtained. HIV-specific T-cell function and expression of the activation markers were evaluated before ART by flow cytometry. Cell-associated HIV DNA and unspliced (US)-RNA were quantified in purified CD4 T cells by real-time polymerase chain reaction. Data were analyzed using nonparametric statistics. RESULTS: Elevated immune activation, dominance of monofunctional CD8 T cells, and skewed distribution of memory profile were observed before ART. After ART initiation, HIV DNA and US-RNA levels rapidly diminished, reaching a plateau by 30 weeks after ART. The proportion of baseline HIV-specific effector memory and terminal effector CD8 T cells directly correlated with HIV DNA levels at 1 year after ART. A strong positive correlation was observed between the proportion of bulk and HIV-specific PD-1 CD8 T cells measured before ART and HIV DNA at 1 year after ART. CONCLUSIONS: A higher proportion of terminally differentiated CD8 T cells and increased PD1 expression were associated with HIV persistence on ART after treatment of primary infection. Thus, the quality of the early CD8 T-cell immune response may serve as a predictor of HIV persistence on ART.

Phenotype, Polyfunctionality, and Antiviral Activity of in vitro Stimulated CD8+ T-Cells From HIV+ Subjects Who Initiated cART at Different Time-Points After Acute Infection.

Since anti-HIV treatment cannot cure the infection, many strategies have been proposed to eradicate the viral reservoir, which still remains as a major challenge. The success of some of these strategies will rely on the ability of HIV-specific CD8+ T-cells (CD8TC) to clear reactivated infected cells. Here, we aimed to investigate the phenotype and function of in vitro expanded CD8TC obtained from HIV+ subjects on combination antiretroviral therapy (cART), either initiated earlier (median = 3 months postinfection, ET: Early treatment) or later (median = 20 months postinfection, DT: Delayed treatment) after infection. Peripheral blood mononuclear cells from 12 DT and 13 ET subjects were obtained and stimulated with Nef and Gag peptide pools plus IL-2 for 14 days. ELISPOT was performed pre- and post-expansion. CD8TC memory/effector phenotype, PD-1 expression, polyfunctionality (CD107a/b, IFN-γ, IL-2, CCL4 (MIP-1ß), and/or TNF-α production) and antiviral activity were evaluated post-expansion. Magnitude of ELISPOT responses increased after expansion by 103 times, in both groups. Expanded cells were highly polyfunctional, regardless of time of cART initiation. The memory/effector phenotype distribution was sharply skewed toward an effector phenotype after expansion in both groups although ET subjects showed significantly higher proportions of stem-cell and central memory CD8TCs. PD-1 expression was clustered in HIV-specific effector memory CD8TCs, subset that also showed the highest proportion of cytokine-producing cells. Moreover, PD-1 expression directly correlated with CD8TC functionality. Expanded CD8TCs from DT and ET subjects were highly capable of mediating antiviral activity, measured by two different assays. Antiviral function directly correlated with the proportion of fully differentiated effector cells (viral inhibition assay) as well as with CD8TC polyfunctionality and PD-1 expression (VITAL assay). In sum, we show that, despite being dampened in subjects on cART, the HIV-specific CD8TC response could be selectively stimulated and expanded in vitro, presenting a high proportion of cells able to carry-out multiple effector functions. Timing of cART initiation had an impact on the memory/effector differentiation phenotype, most likely reflecting how different periods of antigen persistence affected immune function. Overall, these results have important implications for the design and evaluation of strategies aimed at modulating CD8TCs to achieve the HIV functional cure.

Interaction Between Macrophage Migration Inhibitory Factor and CD74 in Human Immunodeficiency Virus Type I Infected Primary Monocyte-Derived Macrophages Triggers the Production of Proinflammatory Mediators and Enhances Infection of Unactivated CD4+ T Cells.

Understanding the mechanisms of human immunodeficiency virus type I (HIV-1) pathogenesis would facilitate the identification of new therapeutic targets to control the infection in face of current antiretroviral therapy limitations. CD74 membrane expression is upregulated in HIV-1-infected cells and the magnitude of its modulation correlates with immune hyperactivation in HIV-infected individuals. In addition, plasma level of the CD74 activating ligand macrophage migration inhibitory factor (MIF) is increased in infected subjects. However, the role played by MIF/CD74 interaction in HIV pathogenesis remains unexplored. Here, we studied the effect of MIF/CD74 interaction on primary HIV-infected monocyte-derived macrophages (MDMs) and its implications for HIV immunopathogenesis. Confocal immunofluorescence analysis of CD74 and CD44 (the MIF signal transduction co-receptor) expression indicated that both molecules colocalized at the plasma membrane specifically in wild-type HIV-infected MDMs. Treatment of infected MDMs with MIF resulted in an MIF-dependent increase in TLR4 expression. Similarly, there was a dose-dependent increase in the production of IL-6, IL-8, TNFα, IL-1ß, and sICAM compared to the no-MIF condition, specifically from infected MDMs. Importantly, the effect observed on IL-6, IL-8, TNFα, and IL-1ß was abrogated by impeding MIF interaction with CD74. Moreover, the use of a neutralizing αMIF antibody or an MIF antagonist reverted these effects, supporting the specificity of the results. Treatment of unactivated CD4+ T-cells with MIF-treated HIV-infected MDM-derived culture supernatants led to enhanced permissiveness to HIV-1 infection. This effect was lost when CD4+ T-cells were treated with supernatants derived from infected MDMs in which CD74/MIF interaction had been blocked. Moreover, the enhanced permissiveness of unactivated CD4+ T-cells was recapitulated by exogenous addition of IL-6, IL-8, IL-1ß, and TNFα, or abrogated by neutralizing its biological activity using specific antibodies. Results obtained with BAL and NL4-3 HIV laboratory strains were reproduced using transmitted/founder primary isolates. This evidence indicated that MIF/CD74 interaction resulted in a higher production of proinflammatory cytokines from HIV-infected MDMs. This caused the generation of an inflammatory microenvironment which predisposed unactivated CD4+ T-cells to HIV-1 infection, which might contribute to viral spreading and reservoir seeding. Overall, these results support a novel role of the MIF/CD74 axis in HIV pathogenesis that deserves further investigation.

