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1.
Methods Mol Biol ; 2380: 201-209, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-34802133

RESUMO

Generation of effective immune protection against viral infection and vaccination depends greatly on a successful engagement and stimulation of adaptive immune B cells and a specialized CD4+ T cell subset called T follicular helper cells (TFH cells). Since TFH cells primarily reside in lymphoid tissues, they can be challenging to study in human settings. However, a counterpart of these cells, circulating TFH (cTFH) cells, can be detected in peripheral blood. Assessment of cTFH cells serves as an informative marker of humoral responses following viral infection and vaccination and can be predictive of antibody titers. Here, we describe a comprehensive flow cytometry detection method for dissecting cTFH subsets and activation, together with the assessment of antibody-secreting cells (ASCs), from a small volume of human whole blood. This approach allows the investigation of cellular events that underpin successful immune responses following influenza and SARS-CoV-2 infection/vaccination in humans and is applicable to other viral disease settings.


Assuntos
Linfócitos B/imunologia , COVID-19 , Influenza Humana , Células T Auxiliares Foliculares/imunologia , COVID-19/imunologia , Humanos , Influenza Humana/imunologia
2.
Proc Natl Acad Sci U S A ; 118(41)2021 Oct 12.
Artigo em Inglês | MEDLINE | ID: mdl-34607957

RESUMO

Morbidity and mortality rates from seasonal and pandemic influenza occur disproportionately in high-risk groups, including Indigenous people globally. Although vaccination against influenza is recommended for those most at risk, studies on immune responses elicited by seasonal vaccines in Indigenous populations are largely missing, with no data available for Indigenous Australians and only one report published on antibody responses in Indigenous Canadians. We recruited 78 Indigenous and 84 non-Indigenous Australians vaccinated with the quadrivalent influenza vaccine into the Looking into InFluenza T cell immunity - Vaccination cohort study and collected blood to define baseline, early (day 7), and memory (day 28) immune responses. We performed in-depth analyses of T and B cell activation, formation of memory B cells, and antibody profiles and investigated host factors that could contribute to vaccine responses. We found activation profiles of circulating T follicular helper type-1 cells at the early stage correlated strongly with the total change in antibody titers induced by vaccination. Formation of influenza-specific hemagglutinin-binding memory B cells was significantly higher in seroconverters compared with nonseroconverters. In-depth antibody characterization revealed a reduction in immunoglobulin G3 before and after vaccination in the Indigenous Australian population, potentially linked to the increased frequency of the G3m21* allotype. Overall, our data provide evidence that Indigenous populations elicit robust, broad, and prototypical immune responses following immunization with seasonal inactivated influenza vaccines. Our work strongly supports the recommendation of influenza vaccination to protect Indigenous populations from severe seasonal influenza virus infections and their subsequent complications.

3.
Clin Transl Immunology ; 10(9): e1336, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34522380

RESUMO

Objectives: Although co-expression of CD38 and HLA-DR reflects T-cell activation during viral infections, high and prolonged CD38+HLA-DR+ expression is associated with severe disease. To date, the mechanism underpinning expression of CD38+HLA-DR+ is poorly understood. Methods: We used mouse models of influenza A/H9N2, A/H7N9 and A/H3N2 infection to investigate mechanisms underpinning CD38+MHC-II+ phenotype on CD8+ T cells. To further understand MHC-II trogocytosis on murine CD8+ T cells as well as the significance behind the scenario, we used adoptively transferred transgenic OT-I CD8+ T cells and A/H3N2-SIINKEKL infection. Results: Analysis of influenza-specific immunodominant DbNP366 +CD8+ T-cell responses showed that CD38+MHC-II+ co-expression was detected on both virus-specific and bystander CD8+ T cells, with increased numbers of both CD38+MHC-II+CD8+ T-cell populations observed in immune organs including the site of infection during severe viral challenge. OT-I cells adoptively transferred into MHC-II-/- mice had no MHC-II after infection, suggesting that MHC-II was acquired via trogocytosis. The detection of CD19 on CD38+MHC-II+ OT-I cells supports the proposition that MHC-II was acquired by trogocytosis sourced from B cells. Co-expression of CD38+MHC-II+ on CD8+ T cells was needed for optimal recall following secondary infection. Conclusions: Overall, our study demonstrates that both virus-specific and bystander CD38+MHC-II+ CD8+ T cells are recruited to the site of infection during severe disease, and that MHC-II presence occurs via trogocytosis from antigen-presenting cells. Our findings highlight the importance of the CD38+MHC-II+ phenotype for CD8+ T-cell recall.

