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1.
Biomedica ; 41(Sp. 2): 86-102, 2021 10 15.
Article in English, Spanish | MEDLINE | ID: covidwho-1529016

ABSTRACT

INTRODUCTION: Immunological markers have been described during COVID-19 and persist after recovery. These immune markers are associated with clinical features among SARSCoV-2 infected individuals. Nevertheless, studies reporting a comprehensive analysis of the immune changes occurring during SARS-CoV-2 infection are still limited. OBJECTIVE: To evaluate the production of proinflammatory cytokines, the antibody response, and the phenotype and function of NK cells and T cells in a Colombian family cluster with SARS-CoV-2 infection. MATERIALS AND METHODS: Proinflammatory cytokines were evaluated by RT-PCR and ELISA. The frequency, phenotype, and function of NK cells (cocultures with K562 cells) and T-cells (stimulated with spike/RdRp peptides) were assessed by flow cytometry. Anti-SARS-CoV-2 antibodies were determined using indirect immunofluorescence and plaque reduction neutralization assay. RESULTS: During COVID-19, we observed a high proinflammatory-cytokine production and a reduced CD56bright-NK cell and cytotoxic response. Compared with healthy controls, infected individuals had a higher frequency of dysfunctional CD8+ T cells CD38+HLA-DR-. During the acute phase, CD8+ T cells stimulated with viral peptides exhibited a monofunctional response characterized by high IL-10 production. However, during recovery, we observed a bifunctional response characterized by the co-expression of CD107a and granzyme B or perforin. CONCLUSION: Although the proinflammatory response is a hallmark of SARS-CoV-2 infection, other phenotypic and functional alterations in NK cells and CD8+ T cells could be associated with the outcome of COVID-19. However, additional studies are required to understand these alterations and to guide future immunotherapy strategies.


Introducción. Se han descrito diferentes marcadores inmunológicos durante la COVID-19, los cuales persisten incluso después de la convalecencia y se asocian con los estadios clínicos de la infección. Sin embargo, aún son pocos los estudios orientados al análisis exhaustivo de las alteraciones del sistema inmunológico en el curso de la infección. Objetivo. Evaluar la producción de citocinas proinflamatorias, la reacción de anticuerpos, y el fenotipo y la función de las células NK y los linfocitos T en una familia colombiana con infección por SARS-CoV-2. Materiales y métodos. Se evaluaron las citocinas proinflamatorias mediante RT-PCR y ELISA; la frecuencia, el fenotipo y la función de las células NK (en cocultivos con células K562) y linfocitos T CD8+ (estimulados con péptidos spike/RdRp) mediante citometría de flujo, y los anticuerpos anti-SARS-CoV-2, mediante inmunofluorescencia indirecta y prueba de neutralización por reducción de placa. Resultados. Durante la COVID-19 hubo una producción elevada de citocinas proinflamatorias, con disminución de las células NK CD56bright y reacción citotóxica. Comparados con los controles sanos, los individuos infectados presentaron con gran frecuencia linfocitos T CD8+ disfuncionales CD38+HLA-DR-. Además, en los linfocitos T CD8+ estimulados con péptidos virales, predominó una reacción monofuncional con gran producción de IL-10 durante la fase aguda y una reacción bifuncional caracterizada por la coexpresión de CD107a y granzima B o perforina durante la convalecencia. Conclusión. Aunque la reacción inflamatoria caracteriza la infección por SARS-CoV-2, hay otras alteraciones fenotípicas y funcionales en células NK y linfocitos T CD8+ que podrían asociarse con la progresión de la infección. Se requieren estudios adicionales para entender estas alteraciones y guiar futuras estrategias de inmunoterapia.


