Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 17.326
Filtrar
1.
BMC Infect Dis ; 20(1): 677, 2020 Sep 17.
Artigo em Inglês | MEDLINE | ID: mdl-32942991

RESUMO

BACKGROUND: Approximately 80% - 90% of individuals infected with latent Mycobacterium tuberculosis (Mtb) remain protected throughout their life-span. The release of unique, latent-phase antigens are known to have a protective role in the immune response against Mtb. Although the BCG vaccine has been administered for nine decades to provide immunity against Mtb, the number of TB cases continues to rise, thereby raising doubts on BCG vaccine efficacy. The shortcomings of BCG have been associated with inadequate processing and presentation of its antigens, an inability to optimally activate T cells against Mtb, and generation of regulatory T cells. Furthermore, BCG vaccination lacks the ability to eliminate latent Mtb infection. With these facts in mind, we selected six immunodominant CD4 and CD8 T cell epitopes of Mtb expressed during latent, acute, and chronic stages of infection and engineered a multi-epitope-based DNA vaccine (C6). RESULT: BALB/c mice vaccinated with the C6 construct along with a BCG vaccine exhibited an expansion of both CD4 and CD8 T cell memory populations and augmented IFN-γ and TNF-α cytokine release. Furthermore, enhancement of dendritic cell and macrophage activation was noted. Consequently, illustrating the elicitation of immunity that helps in the protection against Mtb infection; which was evident by a significant reduction in the Mtb burden in the lungs and spleen of C6 + BCG administered animals. CONCLUSION: Overall, the results suggest that a C6 + BCG vaccination approach may serve as an effective vaccination strategy in future attempts to control TB.


Assuntos
Vacina BCG/imunologia , Epitopos de Linfócito T , Tuberculose/prevenção & controle , Vacinas de DNA/imunologia , Animais , Antígenos de Bactérias/imunologia , Vacina BCG/genética , Vacina BCG/farmacologia , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Epitopos de Linfócito T/genética , Feminino , Memória Imunológica , Interferon gama/metabolismo , Tuberculose Latente/prevenção & controle , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Mycobacterium tuberculosis/imunologia , Tuberculose/imunologia , Fator de Necrose Tumoral alfa/metabolismo , Vacinas de DNA/farmacologia
2.
Nat Commun ; 11(1): 4596, 2020 09 14.
Artigo em Inglês | MEDLINE | ID: mdl-32929083

RESUMO

Earlier studies indicate that either the canonical or non-canonical pathways of inflammasome activation have a limited role on malaria pathogenesis. Here, we report that caspase-8 is a central mediator of systemic inflammation, septic shock in the Plasmodium chabaudi-infected mice and the P. berghei-induced experimental cerebral malaria (ECM). Importantly, our results indicate that the combined deficiencies of caspases-8/1/11 or caspase-8/gasdermin-D (GSDM-D) renders mice impaired to produce both TNFα and IL-1ß and highly resistant to lethality in these models, disclosing a complementary, but independent role of caspase-8 and caspases-1/11/GSDM-D in the pathogenesis of malaria. Further, we find that monocytes from malaria patients express active caspases-1, -4 and -8 suggesting that these inflammatory caspases may also play a role in the pathogenesis of human disease.


Assuntos
Caspase 8/metabolismo , Inflamação/patologia , Malária Cerebral/enzimologia , Animais , Encéfalo/patologia , Caspase 1/metabolismo , Células Dendríticas/metabolismo , Ativação Enzimática , Matriz Extracelular/metabolismo , Regulação da Expressão Gênica , Humanos , Interferon gama/metabolismo , Interleucina-1beta/metabolismo , Lipopolissacarídeos , Malária Cerebral/genética , Camundongos Endogâmicos C57BL , Monócitos/metabolismo , Plasmodium chabaudi/fisiologia , Baço/metabolismo , Receptores Toll-Like/metabolismo
3.
Nat Commun ; 11(1): 4835, 2020 09 24.
Artigo em Inglês | MEDLINE | ID: mdl-32973173

