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1.
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.

2.
Medicine (Baltimore) ; 100(18): e25606, 2021 May 07.
Artigo em Inglês | MEDLINE | ID: mdl-33950937

RESUMO

BACKGROUND: Osteoporosis (OP) is an age-related disease characterized by reduced bone mass and increased bone fragility. It is more common in older people and postmenopausal women. As a new type of exercise training for OP, whole-body vibration (WBV) exercise has been proved to have a good effect on postmenopausal women with OP. It can increase bone density and improve strength and balance in postmenopausal population, which has certain clinical value, but lacks evidence-based medicine evidence. This study aims to systematically study the effectiveness of WBV exercise on postmenopausal women with OP. METHODS: The English databases (PubMed, Embase, Web of Science, The Cochrane Library) and Chinese databases (China National Knowledge Network, Wanfang, Weipu, China Biomedical Database) were searched by computer. From the establishment of the database to February 2021, the randomized controlled clinical studies on WBV exercise on postmenopausal women with OP were conducted. The quality of the included studies was independently extracted by 2 researchers and literature quality was evaluated. Meta-analysis of the included studies was performed using RevMan5.3 software. RESULTS: In this study, the efficacy and safety of WBV exercise on postmenopausal women with OP were evaluated by lumbar spine bone density, femoral neck bone density, pain, incidence of falls, incidence of fractures, and quality of life scale score, etc. CONCLUSION: This study will provide reliable evidences for the clinical application of WBV exercise on postmenopausal women with OP. ETHICS AND DISSEMINATION: Private information from individuals will not be published. This systematic review also does not involve endangering participant rights. Ethical approval will not be required. The results may be published in a peer-reviewed journal or disseminated at relevant conferences. OSF REGISTRATION NUMBER: DOI 10.17605/OSF.IO/WPYT9.


Assuntos
Terapia por Exercício/métodos , Osteoporose Pós-Menopausa/terapia , Fraturas por Osteoporose/epidemiologia , Vibração/uso terapêutico , Densidade Óssea/fisiologia , Medicina Baseada em Evidências/métodos , Terapia por Exercício/efeitos adversos , Feminino , Humanos , Incidência , Metanálise como Assunto , Osteoporose Pós-Menopausa/complicações , Osteoporose Pós-Menopausa/fisiopatologia , Fraturas por Osteoporose/etiologia , Fraturas por Osteoporose/fisiopatologia , Fraturas por Osteoporose/prevenção & controle , Qualidade de Vida , Ensaios Clínicos Controlados Aleatórios como Assunto , Revisões Sistemáticas como Assunto , Resultado do Tratamento
3.
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
4.
ERJ Open Res ; 7(2)2021 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-33829055

RESUMO

Severe COVID-19 patient airways plugged by MUC5AC-containing mucus exhibit hyperplasia of goblet cells, and hypoplasia of multiciliated cells and club cells, as well as significantly reduced CC16 and MUC5B levels, and increased IL-13 levels https://bit.ly/2M2NcdO.

5.
Nat Commun ; 12(1): 1724, 2021 03 19.
Artigo em Inglês | MEDLINE | ID: mdl-33741972

RESUMO

T-cell immunity is important for recovery from COVID-19 and provides heightened immunity for re-infection. However, little is known about the SARS-CoV-2-specific T-cell immunity in virus-exposed individuals. Here we report virus-specific CD4+ and CD8+ T-cell memory in recovered COVID-19 patients and close contacts. We also demonstrate the size and quality of the memory T-cell pool of COVID-19 patients are larger and better than those of close contacts. However, the proliferation capacity, size and quality of T-cell responses in close contacts are readily distinguishable from healthy donors, suggesting close contacts are able to gain T-cell immunity against SARS-CoV-2 despite lacking a detectable infection. Additionally, asymptomatic and symptomatic COVID-19 patients contain similar levels of SARS-CoV-2-specific T-cell memory. Overall, this study demonstrates the versatility and potential of memory T cells from COVID-19 patients and close contacts, which may be important for host protection.