Biomarkers of Progression after HIV Acute/Early Infection: Nothing Compares to CD4⁺ T-cell Count?

Progression of HIV infection is variable among individuals, and definition disease progression biomarkers is still needed. Here, we aimed to categorize the predictive potential of several variables using feature selection methods and decision trees. A total of seventy-five treatment-naïve subjects were enrolled during acute/early HIV infection. CD4⁺ T-cell counts (CD4TC) and viral load (VL) levels were determined at enrollment and for one year. Immune activation, HIV-specific immune response, Human Leukocyte Antigen (HLA) and C-C chemokine receptor type 5 (CCR5) genotypes, and plasma levels of 39 cytokines were determined. Data were analyzed by machine learning and non-parametric methods. Variable hierarchization was performed by Weka correlation-based feature selection and J48 decision tree. Plasma interleukin (IL)-10, interferon gamma-induced protein (IP)-10, soluble IL-2 receptor alpha (sIL-2Rα) and tumor necrosis factor alpha (TNF-α) levels correlated directly with baseline VL, whereas IL-2, TNF-α, fibroblast growth factor (FGF)-2 and macrophage inflammatory protein (MIP)-1ß correlated directly with CD4⁺ T-cell activation (p < 0.05). However, none of these cytokines had good predictive values to distinguish "progressors" from "non-progressors". Similarly, immune activation, HIV-specific immune responses and HLA/CCR5 genotypes had low discrimination power. Baseline CD4TC was the most potent discerning variable with a cut-off of 438 cells/µL (accuracy = 0.93, κ-Cohen = 0.85). Limited discerning power of the other factors might be related to frequency, variability and/or sampling time. Future studies based on decision trees to identify biomarkers of post-treatment control are warrantied.

Evaluation of Different Parameters of Humoral and Cellular Immune Responses in HIV Serodiscordant Heterosexual Couples: Humoral Response Potentially Implicated in Modulating Transmission Rates.

As the HIV/AIDS pandemic still progresses, understanding the mechanisms governing viral transmission as well as protection from HIV acquisition is fundamental. In this context, cohorts of HIV serodiscordant heterosexual couples (SDC) represent a unique tool. The present study was aimed to evaluate specific parameters of innate, cellular and humoral immune responses in SDC. Specifically, plasma levels of cytokines and chemokines, HIV-specific T-cell responses, gp120-specific IgG and IgA antibodies, and HIV-specific antibody-dependent cellular cytotoxicity (ADCC) activity were assessed in nine HIV-exposed seronegative individuals (ESN) and their corresponding HIV seropositive partners (HIV+-P), in eighteen chronically infected HIV subjects (C), nine chronically infected subjects known to be HIV transmitters (CT) and ten healthy HIV- donors (HD). Very low magnitude HIV-specific cellular responses were found in two out of six ESN. Interestingly, HIV+-P had the highest ADCC magnitude, the lowest IgA levels and the highest IgG/IgA ratio, all compared to CT. Positive correlations between CD4+ T-cell counts and both IgG/IgA ratios and %ADCC killing uniquely distinguished HIV+-P. Additionally, evidence of IgA interference with ADCC responses from HIV+-P and CT is provided. These data suggest for the first time a potential role of ADCC and/or gp120-specific IgG/IgA balance in modulating heterosexual transmission. In sum, this study provides key information to understand the host factors that influence viral transmission, which should be considered in both the development of prophylactic vaccines and novel immunotherapies for HIV-1 infection.

Modification of the HIV-specific CD8+ T-cell response in an HIV elite controller after chikungunya virus infection.

OBJECTIVE: To evaluate the impact of chikungunya virus (CHIKV) infection on the quality of the HIV-specific CD8 T-cell (CTL) response in an HIV elite controller. DESIGN: Three blood samples were obtained from an elite controller at 27 days (EC-CHIKV, Sample 1, S1), 41 days (S2) and 1 year (S3) after CHIKV infection. Additionally, samples from another nine elite controllers and nine viremic chronics were obtained. METHODS: CD4 T-cell counts, viral load and immune activation were recorded. Natural killer (NK) cells and HIV-specific CTL quality were evaluated. Data were analyzed using nonparametric statistics. RESULTS: A male HIV elite controller was confirmed for CHIKV infection. At S1, he presented 211 cells/µl CD4 T-cell count, a HIV viral load blip (145 copies/ml) and high T-cell activation. NK cell percentage and activation were higher at S2. All parameters were recovered by S3. CTLs at S1 were exclusively monofunctional with a high proportion (>80%) of degranulating CTLs. By S3, CTL polyfunctionality was more similar to that of a typical elite controller. The distribution of CTL memory subsets also displayed altered profiles. CONCLUSION: The results showed that the phenotype and function of HIV-specific CTLs were modified in temporal association with an HIV viral load blip that followed CHIKV infection. This might have helped to control the transient HIV rebound. Additionally, NK cells could have been involved in this control. These results provide useful information to help understand how elite controllers maintain their status, control HIV infection and alert about the negative impact to the immune function of HIV-infected individuals living in CHIKV endemic areas.