RESUMO

Immune checkpoint blockade therapies have shown clinical promise in a variety of cancers, but how tumor-infiltrating T cells are activated remains unclear. In this study, we explore the functions of PD-L1 on dendritic cells (DCs), which highly express PD-L1. We observe that PD-L1 on DC plays a critical role in limiting T cell responses. Type 1 conventional DCs are essential for PD-L1 blockade and they upregulate PD-L1 upon antigen uptake. Upregulation of PD-L1 on DC is mediated by type II interferon. While DCs are the major antigen presenting cells for cross-presenting tumor antigens to T cells, subsequent PD-L1 upregulation protects them from killing by cytotoxic T lymphocytes, yet dampens the antitumor responses. Blocking PD-L1 in established tumors promotes re-activation of tumor-infiltrating T cells for tumor control. Our study identifies a critical and dynamic role of PD-L1 on DC, which needs to be harnessed for better invigoration of antitumor immune responses.


Assuntos
Antígeno B7-H1/metabolismo , Células Dendríticas/imunologia , Células Dendríticas/metabolismo , Ativação Linfocitária , Animais , Antígenos de Neoplasias/metabolismo , Antígeno B7-H1/genética , Linhagem Celular Tumoral , Modelos Animais de Doenças , Feminino , Células HEK293 , Humanos , Interferon gama/metabolismo , Linfócitos do Interstício Tumoral/imunologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Linfócitos T Citotóxicos/imunologia , Microambiente Tumoral/imunologia , Regulação para Cima
4.
PLoS One ; 15(8): e0237034, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32745117

RESUMO

Production of IFN-γ is a key innate immune mechanism that limits replication of intracellular bacteria such as Francisella tularensis (Ft) until adaptive immune responses develop. Previously, we demonstrated that the host cell types responsible for IFN-γ production in response to murine Francisella infection include not only natural killer (NK) and T cells, but also a variety of myeloid cells. However, production of IFN-γ by mouse dendritic cells (DC) is controversial. Here, we directly demonstrated substantial production of IFN-γ by DC, as well as hybrid NK-DC, from LVS-infected wild type C57BL/6 or Rag1 knockout mice. We demonstrated that the numbers of conventional DC producing IFN-γ increased progressively over the course of 8 days of LVS infection. In contrast, the numbers of conventional NK cells producing IFN-γ, which represented about 40% of non-B/T IFN-γ-producing cells, peaked at day 4 after LVS infection and declined thereafter. This pattern was similar to that of hybrid NK-DC. To further confirm IFN-γ production by infected cells, DC and neutrophils were sorted from naïve and LVS-infected mice and analyzed for gene expression. Quantification of LVS by PCR revealed the presence of Ft DNA not only in macrophages, but also in highly purified, IFN-γ producing DC and neutrophils. Finally, production of IFN-γ by infected DC was confirmed by immunohistochemistry and confocal microscopy. Notably, IFN-γ production patterns similar to those in wild type mice were observed in cells derived from LVS-infected TLR2, TLR4, and TLR2xTLR9 knockout (KO) mice, but not from MyD88 KO mice. Taken together, these studies demonstrate the pivotal roles of DC and MyD88 in IFN-γ production and in initiating innate immune responses to this intracellular bacterium.


Assuntos
Interferon gama/metabolismo , Fator 88 de Diferenciação Mieloide/genética , Receptores Toll-Like/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Animais , Células Dendríticas/imunologia , Células Dendríticas/metabolismo , Feminino , Francisella tularensis/imunologia , Imunidade Inata/imunologia , Células Matadoras Naturais/metabolismo , Macrófagos/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Células Mieloides/metabolismo , Fator 88 de Diferenciação Mieloide/metabolismo , Neutrófilos/metabolismo , Baço/metabolismo , Linfócitos T/imunologia , Receptor 2 Toll-Like/metabolismo , Receptor 4 Toll-Like/metabolismo , Receptor Toll-Like 9/metabolismo , Receptores Toll-Like/imunologia , Tularemia/microbiologia
5.
J Virol ; 94(20)2020 09 29.
Artigo em Inglês | MEDLINE | ID: mdl-32796063