4.
Open Forum Infect Dis ; 8(9): ofab359, 2021 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-34514016

RESUMO

We describe severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-specific immune responses in a patient with lymphoma and recent programmed death 1 (PD-1) inhibitor therapy with late onset of severe coronavirus disease 2019 disease and prolonged SARS-CoV-2 replication, in comparison to age-matched and immunocompromised controls. High levels of HLA-DR+/CD38+ activation, interleukin 6, and interleukin 18 in the absence of B cells and PD-1 expression was observed. SARS-CoV-2-specific antibody responses were absent and SARS-CoV-2-specific T cells were minimally detected. This case highlights challenges in managing immunocompromised hosts who may fail to mount effective virus-specific immune responses.

5.
J Biol Chem ; 297(3): 101065, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-34384783

RESUMO

CD8+ T cells play an important role in vaccination and immunity against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. Although numerous SARS-CoV-2 CD8+ T cell epitopes have been identified, the molecular basis underpinning T cell receptor (TCR) recognition of SARS-CoV-2-specific T cells remains unknown. The T cell response directed toward SARS-CoV-2 spike protein-derived S269-277 peptide presented by the human leukocyte antigen (HLA)-A∗02:01 allomorph (hereafter the HLA-A2S269-277 epitope) is, to date, the most immunodominant SARS-CoV-2 epitope found in individuals bearing this allele. As HLA-A2S269-277-specific CD8+ T cells utilize biased TRAV12 gene usage within the TCR α-chain, we sought to understand the molecular basis underpinning this TRAV12 dominance. We expressed four TRAV12+ TCRs which bound the HLA-A2S269-277 complex with low micromolar affinity and determined the crystal structure of the HLA-A2S269-277 binary complex, and subsequently a ternary structure of the TRAV12+ TCR complexed to HLA-A2S269-277. We found that the TCR made extensive contacts along the entire length of the S269-277 peptide, suggesting that the TRAV12+ TCRs would be sensitive to sequence variation within this epitope. To examine this, we investigated cross-reactivity toward analogous peptides from existing SARS-CoV-2 variants and closely related coronaviruses. We show via surface plasmon resonance and tetramer studies that the TRAV12+ T cell repertoire cross-reacts poorly with these analogous epitopes. Overall, we defined the structural basis underpinning biased TCR recognition of CD8+ T cells directed at an immunodominant epitope and provide a framework for understanding TCR cross-reactivity toward viral variants within the S269-277 peptide.


Assuntos
Epitopos de Linfócito T/química , Antígeno HLA-A2/metabolismo , Epitopos Imunodominantes/metabolismo , Receptores de Antígenos de Linfócitos T/metabolismo , Glicoproteína da Espícula de Coronavírus/metabolismo , Humanos , Conformação Proteica , Receptores de Antígenos de Linfócitos T/química
7.
Res Sq ; 2021 Aug 26.
Artigo em Inglês | MEDLINE | ID: mdl-34462740

RESUMO

Although the respiratory tract is the primary site of SARS-CoV-2 infection and the ensuing immunopathology, respiratory immune responses are understudied and urgently needed to understand mechanisms underlying COVID-19 disease pathogenesis. We collected paired longitudinal blood and respiratory tract samples (endotracheal aspirate, sputum or pleural fluid) from hospitalized COVID-19 patients and non-COVID-19 controls. Cellular, humoral and cytokine responses were analysed and correlated with clinical data. SARS-CoV-2-specific IgM, IgG and IgA antibodies were detected using ELISA and multiplex assay in both the respiratory tract and blood of COVID-19 patients, although a higher receptor binding domain (RBD)-specific IgM and IgG seroconversion level was found in respiratory specimens. SARS-CoV-2 neutralization activity in respiratory samples was detected only when high levels of RBD-specific antibodies were present. Strikingly, cytokine/chemokine levels and profiles greatly differed between respiratory samples and plasma, indicating that inflammation needs to be assessed in respiratory specimens for the accurate assessment of SARS-CoV-2 immunopathology. Diverse immune cell subsets were detected in respiratory samples, albeit dominated by neutrophils. Importantly, we also showed that dexamethasone and/or remdesivir treatment did not affect humoral responses in blood of COVID-19 patients. Overall, our study unveils stark differences in innate and adaptive immune responses between respiratory samples and blood and provides important insights into effect of drug therapy on immune responses in COVID-19 patients.

