Browsing by Author "Turk, Gabriela"
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Item Acute HIV Seroconversion Presenting with Active Tuberculosis and Associated with High Levels of T-Regulatory Cells(2011) Sued, Omar; Quiroga, Maria F.; Socias, Maria E.; Turk, Gabriela; Salomon, Horacio; Cahn, PedroA patient with well-defined acute HIV infection who developed concomitant pulmonary tuberculosis during the retroviral acute syndrome is reported here. In this patient high levels of T-regulatory cells (Tregs) and a low proliferation response to M. tuberculosis were initially detected, which normalized throughout follow-up. This case calls for the consideration of tuberculosis in patients in the early stages of HIV, and emphasizes the need for further study of the potential causal relationship between Treg cells and the risk of TB reactivation in HIV patients.Item Acute retroviral syndrome and high baseline viral load are predictors of rapid HIV progression among untreated Argentinean seroconverters(2011) Socias, Maria E.; Sued, Omar; Laufer, Natalia; Lázaro, Maria E.; Mingrone, Hugo; Remondegui, Claudio; Figueroa, Maria Ines; Cesar, Carina; Gun, Ana; Turk, Gabriela; Bouzas, Maria B.; Kavasery, Rosanna; Krolewiecki, Alejandro J.; Perez, Hector; Salomon, Horacio; Pryluka, DamianBackground Diagnosis of primary HIV infection (PHI) has important clinical and public health implications. HAART initiation at this stage remains controversial. Methods Our objective was to identify predictors of disease progression among Argentinean seroconverters during the first year of infection, within a multicentre registry of PHI-patients diagnosed between 1997 and 2008. Cox regression was used to analyze predictors of progression (LT-CD4 < 350 cells/mm3, B, C events or death) at 12 months among untreated patients. Results Among 134 subjects, 74% presented with acute retroviral syndrome (ARS). Seven opportunistic infections (one death), nine B events, and 10 non-AIDS defining serious events were observed. Among the 92 untreated patients, 24 (26%) progressed at 12 months versus three (7%) in the treated group (p = 0.01). The 12-month progression rate among untreated patients with ARS was 34% (95% CI 22.5-46.3) versus 13% (95% CI 1.1-24.7) in asymptomatic patients (p = 0.04). In univariate analysis, ARS, baseline LT-CD4 < 350 cells/mm3, and baseline and six-month viral load (VL) > 100,000 copies/mL were associated with progression. In multivariate analysis, only ARS and baseline VL > 100,000 copies/mL remained independently associated; HR: 8.44 (95% CI 0.97-73.42) and 9.44 (95% CI 1.38-64.68), respectively. Conclusions In Argentina, PHI is associated with significant morbidity. HAART should be considered in PHI patients with ARS and high baseline VL to prevent disease progression.Item Bioinformatic analysis of post-transmission viral readaptation in Argentine patients with acute HIV-1 infection(2020-07) Damilano, G; Sued, Omar; Satorres, S; Ruiz, Maria; Ghiglione, Y; Guzman, F; Turk, Gabriela ; Quiroga, F; Cahn, Pedro; Salomon, Horacio; Dilernia, DarioDuring the acute phase of HIV-1 infection, a strong readaptation occurs in the viral population. Our objective was to analyze the post-transmission mutations associated with escape to the cytotoxic immune response and its relationship with the progression of the infection. In this study, a total of 17 patients were enrolled during acute/early primary HIV infection and 8 subjects that were the HIV positive partner resulting in 8 transmission pairs. Genotyping of the genetic polymorphisms of HLA class I A and B was performed using PCR-SSOP. Viral RNA extraction was from plasma. 570 single Gag-gene amplifications were obtained by limiting-dilution RT-PCR. Epitope prediction was performed with NetMHC CBS prediction server for the 19 HLA-A and single bondB alleles. Cytotoxic response prediction was performed by using the IEDB Analysis Resource. From our results, we deduce that the transmitted CTL / gag escape frequency in the founder virus was at least double compared to the post-transmission events. Additionally, by means of an algorithm that combines these frequencies, we observed that the founder viruses better adapted to the HLA A / B alleles of the recipient could contribute to a greater progression of the infection. Our results suggest that there is a large adaptation of HIV-1 to the HLA A / B alleles prevalent in our population. However, despite this adaptive advantage, the virus needs to make “readjustments” through new escape and compensatory mutations. Interestingly, according to our results, this readaptation could have a role in the progression of the infection.Item Biomarkers of Progression after HIV Acute/Early Infection: Nothing Compares to CD4+ T-cell Count?