RESUMO

Alpha/beta interferon (IFN-α/ß) signaling through the IFN-α/ß receptor (IFNAR) is essential to limit virus dissemination throughout the central nervous system (CNS) following many neurotropic virus infections. However, the distinct expression patterns of factors associated with the IFN-α/ß pathway in different CNS resident cell populations implicate complex cooperative pathways in IFN-α/ß induction and responsiveness. Here we show that mice devoid of IFNAR1 signaling in calcium/calmodulin-dependent protein kinase II alpha (CaMKIIα) expressing neurons (CaMKIIcre:IFNARfl/fl mice) infected with a mildly pathogenic neurotropic coronavirus (mouse hepatitis virus A59 strain [MHV-A59]) developed severe encephalomyelitis with hind-limb paralysis and succumbed within 7 days. Increased virus spread in CaMKIIcre:IFNARfl/fl mice compared to IFNARfl/fl mice affected neurons not only in the forebrain but also in the mid-hind brain and spinal cords but excluded the cerebellum. Infection was also increased in glia. The lack of viral control in CaMKIIcre:IFNARfl/fl relative to control mice coincided with sustained Cxcl1 and Ccl2 mRNAs but a decrease in mRNA levels of IFNα/ß pathway genes as well as Il6, Tnf, and Il1ß between days 4 and 6 postinfection (p.i.). T cell accumulation and IFN-γ production, an essential component of virus control, were not altered. However, IFN-γ responsiveness was impaired in microglia/macrophages irrespective of similar pSTAT1 nuclear translocation as in infected controls. The results reveal how perturbation of IFN-α/ß signaling in neurons can worsen disease course and disrupt complex interactions between the IFN-α/ß and IFN-γ pathways in achieving optimal antiviral responses.IMPORTANCE IFN-α/ß induction limits CNS viral spread by establishing an antiviral state, but also promotes blood brain barrier integrity, adaptive immunity, and activation of microglia/macrophages. However, the extent to which glial or neuronal signaling contributes to these diverse IFN-α/ß functions is poorly understood. Using a neurotropic mouse hepatitis virus encephalomyelitis model, this study demonstrated an essential role of IFN-α/ß receptor 1 (IFNAR1) specifically in neurons to control virus spread, regulate IFN-γ signaling, and prevent acute mortality. The results support the notion that effective neuronal IFNAR1 signaling compensates for their low basal expression of genes in the IFN-α/ß pathway compared to glia. The data further highlight the importance of tightly regulated communication between the IFN-α/ß and IFN-γ signaling pathways to optimize antiviral IFN-γ activity.


Assuntos
Sistema Nervoso Central/virologia , Interferon Tipo I/metabolismo , Interferon gama/metabolismo , Macrófagos/metabolismo , Microglia/metabolismo , Neurônios/metabolismo , Transdução de Sinais , Animais , Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina/genética , Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina/metabolismo , Sistema Nervoso Central/imunologia , Infecções por Coronavirus/imunologia , Infecções por Coronavirus/virologia , Modelos Animais de Doenças , Encefalomielite/imunologia , Encefalomielite/virologia , Macrófagos/virologia , Camundongos , Camundongos Mutantes , Microglia/virologia , Vírus da Hepatite Murina/fisiologia , Neurônios/virologia , Infiltração de Neutrófilos , Receptor de Interferon alfa e beta/deficiência , Receptor de Interferon alfa e beta/genética , Receptor de Interferon alfa e beta/metabolismo , Replicação Viral
6.
Cells ; 9(9)2020 08 26.
Artigo em Inglês | MEDLINE | ID: mdl-32859121

RESUMO

Natural killer cells are important in the control of viral infections. However, the role of NK cells during severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection has previously not been identified. Peripheral blood NK cells from SARS-CoV and SARS-CoV-2 naïve subjects were evaluated for their activation, degranulation, and interferon-gamma expression in the presence of SARS-CoV and SARS-CoV-2 spike proteins. K562 and lung epithelial cells were transfected with spike proteins and co-cultured with NK cells. The analysis was performed by flow cytometry and immune fluorescence. SARS-CoV and SARS-CoV-2 spike proteins did not alter NK cell activation in a K562 in vitro model. On the contrary, SARS-CoV-2 spike 1 protein (SP1) intracellular expression by lung epithelial cells resulted in NK cell-reduced degranulation. Further experiments revealed a concomitant induction of HLA-E expression on the surface of lung epithelial cells and the recognition of an SP1-derived HLA-E-binding peptide. Simultaneously, there was increased modulation of the inhibitory receptor NKG2A/CD94 on NK cells when SP1 was expressed in lung epithelial cells. We ruled out the GATA3 transcription factor as being responsible for HLA-E increased levels and HLA-E/NKG2A interaction as implicated in NK cell exhaustion. We show for the first time that NK cells are affected by SP1 expression in lung epithelial cells via HLA-E/NKG2A interaction. The resulting NK cells' exhaustion might contribute to immunopathogenesis in SARS-CoV-2 infection.