Subject(s)
COVID-19/immunology , Killer Cells, Natural , SARS-CoV-2/immunology , T-Lymphocytes , Adult , Antibodies, Viral/analysis , CD56 Antigen/immunology , Case-Control Studies , Colombia , Family Health , Granzymes/metabolism , Humans , Interleukin-10/metabolism , Interleukin-1beta/blood , Interleukin-6/blood , Interleukin-8/blood , K562 Cells , Killer Cells, Natural/cytology , Killer Cells, Natural/immunology , Lymphocyte Activation , Male , Middle Aged , Perforin/metabolism , Phenotype , Receptors, CCR7/metabolism , T-Lymphocytes/cytology , T-Lymphocytes/immunology , Tumor Necrosis Factor-alpha/blood , Young Adult
2.
Biomedica ; 41(Sp. 2): 86-102, 2021 10 15.
Article in English, Spanish | MEDLINE | ID: covidwho-1332335

ABSTRACT

INTRODUCTION: Immunological markers have been described during COVID-19 and persist after recovery. These immune markers are associated with clinical features among SARSCoV-2 infected individuals. Nevertheless, studies reporting a comprehensive analysis of the immune changes occurring during SARS-CoV-2 infection are still limited. OBJECTIVE: To evaluate the production of proinflammatory cytokines, the antibody response, and the phenotype and function of NK cells and T cells in a Colombian family cluster with SARS-CoV-2 infection. MATERIALS AND METHODS: Proinflammatory cytokines were evaluated by RT-PCR and ELISA. The frequency, phenotype, and function of NK cells (cocultures with K562 cells) and T-cells (stimulated with spike/RdRp peptides) were assessed by flow cytometry. Anti-SARS-CoV-2 antibodies were determined using indirect immunofluorescence and plaque reduction neutralization assay. RESULTS: During COVID-19, we observed a high proinflammatory-cytokine production and a reduced CD56bright-NK cell and cytotoxic response. Compared with healthy controls, infected individuals had a higher frequency of dysfunctional CD8+ T cells CD38+HLA-DR-. During the acute phase, CD8+ T cells stimulated with viral peptides exhibited a monofunctional response characterized by high IL-10 production. However, during recovery, we observed a bifunctional response characterized by the co-expression of CD107a and granzyme B or perforin. CONCLUSION: Although the proinflammatory response is a hallmark of SARS-CoV-2 infection, other phenotypic and functional alterations in NK cells and CD8+ T cells could be associated with the outcome of COVID-19. However, additional studies are required to understand these alterations and to guide future immunotherapy strategies.


Introducción. Se han descrito diferentes marcadores inmunológicos durante la COVID-19, los cuales persisten incluso después de la convalecencia y se asocian con los estadios clínicos de la infección. Sin embargo, aún son pocos los estudios orientados al análisis exhaustivo de las alteraciones del sistema inmunológico en el curso de la infección. Objetivo. Evaluar la producción de citocinas proinflamatorias, la reacción de anticuerpos, y el fenotipo y la función de las células NK y los linfocitos T en una familia colombiana con infección por SARS-CoV-2. Materiales y métodos. Se evaluaron las citocinas proinflamatorias mediante RT-PCR y ELISA; la frecuencia, el fenotipo y la función de las células NK (en cocultivos con células K562) y linfocitos T CD8+ (estimulados con péptidos spike/RdRp) mediante citometría de flujo, y los anticuerpos anti-SARS-CoV-2, mediante inmunofluorescencia indirecta y prueba de neutralización por reducción de placa. Resultados. Durante la COVID-19 hubo una producción elevada de citocinas proinflamatorias, con disminución de las células NK CD56bright y reacción citotóxica. Comparados con los controles sanos, los individuos infectados presentaron con gran frecuencia linfocitos T CD8+ disfuncionales CD38+HLA-DR-. Además, en los linfocitos T CD8+ estimulados con péptidos virales, predominó una reacción monofuncional con gran producción de IL-10 durante la fase aguda y una reacción bifuncional caracterizada por la coexpresión de CD107a y granzima B o perforina durante la convalecencia. Conclusión. Aunque la reacción inflamatoria caracteriza la infección por SARS-CoV-2, hay otras alteraciones fenotípicas y funcionales en células NK y linfocitos T CD8+ que podrían asociarse con la progresión de la infección. Se requieren estudios adicionales para entender estas alteraciones y guiar futuras estrategias de inmunoterapia.