(2018-01-13) Turk, Gabriela; Ghiglione, Yanina; Hormanstorfer, Macarena; Laufer, Natalia; Coloccini, Romina; Salido, Jimena; Trifone, César; Ruiz, Maria; Falivene, Juliana; Holgado, María Pía; Caruso, María Paula; Figueroa, María Inés; Salomon, Horacio ; Giavedoni, Luis D; De los Ángeles Pando, María; Gherardi, María Magdalena; Rabinovich, Roberto Daniel; Pury, Pedro A; Sued, OmarProgression 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)-1β 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.Item Computational comparison of availability in CTL/gag epitopes among patients with acute and chronic HIV-1 infection(2016) Damilano, Gabriel Dario; Sued, Omar; Ruiz, Maria; Ghiglione, Yanina; Canitano, Flavia; Pando, Maria; Turk, Gabriela; Cahn, Pedro; Salomon, Horacio; Dilernia, DarioBackground Recent studies indicate that there is selection bias for transmission of viral polymorphisms associated with higher viral fitness. Furthermore, after transmission and before a specific immune response is mounted in the recipient, the virus undergoes a number of reversions which allow an increase in their replicative capacity. These aspects, and others, affect the viral population characteristic of early acute infection. Methods 160 single gag-gene amplifications were obtained by limiting-dilution RT-PCR from plasma samples of 8 ARV-naïve patients with early acute infection (<30 days, 22 days average) and 8 ARV-naive patients with approximately a year of infection (10 amplicons per patient). Sanger sequencing and NGS SMRT technology (Pacific Biosciences) were implemented to sequence the amplicons. Phylogenetic analysis was performed by using MEGA 6.06. HLA-I (A and B) typing was performed by SSOP-PCR method. The chromatograms were analyzed with Sequencher 4.10. Epitopes and immune-proteosomal cleavages prediction was performed with CBS prediction server for the 30 HLA-A and -B alleles most prevalent in our population with peptide lengths from 8 to 14 mer. Cytotoxic response prediction was performed by using IEDB Analysis Resource. Results After implementing epitope prediction analysis, we identified a total number of 325 possible viral epitopes present in two or more acute or chronic patients. 60.3% (n = 196) of them were present only in acute infection (prevalent acute epitopes) while 39.7% (n = 129) were present only in chronic infection (prevalent chronic epitopes). Within p24, the difference was equally dramatic with 59.4% (79/133) being acute epitopes (p < 0.05). This is consistent with progressive viral adaptation to immune response in time and further supported by the fact that cytotoxic responses prediction showed that acute epitopes are more likely to generate immune response than chronic epitopes. Interestingly, only 27.5% of acute epitopes match the population-level consensus sequence of the virus. Conclusions Our results indicate that certain non-consensus viral residues might be transmitted more frequently than consensus-residues when located in immunological relevant positions (epitopes). This observation might be relevant to the rationale behind development of an effective vaccine to reduce viral reservoir and induce functional cure of HIV infection based in prevalent acute epitopes. Introduction Following transmission, Cytotoxic CD8+T lymphocytes (CTLs) mount a powerful response to transmitted HIV in the acute phase of infection [1]. However, the vast majority of cases is an inefficient response and directed to a limited number of epitopes [2]. This response is manifested producing a viral set point generally after first month of infection [3], [4], [5] with great specificity on the gag protein, which is strongly associated with control of viral replication [6], [7], [8], [9]. At this point, just after the peak of viremia, the first viral populations with HLA/CTL escape mutations are generated [2], which will increase over time and will have an impact on viral diversity during the chronic phase [10], [11], [12]. Therefore, it is interesting to investigate if