Assuntos
Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , COVID-19/imunologia , Memória Imunológica/imunologia , SARS-CoV-2/imunologia , Viroses/diagnóstico , Anticorpos Antivirais/imunologia , Infecções Assintomáticas , COVID-19/sangue , Estudos de Casos e Controles , Humanos , Interferon gama/biossíntese , Interferon gama/imunologia
6.
J Thorac Dis ; 12(5): 1811-1823, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-32642086

RESUMO

Background: Coronavirus disease 2019 (COVID-19) has been a global pandemic disease, with more than 4 million cases and nearly 300,000 deaths. Little is known about COVID-19 in patients with chronic obstructive pulmonary disease (COPD). We aimed to evaluate the influence of preexisting COPD on the progress and outcomes of COVID-19. Methods: This was a multicenter, retrospective, observational study. We enrolled 1,048 patients aged 40 years and above, including 50 patients with COPD and 998 patients without COPD, and with COVID-19 confirmed via high-throughput sequencing or real-time reverse transcription-polymerase chain reaction, between December 11, 2019 and February 20, 2020. We collected data of demographics, pathologic test results, radiologic imaging, and treatments. The primary outcomes were composite endpoints determined by admission to an intensive care unit, the use of mechanical ventilation, or death. Results: Compared with patients who had COVID-19 but not COPD, those with COPD had higher rates of fatigue (56.0% vs. 40.2%), dyspnea (66.0% vs. 26.3%), diarrhea (16.0% vs. 3.6%), and unconsciousness (8.0% vs. 1.7%) and a significantly higher proportion of increased activated partial thromboplastin time (23.5% vs. 5.2%) and D-dimer (65.9% vs. 29.3%), as well as ground-glass opacities (77.6% vs. 60.3%), local patchy shadowing (61.2% vs. 41.4%), and interstitial abnormalities (51.0% vs. 19.8%) on chest computed tomography. Patients with COPD were more likely to develop bacterial or fungal coinfection (20.0% vs. 5.9%), acute respiratory distress syndrome (ARDS) (20.0% vs. 7.3%), septic shock (14.0% vs. 2.3%), or acute renal failure (12.0% vs. 1.3%). Patients with COPD and COVID-19 had a higher risk of reaching the composite endpoints [hazard ratio (HR): 2.17, 95% confidence interval (CI): 1.40-3.38; P=0.001] or death (HR: 2.28, 95% CI: 1.15-4.51; P=0.019), after adjustment. Conclusions: In this study, patients with COPD who developed COVID-19 showed a higher risk of admission to the intensive care unit, mechanical ventilation, or death.

8.
Sci Immunol ; 5(48)2020 06 26.
Artigo em Inglês | MEDLINE | ID: mdl-32591409

RESUMO

Liver resident-memory CD8+ T cells (TRM cells) can kill liver-stage Plasmodium-infected cells and prevent malaria, but simple vaccines for generating this important immune population are lacking. Here, we report the development of a fully synthetic self-adjuvanting glycolipid-peptide conjugate vaccine designed to efficiently induce liver TRM cells. Upon cleavage in vivo, the glycolipid-peptide conjugate vaccine releases an MHC I-restricted peptide epitope (to stimulate Plasmodium-specific CD8+ T cells) and an adjuvant component, the NKT cell agonist α-galactosylceramide (α-GalCer). A single dose of this vaccine in mice induced substantial numbers of intrahepatic malaria-specific CD8+ T cells expressing canonical markers of liver TRM cells (CD69, CXCR6, and CD101), and these cells could be further increased in number upon vaccine boosting. We show that modifications to the peptide, such as addition of proteasomal-cleavage sequences or epitope-flanking sequences, or the use of alternative conjugation methods to link the peptide to the glycolipid improved liver TRM cell generation and led to the development of a vaccine able to induce sterile protection in C57BL/6 mice against Plasmodium berghei sporozoite challenge after a single dose. Furthermore, this vaccine induced endogenous liver TRM cells that were long-lived (half-life of ~425 days) and were able to maintain >90% sterile protection to day 200. Our findings describe an ideal synthetic vaccine platform for generating large numbers of liver TRM cells for effective control of liver-stage malaria and, potentially, a variety of other hepatotropic infections.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Glicolipídeos/imunologia , Fígado/imunologia , Vacinas Antimaláricas/imunologia , Malária/imunologia , Peptídeos/imunologia , Animais , Linfócitos T CD8-Positivos/patologia , Fígado/patologia , Malária/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Vacinação
9.
Influenza Other Respir Viruses ; 14(6): 678-687, 2020 11.
Artigo em Inglês | MEDLINE | ID: mdl-32588557