8.
PLoS Pathog ; 17(7): e1009759, 2021 07.
Artigo em Inglês | MEDLINE | ID: mdl-34320031

RESUMO

The host response to SARS-CoV-2 infection provide insights into both viral pathogenesis and patient management. The host-encoded microRNA (miRNA) response to SARS-CoV-2 infection, however, remains poorly defined. Here we profiled circulating miRNAs from ten COVID-19 patients sampled longitudinally and ten age and gender matched healthy donors. We observed 55 miRNAs that were altered in COVID-19 patients during early-stage disease, with the inflammatory miR-31-5p the most strongly upregulated. Supervised machine learning analysis revealed that a three-miRNA signature (miR-423-5p, miR-23a-3p and miR-195-5p) independently classified COVID-19 cases with an accuracy of 99.9%. In a ferret COVID-19 model, the three-miRNA signature again detected SARS-CoV-2 infection with 99.7% accuracy, and distinguished SARS-CoV-2 infection from influenza A (H1N1) infection and healthy controls with 95% accuracy. Distinct miRNA profiles were also observed in COVID-19 patients requiring oxygenation. This study demonstrates that SARS-CoV-2 infection induces a robust host miRNA response that could improve COVID-19 detection and patient management.


Assuntos
Teste para COVID-19/métodos , COVID-19/diagnóstico , COVID-19/genética , MicroRNAs/genética , SARS-CoV-2 , Adulto , Idoso , Animais , COVID-19/sangue , Estudos de Casos e Controles , Diagnóstico Diferencial , Modelos Animais de Doenças , Feminino , Furões , Expressão Gênica , Interações entre Hospedeiro e Microrganismos/genética , Humanos , Vírus da Influenza A Subtipo H1N1 , Estudos Longitudinais , Masculino , MicroRNAs/sangue , Pessoa de Meia-Idade , Infecções por Orthomyxoviridae/diagnóstico , Infecções por Orthomyxoviridae/genética , Pandemias , Aprendizado de Máquina Supervisionado
9.
Vaccine ; 39(29): 3940-3951, 2021 06 29.
Artigo em Inglês | MEDLINE | ID: mdl-34090697

RESUMO

Current detergent or ether-disrupted split vaccines (SVs) for influenza do not always induce adequate immune responses, especially in young children. This contrasts with the whole virus particle vaccines (WPVs) originally used against influenza that were immunogenic in both adults and children but were replaced by SV in the 1970s due to concerns with reactogenicity. In this study, we re-evaluated the immunogenicity of WPV and SV, prepared from the same batch of purified influenza virus, in cynomolgus macaques and confirmed that WPV is superior to SV in priming potency. In addition, we compared the ability of WPV and SV to induce innate immune responses, including the maturation of dendritic cells (DCs) in vitro. WPV stimulated greater production of inflammatory cytokines and type-I interferon in immune cells from mice and macaques compared to SV. Since these innate responses are likely triggered by the activation of pattern recognition receptors (PRRs) by viral RNA, the quantity and quality of viral RNA in each vaccine were assessed. Although the quantity of viral RNA was similar in the two vaccines, the amount of viral RNA of a length that can be recognized by PRRs was over 100-fold greater in WPV than in SV. More importantly, 1000-fold more viral RNA was delivered to DCs by WPV than by SV when exposed to preparations containing the same amount of HA protein. Furthermore, WPV induced up-regulation of the DC maturation marker CD86 on murine DCs, while SV did not. The present results suggest that the activation of antigen-presenting DCs, by PRR-recognizable viral RNA contained in WPV is responsible for the effective priming potency of WPV observed in naïve mice and macaques. WPV is thus recommended as an alternative option for seasonal influenza vaccines, especially for children.


Assuntos
Vacinas contra Influenza , Infecções por Orthomyxoviridae , Orthomyxoviridae , Animais , Anticorpos Antivirais , Células Apresentadoras de Antígenos , Camundongos , Infecções por Orthomyxoviridae/prevenção & controle , RNA Viral , Vacinas de Produtos Inativados , Vírion
10.
Immunol Cell Biol ; 99(9): 990-1000, 2021 10.
Artigo em Inglês | MEDLINE | ID: mdl-34086357

