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2.
Science ; 364(6436)2019 04 12.
Artigo em Inglês | MEDLINE | ID: mdl-30975860

RESUMO

To understand the health impact of long-duration spaceflight, one identical twin astronaut was monitored before, during, and after a 1-year mission onboard the International Space Station; his twin served as a genetically matched ground control. Longitudinal assessments identified spaceflight-specific changes, including decreased body mass, telomere elongation, genome instability, carotid artery distension and increased intima-media thickness, altered ocular structure, transcriptional and metabolic changes, DNA methylation changes in immune and oxidative stress-related pathways, gastrointestinal microbiota alterations, and some cognitive decline postflight. Although average telomere length, global gene expression, and microbiome changes returned to near preflight levels within 6 months after return to Earth, increased numbers of short telomeres were observed and expression of some genes was still disrupted. These multiomic, molecular, physiological, and behavioral datasets provide a valuable roadmap of the putative health risks for future human spaceflight.


Assuntos
Adaptação Fisiológica , Astronautas , Voo Espacial , Imunidade Adaptativa , Peso Corporal , Artérias Carótidas/diagnóstico por imagem , Espessura Intima-Media Carotídea , Dano ao DNA , Metilação de DNA , Microbioma Gastrointestinal , Instabilidade Genômica , Humanos , Masculino , Homeostase do Telômero , Fatores de Tempo , Estados Unidos , United States National Aeronautics and Space Administration
4.
Nat Commun ; 9(1): 5229, 2018 Dec 06.
Artigo em Inglês | MEDLINE | ID: mdl-30523329

RESUMO

Analysis of sleep for the diagnosis of sleep disorders such as Type-1 Narcolepsy (T1N) currently requires visual inspection of polysomnography records by trained scoring technicians. Here, we used neural networks in approximately 3,000 normal and abnormal sleep recordings to automate sleep stage scoring, producing a hypnodensity graph-a probability distribution conveying more information than classical hypnograms. Accuracy of sleep stage scoring was validated in 70 subjects assessed by six scorers. The best model performed better than any individual scorer (87% versus consensus). It also reliably scores sleep down to 5 s instead of 30 s scoring epochs. A T1N marker based on unusual sleep stage overlaps achieved a specificity of 96% and a sensitivity of 91%, validated in independent datasets. Addition of HLA-DQB1*06:02 typing increased specificity to 99%. Our method can reduce time spent in sleep clinics and automates T1N diagnosis. It also opens the possibility of diagnosing T1N using home sleep studies.

5.
Proc Natl Acad Sci U S A ; 115(52): E12323-E12332, 2018 12 26.
Artigo em Inglês | MEDLINE | ID: mdl-30541895

RESUMO

Type 1 narcolepsy (T1N) is caused by hypocretin/orexin (HCRT) neuronal loss. Association with the HLA DQB1*06:02/DQA1*01:02 (98% vs. 25%) heterodimer (DQ0602), T cell receptors (TCR) and other immune loci suggest autoimmunity but autoantigens are unknown. Onset is seasonal and associated with influenza A, notably pandemic 2009 H1N1 (pH1N1) infection and vaccination (Pandemrix). Peptides derived from HCRT and influenza A, including pH1N1, were screened for DQ0602 binding and presence of cognate DQ0602 tetramer-peptide-specific CD4+ T cells tested in 35 T1N cases and 22 DQ0602 controls. Higher reactivity to influenza pHA273-287 (pH1N1 specific), PR8 (H1N1 pre-2009 and H2N2)-specific NP17-31 and C-amidated but not native version of HCRT54-66 and HCRT86-97 (HCRTNH2) were observed in T1N. Single-cell TCR sequencing revealed sharing of CDR3ß TRBV4-2-CASSQETQGRNYGYTF in HCRTNH2 and pHA273-287-tetramers, suggesting molecular mimicry. This public CDR3ß uses TRBV4-2, a segment modulated by T1N-associated SNP rs1008599, suggesting causality. TCR-α/ß CDR3 motifs of HCRT54-66-NH2 and HCRT86-97-NH2 tetramers were extensively shared: notably public CDR3α, TRAV2-CAVETDSWGKLQF-TRAJ24, that uses TRAJ24, a chain modulated by T1N-associated SNPs rs1154155 and rs1483979. TCR-α/ß CDR3 sequences found in pHA273-287, NP17-31, and HCRTNH2 tetramer-positive CD4+ cells were also retrieved in single INF-γ-secreting CD4+ sorted cells stimulated with Pandemrix, independently confirming these results. Our results provide evidence for autoimmunity and molecular mimicry with flu antigens modulated by genetic components in the pathophysiology of T1N.