RESUMO

BACKGROUND: Influenza viruses cause significant morbidity and mortality, especially in young children, elderly, pregnant women and individuals with co-morbidities. Patients with severe influenza disease are typically treated with one neuraminidase inhibitor, oseltamivir or zanamivir. These antivirals need to be taken early to be most effective and often lead to the emergence of drug resistance and/or decreased drug susceptibility. Combining oseltamivir with another antiviral with an alternative mode of action has the potential to improve clinical effectiveness and reduce drug resistance. METHODS: In this study, we utilized a host-targeting molecule RM-5061, a second-generation thiazolide, in combination with oseltamivir to determine whether these compounds could reduce viral burden and understand their effects on the immune response to influenza virus infection in mice, compared with either monotherapy or placebo. RESULTS: The combination of RM-5061 and OST administered for 5 days after influenza infection reduced viral burden at day 5 post-infection, when compared to placebo and RM-5061 monotherapy, but was not significantly different from oseltamivir monotherapy. The inflammatory cytokine milieu was also reduced in animals which received a combination therapy when compared to RM-5061 and placebo-treated animals. Antiviral treatment in all groups led to a reduction in CD8+ T-cell responses in the BAL when compared to placebo. CONCLUSIONS: To our knowledge, this is the first time a combination of a host-targeting compound, RM-5061, and neuraminidase inhibitor, OST, has been tested in vivo. This antiviral combination was safe in mice and led to reduced inflammatory responses following viral infection when compared to untreated animals.

10.
J Virol ; 94(15)2020 07 16.
Artigo em Inglês | MEDLINE | ID: mdl-32434886

RESUMO

Middle East respiratory syndrome coronavirus (MERS-CoV) causes severe acute respiratory disease in humans. MERS-CoV strains from early epidemic clade A and contemporary epidemic clade B have not been phenotypically characterized to compare their abilities to infect cells and mice. We isolated the clade B MERS-CoV ChinaGD01 strain from a patient infected during the South Korean MERS outbreak in 2015 and compared the phylogenetics and pathogenicity of MERS-CoV EMC/2012 (clade A) and ChinaGD01 (clade B) in vitro and in vivo Genome alignment analysis showed that most clade-specific mutations occurred in the orf1ab gene, including mutations that were predicted to be potential glycosylation sites. Minor differences in viral growth but no significant differences in plaque size or sensitivity to beta interferon (IFN-ß) were detected between these two viruses in vitro ChinaGD01 virus infection induced more weight loss and inflammatory cytokine production in human DPP4-transduced mice. Viral titers were higher in the lungs of ChinaGD01-infected mice than with EMC/2012 infection. Decreased virus-specific CD4+ and CD8+ T cell numbers were detected in the lungs of ChinaGD01-infected mice. In conclusion, MERS-CoV evolution induced changes to reshape its pathogenicity and virulence in vitro and in vivo and to evade adaptive immune response to hinder viral clearance.IMPORTANCE MERS-CoV is an important emerging pathogen and causes severe respiratory infection in humans. MERS-CoV strains from early epidemic clade A and contemporary epidemic clade B have not been phenotypically characterized to compare their abilities to infect cells and mice. In this study, we showed that a clade B virus ChinaGD01 strain caused more severe disease in mice, with delayed viral clearance, increased inflammatory cytokines, and decreased antiviral T cell responses, than the early clade A virus EMC/2012. Given the differences in pathogenicity of different clades of MERS-CoV, periodic assessment of currently circulating MERS-CoV is needed to monitor potential severity of zoonotic disease.