these HLA-dependent reversions that occur during the acute phase are associated with a greater availability of epitopes in this stage. Reversions are another important aspect to be considered among viral factors shaping the viral diversity during the acute stage of infection [13], [14]. While their impact on viral fitness on transmitted/founder virus has not yet been elucidated, it is believes reversal mutations make the virus more fit [13], [15], [16].These post-transmission reversions may range from those associated with HLA alleles present on the donor [17], to the CD8 T-cell TCR receptor [13], [18], [19], to the proteasome [20], [21], or reversions that arise at random and increase the viral fitness that end up being established in the major viral population [22], [24]. Also, these reversions could occur prior to transmission, in a process of viral compartmentalization in mucosal associated tissues [25], [26], [27]. In this research, our objective was to determine, in patients under study, whether the viral sequences approach phylogenetically in acute patients compared with chronic patients. In turn, we evaluated whether possible reversions in acute viral sequences could significantly influence to decrease viral phylogenetic distance in these patients. Also, using computational prediction analysis, we evaluated the relationship of phylogenetically relevant amino acid positions (possibly reversions), with epitopes associated with cytotoxic immune response mediated by HLA I. Finally, we evaluated the distribution and characteristics of CTL epitopes, in conserved viral segments. We did an availability and frequency analysis of possible CTL epitopes found in the early acute phase compared to those found in the chronic phase.Item Correction: HLA-Driven Convergence of HIV-1 Viral Subtypes B and F Toward the Adaptation to Immune Responses in Human Populations(2008-11-04) Dilernia, Dario; Jones, Leandro; Rodriguez, Sabrina; Turk, Gabriela; Rubio, Andrea E; Pampuro, Sandra; Gomez-Carrillo, Manuel; Bautista, Christian T; Deluchi, Gabriel; Benetucci, Jorge; Lasala, María Beatriz; Lourtau, Leonardo; Losso, Marcelo Horacio; Perez, Héctor; Cahn, Pedro; Salomon, HoracioThe eighth author's name was displayed incorrectly. It should be: Christian T. Bautista.Item Distribution of Bulk and HIV-specific CD8 + T Cell Memory Phenotypes during Acute/Early HIV Infection Is Related to Reduced Antiviral Activity(2014) Ghiglione, Yanina; Falivene, Juliana; Ruiz, Maria; Laufer, Natalia; Socias, Maria E.; Cahn, Pedro; Sued, Omar; Salomon, Horacio; Gherardi, María Magdalena; Turk, GabrielaBackground: Memory CD8+ T-cells are important components of protective immunity. Understanding their development during primary HIV infection (PHI) may contribute to optimal vaccine design. Aim: To analyze the distribution of memory subsets during PHI and their correlation with functionality and clinical parameters. Methods: 19 samples from acutely infected subjects were obtained at baseline and 12 months post-infection (mpi). Phenotypic (CD45RO, CCR7, PD-1) and functional markers (cytokines) were used to identify bulk and HIV-specific CD8+ memory populations. CD8 virus inhibitory assay (VIA) was performed. Data was compared intra-group and correlated to clinical parameters, PD-1 analysis and CD8 antiviral activity, using non-parametric statistics. Results: Bulk and HIV-specific CD8+ profile was terminal effectors (TE)>naïve>effector memory (TEM)>central memory. Spearman's correlation showed that baseline CD8+ VIA inversely correlated with the concurrent proportion of HIV-specific CD8+ TEM cells (r=-0.593, p=0.009) and directly correlated with the proportion of HIV-specific CD8+ TE cells (r=0.718, p=0.0008). Identical correlations were observed between baseline CD8+ T cell phenotype and CD8+ VIA at 12 mpi. Also, percentage of PD-1high CD8+ T cells negatively correlated with bulk and HIV-specific CD8+ TEM cells (r=−0.501, p=0.034 and r=−0.668, p=0.004, respectively). Conversely, positive correlations were observed with the proportion of bulk and HIV-specific CD8+ TE cells (r=-0.510, p=0.0308 and r=−0.564, p=0.022, respectively). Conclusions: A higher proportion of fully differentiated HIV-specific cells are related to the magnitude of CD8+ antiviral activity (rapidly able to exert effector functions) and to a higher PD-1 expression (related to T cell differentiation stage and