Assuntos
Narcolepsia/imunologia , Orexinas/imunologia , Orexinas/metabolismo , Adolescente , Adulto , Autoantígenos/metabolismo , Autoimunidade/imunologia , Linfócitos T CD4-Positivos/imunologia , Criança , Epitopos/imunologia , Feminino , Cadeias beta de HLA-DQ , Hemaglutininas , Humanos , Vírus da Influenza A Subtipo H1N1/imunologia , Vírus da Influenza A/imunologia , Vírus da Influenza A/patogenicidade , Influenza Humana/imunologia , Peptídeos e Proteínas de Sinalização Intracelular/genética , Masculino , Pessoa de Meia-Idade , Mimetismo Molecular/imunologia , Orexinas/genética , Peptídeos/genética , Receptores de Antígenos de Linfócitos T/genética , Vacinação
6.
Drug Des Devel Ther ; 12: 2915-2921, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30254421

RESUMO

Background: Lyme disease accounts for >90% of all vector-borne disease cases in the United States and affect ~300,000 persons annually in North America. Though traditional tetracycline antibiotic therapy is generally prescribed for Lyme disease, still 10%-20% of patients treated with current antibiotic therapy still show lingering symptoms. Methods: In order to identify new drugs, we have evaluated four cephalosporins as a therapeutic alternative to commonly used antibiotics for the treatment of Lyme disease by using microdilution techniques like minimum inhibitory concentration (MIC) and the minimum bactericidal concentration (MBC). We have determined the MIC and MBC of four drugs for three Borrelia burgdorferi s.s strains namely CA8, JLB31 and NP40. The binding studies were performed using in silico analysis. Results: The MIC order of the four drugs tested is cefoxitin (1.25 µM/mL) > cefamandole (2.5 µM/mL), > cefuroxime (5 µM/mL) > cefapirin (10 µM/mL). Among the drugs that are tested in this study using in vivo C3H/HeN mouse model, cefoxitin effectively kills B. burgdorferi. The in silico analysis revealed that all four cephalosporins studied binds effectively to B. burgdorferi proteins, SecA subunit penicillin-binding protein (PBP) and Outer surface protein E (OspE). Conclusion: Based on the data obtained, cefoxitin has shown high efficacy killing B. burgdorferi at concentration of 1.25 µM/mL. In addition to it, cefoxitin cleared B. burgdorferi infection in C3H/HeN mice model at 20 mg/kg.

7.
PLoS One ; 12(12): e0187305, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-29220370

RESUMO

BACKGROUND: A recent publication suggested molecular mimicry of a nucleoprotein (NP) sequence from A/Puerto Rico/8/1934 (PR8) strain, the backbone used in the construction of the reassortant strain X-179A that was used in Pandemrix® vaccine, and reported on anti-hypocretin (HCRT) receptor 2 (anti-HCRTR2) autoantibodies in narcolepsy, mostly in post Pandemrix® narcolepsy cases (17 of 20 sera). In this study, we re-examined this hypothesis through mass spectrometry (MS) characterization of Pandemrix®, and two other pandemic H1N1 (pH1N1)-2009 vaccines, Arepanrix® and Focetria®, and analyzed anti-HCRTR2 autoantibodies in narcolepsy patients and controls using three independent strategies. METHODS: MS characterization of Pandemrix® (2 batches), Arepanrix® (4 batches) and Focetria® (1 batch) was conducted with mapping of NP 116I or 116M spectrogram. Two sets of narcolepsy cases and controls were used: 40 post Pandemrix® narcolepsy (PP-N) cases and 18 age-matched post Pandemrix® controls (PP-C), and 48 recent (≤6 months) early onset narcolepsy (EO-N) cases and 70 age-matched other controls (O-C). Anti-HCRTR2 autoantibodies were detected using three strategies: (1) Human embryonic kidney (HEK) 293T cells with transient expression of HCRTR2 were stained with human sera and then analyzed by flow cytometer; (2) In vitro translation of [35S]-radiolabelled HCRTR2 was incubated with human sera and immune complexes of autoantibody and [35S]-radiolabelled HCRTR2 were quantified using a radioligand-binding assay; (3) Optical density (OD) at 450 nm (OD450) of human serum immunoglobulin G (IgG) binding to HCRTR2 stably expressed in Chinese hamster ovary (CHO)-K1 cell line was measured using an in-cell enzyme-linked immunosorbent assay (ELISA). RESULTS: NP 116M mutations were predominantly present in all batches of Pandemrix®, Arepanrix® and Focetria®. The wild-type NP109-123 (ILYDKEEIRRIWRQA), a mimic to HCRTR234-45 (YDDEEFLRYLWR), was not found to bind to DQ0602. Three or four subjects were found positive for anti-HCRTR2 autoantibodies using two strategies or the third one, respectively. None of the post Pandemrix® narcolepsy cases (0 of 40 sera) was found positive with all three strategies. CONCLUSION: Anti-HCRTR2 autoantibody is not a significant biological feature of narcolepsy or of post Pandemrix® autoimmune responses.