Assuntos
Infecções por Coronavirus/virologia , Genótipo , Interações Hospedeiro-Patógeno , Coronavírus da Síndrome Respiratória do Oriente Médio/fisiologia , Adulto , Animais , Modelos Animais de Doenças , Genoma Viral , Interações Hospedeiro-Patógeno/imunologia , Humanos , Interferon Tipo I/farmacologia , Masculino , Camundongos , Coronavírus da Síndrome Respiratória do Oriente Médio/classificação , Coronavírus da Síndrome Respiratória do Oriente Médio/isolamento & purificação , Coronavírus da Síndrome Respiratória do Oriente Médio/patogenicidade , Filogenia , RNA Viral , Linfócitos T/imunologia , Linfócitos T/metabolismo , Virulência , Replicação Viral/efeitos dos fármacos , Replicação Viral/genética , Sequenciamento Completo do Genoma
11.
Clin Transl Immunology ; 8(9): e1079, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31559018

RESUMO

Background: Although γδ T cells comprise up to 10% of human peripheral blood T cells, questions remain regarding their role in disease states and T-cell receptor (TCR) clonal expansions. We dissected anti-viral functions of human γδ T cells towards influenza viruses and defined influenza-reactive γδ TCRs in the context of γδ-TCRs across the human lifespan. Methods: We performed 51Cr-killing assay and single-cell time-lapse live video microscopy to define mechanisms underlying γδ T-cell-mediated killing of influenza-infected targets. We assessed cytotoxic profiles of γδ T cells in influenza-infected patients and IFN-γ production towards influenza-infected lung epithelial cells. Using single-cell RT-PCR, we characterised paired TCRγδ clonotypes for influenza-reactive γδ T cells in comparison with TCRs from healthy neonates, adults, elderly donors and tissues. Results: We provide the first visual evidence of γδ T-cell-mediated killing of influenza-infected targets and show distinct features to those reported for CD8+ T cells. γδ T cells displayed poly-cytotoxic profiles in influenza-infected patients and produced IFN-γ towards influenza-infected cells. These IFN-γ-producing γδ T cells were skewed towards the γ9δ2 TCRs, particularly expressing the public GV9-TCRγ, capable of pairing with numerous TCR-δ chains, suggesting their significant role in γδ T-cell immunity. Neonatal γδ T cells displayed extensive non-overlapping TCRγδ repertoires, while adults had enriched γ9δ2-pairings with diverse CDR3γδ regions. Conversely, the elderly showed distinct γδ-pairings characterised by large clonal expansions, a profile also prominent in adult tissues. Conclusion: Human TCRγδ repertoire is shaped by age, tissue compartmentalisation and the individual's history of infection, suggesting that these somewhat enigmatic γδ T cells indeed respond to antigen challenge.

12.
Nat Commun ; 9(1): 4706, 2018 11 09.
Artigo em Inglês | MEDLINE | ID: mdl-30413689

RESUMO

Mucosal associated invariant T (MAIT) cells are evolutionarily-conserved, innate-like lymphocytes which are abundant in human lungs and can contribute to protection against pulmonary bacterial infection. MAIT cells are also activated during human viral infections, yet it remains unknown whether MAIT cells play a significant protective or even detrimental role during viral infections in vivo. Using murine experimental challenge with two strains of influenza A virus, we show that MAIT cells accumulate and are activated early in infection, with upregulation of CD25, CD69 and Granzyme B, peaking at 5 days post-infection. Activation is modulated via cytokines independently of MR1. MAIT cell-deficient MR1-/- mice show enhanced weight loss and mortality to severe (H1N1) influenza. This is ameliorated by prior adoptive transfer of pulmonary MAIT cells in both immunocompetent and immunodeficient RAG2-/-γC-/- mice. Thus, MAIT cells contribute to protection during respiratory viral infections, and constitute a potential target for therapeutic manipulation.


Assuntos
Influenza Humana/patologia , Influenza Humana/virologia , Células T Invariantes Associadas à Mucosa/virologia , Infecções por Orthomyxoviridae/patologia , Infecções por Orthomyxoviridae/virologia , Transferência Adotiva , Animais , Citocinas/metabolismo , Antígenos de Histocompatibilidade Classe I/metabolismo , Humanos , Pulmão/patologia , Camundongos Endogâmicos C57BL , Antígenos de Histocompatibilidade Menor/metabolismo
13.
Front Immunol ; 9: 1453, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29997621