activation status). This is the first report were a relation between CD8+ T cell memory differentiation hierarchy and antiviral function is reported during acute infection, providing information potentially useful for vaccine design.Item Early Gag Immunodominance of the HIV-Specific T-Cell Response during Acute/Early Infection Is Associated with Higher CD8(+) T-Cell Antiviral Activity and Correlates with Preservation of the CD4(+) T-Cell Compartment(2013) Turk, Gabriela; Ghiglione, Yanina; Falivene, Juliana; Socias, Maria E.; Laufer, Natalia; Coloccini, Romina.; Rodriguez, Ana María; Ruiz, Maria; Pando, María; Giavedoni, Luis; Cahn, Pedro; Sued, Omar; Salomon, Horacio; Gherardi, MaríaThe important role of the CD8+ T-cell response on HIV control is well established. Moreover, the acute phase of infection represents a proper scenario to delineate the antiviral cellular functions that best correlate with control. Here, multiple functional aspects (specificity, ex vivo viral inhibitory activity [VIA] and polyfunctionality) of the HIV-specific CD8+ T-cell subset arising early after infection, and their association with disease progression markers, were examined. Blood samples from 44 subjects recruited within 6 months from infection (primary HIV infection [PHI] group), 16 chronically infected subjects, 11 elite controllers (EC), and 10 healthy donors were obtained. Results indicated that, although Nef dominated the anti-HIV response during acute/early infection, a higher proportion of early anti-Gag T cells correlated with delayed progression. Polyfunctional HIV-specific CD8+ T cells were detected at early time points but did not associate with virus control. Conversely, higher CD4+ T-cell set points were observed in PHI subjects with higher HIV-specific CD8+ T-cell VIA at baseline. Importantly, VIA levels correlated with the magnitude of the anti-Gag cellular response. The advantage of Gag-specific cells may result from their enhanced ability to mediate lysis of infected cells (evidenced by a higher capacity to degranulate and to mediate VIA) and to simultaneously produce IFN-γ. Finally, Gag immunodominance was associated with elevated plasma levels of interleukin 2 (IL-2) and macrophage inflammatory protein 1β (MIP-1β). All together, this study underscores the importance of CD8+ T-cell specificity in the improved control of disease progression, which was related to the capacity of Gag-specific cells to mediate both lytic and nonlytic antiviral mechanisms at early time points postinfection.Item Early skewed differentiation and PD-1 expression in CD4+ cells relate to immune dysfunction and viral persistence in individuals living with HIV 1 year post-cART initiationSalido, Jorge; Czernikier, Ana; Trifone, Carolina; Figueroa, María Isabel; Salomon, Horacio; Cahn, Pedro; Sued, Omar; Laufer, Natalia; Ghiglione, Yanina; Turk, GabrielaAchieving HIV functional cure is a priority. Strategies such as adoptive cell transfer have been assayed, without success yet mainly due to immune dysfunctions observed among individuals. Samples from 25 HIV+ subjects were collected at diagnosis (baseline sample, BSL) and one year post-cART initiation (post- cART ). At BSL, bulk and HIV-specific CD4 phenotype (CD45RO , CCR 7, CD95 and PD1 expression) was assessed by flow cytometry after a short stimulation with HIV peptides. Also, proportion of CD4+/HLA-DR+/ CD38+ cells was measured. At post-cART , HIV-specific CD8TC s were obtained after 2-week expansion with peptides. Phenotype and antiviral activity (VIA and VITA L assays) were evaluated post-expansion. Plasma CXCL10 (IP-10) was assessed by ELISA. Cell-associated HIV DNA (total and integrated) and unspliced (US) and multiply-spliced (MS) RNA were quantified by real-time PCR. Non-parametric statistics were applied. Early CD4TC exhaustion, elevated activation and inadequate differentiation seem to be associated with viral persistence, inflammation, as well as with the phenotype and antiviral capacity of HIV-specific CD8TC s that persist one year after cART is initiated. These parameters could serve as predictors of CD8TC function on treated subjects.Item Early Skewed Distribution of Total and HIV-Specific CD8 T-Cell Memory Phenotypes during Primary HIV Infection Is Related to Reduced Antiviral Activity and Faster Disease Progression(2014-8) Ghiglione, Yanina; Falivene, Juliana; Ruiz, Maria; Laufer, Natalia; Socias, Maria