Assuntos
Autoanticorpos/sangue , Narcolepsia/sangue , Receptores de Orexina/imunologia , Adolescente , Estudos de Casos e Controles , Criança , Ensaio de Imunoadsorção Enzimática , Feminino , Citometria de Fluxo , Células HEK293 , Humanos , Masculino , Espectrometria de Massas
8.
Curr Opin Pulm Med ; 23(6): 522-529, 2017 11.
Artigo em Inglês | MEDLINE | ID: mdl-28991006

RESUMO

PURPOSE OF REVIEW: Summarize the recent findings in narcolepsy focusing on the environmental and genetic risk factors in disease development. RECENT FINDINGS: Both genetic and epidemiological evidence point towards an autoimmune mechanism in the destruction of orexin/hypocretin neurons. Recent studies suggest both humoral and cellular immune responses in the disease development. SUMMARY: Narcolepsy is a severe sleep disorder, in which neurons producing orexin/hypocretin in the hypothalamus are destroyed. The core symptoms of narcolepsy are debilitating, extreme sleepiness, cataplexy, and abnormalities in the structure of sleep. Both genetic and epidemiological evidence point towards an autoimmune mechanism in the destruction of orexin/hypocretin neurons. Importantly, the highest environmental risk is seen with influenza-A infection and immunization. However, how the cells are destroyed is currently unknown. In this review we summarize the disease symptoms, and focus on the immunological findings in narcolepsy. We also discuss the environmental and genetic risk factors as well as propose a model for disease development.


Assuntos
Autoimunidade , Narcolepsia/imunologia , Predisposição Genética para Doença , Humanos , Imunidade Celular , Imunidade Humoral , Vírus da Influenza A , Vacinas contra Influenza/efeitos adversos , Vacinas contra Influenza/imunologia , Influenza Humana/complicações , Narcolepsia/diagnóstico , Narcolepsia/genética , Fatores de Risco
9.
Viral Immunol ; 30(8): 590-600, 2017 10.
Artigo em Inglês | MEDLINE | ID: mdl-28796576

RESUMO

Increased narcolepsy incidence was observed in Sweden following the 2009 influenza vaccination with Pandemrix®. A substitution of the 2009 nucleoprotein for the 1934 variant has been implicated in narcolepsy development. The aims were to determine (a) antibody levels toward wild-type A/H1N1-2009[A/California/04/2009(H1N1)] (NP-CA2009) and Pandemrix-[A/Puerto Rico/8/1934(H1N1)] (NP-PR1934) nucleoproteins in 43 patients and 64 age-matched controls; (b) antibody affinity in reciprocal competitive assays in 11 childhood narcolepsy patients compared with 21 age-matched controls; and (c) antibody levels toward wild-type A/H1N1-2009[A/California/04/2009(H1N1)] (H1N1 NS1), not a component of the Pandemrix vaccine. In vitro transcribed and translated 35S-methionine-labeled H1N1 influenza A virus proteins were used in radiobinding reciprocal competition assays to estimate antibody levels and affinity (Kd). Childhood patients had higher NP-CA2009 (p = 0.0339) and NP-PR1934 (p = 0.0246) antibody levels compared with age-matched controls. These childhood controls had lower NP-CA2009 (p = 0.0221) and NP-PR1934 (p = 0.00619) antibodies compared with controls 13 years or older. In contrast, in patients 13 years or older, the levels of NP-PR1934 (p = 0.279) and NP-CA2009 (p = 0.0644) antibodies did not differ from the older controls. Childhood antibody affinity (Kd) against NP-CA2009 was comparable between controls (68 ng/mL) and patients (74 ng/mL; p = 0.21) with NP-CA2009 and NP-PR1934 displacement (controls: 165 ng/mL; patients: 199 ng/mL; p = 0.48). In contrast, antibody affinity against NP-PR1934 was higher in controls with either NP-PR1934 (controls: 9 ng/mL; patients: 20 ng/mL; p = 0.0031) or NP-CA2009 (controls: 14 ng/mL; patients: 23 ng/mL; p = 0.0048). A/H1N1-NS1 antibodies were detected in 0/43 of the narcolepsy patients compared with 3/64 (4.7%) controls (p = 0.272). Similarly, none (0/11) of the childhood patients and 1/21 (4.8%) of the childhood controls had A/H1N1-NS1 antibodies. The higher antibody affinities against NP-PR1934 in controls suggest better protection against wild-type virus. In contrast, the reduced NP-PR1934 antibody affinities among childhood narcolepsy patients suggest poor protection from the wild-type A/H1N1 virus and possibly increased risk for viral damage.