Assuntos
Betacoronavirus/química , Infecções por Coronavirus/imunologia , Antígenos de Histocompatibilidade Classe I/metabolismo , Células Matadoras Naturais/imunologia , Ativação Linfocitária/genética , Subfamília C de Receptores Semelhantes a Lectina de Células NK/metabolismo , Pneumonia Viral/imunologia , Glicoproteína da Espícula de Coronavírus/metabolismo , Doadores de Sangue , Brônquios/citologia , Degranulação Celular/genética , Técnicas de Cocultura , Infecções por Coronavirus/metabolismo , Infecções por Coronavirus/virologia , Células Epiteliais/metabolismo , Humanos , Interferon gama/metabolismo , Células K562 , Pandemias , Pneumonia Viral/metabolismo , Pneumonia Viral/virologia , RNA Viral/genética , Vírus da SARS/química , Síndrome Respiratória Aguda Grave/imunologia , Síndrome Respiratória Aguda Grave/metabolismo , Síndrome Respiratória Aguda Grave/virologia , Glicoproteína da Espícula de Coronavírus/genética , Transfecção
7.
PLoS Pathog ; 16(8): e1008327, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32853276

RESUMO

Host resistance to Toxoplasma gondii relies on CD8 T cell IFNγ responses, which if modulated by the host or parasite could influence chronic infection and parasite transmission between hosts. Since host-parasite interactions that govern this response are not fully elucidated, we investigated requirements for eliciting naïve CD8 T cell IFNγ responses to a vacuolar resident antigen of T. gondii, TGD057. Naïve TGD057 antigen-specific CD8 T cells (T57) were isolated from transnuclear mice and responded to parasite-infected bone marrow-derived macrophages (BMDMs) in an antigen-dependent manner, first by producing IL-2 and then IFNγ. T57 IFNγ responses to TGD057 were independent of the parasite's protein export machinery ASP5 and MYR1. Instead, host immunity pathways downstream of the regulatory Immunity-Related GTPases (IRG), including partial dependence on Guanylate-Binding Proteins, are required. Multiple T. gondii ROP5 isoforms and allele types, including 'avirulent' ROP5A from clade A and D parasite strains, were able to suppress CD8 T cell IFNγ responses to parasite-infected BMDMs. Phenotypic variance between clades B, C, D, F, and A strains suggest T57 IFNγ differentiation occurs independently of parasite virulence or any known IRG-ROP5 interaction. Consistent with this, removal of ROP5 is not enough to elicit maximal CD8 T cell IFNγ production to parasite-infected cells. Instead, macrophage expression of the pathogen sensors, NLRP3 and to a large extent NLRP1, were absolute requirements. Other members of the conventional inflammasome cascade are only partially required, as revealed by decreased but not abrogated T57 IFNγ responses to parasite-infected ASC, caspase-1/11, and gasdermin D deficient cells. Moreover, IFNγ production was only partially reduced in the absence of IL-12, IL-18 or IL-1R signaling. In summary, T. gondii effectors and host machinery that modulate parasitophorous vacuolar membranes, as well as NLR-dependent but inflammasome-independent pathways, determine the full commitment of CD8 T cells IFNγ responses to a vacuolar antigen.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Inflamassomos/imunologia , Interferon gama/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Proteínas de Protozoários/metabolismo , Transdução de Sinais , Toxoplasma/imunologia , Toxoplasmose Animal/imunologia , Animais , Linfócitos T CD8-Positivos/parasitologia , Feminino , Macrófagos/imunologia , Macrófagos/parasitologia , Camundongos , Proteína 3 que Contém Domínio de Pirina da Família NLR/genética , Proteínas de Protozoários/genética , Toxoplasmose Animal/parasitologia , Vacúolos/imunologia , Vacúolos/metabolismo , Vacúolos/parasitologia , Virulência/imunologia
8.
BMC Infect Dis ; 20(1): 469, 2020 Jul 02.
Artigo em Inglês | MEDLINE | ID: mdl-32615981