RESUMO

CD8+ T cells recognizing antigenic peptides derived from conserved internal viral proteins confer broad protection against distinct influenza viruses. As memory CD8+ T cells change throughout the human lifetime and across tissue compartments, we investigated how T cell receptor (TCR) composition and diversity relate to memory CD8+ T cells across anatomical sites and immunological phases of human life. We used ex vivo peptide-HLA tetramer magnetic enrichment, single-cell multiplex RT-PCR for both the TCR-alpha (TCRα) and TCR-beta (TCRß) chains, and new TCRdist and grouping of lymphocyte interactions by paratope hotspots (GLIPH) algorithms to compare TCRs directed against the most prominent human influenza epitope, HLA-A*02:01-M158-66 (A2+M158). We dissected memory TCR repertoires directed toward A2+M158 CD8+ T cells within human tissues and compared them to human peripheral blood of young and elderly adults. Furthermore, we compared these memory CD8+ T cell repertoires to A2+M158 CD8+ TCRs during acute influenza disease in patients hospitalized with avian A/H7N9 virus. Our study provides the first ex vivo comparative analysis of paired antigen-specific TCR-α/ß clonotypes across different tissues and peripheral blood across different age groups. We show that human A2+M158 CD8+ T cells can be readily detected in human lungs, spleens, and lymph nodes, and that tissue A2+M158 TCRαß repertoires reflect A2+M158 TCRαß clonotypes derived from peripheral blood in healthy adults and influenza-infected patients. A2+M158 TCRαß repertoires displayed distinct features only in elderly adults, with large private TCRαß clonotypes replacing the prominent and public TRBV19/TRAV27 TCRs. Our study provides novel findings on influenza-specific TCRαß repertoires within human tissues, raises the question of how we can prevent the loss of optimal TCRαß signatures with aging, and provides important insights into the rational design of T cell-mediated vaccines and immunotherapies.

14.
Nat Commun ; 9(1): 824, 2018 02 26.
Artigo em Inglês | MEDLINE | ID: mdl-29483513

RESUMO

Severe influenza A virus (IAV) infection is associated with immune dysfunction. Here, we show circulating CD8+ T-cell profiles from patients hospitalized with avian H7N9, seasonal IAV, and influenza vaccinees. Patient survival reflects an early, transient prevalence of highly activated CD38+HLA-DR+PD-1+ CD8+ T cells, whereas the prolonged persistence of this set is found in ultimately fatal cases. Single-cell T cell receptor (TCR)-αß analyses of activated CD38+HLA-DR+CD8+ T cells show similar TCRαß diversity but differential clonal expansion kinetics in surviving and fatal H7N9 patients. Delayed clonal expansion associated with an early dichotomy at a transcriptome level (as detected by single-cell RNAseq) is found in CD38+HLA-DR+CD8+ T cells from patients who succumbed to the disease, suggesting a divergent differentiation pathway of CD38+HLA-DR+CD8+ T cells from the outset during fatal disease. Our study proposes that effective expansion of cross-reactive influenza-specific TCRαß clonotypes with appropriate transcriptome signatures is needed for early protection against severe influenza disease.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Seleção Clonal Mediada por Antígeno/genética , Subtipo H7N9 do Vírus da Influenza A/patogenicidade , Influenza Humana/imunologia , Receptores de Antígenos de Linfócitos T alfa-beta/imunologia , Subpopulações de Linfócitos T/imunologia , Transcriptoma/imunologia , ADP-Ribosil Ciclase 1/genética , ADP-Ribosil Ciclase 1/imunologia , Linfócitos T CD8-Positivos/patologia , Linfócitos T CD8-Positivos/virologia , Estudos de Coortes , Estado Terminal , Regulação da Expressão Gênica , Antígenos HLA-DR/genética , Antígenos HLA-DR/imunologia , Hospitalização , Humanos , Subtipo H7N9 do Vírus da Influenza A/imunologia , Influenza Humana/genética , Influenza Humana/mortalidade , Influenza Humana/virologia , Ativação Linfocitária , Glicoproteínas de Membrana/genética , Glicoproteínas de Membrana/imunologia , Receptor de Morte Celular Programada 1/genética , Receptor de Morte Celular Programada 1/imunologia , Receptores de Antígenos de Linfócitos T alfa-beta/genética , Análise de Sobrevida , Subpopulações de Linfócitos T/patologia , Subpopulações de Linfócitos T/virologia
15.
ACS Chem Biol ; 12(11): 2898-2905, 2017 11 17.
Artigo em Inglês | MEDLINE | ID: mdl-29043774