Assuntos
Afinidade de Anticorpos/imunologia , Vírus da Influenza A Subtipo H1N1/imunologia , Vacinas contra Influenza/efeitos adversos , Vacinas contra Influenza/imunologia , Influenza Humana/terapia , Narcolepsia/imunologia , Proteínas do Nucleocapsídeo/imunologia , Nucleoproteínas/imunologia , Vacinação/efeitos adversos , Adolescente , Anticorpos Antivirais/sangue , Criança , Feminino , Humanos , Vacinas contra Influenza/uso terapêutico , Masculino , Narcolepsia/epidemiologia , Proteínas do Nucleocapsídeo/genética , Nucleoproteínas/genética , Ensaio Radioligante , Suécia/epidemiologia , Proteínas não Estruturais Virais/genética
10.
Int J Infect Dis ; 59: 29-36, 2017 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-28279736

RESUMO

OBJECTIVES: The 6-exon-spanning 'canonical' Interleukin-7 (IL-7c) is a non-redundant cytokine in human T-cell homeostasis that undergoes extensive alternative pre-mRNA splicing. The IL-7 gene variant lacking, exon 5 (IL-7δ5), exhibits agonistic effects as compared to IL-7c. We studied in this report for the first time the protein expression of IL-7δ5 variant in tissues and its role in monocyte activation. METHODS: We visualized the expression of IL-7δ5 protein by immunohistochemistry in both healthy and malignant (human) tissues and investigated the impact of IL-7δ5 stimulation on CD14+ monocytes using gene expression analysis and flow cytometry. RESULTS: IL-7δ5 is largely expressed by human epithelial cells, yet also by stromal cells in malignant lesions. Gene expression analysis in CD14+ monocytes, induced by the 6-exon spanning IL-7 or IL-7δ5 showed similar changes resulting in a pro-inflammatory phenotype and increased expression of genes involved in lipid metabolism. IL7δ5 was superior in inducing upregulation of the oxidised low density lipoprotein receptor (OLR), measured by flow cytometry, in CD14+ cells. CONCLUSION: IL-7δ5, produced from non-transformed and transformed cells, may contribute to chronic inflammatory responses and development of 'foamy' cells by increased OLR1 expression that mediates increased oxLDL uptake.


Assuntos
Citocinas/análise , Interleucina-7/metabolismo , Receptores Depuradores Classe E/metabolismo , Citocinas/imunologia , Perfilação da Expressão Gênica , Regulação da Expressão Gênica , Humanos , Interleucina-7/imunologia , Receptores de Lipopolissacarídeos/metabolismo , Lipoproteínas LDL/metabolismo , Monócitos/imunologia , Análise de Sequência com Séries de Oligonucleotídeos , Receptores Depuradores Classe E/genética
11.
Oncotarget ; 7(46): 74686-74700, 2016 11 15.
Artigo em Inglês | MEDLINE | ID: mdl-27732960

RESUMO

Selective targeting of the PML/RARα oncoprotein demonstrates a successful molecular targeted therapy in acute promyelocytic leukemia (APL) with a typical t(15:17) chromosomal translocation. The zinc-thiolate coordination is critical for structural stability of zinc finger proteins, including the PML moiety of PML/RARα. Based on the known interaction of redox-active selenium compounds with thiolate ligands of zinc, we herein have investigated the abrogatory effects of selenite alone or in combination with all-trans retinoic acid on PML/RARα and the possible effects on differentiation in these cells. At pharmacological concentrations, selenite inhibited the proliferation and survival of APL originated NB4 cells. In combination with ATRA, it potentiated the differentiation of NB4 cells without any differentiating effects of its own as a single agent. Concordant with our hypothesis, PML/RARα oncoprotein expression was completely abrogated by selenite. Increased expression of RARα, PU.1 and FOXO3A transcription factors in the combined treatment suggested the plausible basis for increased differentiation in these cells. We show that selenite at clinically achievable dose targets PML/RARα oncoprotein for degradation and potentiates differentiation of promyelocytic leukemic cells in combination with ATRA. The present investigation reveals the hitherto unknown potential of selenite in targeted abrogation of PML/RARα in APL cells with prospective therapeutic value.