RESUMO

BACKGROUND: Interferon-γ release assays (IGRA) with Resuscitation promoting factor (Rpf) proteins enhanced tuberculosis (TB) screening and diagnosis in adults but have not been evaluated in children. Children often develop paucibacillary TB and their immune response differs from that of adults, which together affect TB disease diagnostics and immunodiagnostics. We assessed the ability of Rpf to identify infection among household TB-exposed children in The Gambia and investigated their ability to discriminate Mycobacterium tuberculosis complex (MTBC) infection from active TB disease in children. METHODS: Detailed clinical investigations were done on 93 household TB-exposed Gambian children and a tuberculin skin test (TST) was administered to asymptomatic children. Venous blood was collected for overnight stimulation with ESAT-6/CFP-10-fusion protein (EC), purified protein derivative and RpfA, B, C, D and E. Interferon gamma (IFN-γ) production was measured by ELISA in supernatants and corrected for the background level. Infection status was defined by IGRA with EC and TB disease by mycobacterial confirmation and/or clinical diagnosis. We compared IFN-γ levels between infected and uninfected children and between infected and TB diseased children using a binomial logistic regression model while correcting for age and sex. A Receiver Operating Characteristics analysis was done to find the best cut-off for IFN-γ level and calculate sensitivity and specificity. RESULTS: Interferon gamma production was significantly higher in infected (IGRA+, n = 45) than in uninfected (IGRA-, n = 20) children after stimulation with RpfA, B, C, and D (P = 0.03; 0.007; 0.03 and 0.003, respectively). Using RpfB and D-specific IFN-γ cut-offs (33.9 pg/mL and 67.0 pg/mL), infection was classified with a sensitivity-specificity combination of 73-92% and 77-72% respectively, which was similar to and better than 65-75% for TST. Moreover, IFN-γ production was higher in infected than in TB diseased children (n = 28, 5 bacteriologically confirmed, 23 clinically diagnosed), following RpfB and D stimulation (P = 0.02 and 0.03, respectively). CONCLUSION: RpfB and RpfD show promising results for childhood MTBC infection screening, and both performed similar to and better than the TST in our study population. Additionally, both antigens appear to discriminate between infection and disease in children and thus warrant further investigation as screening and diagnostic antigens for childhood TB.


Assuntos
Antígenos de Bactérias/imunologia , Proteínas de Bactérias/imunologia , Citocinas/imunologia , Testes de Liberação de Interferon-gama/métodos , Tuberculose Latente/diagnóstico , Tuberculose Latente/epidemiologia , Programas de Rastreamento/métodos , Mycobacterium tuberculosis/imunologia , Criança , Pré-Escolar , Ensaio de Imunoadsorção Enzimática , Características da Família , Feminino , Gâmbia/epidemiologia , Humanos , Interferon gama/imunologia , Interferon gama/metabolismo , Tuberculose Latente/microbiologia , Masculino , Sensibilidade e Especificidade , Teste Tuberculínico
9.
JCI Insight ; 5(17)2020 09 03.
Artigo em Inglês | MEDLINE | ID: mdl-32687484

RESUMO

COVID-19-associated morbidity and mortality have been attributed to a pathologic host response. Two divergent hypotheses have been proposed: hyperinflammatory cytokine storm; and failure of host protective immunity that results in unrestrained viral dissemination and organ injury. A key explanation for the inability to address this controversy has been the lack of diagnostic tools to evaluate immune function in COVID-19 infections. ELISpot, a highly sensitive, functional immunoassay, was employed in 27 patients with COVID-19, 51 patients with sepsis, 18 critically ill nonseptic (CINS) patients, and 27 healthy control volunteers to evaluate adaptive and innate immune status by quantitating T cell IFN-É£ and monocyte TFN-α production. Circulating T cell subsets were profoundly reduced in COVID-19 patients. Additionally, stimulated blood mononuclear cells produced less than 40%-50% of the IFN-É£ and TNF-α observed in septic and CINS patients, consistent with markedly impaired immune effector cell function. Approximately 25% of COVID-19 patients had increased IL-6 levels that were not associated with elevations in other canonical proinflammatory cytokines. Collectively, these findings support the hypothesis that COVID-19 suppresses host functional adaptive and innate immunity. Importantly, IL-7 administered ex vivo restored T cell IFN-É£ production in COVID-19 patients. Thus, ELISpot may functionally characterize host immunity in COVID-19 and inform prospective therapies.