RESUMO

In-depth understanding of human T-cell-mediated immunity in coronavirus disease 2019 (COVID-19) is needed if we are to optimize vaccine strategies and immunotherapies. Identification of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) T-cell epitopes and generation of peptide-human leukocyte antigen (peptide-HLA) tetramers facilitate direct ex vivo analyses of SARS-CoV-2-specific T cells and their T-cell receptor (TCR) repertoires. We utilized a combination of peptide prediction and in vitro peptide stimulation to validate novel SARS-CoV-2 epitopes restricted by HLA-A*24:02, one of the most prominent HLA class I alleles, especially in Indigenous and Asian populations. Of the peptides screened, three spike-derived peptides generated CD8+ IFNγ+ responses above background, S1208-1216 (QYIKWPWYI), S448-456 (NYNYLYRLF) and S193-201 (VFKNIDGYF), with S1208 generating immunodominant CD8+ IFNγ+ responses. Using peptide-HLA-I tetramers, we performed direct ex vivo tetramer enrichment for HLA-A*24:02-restricted CD8+ T cells in COVID-19 patients and prepandemic controls. The precursor frequencies for HLA-A*24:02-restricted epitopes were within the range previously observed for other SARS-CoV-2 epitopes for both COVID-19 patients and prepandemic individuals. Naïve A24/SARS-CoV-2-specific CD8+ T cells increased nearly 7.5-fold above the average precursor frequency during COVID-19, gaining effector and memory phenotypes. Ex vivo single-cell analyses of TCRαß repertoires found that the A24/S448 + CD8+ T-cell TCRαß repertoire was driven by a common TCRß chain motif, whereas the A24/S1208 + CD8+ TCRαß repertoire was diverse across COVID-19 patients. Our study provides an in depth characterization and important insights into SARS-CoV-2-specific CD8+ T-cell responses associated with a prominent HLA-A*24:02 allomorph. This contributes to our knowledge on adaptive immune responses during primary COVID-19 and could be exploited in vaccine or immunotherapeutic approaches.


Assuntos
Linfócitos T CD8-Positivos/imunologia , COVID-19 , Antígeno HLA-A24 , Receptores de Antígenos de Linfócitos T/imunologia , COVID-19/imunologia , Humanos , SARS-CoV-2
11.
Immunity ; 54(5): 1066-1082.e5, 2021 05 11.
Artigo em Inglês | MEDLINE | ID: mdl-33951417

RESUMO

To better understand primary and recall T cell responses during coronavirus disease 2019 (COVID-19), it is important to examine unmanipulated severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-specific T cells. By using peptide-human leukocyte antigen (HLA) tetramers for direct ex vivo analysis, we characterized CD8+ T cells specific for SARS-CoV-2 epitopes in COVID-19 patients and unexposed individuals. Unlike CD8+ T cells directed toward subdominant epitopes (B7/N257, A2/S269, and A24/S1,208) CD8+ T cells specific for the immunodominant B7/N105 epitope were detected at high frequencies in pre-pandemic samples and at increased frequencies during acute COVID-19 and convalescence. SARS-CoV-2-specific CD8+ T cells in pre-pandemic samples from children, adults, and elderly individuals predominantly displayed a naive phenotype, indicating a lack of previous cross-reactive exposures. T cell receptor (TCR) analyses revealed diverse TCRαß repertoires and promiscuous αß-TCR pairing within B7/N105+CD8+ T cells. Our study demonstrates high naive precursor frequency and TCRαß diversity within immunodominant B7/N105-specific CD8+ T cells and provides insight into SARS-CoV-2-specific T cell origins and subsequent responses.


Assuntos
Linfócitos T CD8-Positivos/imunologia , COVID-19/imunologia , Proteínas do Nucleocapsídeo de Coronavírus/imunologia , Epitopos Imunodominantes/imunologia , Receptores de Antígenos de Linfócitos T/imunologia , SARS-CoV-2/imunologia , Adulto , Idoso , Motivos de Aminoácidos , Linfócitos T CD4-Positivos , Criança , Convalescença , Proteínas do Nucleocapsídeo de Coronavírus/química , Epitopos de Linfócito T/química , Epitopos de Linfócito T/imunologia , Feminino , Humanos , Epitopos Imunodominantes/química , Masculino , Pessoa de Meia-Idade , Fenótipo , Fosfoproteínas/química , Fosfoproteínas/imunologia , Receptores de Antígenos de Linfócitos T/química , Receptores de Antígenos de Linfócitos T/genética , Receptores de Antígenos de Linfócitos T alfa-beta/química , Receptores de Antígenos de Linfócitos T alfa-beta/genética , Receptores de Antígenos de Linfócitos T alfa-beta/imunologia , Glicoproteína da Espícula de Coronavírus/química , Glicoproteína da Espícula de Coronavírus/imunologia
13.
Nat Commun ; 12(1): 2931, 2021 05 18.
Artigo em Inglês | MEDLINE | ID: mdl-34006841