E.; Cahn, Pedro; Giavedoni, Luis; Sued, Omar; Gherardi, María Magdalena; Salomon, Horacio; Turk, GabrielaThe important role of the CD8+ T-cells on HIV control is well established. However, correlates of immune protection remain elusive. Although the importance of CD8+ T-cell specificity and functionality in virus control has been underscored, further unraveling the link between CD8+ T-cell differentiation and viral control is needed. Here, an immunophenotypic analysis (in terms of memory markers and Programmed cell death 1 (PD-1) expression) of the CD8+ T-cell subset found in primary HIV infection (PHI) was performed. The aim was to seek for associations with functional properties of the CD8+ T-cell subsets, viral control and subsequent disease progression. Also, results were compared with samples from Chronics and Elite Controllers. It was found that normal maturation of total and HIV-specific CD8+ T-cells into memory subsets is skewed in PHI, but not at the dramatic level observed in Chronics. Within the HIV-specific compartment, this alteration was evidenced by an accumulation of effector memory CD8+ T (TEM) cells over fully differentiated terminal effector CD8+ T (TTE) cells. Furthermore, higher proportions of total and HIV-specific CD8+ TEM cells and higher HIV-specific TEM/(TEM+TTE) ratio correlated with markers of faster progression. Analysis of PD-1 expression on total and HIV-specific CD8+ T-cells from PHI subjects revealed not only an association with disease progression but also with skewed memory CD8+ T-cell differentiation. Most notably, significant direct correlations were obtained between the functional capacity of CD8+ T-cells to inhibit viral replication in vitro with higher proportions of fully-differentiated HIV-specific CD8+ TTE cells, both at baseline and at 12 months post-infection. Thus, a relationship between preservation of CD8+ T-cell differentiation pathway and cell functionality was established. This report presents evidence concerning the link among CD8+ T-cell function, phenotype and virus control, hence supporting the instauration of early interventions to prevent irreversible immune damage.Item Env-Specific IgA from Viremic HIV-Infected Subjects Compromises Antibody-Dependent Cellular Cytotoxicity(2016) Ruiz, Maria; Ghiglione, Yanina; Falivene, Juliana; Laufer, Natalia; Holgado, Maria Pia; Socias, Maria E.; Cahn, Pedro; Sued, Omar; Giavedoni, Luis; Salomon, Horacio; Gherardi, María Magdalena; Rodriguez, Ana María; Turk, GabrielaElucidating the factors that modulate HIV-specific antibody-dependent cellular cytotoxicity (ADCC) will help in understanding its role in HIV immunity. The aim of this study was to determine whether IgA could modify the magnitude of ADCC in HIV infection, abrogating its protective role. Plasma samples from 20 HIV-positive (HIV(+)) subjects enrolled during primary HIV infection (PHI), 10 chronically infected subjects (chronic), and 7 elite controllers (EC) were used. ADCC was determined by using a fluorometric ADCC assay, before and after removal of plasma IgA. Data were analyzed by using nonparametric statistics. ADCC was documented in 80% of PHI enrollment samples and in 100% of PHI 12-month, chronic, and EC samples; it peaked after acute infection, reached a plateau in chronic infection, and decreased after initiation of antiretroviral treatment (ART). Significant associations between ADCC and disease progression were found only after removal of plasma IgA from 12-month PHI samples: the magnitude of ADCC not only increased after IgA removal but also correlated with CD4(+) T-cell preservation. This work provides evidence that gp120-specific IgA was capable of modifying ADCC responses during natural HIV infection for the first time and adds to similar evidence provided in other settings. Furthermore, it underscores the complexity of the ADCC phenomenon and will help in an understanding of its underlying mechanisms. Importance: Although the induction of ADCC-mediating antibodies in HIV-infected subjects has been extensively documented, the association of these antibodies with protection from disease progression is poorly understood. Here, we demonstrate that plasma IgA is a factor capable of modifying the magnitude of IgG-mediated ADCC in HIV infection, mitigating its beneficial effect. These results help in understanding why previous studies failed to demonstrate