Assuntos
Antineoplásicos/farmacologia , Diferenciação Celular/efeitos dos fármacos , Leucemia Promielocítica Aguda/patologia , Ácido Selenioso/farmacologia , Tretinoína/farmacologia , Diferenciação Celular/genética , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Sinergismo Farmacológico , Regulação Leucêmica da Expressão Gênica/efeitos dos fármacos , Glutarredoxinas/genética , Glutarredoxinas/metabolismo , Humanos , Leucemia Promielocítica Aguda/genética , Proteínas de Fusão Oncogênica/genética , Proteínas de Fusão Oncogênica/metabolismo , Tiorredoxinas/genética , Tiorredoxinas/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Células Tumorais Cultivadas
12.
J Immunother ; 39(2): 81-9, 2016 Feb-Mar.
Artigo em Inglês | MEDLINE | ID: mdl-26849077

RESUMO

Generation of T lymphocytes with reactivity against cancer is a prerequisite for effective adoptive cellular therapies. We established a protocol for tumor-infiltrating lymphocytes (TILs) from patients with pancreatic ductal adenocarcinoma. Tumor samples from 17 pancreatic cancer specimens were cultured with cytokines (IL-2, IL-15, and IL-21) to expand TILs. After 10 days of culture, TILs were stimulated with an anti-CD3 antibody (OKT3) and irradiated allogeneic peripheral blood mononuclear cells. Reactivity of TILs against tumor-associated antigens (mesothelin, survivin, or NY-ESO-1) was detected by intracellular cytokine production by flow cytometry. Cytotoxicity was measured using a Chromium 51 release assay, and reactivity of TILs against autologous tumor cells was detected by INF-[gamma] production (ELISA). TIL composition was tested by CD45RA, CCR7, 4-1BB, LAG-3, PD-1, TIM3, and CTLA-4 marker analysis. TCR V[beta] was determined by flow cytometry and TCR clonality was gauged measuring the CDR3 region length by PCR analysis and subsequent sequencing. We could reliably obtain TILs from 17/17 patients with a majority of CD8(+) T cells. CD3(+)CD8(+), CD3(+)CD4(+), and CD3(+)CD4(-)CD8(-)[double-negative (DN) T cells] resided predominantly in central (CD45RA(-)CCR7(+)) and effector (CD45RA-CCR7-) memory subsets. CD8(+) TILs tested uniformly positive for LAG-3 (about 100%), whereas CD4(+) TILs showed only up to 12% LAG-3(+) staining and PD-1 showed a broad expression pattern in TILs from different patients. TILs from individual patients recognized strongly (up to 11.9% and 8.2% in CD8(+)) NY-ESO-1, determined by ICS, or mesothelin, determined respectively by TNF-[alpha] and IFN-[gamma] production. Twelve of 17 of CD8(+) TILs showed preferential expansion of certain TCR V[beta] families (eg, 99.2% V[beta]13.2 in CD8(+) TILs, 77% in the V[beta]1, 65.9% in the V[beta]22, and 63.3% in the V[beta]14 family). TCR CDR3 analysis exhibited monoclonal or oligoclonal TCRs, some of them (eg, CD8(+) V[beta]13.2) reacting strongly against autologous tumor defined by INF-[gamma] production or by cytotoxicity. We have optimized methods for generating pancreatic cancer­specific TILs that can be used for adoptive cellular therapy of patients with pancreatic cancer.