RESUMO

Indigenous people worldwide are at high risk of developing severe influenza disease. HLA-A*24:02 allele, highly prevalent in Indigenous populations, is associated with influenza-induced mortality, although the basis for this association is unclear. Here, we define CD8+ T-cell immune landscapes against influenza A (IAV) and B (IBV) viruses in HLA-A*24:02-expressing Indigenous and non-Indigenous individuals, human tissues, influenza-infected patients and HLA-A*24:02-transgenic mice. We identify immunodominant protective CD8+ T-cell epitopes, one towards IAV and six towards IBV, with A24/PB2550-558-specific CD8+ T cells being cross-reactive between IAV and IBV. Memory CD8+ T cells towards these specificities are present in blood (CD27+CD45RA- phenotype) and tissues (CD103+CD69+ phenotype) of healthy individuals, and effector CD27-CD45RA-PD-1+CD38+CD8+ T cells in IAV/IBV patients. Our data show influenza-specific CD8+ T-cell responses in Indigenous Australians, and advocate for T-cell-mediated vaccines that target and boost the breadth of IAV/IBV-specific CD8+ T cells to protect high-risk HLA-A*24:02-expressing Indigenous and non-Indigenous populations from severe influenza disease.


Assuntos
Linfócitos T CD8-Positivos/metabolismo , Epitopos de Linfócito T/genética , Antígeno HLA-A24/genética , Povos Indígenas/genética , Adulto , Alelos , Sequência de Aminoácidos , Animais , Austrália , Linfócitos T CD8-Positivos/imunologia , Células Cultivadas , Cães , Epitopos de Linfócito T/imunologia , Feminino , Frequência do Gene , Antígeno HLA-A24/imunologia , Humanos , Vírus da Influenza A/imunologia , Vírus da Influenza A/fisiologia , Vírus da Influenza B/imunologia , Vírus da Influenza B/fisiologia , Influenza Humana/imunologia , Influenza Humana/virologia , Masculino , Camundongos Transgênicos , Pessoa de Meia-Idade
14.
Nat Commun ; 12(1): 3073, 2021 05 24.
Artigo em Inglês | MEDLINE | ID: mdl-34031386

RESUMO

Follicular helper T (TFH) cells control antibody responses by supporting antibody affinity maturation and memory formation. Inadequate TFH function has been found in individuals with ineffective responses to vaccines, but the mechanism underlying TFH regulation in vaccination is not understood. Here, we report that lower serum levels of the metabolic hormone leptin associate with reduced vaccine responses to influenza or hepatitis B virus vaccines in healthy populations. Leptin promotes mouse and human TFH differentiation and IL-21 production via STAT3 and mTOR pathways. Leptin receptor deficiency impairs TFH generation and antibody responses in immunisation and infection. Similarly, leptin deficiency induced by fasting reduces influenza vaccination-mediated protection for the subsequent infection challenge, which is mostly rescued by leptin replacement. Our results identify leptin as a regulator of TFH cell differentiation and function and indicate low levels of leptin as a risk factor for vaccine failure.


Assuntos
Formação de Anticorpos/imunologia , Vacinas contra Influenza/imunologia , Leptina/metabolismo , Animais , Anticorpos Antivirais/imunologia , Diferenciação Celular , Feminino , Homeostase , Humanos , Imunização , Influenza Humana/prevenção & controle , Leptina/deficiência , Ativação Linfocitária/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Vacinação/métodos
15.
J Virol ; 95(14): e0011121, 2021 06 24.
Artigo em Inglês | MEDLINE | ID: mdl-33952646