correlations between ADCC and disease progression, and they also contribute to the notion that an HIV vaccine should stimulate the production of ADCC-mediating IgG antibodies but not IgA.Item Evaluation of Different Parameters of Humoral and Cellular Immune Responses in HIV Serodiscordant Heterosexual Couples: Humoral Response Potentially Implicated in Modulating Transmission Rates(2017-11-03) Ruiz, Maria; Salido, Jimena; Abusamra, Lorena; Ghiglione, Yanina; Cevallos, Cintia; Damilano, Gabriel; Rodriguez, Ana María; Trifone, César; Laufer, Natalia; Giavedoni, Luis D; Sued, Omar; Salomon, Horacio ; Gherardi, María Magdalena; Turk, GabrielaAs 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.Item Fostemsavir: a new CD4 attachment inhibitor(2018-10-23) Salido, Jimena; Ruiz, Maria; Trifone, César; Figueroa, María Inés; Caruso, María Paula; Gherardi, María Magdalena; Sued, Omar; Salomon, Horacio; Laufer, Natalia; Ghiglione, Yanina; Turk, GabrielaSince 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-1β), 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.Item Functionality of CD8+ T-cells in subjects under cART: implications on cure strategies(2018-08) Salido, J; Ruiz, Maria; Trifone, C; Figueroa, MI; Caruso, MP; Gherardi, MM; Sued, Omar; Horacio, S; Natalia, L; Ghiglione, Y; Turk, GabrielaBackground: Reaching HIV cure will largely depend on the capacity of HIV-specific memory CD8+ T-cells (CD8TC) to eliminate the viral reservoir. However, CD8TC response is limited in subjects on cART. Here, we aimed to investigate the phenotype and function of in vitro expanded CD8TCs in HIV+ subjects and the impact of ART initiation timing on these parameters. Methods & Materials: PBMCs from 28 HIV+ subjects on cART for 1 year were obtained. Twelve initiated treatment during chronic infection (Delayed Treatment, DT) and 16 within four months post-infection (Early Treatment, ET). PBMCs were stimulated with peptides spanning Nef and Gag plus IL-2 during 14 days. ELISPOT (pre and post-expansion) and Flow Cytometry (FC, post-expansion) were performed to assess expanded CD8TC function (CD107a/b, IFN-g, IL-2, MIP-1b and TNF-a) and phenotype (CD45RO, CCR7, CD95 and PD1). Data was analyzed using non-parametric statistics. Results: Magnitude of ELISPOT responses increased after expansion by 103 times (p < 0.002), in both groups, being this effect more pronounced in CD8TCs, compared to CD4TCs (p < 0.0001), as confirmed by FC. Cells showed higher avidity after stimulation (evidenced by greater spot sizes, p < 0.002). DT subjects displayed a broader response to HIV than ET, after expansion. ET group had a significantly higher proportion of monofunctional degranulating CD8TCs (CD107a/b+), when challenged against Gag peptides (p = 0.037) compared to DT. Contrary, DT group showed higher polyfunctionality (p = 0.009). In both groups, CD4TC responses were of lesser magnitude compared to CD8TCs and predominantly monofunctional. Bulk and HIV-specific CD8TC phenotype varied significantly between groups: ET subjects showed a preservation of stem and central memory cells while DT showed a fully-differentiated profile (p < 0.005). When analyzing memory distribution within PD1+CD8+ cells, terminal effector were the most frequent subpopulation in DT and effector memory cells in ET individuals; evidencing a differential cell exhaustion profile in both groups. Conclusion: We demonstrated that HIV-specific CD8TCs could be selectively stimulated and expanded in subjects under ART. We also showed that ART initiation timing has an impact on phenotype and function of CD8TCs, reflecting consequences of longer antigen persistence on immune function. Overall, results presented in this work have important implications for the development of cure strategies aim at boosting CD8TC responses.Item 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(2020-04-02) Ghiglione, Yanina; Polo, María Laura; Urioste, Alejandra; Rhodes, Ajantha; Czernikier, Alejandro; Trifone, César; Florencia Quiroga, María; Sisto, Alicia; Patterson, Patricia; Salomon, Horacio ; Rolón, María José; Bakkour, Sonia; Lewin, Sharon R; Turk, Gabriela; Laufer, Nataliahttps://doi.org/10.1093/ofid/ofaa115Item HIV-TB co-infection impairs CD8+ T-cell differentiation and function while dehydroepiandrosterone improves cytotoxic anti-tubercular immune responses(2015-9) Suarez, Guadalupe V.; Angerami, Matias; Vecchione, María