Assuntos
Adenocarcinoma/terapia , Linfócitos T CD8-Positivos/imunologia , Imunoterapia Adotiva/métodos , Linfócitos do Interstício Tumoral/imunologia , Neoplasias Pancreáticas/terapia , Receptores de Antígenos de Linfócitos T/metabolismo , Adenocarcinoma/imunologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Antígenos de Neoplasias/imunologia , Proliferação de Células , Células Cultivadas , Citocinas/metabolismo , Citotoxicidade Imunológica , Feminino , Humanos , Memória Imunológica , Ativação Linfocitária , Masculino , Pessoa de Meia-Idade , Neoplasias Pancreáticas/imunologia , Receptores de Antígenos de Linfócitos T/genética
13.
Clin Infect Dis ; 61Suppl 3: S217-24, 2015 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-26409284

RESUMO

The emergence of drug-resistant tuberculosis is challenging tuberculosis control worldwide. In the absence of an effective vaccine to prevent primary infection with Mycobacterium tuberculosis and tuberculosis disease, host-directed therapies may offer therapeutic options, particularly for patients with multidrug-resistant and extensively drug-resistant tuberculosis where prognosis is often limited. CD8(+) and CD4(+) T cells mediate antigen-specific adaptive cellular immune responses. Their use in precision immunotherapy in clinical conditions, especially in treating cancer as well as for prevention of life-threatening viral infections in allogeneic transplant recipients, demonstrated safety and clinical efficacy. We review key achievements in T-cell therapy, including the use of recombinant immune recognition molecules (eg, T-cell receptors and CD19 chimeric antigen receptors), and discuss its potential in the clinical management of patients with drug-resistant and refractory tuberculosis failing conventional therapy.


Assuntos
Transferência Adotiva , Linfócitos T/imunologia , Linfócitos T/transplante , Tuberculose Resistente a Múltiplos Medicamentos/terapia , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Citocinas/uso terapêutico , Tuberculose Extensivamente Resistente a Medicamentos/terapia , Humanos , Imunidade Celular , Mycobacterium tuberculosis/imunologia , Receptores de Antígenos de Linfócitos T/imunologia , Proteínas Recombinantes de Fusão
14.
Immunology ; 145(3): 357-66, 2015 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-25639813

RESUMO

A high content peptide microarray containing the entire influenza A virus [A/California/08/2009(H1N1)] proteome and haemagglutinin proteins from 12 other influenza A subtypes, including the haemagglutinin from the [A/South Carolina/1/1918(H1N1)] strain, was used to gauge serum IgG epitope signatures before and after Pandemrix(®) vaccination or H1N1 infection in a Swedish cohort during the pandemic influenza season 2009. A very narrow pattern of pandemic flu-specific IgG epitope recognition was observed in the serum from individuals who later contracted H1N1 infection. Moreover, the pandemic influenza infection generated IgG reactivity to two adjacent epitopes of the neuraminidase protein. The differential serum IgG recognition was focused on haemagglutinin 1 (H1) and restricted to classical antigenic sites (Cb) in both the vaccinated controls and individuals with flu infections. We further identified a novel epitope VEPGDKITFEATGNL on the Ca antigenic site (251-265) of the pandemic flu haemagglutinin, which was exclusively recognized in serum from individuals with previous vaccinations and never in serum from individuals with H1N1 infection (confirmed by RNA PCR analysis from nasal swabs). This epitope was mapped to the receptor-binding domain of the influenza haemagglutinin and could serve as a correlate of immune protection in the context of pandemic flu. The study shows that unbiased epitope mapping using peptide microarray technology leads to the identification of biologically and clinically relevant target structures. Most significantly an H1N1 infection induced a different footprint of IgG epitope recognition patterns compared with the pandemic H1N1 vaccine.


Assuntos
Vírus da Influenza A Subtipo H1N1/imunologia , Vacinas contra Influenza/imunologia , Influenza Humana/imunologia , Proteoma/imunologia , Sequência de Aminoácidos , Estudos de Coortes , Mapeamento de Epitopos/métodos , Epitopos/imunologia , Hemaglutininas Virais/imunologia , Hemaglutininas Virais/metabolismo , Interações Hospedeiro-Patógeno/imunologia , Humanos , Imunoglobulina G/sangue , Imunoglobulina G/imunologia , Vírus da Influenza A Subtipo H1N1/metabolismo , Vírus da Influenza A Subtipo H1N1/fisiologia , Influenza Humana/diagnóstico , Influenza Humana/virologia , Dados de Sequência Molecular , Neuraminidase/imunologia , Neuraminidase/metabolismo , Prognóstico , Proteoma/metabolismo , Proteômica/métodos , Reprodutibilidade dos Testes , Fatores de Risco , Vacinação/métodos
15.
J Immunother ; 37(8): 416-25, 2014 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-25198529