RESUMO

The current fears of a future influenza pandemic have resulted in an increased emphasis on the development and testing of novel therapeutic strategies against the virus. Fundamental to this is the ferret model of influenza infection, which is critical in examining pathogenesis and treatment. Nevertheless, a precise evaluation of the efficacy of any treatment strategy in ferrets is reliant on understanding the immune response in this model. Interferon-inducible transmembrane proteins (IFITMs) are interferon-stimulated proteins shown to be critically important in the host immune response against viral infections. These proteins confer intrinsic innate immunity to pH-dependent viruses such as influenza viruses and can inhibit cytosolic entry of such viruses to limit the severity of infection following interferon upregulation. Mutations in IFITM genes in humans have been identified as key risk factors for worsened disease progression, particularly in the case of avian influenza viruses such as H7N9. While the IFITM genes of humans and mice have been well characterized, no studies have been conducted to classify the IFITM locus and interferon-driven upregulation of IFITMs in ferrets. Here, we show the architecture of the ferret IFITM locus and its synteny to the IFITM locus of other mammalian and avian species. Furthermore, we show that ferret IFITM1, -2, and -3 are functionally responsive to both interferon-α (IFN-α) and influenza virus stimulation. Thus, we show that ferret IFITMs exhibit interferon-stimulated properties similar to those shown in other species, furthering our knowledge of the innate immune response in the ferret model of human influenza virus infections. IMPORTANCE IFITM proteins can prevent the entry of several pH-dependent viruses, including high-consequence viruses such as HIV, influenza viruses, and SARS-coronaviruses. Mutations in these genes have been associated with worsened disease outcomes with mutations in their IFITM genes, highlighting these genes as potential disease risk factors. Ferrets provide a valuable tool to model infectious diseases; however, there is a critical shortage of information regarding their interferon-stimulated genes. We identified the putative ferret IFITM genes and mapped their complete gene locus. Thus, our study fills a critical gap in knowledge and supports the further use of the ferret model to explore the importance of IFITMs in these important diseases.


Assuntos
Furões , Vírus da Influenza A Subtipo H1N1 , Interferon-alfa/metabolismo , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Infecções por Orthomyxoviridae/imunologia , Animais , Linhagem Celular , Sequência Conservada , Modelos Animais de Doenças , Furões/imunologia , Furões/metabolismo , Furões/virologia , Humanos , Modelos Moleculares , Infecções por Orthomyxoviridae/genética , Infecções por Orthomyxoviridae/metabolismo , Reação em Cadeia da Polimerase , Análise de Sequência de Proteína , Regulação para Cima
16.
Nat Commun ; 12(1): 2691, 2021 05 11.
Artigo em Inglês | MEDLINE | ID: mdl-33976217

RESUMO

How innate and adaptive immune responses work in concert to resolve influenza disease is yet to be fully investigated in one single study. Here, we utilize longitudinal samples from patients hospitalized with acute influenza to understand these immune responses. We report the dynamics of 18 important immune parameters, related to clinical, genetic and virological factors, in influenza patients across different severity levels. Influenza disease correlates with increases in IL-6/IL-8/MIP-1α/ß cytokines and lower antibody responses. Robust activation of circulating T follicular helper cells correlates with peak antibody-secreting cells and influenza heamaglutinin-specific memory B-cell numbers, which phenotypically differs from vaccination-induced B-cell responses. Numbers of influenza-specific CD8+ or CD4+ T cells increase early in disease and retain an activated phenotype during patient recovery. We report the characterisation of immune cellular networks underlying recovery from influenza infection which are highly relevant to other infectious diseases.


Assuntos
Formação de Anticorpos/imunologia , Linfócitos B/imunologia , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Citocinas/imunologia , Influenza Humana/imunologia , Linfócitos T Auxiliares-Indutores/imunologia , Estudos de Coortes , Citocinas/metabolismo , Hospitalização/estatística & dados numéricos , Humanos , Vírus da Influenza A/classificação , Vírus da Influenza A/genética , Vírus da Influenza A/fisiologia , Vacinas contra Influenza/imunologia , Influenza Humana/virologia , Pessoa de Meia-Idade , Filogenia , Vacinação/métodos
17.
Sci Immunol ; 6(58)2021 Apr 23.
Artigo em Inglês | MEDLINE | ID: mdl-33893172

RESUMO

Human cytomegalovirus (CMV) infection can stimulate robust human leukocyte antigen (HLA)-E-restricted CD8+ T cell responses. These T cells recognize a peptide from UL40, which differs by as little as a single methyl group from self-peptides that also bind HLA-E, challenging their capacity to avoid self-reactivity. Unexpectedly, we showed that the UL40/HLA-E T cell receptor (TCR) repertoire included TCRs that had high affinities for HLA-E/self-peptide. However, paradoxically, lower cytokine responses were observed from UL40/HLA-E T cells bearing TCRs with high affinity for HLA-E. RNA sequencing and flow cytometric analysis revealed that these T cells were marked by the expression of inhibitory natural killer cell receptors (NKRs) KIR2DL1 and KIR2DL2/L3. On the other hand, UL40/HLA-E T cells bearing lower-affinity TCRs expressed the activating receptor NKG2C. Activation of T cells bearing higher-affinity TCRs was regulated by the interaction between KIR2D receptors and HLA-C. These findings identify a role for NKR signaling in regulating self/non-self discrimination by HLA-E-restricted T cells, allowing for antiviral responses while avoiding contemporaneous self-reactivity.