RESUMO

The development of a universal vaccine for influenza A virus (IAV) that does not require seasonal modification is a long-standing health goal, particularly in the context of the increasing threat of new global pandemics. Vaccines that specifically induce T cell responses are of considerable interest because they can target viral proteins that are more likely to be shared between different virus strains and subtypes and hence provide effective cross-reactive IAV immunity. From a practical perspective, such vaccines should induce T cell responses with long-lasting memory, while also being simple to manufacture and cost-effective. Here we describe the synthesis and evaluation of a vaccine platform based on solid phase peptide synthesis and bio-orthogonal conjugation methodologies. The chemical approach involves covalently attaching synthetic long peptides from a virus-associated protein to a powerful adjuvant molecule, α-galactosylceramide (α-GalCer). Strain-promoted azide-alkyne cycloaddition is used as a simple and efficient method for conjugation, and pseudoproline methodology is used to increase the efficiency of the peptide synthesis. α-GalCer is a glycolipid that stimulates NKT cells, a population of lymphoid-resident immune cells that can provide potent stimulatory signals to antigen-presenting cells engaged in driving proliferation and differentiation of peptide-specific T cells. When used in mice, the vaccine induced T cell responses that provided effective prophylactic protection against IAV infection, with the speed of viral clearance greater than that seen from previous viral exposure. These findings are significant because the vaccines are highly defined, quick to synthesize, and easily characterized and are therefore appropriate for large scale affordable manufacture.


Assuntos
Adjuvantes Imunológicos/uso terapêutico , Galactosilceramidas/uso terapêutico , Vírus da Influenza A/imunologia , Vacinas contra Influenza/uso terapêutico , Infecções por Orthomyxoviridae/prevenção & controle , Peptídeos/uso terapêutico , Adjuvantes Imunológicos/síntese química , Adjuvantes Imunológicos/farmacologia , Animais , Linfócitos T CD8-Positivos/imunologia , Reação de Cicloadição , Feminino , Galactosilceramidas/síntese química , Galactosilceramidas/imunologia , Humanos , Vírus da Influenza A/química , Vacinas contra Influenza/síntese química , Influenza Humana/imunologia , Influenza Humana/prevenção & controle , Camundongos Endogâmicos C57BL , Células T Matadoras Naturais/imunologia , Infecções por Orthomyxoviridae/imunologia , Peptídeos/síntese química , Peptídeos/imunologia , Técnicas de Síntese em Fase Sólida
17.
Proc Natl Acad Sci U S A ; 114(20): 5225-5230, 2017 05 16.
Artigo em Inglês | MEDLINE | ID: mdl-28461487

RESUMO

The lymphoid tissue that drains the upper respiratory tract represents an important induction site for cytotoxic T lymphocyte (CTL) immunity to airborne pathogens and intranasal vaccines. Here, we investigated the role of the nasal-associated lymphoid tissues (NALTs), which are mucosal-associated lymphoid organs embedded in the submucosa of the nasal passage, in the initial priming and recall expansion of CD8+ T cells following an upper respiratory tract infection with a pathogenic influenza virus and immunization with a live attenuated influenza virus vaccine. Whereas NALTs served as the induction site for the recall expansion of memory CD8+ T cells following influenza virus infection or vaccination, they failed to support activation of naïve CD8+ T cells. Strikingly, NALTs, unlike other lymphoid tissues, were not routinely surveyed during the steady state by circulating T cells. The selective recruitment of memory T cells into these lymphoid structures occurred in response to infection-induced elevation of the chemokine CXCL10, which attracted CXCR3+ memory CD8+ T cells. These results have significant implications for intranasal vaccines, which deliver antigen to mucosal-associated lymphoid tissue and aim to elicit protective CTL-mediated immunity.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Imunidade nas Mucosas/imunologia , Linfócitos T Citotóxicos/imunologia , Administração Intranasal , Animais , Imunização , Vírus da Influenza A/imunologia , Vacinas contra Influenza/imunologia , Linfonodos/fisiologia , Tecido Linfoide/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Mucosa Nasal/metabolismo , Mucosa Nasal/fisiologia , Infecções por Orthomyxoviridae/imunologia , Infecções Respiratórias , Vacinação
18.
J Gen Virol ; 97(12): 3205-3214, 2016 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-27902386