RESUMO

Human TCRαß(+) CD4(-)CD8(-) double-negative (DN) T cells represent a minor subset in peripheral blood, yet are important in infectious diseases and autoimmune responses. We examined the frequency of DN T cells in 17 patients after allogeneic hematopoietic stem cell transplantation (aHSCT) at 1, 2, 3, 6, and 12 months post-aHSCT and show that these cells increase early after aHSCT and decrease with time after aHSCT. DN T cells reside in the terminally differentiated effector (CD45RA(+)CCR7(-)) T-cell population and are polyclonal, determined by T-cell receptor Vß CDR3 analysis. Gene expression analysis of ex vivo sorted DN T cells showed a distinct set of gene expression, including interleukin-8, as compared with CD4(+) or CD8(+) T cells. DN T cells contributed to MHC class I-restricted EBV-directed immune responses, defined by antigen-specific cytokine production and by detection of HLA-A*02:01-restricted EBV BMLF-1 (GLCTLVAML), LMP-2A (CLGGLLTMV), and HLA-A*24:02-restricted EBV BRLF-1 (DYCNVLNKEF) and EBNA3 (RYSIFFDY)-specific T cells. We created retroviral-transfected Jurkat cell lines with a Melan-A/MART-1-specific TCR(+) and the CD8α chain to study TCR(+) DN T cells in response to their nominal MHC class I/peptide ligand. We show that DN T cells exhibit increased TCRζ chain phosphorylation as compared with the TCR(+)CD8(+) transgenic T-cell line. DN T cells contribute to antigen-specific T-cell responses and represent an effector T-cell population that may be explored in immunotherapeutic approaches against viral infections or transformed cells.


Assuntos
Infecções por Vírus Epstein-Barr/imunologia , Transplante de Células-Tronco Hematopoéticas , Herpesvirus Humano 4/imunologia , Receptores de Antígenos de Linfócitos T alfa-beta/metabolismo , Linfócitos T/imunologia , Adolescente , Adulto , Antígenos Virais/imunologia , Antígenos CD4/metabolismo , Antígenos CD8/metabolismo , Feminino , Seguimentos , Antígeno HLA-A2/metabolismo , Antígeno HLA-A24/metabolismo , Humanos , Interleucina-8/metabolismo , Células Jurkat , Masculino , Pessoa de Meia-Idade , Receptores de Antígenos de Linfócitos T alfa-beta/genética , Transcriptoma , Transplante Homólogo , Adulto Jovem
16.
BMC Infect Dis ; 14: 319, 2014 Jun 11.
Artigo em Inglês | MEDLINE | ID: mdl-24916787

RESUMO

BACKGROUND: Previous exposures to flu and subsequent immune responses may impact on 2009/2010 pandemic flu vaccine responses and clinical symptoms upon infection with the 2009 pandemic H1N1 influenza strain. Qualitative and quantitative differences in humoral and cellular immune responses associated with the flu vaccination in 2009/2010 (pandemic H1N1 vaccine) and natural infection have not yet been described in detail. We designed a longitudinal study to examine influenza- (flu-) specific immune responses and the association between pre-existing flu responses, symptoms of influenza-like illness (ILI), impact of pandemic flu infection, and pandemic flu vaccination in a cohort of 2,040 individuals in Sweden in 2009-2010. METHODS: Cellular flu-specific immune responses were assessed by whole-blood antigen stimulation assay, and humoral responses by a single radial hemolysis test. RESULTS: Previous seasonal flu vaccination was associated with significantly lower flu-specific IFN-γ responses (using a whole-blood assay) at study entry. Pandemic flu vaccination induced long-lived T-cell responses (measured by IFN-γ production) to influenza A strains, influenza B strains, and the matrix (M1) antigen. In contrast, individuals with pandemic flu infection (PCR positive) exhibited increased flu-specific T-cell responses shortly after onset of ILI symptoms but the immune response decreased after the flu season (spring 2010). We identified non-pandemic-flu vaccinated participants without ILI symptoms who showed an IFN-γ production profile similar to pandemic-flu infected participants, suggesting exposure without experiencing clinical symptoms. CONCLUSIONS: Strong and long-lived flu-M1 specific immune responses, defined by IFN-γ production, in individuals after vaccination suggest that M1-responses may contribute to protective cellular immune responses. Silent flu infections appeared to be frequent in 2009/2010. The pandemic flu vaccine induced qualitatively and quantitatively different humoral and cellular immune responses as compared to infection with the 2009 H1N1 pandemic H1N1 influenza strain.