Assuntos
Imunidade Adaptativa/imunologia , Infecções por Coronavirus/imunologia , Síndrome da Liberação de Citocina/imunologia , Tolerância Imunológica/imunologia , Imunidade Inata/imunologia , Pneumonia Viral/imunologia , Sepse/imunologia , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Betacoronavirus , Estudos de Casos e Controles , Estado Terminal , ELISPOT , Feminino , Voluntários Saudáveis , Humanos , Interferon gama/imunologia , Interferon gama/metabolismo , Interleucina-6/imunologia , Masculino , Pessoa de Meia-Idade , Monócitos/imunologia , Monócitos/metabolismo , Pandemias , Subpopulações de Linfócitos T/imunologia , Subpopulações de Linfócitos T/metabolismo , Linfócitos T/imunologia , Linfócitos T/metabolismo , Fator de Necrose Tumoral alfa/imunologia , Fator de Necrose Tumoral alfa/metabolismo , Adulto Jovem
10.
Cells ; 9(8)2020 07 22.
Artigo em Inglês | MEDLINE | ID: mdl-32707842

RESUMO

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


Assuntos
5'-Nucleotidase/metabolismo , Apirase/metabolismo , Infecções por Coronavirus/imunologia , Células Matadoras Naturais/imunologia , Células T Matadoras Naturais/imunologia , Pneumonia Viral/imunologia , Linfócitos T Citotóxicos/imunologia , Adenosina/metabolismo , Adulto , Idoso , Betacoronavirus , Infecções por Coronavirus/enzimologia , Feminino , Proteínas Ligadas por GPI/metabolismo , Granzimas/metabolismo , Humanos , Inflamação/enzimologia , Inflamação/imunologia , Interferon gama/metabolismo , Masculino , Pessoa de Meia-Idade , Pandemias , Perforina/metabolismo , Pneumonia Viral/enzimologia , Transdução de Sinais/imunologia , Linfócitos T Citotóxicos/metabolismo , Fator de Necrose Tumoral alfa/metabolismo
11.
Proc Natl Acad Sci U S A ; 117(32): 19388-19398, 2020 08 11.
Artigo em Inglês | MEDLINE | ID: mdl-32727906

RESUMO

CD8+ T cells play pivotal roles in eradicating pathogens and tumor cells. T cell receptor (TCR) signaling is vital for the optimal activation of CD8+ T cells. Upon TCR engagement, the transmembrane adapter protein LAT (linker for activation of T cells) recruits other key signaling molecules and forms the "LAT signalosome" for downstream signal transduction. However, little is known about which functional partners could restrain the formation of the LAT signalosome and inhibit CD8+ cytotoxic T lymphocyte (CTL)-mediated cytotoxicity. Here we have demonstrated that LRCH1 (leucine-rich repeats and calponin homology domain containing 1) directly binds LAT, reduces LAT phosphorylation and interaction with GRB2, and also promotes the endocytosis of LAT. Lrch1 -/- mice display better protection against influenza virus and Listeria infection, with enhanced CD8+ T cell proliferation and cytotoxicity. Adoptive transfer of Lrch1 -/- CD8+ CTLs leads to increased B16-MO5 tumor clearance in vivo. Furthermore, knockout of LRCH1 in human chimeric antigen receptor (CAR) T cells that recognize the liver tumor-associated antigen glypican-3 could improve CAR T cell migration and proliferation in vitro. These findings suggest LRCH1 as a potential translational target to improve T cell immunotherapy against infection and tumors.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Linfócitos T CD8-Positivos/imunologia , Proteínas de Membrana/metabolismo , Proteínas dos Microfilamentos/deficiência , Transdução de Sinais , Animais , Linfócitos T CD8-Positivos/metabolismo , Linfócitos T CD8-Positivos/transplante , Movimento Celular , Células Cultivadas , Citotoxicidade Imunológica , Endocitose , Proteína Adaptadora GRB2/metabolismo , Humanos , Imunoterapia Adotiva , Infecções/imunologia , Infecções/microbiologia , Infecções/virologia , Interferon gama/metabolismo , Neoplasias Pulmonares/terapia , Ativação Linfocitária , Camundongos , Camundongos Knockout , Proteínas dos Microfilamentos/genética , Proteínas dos Microfilamentos/metabolismo , Fosforilação , Ligação Proteica , Receptores de Antígenos de Linfócitos T/metabolismo , Receptores de Antígenos Quiméricos/metabolismo
12.
Cell ; 182(3): 734-743.e5, 2020 08 06.
Artigo em Inglês | MEDLINE | ID: mdl-32643603