Subject(s)
COVID-19/immunology , Killer Cells, Natural , SARS-CoV-2/immunology , T-Lymphocytes , Adult , Antibodies, Viral/analysis , CD56 Antigen/immunology , Case-Control Studies , Colombia , Family Health , Granzymes/metabolism , Humans , Interleukin-10/metabolism , Interleukin-1beta/blood , Interleukin-6/blood , Interleukin-8/blood , K562 Cells , Killer Cells, Natural/cytology , Killer Cells, Natural/immunology , Lymphocyte Activation , Male , Middle Aged , Perforin/metabolism , Phenotype , Receptors, CCR7/metabolism , T-Lymphocytes/cytology , T-Lymphocytes/immunology , Tumor Necrosis Factor-alpha/blood , Young Adult
3.
Sci Rep ; 11(1): 14090, 2021 07 08.
Article in English | MEDLINE | ID: covidwho-1303787

ABSTRACT

MAIT cells have been shown to be activated upon several viral infections in a TCR-independent manner by responding to inflammatory cytokines secreted by antigen-presenting cells. Recently, a few studies have shown a similar activation of MAIT cells in response to severe acute respiratory coronavirus 2 (SARS-CoV-2) infection. In this study, we investigate the effect of SARS-CoV-2 infection on the frequency and phenotype of MAIT cells by flow cytometry, and we test in vitro stimulation conditions on the capacity to enhance or rescue the antiviral function of MAIT cells from patients with coronavirus disease 2019 (COVID-19). Our study, in agreement with recently published studies, confirmed the decline in MAIT cell frequency of hospitalized donors in comparison to healthy donors. MAIT cells of COVID-19 patients also had lower expression levels of TNF-alpha, perforin and granzyme B upon stimulation with IL-12 + IL-18. 24 h' incubation with IL-7 successfully restored perforin expression levels in COVID-19 patients. Combined, our findings support the growing evidence that SARS-CoV-2 is dysregulating MAIT cells and that IL-7 treatment might improve their function, rendering them more effective in protecting the body against the virus.


Subject(s)
COVID-19/prevention & control , COVID-19/virology , Interleukin-7/pharmacology , Mucosal-Associated Invariant T Cells/physiology , Mucosal-Associated Invariant T Cells/virology , SARS-CoV-2/pathogenicity , Cells, Cultured , Female , Granzymes/metabolism , Humans , Male , Mucosal-Associated Invariant T Cells/metabolism , Perforin/metabolism , Tumor Necrosis Factor-alpha/metabolism
4.
Int Immunopharmacol ; 97: 107685, 2021 Aug.
Article in English | MEDLINE | ID: covidwho-1188659

ABSTRACT

BACKGROUND: The 2019 Coronavirus (COVID-19) pandemic poses a huge threat internationally; however, the role of the host immune system in the pathogenesis of COVID-19 is not well understood. METHODS: Cytokine and chemokine levels and characterisation of immune cell subsets from 20 COVID-19 cases after hospital admission (17 critically ill and 3 severe patients) and 16 convalescent patients were determined using a multiplex immunoassay and flow cytometry, respectively. RESULTS: IP-10, MCP-1, MIG, IL-6, and IL-10 levels were significantly higher in acute severe/critically ill patients with COVID-19, whereas were normal in patients who had reached convalescence. CD8 T cells in severe and critically ill COVID-19 patients expressed high levels of cytotoxic granules (granzyme B and perforin)and was hyperactivated as evidenced by the high proportions of CD38. Furthermore, the cytotoxic potential of natural killer (NK) cells, and the frequencies of myeloid dendritic cells and plasmacytoid dendritic cells was reduced in patients with severe and critical COVID-19; however, these dysregulations were found to be restored in convalescent phases. CONCLUSION: Thus, elicitation of the hyperactive cytokine-mediated inflammatory response, dysregulation of CD8 T and NK cells, and deficiency of host myeloid and plasmacytoid DCs, may contribute to COVID-19 pathogenesis and provide insights into potential therapeutic targets and strategies.