18.
Nat Commun ; 12(1): 2037, 2021 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-33795692

RESUMO

The hallmarks of COVID-19 are higher pathogenicity and mortality in the elderly compared to children. Examining baseline SARS-CoV-2 cross-reactive immunological responses, induced by circulating human coronaviruses (hCoVs), is needed to understand such divergent clinical outcomes. Here we show analysis of coronavirus antibody responses of pre-pandemic healthy children (n = 89), adults (n = 98), elderly (n = 57), and COVID-19 patients (n = 50) by systems serology. Moderate levels of cross-reactive, but non-neutralizing, SARS-CoV-2 antibodies are detected in pre-pandemic healthy individuals. SARS-CoV-2 antigen-specific Fcγ receptor binding accurately distinguishes COVID-19 patients from healthy individuals, suggesting that SARS-CoV-2 infection induces qualitative changes to antibody Fc, enhancing Fcγ receptor engagement. Higher cross-reactive SARS-CoV-2 IgA and IgG are observed in healthy elderly, while healthy children display elevated SARS-CoV-2 IgM, suggesting that children have fewer hCoV exposures, resulting in less-experienced but more polyreactive humoral immunity. Age-dependent analysis of COVID-19 patients, confirms elevated class-switched antibodies in elderly, while children have stronger Fc responses which we demonstrate are functionally different. These insights will inform COVID-19 vaccination strategies, improved serological diagnostics and therapeutics.


Assuntos
Anticorpos Antivirais/sangue , Anticorpos Antivirais/imunologia , Formação de Anticorpos/imunologia , SARS-CoV-2/imunologia , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , COVID-19/imunologia , Vacinas contra COVID-19/imunologia , Criança , Pré-Escolar , Reações Cruzadas/imunologia , Humanos , Imunoglobulina A/sangue , Imunoglobulina A/imunologia , Imunoglobulina G/sangue , Imunoglobulina G/imunologia , Imunoglobulina M/sangue , Imunoglobulina M/imunologia , Pessoa de Meia-Idade , Receptores de IgG/imunologia , Glicoproteína da Espícula de Coronavírus/imunologia , Adulto Jovem
19.
Lancet Infect Dis ; 21(10): 1383-1394, 2021 10.
Artigo em Inglês | MEDLINE | ID: mdl-33887208

RESUMO

BACKGROUND: Given the scale of the ongoing COVID-19 pandemic, the development of vaccines based on different platforms is essential, particularly in light of emerging viral variants, the absence of information on vaccine-induced immune durability, and potential paediatric use. We aimed to assess the safety and immunogenicity of an MF59-adjuvanted subunit vaccine for COVID-19 based on recombinant SARS-CoV-2 spike glycoprotein stabilised in a pre-fusion conformation by a novel molecular clamp (spike glycoprotein-clamp [sclamp]). METHODS: We did a phase 1, double-blind, placebo-controlled, block-randomised trial of the sclamp subunit vaccine in a single clinical trial site in Brisbane, QLD, Australia. Healthy adults (aged ≥18 to ≤55 years) who had tested negative for SARS-CoV-2, reported no close contact with anyone with active or previous SARS-CoV-2 infection, and tested negative for pre-existing SARS-CoV-2 immunity were included. Participants were randomly assigned to one of five treatment groups and received two doses via intramuscular injection 28 days apart of either placebo, sclamp vaccine at 5 µg, 15 µg, or 45 µg, or one dose of sclamp vaccine at 45 µg followed by placebo. Participants and study personnel, except the dose administration personnel, were masked to treatment. The primary safety endpoints included solicited local and systemic adverse events in the 7 days after each dose and unsolicited adverse events up to 12 months after dosing. Here, data are reported up until day 57. Primary immunogenicity endpoints were antigen-specific IgG ELISA and SARS-CoV-2 microneutralisation assays assessed at 28 days after each dose. The study is ongoing and registered with ClinicalTrials.gov, NCT04495933. FINDINGS: Between June 23, 2020, and Aug 17, 2020, of 314 healthy volunteers screened, 120 were randomly assigned (n=24 per group), and 114 (95%) completed the study up to day 57 (mean age 32·5 years [SD 10·4], 65 [54%] male, 55 [46%] female). Severe solicited reactions were infrequent and occurred at similar rates in participants receiving placebo (two [8%] of 24) and the SARS-CoV-2 sclamp vaccine at any dose (three [3%] of 96). Both solicited reactions and unsolicited adverse events occurred at a similar frequency in participants receiving placebo and the SARS-CoV-2 sclamp vaccine. Solicited reactions occurred in 19 (79%) of 24 participants receiving placebo and 86 (90%) of 96 receiving the SARS-CoV-2 sclamp vaccine at any dose. Unsolicited adverse events occurred in seven (29%) of 24 participants receiving placebo and 35 (36%) of 96 participants receiving the SARS-CoV-2 sclamp vaccine at any dose. Vaccination with SARS-CoV-2 sclamp elicited a similar antigen-specific response irrespective of dose: 4 weeks after the initial dose (day 29) with 5 µg dose (geometric mean titre [GMT] 6400, 95% CI 3683-11 122), with 15 µg dose (7492, 4959-11 319), and the two 45 µg dose cohorts (8770, 5526-13 920 in the two-dose 45 µg cohort; 8793, 5570-13 881 in the single-dose 45 µg cohort); 4 weeks after the second dose (day 57) with two 5 µg doses (102 400, 64 857-161 676), with two 15 µg doses (74 725, 51 300-108 847), with two 45 µg doses (79 586, 55 430-114 268), only a single 45 µg dose (4795, 2858-8043). At day 57, 67 (99%) of 68 participants who received two doses of sclamp vaccine at any concentration produced a neutralising immune response, compared with six (25%) of 24 who received a single 45 µg dose and none of 22 who received placebo. Participants receiving two doses of sclamp vaccine elicited similar neutralisation titres, irrespective of dose: two 5 µg doses (GMT 228, 95% CI 146-356), two 15 µg doses (230, 170-312), and two 45 µg doses (239, 187-307). INTERPRETATION: This first-in-human trial shows that a subunit vaccine comprising mammalian cell culture-derived, MF59-adjuvanted, molecular clamp-stabilised recombinant spike protein elicits strong immune responses with a promising safety profile. However, the glycoprotein 41 peptide present in the clamp created HIV diagnostic assay interference, a possible barrier to widespread use highlighting the criticality of potential non-spike directed immunogenicity during vaccine development. Studies are ongoing with alternative molecular clamp trimerisation domains to ameliorate this response. FUNDING: Coalition for Epidemic Preparedness Innovations, National Health and Medical Research Council, Queensland Government, and further philanthropic sources listed in the acknowledgments.