RESUMO

FluMist has been used in children and adults for more than 10 years. As pre-existing CD8+ T cell memory pools can provide heterologous immunity against distinct influenza viruses, it is important to understand influenza-specific CD8+ T cell responses elicited by different live attenuated influenza virus (LAIV) regimens. In this study, we immunized mice intranasally with two different doses of live-attenuated PR8 virus (PR8 ts, H1N1), low and high, and then assessed protective efficacy by challenging animals with heterosubtypic X31-H3N2 virus at 6 weeks post-vaccination. Different LAIV doses elicited influenza-specific CD8+ T cell responses in lungs and spleen, but unexpectedly not in bronchoalveolar lavage. Interestingly, the immunodominance hierarchy at the acute phase after immunization varied depending on the LAIV dose; however, these differences disappeared at 6 weeks post-vaccination, resulting in generation of comparable CD8+ T cell memory pools. After vaccination with either dose, sufficient numbers of specific CD8+ T cells were generated for recall and protection of mice against heterosubtypic H1N1→H3N2 challenge. As a result, immunized mice displayed reduced weight loss, diminished inflammatory responses and lower viral titres in lungs, when compared to unvaccinated animals. Interestingly, the higher dose led to enhanced viral clearance on day 5 post-challenge, though this was not associated with increased CD8+ T cell responses, but with higher levels of non-neutralizing antibodies against the priming virus. Our study suggests that, while different LAIV doses result in distinct immune profiles, even a low dose produces sufficient protective CD8+ T cell memory against challenge infection, though the high dose results in more rapid viral clearance and reduced inflammation.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Vírus da Influenza A Subtipo H1N1/imunologia , Vírus da Influenza A Subtipo H3N2/imunologia , Vacinas contra Influenza/administração & dosagem , Influenza Humana/imunologia , Vacinas Atenuadas/administração & dosagem , Animais , Anticorpos Antivirais/imunologia , Humanos , Memória Imunológica , Vírus da Influenza A Subtipo H1N1/genética , Vírus da Influenza A Subtipo H3N2/genética , Vacinas contra Influenza/imunologia , Influenza Humana/prevenção & controle , Influenza Humana/virologia , Pulmão/virologia , Camundongos , Camundongos Endogâmicos C57BL , Vacinas Atenuadas/imunologia
19.
Proc Natl Acad Sci U S A ; 113(36): 10133-8, 2016 09 06.
Artigo em Inglês | MEDLINE | ID: mdl-27543331

RESUMO

Mucosal-associated invariant T (MAIT) cells are innate-like T lymphocytes known to elicit potent immunity to a broad range of bacteria, mainly via the rapid production of inflammatory cytokines. Whether MAIT cells contribute to antiviral immunity is less clear. Here we asked whether MAIT cells produce cytokines/chemokines during severe human influenza virus infection. Our analysis in patients hospitalized with avian H7N9 influenza pneumonia showed that individuals who recovered had higher numbers of CD161(+)Vα7.2(+) MAIT cells in peripheral blood compared with those who succumbed, suggesting a possible protective role for this lymphocyte population. To understand the mechanism underlying MAIT cell activation during influenza, we cocultured influenza A virus (IAV)-infected human lung epithelial cells (A549) and human peripheral blood mononuclear cells in vitro, then assayed them by intracellular cytokine staining. Comparison of influenza-induced MAIT cell activation with the profile for natural killer cells (CD56(+)CD3(-)) showed robust up-regulation of IFNγ for both cell populations and granzyme B in MAIT cells, although the individual responses varied among healthy donors. However, in contrast to the requirement for cell-associated factors to promote NK cell activation, the induction of MAIT cell cytokine production was dependent on IL-18 (but not IL-12) production by IAV-exposed CD14(+) monocytes. Overall, this evidence for IAV activation via an indirect, IL-18-dependent mechanism indicates that MAIT cells are protective in influenza, and also possibly in any human disease process in which inflammation and IL-18 production occur.