Subject(s)
COVID-19/blood , COVID-19/immunology , Convalescence , Inflammation/etiology , ADP-ribosyl Cyclase 1/blood , Acute Disease , Adult , Aged , CD4-Positive T-Lymphocytes/immunology , CD8-Positive T-Lymphocytes/enzymology , CD8-Positive T-Lymphocytes/immunology , Chemokine CCL2/blood , Chemokine CXCL10/blood , Chemokine CXCL9/blood , Critical Illness , Cytokine Release Syndrome/blood , Cytokine Release Syndrome/immunology , Dendritic Cells/immunology , Female , Granzymes/metabolism , Humans , Interleukin-10/blood , Interleukin-6/blood , Killer Cells, Natural/enzymology , Killer Cells, Natural/immunology , Male , Membrane Glycoproteins/blood , Middle Aged , Perforin/metabolism
5.
J Infect Dis ; 223(9): 1555-1563, 2021 05 20.
Article in English | MEDLINE | ID: covidwho-1069262

ABSTRACT

Repeated infections with endemic human coronaviruses (hCoV) are thought to reflect lack of long-lasting protective immunity. We evaluated circulating human CD4 T cells collected prior to 2020 for reactivity towards hCoV spike proteins, probing for the ability to produce interferon-γ, interleukin-2, or granzyme B. We found robust reactivity to spike-derived epitopes, comparable to influenza, but highly variable abundance and functional potential across subjects, depending on age and viral antigen specificity. To explore potential of these memory cells to be recruited in severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, we examined the subjects for cross-reactive recognition of epitopes from SARS-CoV-2 nucleocapsid, membrane/envelope, and spike. Functional potential of these cross-reactive CD4 T cells was highly variable; nucleocapsid-specific CD4 T cells but not spike-reactive cells showed exceptionally high levels of granzyme production upon stimulation. These results are considered in light of recruitment of hCoV-reactive cells into responses to SARS-CoV infections or vaccinations.


Subject(s)
CD4-Positive T-Lymphocytes/virology , COVID-19/immunology , Coronavirus Infections/immunology , Epitopes, T-Lymphocyte/immunology , SARS-CoV-2/immunology , Adult , Aged , Coronavirus Envelope Proteins/immunology , Coronavirus M Proteins/immunology , Coronavirus Nucleocapsid Proteins/immunology , Cross Reactions , Granzymes/metabolism , Humans , Immunologic Memory , Interferon-gamma/metabolism , Interleukin-2/metabolism , Middle Aged , Spike Glycoprotein, Coronavirus/immunology
6.
Viruses ; 13(2)2021 02 03.
Article in English | MEDLINE | ID: covidwho-1060774

ABSTRACT

Coronavirus disease 2019 (COVID-19), caused by infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), comprises mild courses of disease as well as progression to severe disease, characterised by lung and other organ failure. The immune system is considered to play a crucial role for the pathogenesis of COVID-19, although especially the contribution of innate-like T cells remains poorly understood. Here, we analysed the phenotype and function of mucosal-associated invariant T (MAIT) cells, innate-like T cells with potent antimicrobial effector function, in patients with mild and severe COVID-19 by multicolour flow cytometry. Our data indicate that MAIT cells are highly activated in patients with COVID-19, irrespective of the course of disease, and express high levels of proinflammatory cytokines such as IL-17A and TNFα ex vivo. Of note, expression of the activation marker HLA-DR positively correlated with SAPS II score, a measure of disease severity. Upon MAIT cell-specific in vitro stimulation, MAIT cells however failed to upregulate expression of the cytokines IL-17A and TNFα, as well as cytolytic proteins, that is, granzyme B and perforin. Thus, our data point towards an altered cytokine expression profile alongside an impaired antibacterial and antiviral function of MAIT cells in COVID-19 and thereby contribute to the understanding of COVID-19 immunopathogenesis.


Subject(s)
COVID-19/immunology , Lymphocyte Activation , Mucosal-Associated Invariant T Cells/immunology , Adaptive Immunity , COVID-19/physiopathology , Cytokines/metabolism , Female , Granzymes/metabolism , HLA-DR Antigens , Humans , Interleukin-17/metabolism , Killer Cells, Natural/immunology , Male , Mucosal-Associated Invariant T Cells/metabolism , Severity of Illness Index , T-Lymphocyte Subsets/immunology , Tumor Necrosis Factor-alpha/metabolism
7.
APMIS ; 129(2): 91-102, 2021 Feb.
Article in English | MEDLINE | ID: covidwho-900954

