Markers of Natural Killer Cell Exhaustion in HIV/HCV Coinfection and Their Dynamics After HCV Clearance Mediated by Direct-Acting Antivirals.

Background: Liver fibrosis is a leading cause of morbimortality in people with HIV/hepatitis C virus (HCV). Natural killer (NK) cells are linked with amelioration of liver fibrosis; however, NK cells from individuals coinfected with HIV/HCV with cirrhosis display impaired functionality and high PD-1 expression. Here, we aimed to study PD-1, TIGIT, and Tim3 as potential exhaustion markers in NK cells from persons coinfected with HIV/HCV with mild and advanced liver fibrosis. We also evaluated the role of PD-1 expression on NK cells after HCV clearance by direct-acting antivirals (DAAs). Methods: Peripheral blood mononuclear cells were isolated from individuals coinfected with HIV/HCV (N = 54; METAVIR F0/F1, n = 27; F4, evaluated by transient elastography, n = 27). In 26 participants, samples were collected before, at the end of, and 12 months after successful DAA treatment. The frequency, immunophenotype (PD-1, TIGIT, and Tim3 expression), and degranulation capacity (CD107a assay) of NK cells were determined by flow cytometry. Results: Unlike PD-1, Tim3 and TIGIT were comparably expressed between persons with mild and advanced fibrosis. Degranulation capacity was diminished in NK/TIGIT+ cells in both fibrosis stages, while NK/PD-1+ cells showed a lower CD107a expression in cirrhotic cases. Twelve months after DAA treatment, those with advanced fibrosis showed an improved NK cell frequency and reduced NK/PD-1+ cell frequency but no changes in CD107a expression. In individuals with mild fibrosis, neither PD-1 nor NK cell frequency was modified, although the percentage of NK/CD107a+ cells was improved at 12 months posttreatment. Conclusions: Although DAA improved exhaustion and frequency of NK cells in cirrhotic cases, functionality was reverted only in mild liver fibrosis, remarking the importance of an early DAA treatment.

Actualización en marcadores inmunológicos en el estudio de la infección por HIV: aportes desde el laboratorio clínico / Immunological markers in the study of HIV infection: contributions from the clinical laboratory. An update / Atualização em marcadores imunológicos no estudo da infecção pelo HIV: contribuições do laboratório clínico

Resumen Si bien la buena adherencia al tratamiento antirretroviral en la infección por el virus de la inmunodeficiencia humana (HIV por Human Immunodeficiency virus en inglés o VIH, por su sigla en español) demostró una sustancial mejoría clínica en las personas que viven con él, lograr el acceso al tratamiento continúa siendo un desafío en los tiempos que corren. Los estudios de laboratorio para diagnóstico y seguimiento de la infección por HIV se centran en estudios virológicos y recuento de linfocitos CD4+ y CD8+. Sin embargo, no se evalúa de forma detallada el perfil inmunológico de las personas que viven con HIV, ni la reconstitución inmune luego de recibido el tratamiento. Es por ello que el objetivo de esta revisión fue describir, por un lado, los diversos estudios realizados desde el laboratorio para el diagnóstico y seguimiento de la infección por HIV. Por otro lado, describir los aportes del laboratorio bioquímico en el estudio del perfil inmunológico de las personas que viven con HIV, el cual incluye desde determinaciones actualmente en uso, hasta nuevos biomarcadores inflamatorios solubles o de membrana celular, como así también las subpoblaciones linfocitarias. Dichos biomarcadores podrían ser herramientas valiosas como descriptores del estado inmunológico de las personas e, incluso, predictores de patologías asociadas a la infección por HIV.

Abstract Although a correct adherence rate to antiretroviral treatment in human immunodeficiency virus (HIV) infection reveals a substantial clinical improvement in people living with HIV, achieving access to treatment is still challenging. Laboratory studies for diagnosis and follow-up are mainly focused on virological status, CD4+ and CD8+ T cells counts. However, the immunological profile of people living with HIV is not evaluated in detail, neither is the immune reconstitution after treatment. Consequently, the aim of this review was is to describe, on the one hand, the studies carried out in the laboratory for the diagnosis and monitoring of HIV infection, and on the other hand, is to describe the contributions of the biochemical laboratory in the study of the immunological profile of people living with HIV. This study includes determinations currently in use, and the determination of new soluble or cell membrane inflammatory biomarkers, as well as T cell subsets. These biomarkers could be valuable tools as descriptors of the impervious state of people, and even predictors of pathologies associated with HIV infection.