RESUMO

COVID-19, caused by SARS-CoV-2, is a virulent pneumonia, with >4,000,000 confirmed cases worldwide and >290,000 deaths as of May 15, 2020. It is critical that vaccines and therapeutics be developed very rapidly. Mice, the ideal animal for assessing such interventions, are resistant to SARS-CoV-2. Here, we overcome this difficulty by exogenous delivery of human ACE2 with a replication-deficient adenovirus (Ad5-hACE2). Ad5-hACE2-sensitized mice developed pneumonia characterized by weight loss, severe pulmonary pathology, and high-titer virus replication in lungs. Type I interferon, T cells, and, most importantly, signal transducer and activator of transcription 1 (STAT1) are critical for virus clearance and disease resolution in these mice. Ad5-hACE2-transduced mice enabled rapid assessments of a vaccine candidate, of human convalescent plasma, and of two antiviral therapies (poly I:C and remdesivir). In summary, we describe a murine model of broad and immediate utility to investigate COVID-19 pathogenesis and to evaluate new therapies and vaccines.


Assuntos
Betacoronavirus/imunologia , Infecções por Coronavirus/patologia , Infecções por Coronavirus/prevenção & controle , Modelos Animais de Doenças , Pandemias/prevenção & controle , Pneumonia Viral/patologia , Pneumonia Viral/prevenção & controle , Vacinação , Animais , Chlorocebus aethiops , Infecções por Coronavirus/virologia , Avaliação Pré-Clínica de Medicamentos/métodos , Feminino , Humanos , Interferon gama/genética , Interferon gama/metabolismo , Pulmão/patologia , Pulmão/virologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Knockout , Peptidil Dipeptidase A/genética , Peptidil Dipeptidase A/metabolismo , Pneumonia Viral/virologia , Receptor de Interferon alfa e beta/genética , Receptor de Interferon alfa e beta/metabolismo , Fator de Transcrição STAT1/genética , Fator de Transcrição STAT1/metabolismo , Organismos Livres de Patógenos Específicos , Transdução Genética , Células Vero , Carga Viral , Replicação Viral
13.
PLoS One ; 15(7): e0235986, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32678856

RESUMO

Health Care Workers (HCW) may have an occupational risk of latent tuberculosis infection (LTBI) and TB disease. The objective of this study was to evaluate the performance of the 2-step strategy: tuberculin skin test (TST) followed by confirmation with Interferon (IFN)-γ- release assays (IGRAs) in HCW. A secondary objective was to determine the factors related to conversions and reversions. HCW at risk of occupational exposure who attended the Occupational Department of the Hospital Germans Trias i Pujol were included during the study period (2013-2016). All professionals testing negative for LTBI were included in a cohort study. These workers were followed up with the administration of a TST and an IGRA quantification at least one year after inclusion in the study. Workers with positive TST, regardless of the results of the IGRA tests, were followed-up with an IGRA. 255 workers were enrolled in the study and 108 workers from the same cohort were followed up. During the follow-up period, seven workers presented TST test conversion. One of these conversions was also confirmed by an IGRA test. There were 2 conversions of cases only testing positive with the IGRA. There have been only 2 reversions of cases testing negative with the IGRA. In this study, not all TST conversions were confirmed when using the IGRA test, which highlights the importance of the 2-step strategy. We have detected a low number of conversions and reversions. Our conclusions should be confirmed in studies with a longer follow-up time.