Assuntos
Vacinas contra Influenza/imunologia , Influenza Humana/imunologia , Adolescente , Adulto , Idoso , Anticorpos Antivirais/sangue , Feminino , Humanos , Imunidade Celular/imunologia , Imunidade Humoral/imunologia , Vírus da Influenza A Subtipo H1N1/imunologia , Vírus da Influenza A Subtipo H1N1/patogenicidade , Influenza Humana/epidemiologia , Interferon gama/metabolismo , Estudos Longitudinais , Masculino , Pessoa de Meia-Idade , Estudos Prospectivos , Suécia/epidemiologia , Linfócitos T/imunologia , Vacinação , Adulto Jovem
17.
J Autoimmun ; 50: 99-106, 2014 May.
Artigo em Inglês | MEDLINE | ID: mdl-24485154

RESUMO

Narcolepsy is a lifelong sleep disorder related to hypocretin deficiency resulting from a specific loss of hypocretin-producing neurons in the lateral hypothalamic area. The disease is thought to be autoimmune due to a strong association with HLA-DQB1*06:02. In 2009 the World Health Organization (WHO) declared the H1N1 2009 flu pandemic (A/H1N1PDM09). In response to this, the Swedish vaccination campaign began in October of the same year, using the influenza vaccine Pandemrix(®). A few months later an excess of narcolepsy cases was observed. It is still unclear to what extent the vaccination campaign affected humoral autoimmunity associated with narcolepsy. We studied 47 patients with narcolepsy (6-69 years of age) and 80 healthy controls (3-61 years of age) selected after the Pandemrix vaccination campaign. The first aim was to determine antibodies against A/H1N1 and autoantibodies to Tribbles homolog 2 (TRIB2), a narcolepsy autoantigen candidate as well as to GAD65 and IA-2 as disease specificity controls. The second aim was to test if levels and frequencies of these antibodies and autoantibodies were associated with HLA-DQB1*06:02. In vitro transcribed and translated [(35)S]-methionine and -cysteine-labeled influenza A virus (A/California/04/2009/(H1N1)) segment 4 hemagglutinin was used to detect antibodies in a radiobinding assay. Autoantibodies to TRIB2, GAD65 and IA-2 were similarly detected in standard radiobinding assays. The narcolepsy patients had higher median levels of A/H1N1 antibodies than the controls (p = 0.006). A/H1N1 antibody levels were higher among the <13 years old (n = 12) compared to patients who were older than 30 years (n = 12, p = 0.014). Being HLA-DQB1*06:02 positive was associated with higher A/H1N1 antibody levels in both patients and controls (p = 0.026). Serum autoantibody levels to TRIB2 were low overall and high binders did not differ between patients and controls. We observed an association between levels of A/H1N1 antibodies and TRIB2 autoantibody levels particularly among the youngest narcolepsy patients (r = 0.819, p < 0.001). In conclusion, following the 2009 influenza pandemic vaccination, A/H1N1 antibody levels were associated with young age-at-onset narcolepsy patients positive for HLA-DQB1*06:02. The possibility that TRIB2 is an autoantigen in narcolepsy remains to be clarified. We could verify autoantibody responses against TRIB2 which needs to be determined in larger patient cohorts and control populations.


Assuntos
Anticorpos Antivirais/sangue , Autoanticorpos/sangue , Vírus da Influenza A Subtipo H1N1/imunologia , Vacinas contra Influenza/efeitos adversos , Influenza Humana/prevenção & controle , Peptídeos e Proteínas de Sinalização Intracelular/imunologia , Narcolepsia/induzido quimicamente , Adolescente , Adulto , Idoso , Proteínas Quinases Dependentes de Cálcio-Calmodulina , Criança , Pré-Escolar , Feminino , Expressão Gênica , Glutamato Descarboxilase/antagonistas & inibidores , Glutamato Descarboxilase/genética , Glutamato Descarboxilase/imunologia , Cadeias beta de HLA-DQ/genética , Cadeias beta de HLA-DQ/imunologia , Humanos , Vacinas contra Influenza/imunologia , Influenza Humana/imunologia , Peptídeos e Proteínas de Sinalização Intracelular/antagonistas & inibidores , Peptídeos e Proteínas de Sinalização Intracelular/genética , Masculino , Pessoa de Meia-Idade , Narcolepsia/diagnóstico , Narcolepsia/genética , Narcolepsia/imunologia , Pandemias/prevenção & controle , Suécia , Vacinação/efeitos adversos
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