Resumo Embora a boa adesão ao tratamento antirretroviral na infecção pelo vírus da imunodeficiência humana (HIV) tenha demonstrado melhora clínica substancial em pessoas que vivem com HIV, conseguir o acesso ao tratamento continua sendo um desafio nos momentos em que correm. Os estudos laboratoriais para diagnóstico e monitoramento da infecção pelo HIV concentram-se em estudos virológicos e contagem de linfócitos CD4+ e CD8+. No entanto, o perfil imunológico das pessoas que vivem com HIV não é avaliado detalhadamente, nem a reconstituição imunológica após o tratamento. É por isso que o objetivo desta revisão foi descrever, por um lado, os vários estudos realizados em laboratório para o diagnóstico e monitoramento da infecção pelo HIV. Por outro lado, descrever as contribuições do laboratório bioquímico no estudo do perfil imunológico de pessoas que vivem com HIV, que inclui desde determinações atualmente em uso, até novos biomarcadores inflamatórios solúveis ou de membrana celular, bem como subpopulações de linfócitos. Esses biomarcadores podem ser ferramentas valiosas como descritores do estado imunológico das pessoas e até mesmo como preditores de patologias associadas à infecção pelo HIV.

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.

Restoration of recent thymic emigrant CD4+ T cells is associated with sustained adherence to antiretroviral treatment in HIV-infected children.

To evaluate the levels of recent thymic emigrant (RTE) CD4+ T cells in HIV-infected children and to explore the associations among their frequency, antiretroviral treatment (ART) adherence, and CD4+ T cell restoration. The group evaluated comprised 85 HIV-infected patients classified as subjects with moderate or severe immunosuppression or as those with no evidence of immunosuppression. To evaluate the association between the frequency of RTE CD4+ T cells and ART adherence, 23 of the 85 patients were evaluated at two different time points during a one-year follow-up period. Children with severe immunosuppression had lower frequencies of RTE CD4+ T cells compared with children without evidence of immunosuppression (P < .001). The frequency of RTE CD4+ T cells in children with a high rate of adherence was significantly higher (P < .05) than that observed among those with suboptimal adherence. The latter group presented with infectious intercurrences on admission that decreased after initiation of treatment along with improved CD4+ and RTE naïve CD4+ T cells counts. The adequate ART adherence is essential for immune reconstitution, which might be reflected by the levels of RTE CD4+ T cells.

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

Liver cirrhosis in HIV/HCV-coinfected individuals is related to NK cell dysfunction and exhaustion, but not to an impaired NK cell modulation by CD4+ T-cells.

INTRODUCTION: HIV worsens HCV-related liver disease by accelerating fibrosis progression; however, progression rates are extremely variable among HIV/HCV-coinfected individuals. NK cells are associated with modulation of liver fibrosis and are profoundly altered during HCV and HIV infections. CD4+ T-cells modulate NK cell function, and are also affected by HIV infection. Here, we aim to characterize the association of hepatic fibrosis with both the phenotype and function of peripheral NK cells and their regulation by CD4+ T-cells, in HIV/HCV-coinfected individuals. METHODS: Thirty-four HIV/HCV-coinfected individuals with minimal (n = 16) and advanced (n = 18) fibrosis (METAVIR F0/F1 and F4 scores respectively) and 20 healthy volunteers were enrolled. PBMC were obtained from peripheral blood samples and NK and CD4+ T-cells were isolated and analysed. NK cell phenotype (CD25, CD69, Nkp46, NKG2D, PD-1), degranulation (CD107a) and IFN-γ and TNF-α production, as well as CD4+ T-cell activation (CD69, CD25 and CD38) were measured by flow cytometry. CD4+ T-cell conditioned medium (CM) derived from F0/F1 or F4 individuals was assessed for IL-2 levels by ELISA. Modulation of NK cell functionality by these CMs was also analysed. RESULTS: When comparing to NK cells from individuals with minimal fibrosis, degranulation and cytokine secretion by NK cells from subjects with F4 scores was significantly impaired, while PD-1 expression was augmented. On the one hand, neither the expression of activation markers nor IL-2 secretion was distinctly induced in CD4+ T-cells from subjects with F0/F1 or F4 METAVIR scores. Finally, NK cell degranulation and cytokine secretion were not differentially modulated by CD4+ T-cell CM, whether CD4+ T-cells derived from subjects with minimal or advanced fibrosis. CONCLUSIONS: Low levels of NK and CD4+ T-cells in HIV/HCV-coinfected individuals with advanced liver fibrosis have been previously described. Here, we show that advanced liver fibrosis in coinfected individuals is associated to a defective function of NK cells and an increased expression of the exhaustion/senescence marker PD-1. This NK signature could not be attributed to changes in the ability of CD4+ T-cells to modulate NK cell function.