Assuntos
Imunidade nas Mucosas , Subtipo H7N9 do Vírus da Influenza A/patogenicidade , Influenza Humana/imunologia , Interleucina-18/imunologia , Células T Invariantes Associadas à Mucosa/imunologia , Pneumonia Viral/imunologia , Células A549 , Comunicação Celular/imunologia , Técnicas de Cocultura , Expressão Gênica , Granzimas/genética , Granzimas/imunologia , Humanos , Imunidade Inata , Imunofenotipagem , Subtipo H7N9 do Vírus da Influenza A/imunologia , Influenza Humana/mortalidade , Influenza Humana/patologia , Influenza Humana/virologia , Interferon gama/genética , Interferon gama/imunologia , Interleucina-18/genética , Células Matadoras Naturais/imunologia , Células Matadoras Naturais/virologia , Leucócitos Mononucleares/imunologia , Leucócitos Mononucleares/virologia , Ativação Linfocitária , Monócitos/imunologia , Monócitos/virologia , Células T Invariantes Associadas à Mucosa/virologia , Subfamília B de Receptores Semelhantes a Lectina de Células NK/genética , Subfamília B de Receptores Semelhantes a Lectina de Células NK/imunologia , Pneumonia Viral/mortalidade , Pneumonia Viral/patologia , Pneumonia Viral/virologia , Transdução de Sinais , Análise de Sobrevida
20.
J Virol ; 90(15): 6936-6947, 2016 08 01.
Artigo em Inglês | MEDLINE | ID: mdl-27226365

RESUMO

UNLABELLED: Novel influenza viruses often cause differential infection patterns across different age groups, an effect that is defined as heterogeneous demographic susceptibility. This occurred during the A/H2N2 pandemic, when children experienced higher influenza attack rates than adults. Since the recognition of conserved epitopes across influenza subtypes by CD8(+) cytotoxic T lymphocytes (CTLs) limit influenza disease, we hypothesized that conservation of CTL antigenic peptides (Ag-p) in viruses circulating before the pH2N2-1957 may have resulted in differential CTL immunity. We compared viruses isolated in the years preceding the pandemic (1941 to 1957) to which children and adults were exposed to viruses circulating decades earlier (1918 to 1940), which could infect adults only. Consistent with phylogenetic models, influenza viruses circulating from 1941 to 1957, which infected children, shared with pH2N2 the majority (∼89%) of the CTL peptides within the most immunogenic nucleoprotein, matrix 1, and polymerase basic 1, thus providing evidence for minimal pH2N2 CTL escape in children. Our study, however, identified potential CTL immune evasion from pH2N2 irrespective of age, within HLA-A*03:01(+) individuals for PB1471-L473V/N476I variants and HLA-B*15:01(+) population for NP404-414-V408I mutant. Further experiments using the murine model of B-cell-deficient mice showed that multiple influenza infections resulted in superior protection from influenza-induced morbidity, coinciding with accumulation of tissue-resident memory CD8(+) T cells in the lung. Our study suggests that protection against H2N2-1957 pandemic influenza was most likely linked to the number of influenza virus infections prior to the pandemic challenge rather than differential preexisting CTL immunity. Thus, the regimen of a CTL-based vaccine/vaccine-component may benefit from periodic boosting to achieve fully protective, asymptomatic influenza infection. IMPORTANCE: Due to a lack of cross-reactive neutralizing antibodies, children are particularly susceptible to influenza infections caused by novel viral strains. Preexisting T cell immunity directed at conserved viral regions, however, can provide protection against influenza viruses, promote rapid recovery and better clinical outcomes. When we asked whether high susceptibility of children (compared to adults) to the pandemic H2N2 influenza strain was associated with immune evasion from T-cell immunity, we found high conservation within T-cell antigenic regions in pandemic H2N2. However, the number of influenza infections prior to the challenge was linked to protective, asymptomatic infections and establishment of tissue-resident memory T cells. Our study supports development of vaccines that prime and boost T cells to elicit cross-strain protective T cells, especially tissue-resident memory T cells, for lifelong immunity against distinct influenza viruses.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Epitopos de Linfócito T/imunologia , Influenza Humana/imunologia , Infecções por Orthomyxoviridae/imunologia , Orthomyxoviridae/imunologia , Pandemias , Linfócitos T Citotóxicos/imunologia , Adulto , Animais , Linfócitos B/imunologia , Criança , Proteção Cruzada , Evolução Molecular , Feminino , Humanos , Influenza Humana/virologia , Camundongos , Infecções por Orthomyxoviridae/virologia , Filogenia
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