ABSTRACT

T cells play vital roles in the development and progression of acute coronary syndromes (ACS), including cytotoxicity mediated by CD8+ T cells and immunoregulatory activity mediated by CD4+ T cells. Interleukin (IL)-9-secreting CD4+ T cells (Th9 cells) were recently found to be involved in the onset of ACS. We investigated regulatory role of Th9 cells to CD8+ T cells in patients with stable angina pectoris, unstable angina pectoris, and acute myocardial infarction (AMI). Circulating Th9 cells percentage, plasma IL-9 level, and PU.1 mRNA relative level was up-regulated in AMI patients compared with controls. There was no significant difference of IL-9-secreting CD8+ T cells percentage among groups. CD8+ T cells from AMI patients revealed increased cytotoxicity than those from controls, which presented as enhanced cytotolytic activity to target cells, increased interferon-γ and tumor necrosis factor-α secretion, elevated perforin and granzyme B production, and reduced programmed death-1 and cytotoxic T lymphocyte-associated protein 4. IL-9 stimulation did not affect proliferation, but promoted CD8+ T-cell cytotoxicity from both controls and AMI patients. IL-9-secreting CD4+ T cells were enriched in CD4+ CCR4- CCR6- CXCR3- cells. The enhancement of CD8+ T-cell cytotoxicity induced by CD4+ CCR4- CCR6- CXCR3- cells was dependent on IL-9 secretion. The present results indicated that up-regulation of IL-9-secreting CD4+ T cells may contribute to pathogenesis of AMI through enhancement of CD8+ T-cell cytotoxicity.


Subject(s)
Acute Coronary Syndrome/pathology , CD4-Positive T-Lymphocytes/immunology , Interleukin-9/blood , T-Lymphocytes, Cytotoxic/immunology , Acute Coronary Syndrome/immunology , CTLA-4 Antigen/metabolism , Cells, Cultured , Female , Granzymes/metabolism , Humans , Interleukin-9/metabolism , Male , Middle Aged , Programmed Cell Death 1 Receptor/metabolism
8.
Cells ; 9(8)2020 07 22.
Article in English | MEDLINE | ID: covidwho-669617

ABSTRACT

The ectonucleotidases CD39 and CD73 regulate immune responses by balancing extracellular ATP and adenosine in inflammation and are likely to be involved in the pathophysiology of COVID-19. Here, we analyzed CD39 and CD73 on different lymphocyte populations in a small cohort of COVID-19 patients and in healthy individuals. We describe a significantly lower level of expression of CD73 on cytotoxic lymphocyte populations, including CD8+ T, natural killer T (NKT), and natural killer (NK) cells, during COVID-19. Interestingly, the decrease of CD73 on CD8+ T cells and NKT cells correlated with serum ferritin levels. Furthermore, we observed distinct functional differences between the CD73+ and CD73- subsets of CD8+ T cells and NKT cells with regard to cytokine/toxin secretion. In COVID-19 patients, the majority of the CD73-CD8+ T cells were capable of secreting granzyme B, perforin, tumor necrosis factor (TNF-α) or interferon-gamma (IFN-γ). To conclude, in this first study of CD39 and CD73 expression of lymphocytes in COVID-19, we show that CD8+ T cells and NKT cells lacking CD73 possess a significantly higher cytotoxic effector functionality compared to their CD73+ counterparts. Future studies should investigate differences of cellular CD39 and CD73 expression in patients at different disease stages and their potential as prognostic markers or targets for immunomodulatory therapies.


Subject(s)
5'-Nucleotidase/metabolism , Apyrase/metabolism , Coronavirus Infections/immunology , Killer Cells, Natural/immunology , Natural Killer T-Cells/immunology , Pneumonia, Viral/immunology , T-Lymphocytes, Cytotoxic/immunology , Adenosine/metabolism , Adult , Aged , Betacoronavirus , COVID-19 , Coronavirus Infections/enzymology , Female , GPI-Linked Proteins/metabolism , Granzymes/metabolism , Humans , Inflammation/enzymology , Inflammation/immunology , Interferon-gamma/metabolism , Male , Middle Aged , Pandemics , Perforin/metabolism , Pneumonia, Viral/enzymology , SARS-CoV-2 , Signal Transduction/immunology , T-Lymphocytes, Cytotoxic/metabolism , Tumor Necrosis Factor-alpha/metabolism
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