B.; Laufer, Natalia; Turk, Gabriela; Ruiz, Maria; Mesch, Viviana; Fabre, Bibiana; Maidana, Patricia; Ameri, Diego; Cahn, Pedro; Sued, Omar; Salomon, Horacio; Bottasso, Oscar A.; Quiroga, María F.Tuberculosis (TB) is the leading cause of death among HIV-positive patients. The decreasing frequencies of terminal effector (TTE) CD8+T cells may increase reactivation risk in persons latently infected with Mycobacterium tuberculosis (Mtb). We have previously shown that dehydroepiandrosterone (DHEA) increases the protective antitubercular immune responses in HIV–TB patients. Here, we aimed to study Mtb-specific cytotoxicity, IFN-γ secretion, memory status of CD8+T cells, and their modulation by DHEA during HIV–TB coinfection. CD8+T cells from HIV–TB patients showed a more differentiated phenotype with diminished naïve and higher effector memory and TTE T-cell frequencies compared to healthy donors both in total and Mtb-specific CD8+T cells. Notably, CD8+T cells from HIV–TB patients displayed higher Terminal Effector (TTE) CD45RAdim proportions with lower CD45RA expression levels, suggesting a not fully differentiated phenotype. Also, PD-1 expression levels on CD8+T cells from HIV–TB patients increased although restricted to the CD27+ population. Interestingly, DHEA plasma levels positively correlated with TTE in CD8+T cells and in vitro DHEA treatment enhanced Mtb-specific cytotoxic responses and terminal differentiation in CD8+T cells from HIV–TB patients. Our data suggest that HIV–TB coinfection promotes a deficient CD8+ T-cell differentiation, whereas DHEA may contribute to improving antitubercular immunity by enhancing CD8+T-cell functions during HIV–TB coinfection.Item HLA-Driven Convergence of HIV-1 Viral Subtypes B and F Toward the Adaptation to Immune Responses in Human Populations(2008-10-21) Dilernia, Dario; Jones, Leandro; Rodriguez, Sabrina; Turk, Gabriela; Rubio, Andrea E; Pampuro, Sandra; Gomez-Carrillo, Manuel; Bautista, Christian; Deluchi, Gabriel; Benetucci, Jorge; Lasala, María Beatriz; Lourtau, Leonardo; Losso, Marcelo Horacio; Perez, Héctor; Cahn, Pedro; Salomon, HoracioFil: Cahn P. Fundación Huésped, Buenos Aires; ArgentinaItem Host Genetic Factors Associated with Symptomatic Primary HIV Infection and Disease Progression among Argentinean Seroconverters(2014) Coloccini, Romina; Dilernia, Dario; Ghiglione, Yanina; Turk, Gabriela; Laufer, Natalia; Rubio, Andrea; Socias, Maria E.; Figueroa, Maria Ines; Sued, Omar; Cahn, Pedro; Salomon, Horacio; Mangano, Andrea; Pando, María AngelesBackground Variants in HIV-coreceptor C-C chemokine receptor type 5 (CCR5) and Human leukocyte antigen (HLA) genes are the most important host genetic factors associated with HIV infection and disease progression. Our aim was to analyze the association of these genetic factors in the presence of clinical symptoms during Primary HIV Infection (PHI) and disease progression within the first year. Methods Seventy subjects diagnosed during PHI were studied (55 symptomatic and 15 asymptomatic). Viral load (VL) and CD4 T-cell count were evaluated. HIV progression was defined by presence of B or C events and/or CD4 T-cell counts <350 cell/mm3. CCR5 haplotypes were characterized by polymerase chain reaction and SDM-PCR-RFLP. HLA-I characterization was performed by Sequencing. Results Symptoms during PHI were significantly associated with lower frequency of CCR5-CF1 (1.8% vs. 26.7%, p = 0.006). Rapid progression was significantly associated with higher frequency of CCR5-CF2 (16.7% vs. 0%, p = 0.024) and HLA-A*11 (16.7% vs. 1.2%, p = 0.003) and lower frequency of HLA-C*3 (2.8% vs. 17.5%, p = 0.035). Higher baseline VL was significantly associated with presence of HLA-A*11, HLA-A*24, and absence of HLA-A*31 and HLA-B*57. Higher 6-month VL was significantly associated with presence of CCR5-HHE, HLA-A*24, HLA-B*53, and absence of HLA-A*31 and CCR5-CF1. Lower baseline CD4 T-cell count was significantly associated with presence of HLA-A*24/*33, HLA-B*53, CCR5-CF2 and absence of HLA-A*01/*23 and CCR5-HHA. Lower 6-month CD4 T-cell count was associated with presence of HLA-A*24 and HLA-B*53, and absence of HLA-A*01 and HLA-B*07/*39. Moreover, lower 12-month CD4 T-cell count was significantly associated with presence of HLA-A*33, HLA-B*14, HLA-C*08, CCR5-CF2, and absence of HLA-B*07 and HLA-C*07. Conclusion Several host factors were significantly associated