Assuntos
Adjuvantes Imunológicos/farmacologia , Vacinas contra COVID-19/administração & dosagem , COVID-19/prevenção & controle , Glicoproteína da Espícula de Coronavírus/imunologia , Esqualeno/imunologia , Adulto , Anticorpos Neutralizantes/sangue , Anticorpos Antivirais/sangue , Austrália , Feminino , Voluntários Saudáveis , Humanos , Masculino , Pandemias/prevenção & controle , Polissorbatos , Vacinação/efeitos adversos , Adulto Jovem
20.
Clin Transl Immunology ; 10(3): e1258, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33680466

RESUMO

Objectives: As the world transitions into a new era of the COVID-19 pandemic in which vaccines become available, there is an increasing demand for rapid reliable serological testing to identify individuals with levels of immunity considered protective by infection or vaccination. Methods: We used 34 SARS-CoV-2 samples to perform a rapid surrogate virus neutralisation test (sVNT), applicable to many laboratories as it circumvents the need for biosafety level-3 containment. We correlated results from the sVNT with five additional commonly used SARS-CoV-2 serology techniques: the microneutralisation test (MNT), in-house ELISAs, commercial Euroimmun- and Wantai-based ELISAs (RBD, spike and nucleoprotein; IgG, IgA and IgM), antigen-binding avidity, and high-throughput multiplex analyses to profile isotype, subclass and Fc effector binding potential. We correlated antibody levels with antibody-secreting cell (ASC) and circulatory T follicular helper (cTfh) cell numbers. Results: Antibody data obtained with commercial ELISAs closely reflected results using in-house ELISAs against RBD and spike. A correlation matrix across ten measured ELISA parameters revealed positive correlations for all factors. The frequency of inhibition by rapid sVNT strongly correlated with spike-specific IgG and IgA titres detected by both commercial and in-house ELISAs, and MNT titres. Multiplex analyses revealed strongest correlations between IgG, IgG1, FcR and C1q specific to spike and RBD. Acute cTfh-type 1 cell numbers correlated with spike and RBD-specific IgG antibodies measured by ELISAs and sVNT. Conclusion: Our comprehensive analyses provide important insights into SARS-CoV-2 humoral immunity across distinct serology assays and their applicability for specific research and/or diagnostic questions to assess SARS-CoV-2-specific humoral responses.

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