with disease progression in PHI subjects. Most results agree with previous studies performed in other groups. However, some genetic factor associations are being described for the first time, highlighting the importance of genetic studies at a local level.Item Immune variations throughout the course of tuberculosis treatment and its relationship with adrenal hormone changes in HIV-1 patients co-infected with Mycobacterium tuberculosis(2022-03) Vecchione, María Belén; Angerami, Matias; Suarez, Guadalupe Verónica; Turk, Gabriela; Laufer, Natalia; Ben, Graciela; Ameri, Diego; Gonzalez, Diego; Parodi, Laura M.; Giavedoni, Luis D.; Maidana, Patricia; Fabre, Bibiana; Mesch, Viviana; Sued, Omar; Quiroga, Maria FlorenciaHIV infection is a major risk factor predisposing for Mycobacterium tuberculosis infection and progression to active tuberculosis (TB). As host immune response defines the course of infection, we aimed to identify immuno-endocrine changes over six-months of anti-TB chemotherapy in HIV+ people. Plasma levels of cortisol, DHEA and DHEA-S, percentages of CD4+ regulatory T cell subsets and number of IFN-γ-secreting cells were determined. Several cytokines, chemokines and C-reactive protein levels were measured. Results were correlated with clinical parameters as predictors of infection resolution and compared to similar data from HIV+ individuals, HIV-infected persons with latent TB infection and healthy donors. Throughout the course of anti-TB/HIV treatment, DHEA and DHEA-S plasma levels raised while cortisol diminished, which correlated to predictive factors of infection resolution. Furthermore, the balance between cortisol and DHEA, together with clinical assessment, may be considered as an indicator of clinical outcome after anti-TB treatment in HIV+ individuals. Clinical improvement was associated with reduced frequency of unconventional Tregs, increment in IFN-γ-secreting cells, diminution of systemic inflammation and changes of circulating cytokines and chemokines. This study suggests that the combined anti-HIV/TB therapies result in partial restoration of both, immune function and adrenal hormone plasma levels.Item Magnitude, breadth, and functional profile of T-cell responses during human immunodeficiency virus primary infection with B and BF viral variants(2008-03-15) Turk, Gabriela; Gherardi, María Magdalena; Laufer, Natalia; Saracco, Mónica; Luzzi, Renata; Cox, Josephine H; Cahn, Pedro; Salomon, HoracioThe molecular pattern of the human immunodeficiency virus (HIV) epidemic in Argentina provides an appropriate scenario to study cellular immune responses in patients with non-clade B infection. We aimed to map T-cell responses in patients infected with BF recombinant variants and compare them with those of clade B patients. Sixteen recently infected patients were enrolled and grouped by viral subtype. Nef-specific responses were evaluated with a peptide matrix-based gamma interferon (IFN-γ) enzyme-linked immunospot (ELISPOT) assay using B and BF overlapping peptides. Cross-clade and clade-specific responses were found. A correlation between B versus BF Nef-specific responses was identified. Detailed analysis at the single-peptide level revealed that BF patients show a narrower response but greater magnitude. Nef immunodominant responses agreed with previous publications, although the B loop was targeted at an unexpectedly high frequency. The putative HLA allele(s) restricting each positive response was determined. Single-peptide level screening with two different peptide sets uncovered discordant responses (mostly caused by peptide offsetting) and allowed detection of increased breadth. Positive responses identified by ELISPOT assay were further studied by intracellular cytokine staining. These were almost exclusively mediated by CD8 T cells. Characterization of concordant responses revealed that cells show distinct functional profiles, depending on the peptide presented. Last, quality (in terms of polyfunctionality) of T cells was associated with better viral replication containment. Overall, interclade differences in the frequency of epitopes recognized, structural domains targeted, and magnitude of responses were identified. Screening T-cell responses with multiple sets increased sensitivity. Further support for the notion of polyfunctional CD8+ T-cell requirement to better control viral replication is also provided.