Assuntos
Pessoal de Saúde/estatística & dados numéricos , Programas de Rastreamento , Tuberculose/diagnóstico , Adulto , Estudos de Coortes , Feminino , Seguimentos , Humanos , Interferon gama/metabolismo , Masculino , Prevalência , Estudos Prospectivos , Teste Tuberculínico , Tuberculose/metabolismo
15.
PLoS One ; 15(7): e0235518, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32614928

RESUMO

Interruption of the programmed death 1 (PD-1) / programmed death ligand 1 (PD-L1) pathway is an established and effective therapeutic strategy in human oncology and holds promise for veterinary oncology. We report the generation and characterization of monoclonal antibodies specific for canine PD-1 and PD-L1. Antibodies were initially assessed for their capacity to block the binding of recombinant canine PD-1 to recombinant canine PD-L1 and then ranked based on efficiency of binding as judged by flow cytometry. Selected antibodies were capable of detecting PD-1 and PD-L1 on canine tissues by flow cytometry and Western blot. Anti-PD-L1 worked for immunocytochemistry and anti-PD-1 worked for immunohistochemistry on formalin-fixed paraffin embedded canine tissues, suggesting the usage of this antibody with archived tissues. Additionally, anti-PD-L1 (JC071) revealed significantly increased PD-L1 expression on canine monocytes after stimulation with peptidoglycan or lipopolysaccharide. Together, these antibodies display specificity for the natural canine ligand using a variety of potential diagnostic applications. Importantly, multiple PD-L1-specific antibodies amplified IFN-γ production in a canine peripheral blood mononuclear cells (PBMC) concanavlin A (Con A) stimulation assay, demonstrating functional activity.


Assuntos
Anticorpos Monoclonais/imunologia , Antígeno B7-H1/imunologia , Receptor de Morte Celular Programada 1/imunologia , Linfócitos T/imunologia , Animais , Antígeno B7-H1/genética , Antígeno B7-H1/metabolismo , Cães , Interferon gama/metabolismo , Leucócitos Mononucleares/citologia , Leucócitos Mononucleares/metabolismo , Lipopolissacarídeos/farmacologia , Camundongos , Camundongos Endogâmicos BALB C , Monócitos/efeitos dos fármacos , Monócitos/imunologia , Monócitos/metabolismo , Peptidoglicano/farmacologia , Receptor de Morte Celular Programada 1/genética , Receptor de Morte Celular Programada 1/metabolismo , Linfócitos T/metabolismo
16.
PLoS One ; 15(7): e0235706, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32639988

RESUMO

During type 1 immune responses, CD4 T helper 1 (Th1) cells and CD8 T cells are activated via IL-12 and contribute to the elimination of intracellular pathogens through interferon gamma (IFNγ) production. In this study, we identified Placenta-specific 8 (Plac8) as a gene that is uniquely expressed in Th1 CD4 T cells relative to other CD4 T cell subsets and hypothesized that Plac8 may represent a novel therapeutic target in Th1 CD4 T cells. First, we determined that Plac8 mRNA in CD4 T cells was induced following IL-12 stimulation via an indirect route that required new protein synthesis. Upon evaluating the functional relevance of Plac8 expression in Th1 CD4 T cells, we discovered that Plac8 was important for suppressing IFNγ mRNA and protein production by CD4 T cells 24 hours after IL-12 stimulation, however Plac8 did not contribute to pathogenic CD4 T cell function during two models of intestinal inflammation. We also noted relatively high basal expression of Plac8 in CD8 T cells which could be further induced following IL-12 stimulation in CD8 T cells. Furthermore, Plac8 expression was important for establishing an optimal CD8 T cell response against influenza A virus via a T cell-intrinsic manner. Altogether, these results implicate Plac8 as a potential regulator of Th1 CD4 and CD8 T cell responses during Th1 T cell-driven inflammation.


Assuntos
Linfócitos T CD4-Positivos/metabolismo , Linfócitos T CD8-Positivos/metabolismo , Interferon gama/metabolismo , Proteínas/metabolismo , Animais , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Colite/imunologia , Colite/patologia , Modelos Animais de Doenças , Interferon gama/genética , Interleucina-12/farmacologia , Antígenos Comuns de Leucócito/genética , Antígenos Comuns de Leucócito/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Infecções por Orthomyxoviridae/imunologia , Infecções por Orthomyxoviridae/patologia , Proteínas/genética , RNA Mensageiro/metabolismo , Regulação para Cima/efeitos dos fármacos
17.
Zhejiang Da Xue Xue Bao Yi Xue Ban ; 40(7): 930-935, 2020 May 25.
Artigo em Chinês | MEDLINE | ID: mdl-32701231

RESUMO

OBJECTIVE: To assess the effect of neutralizing CD96 on natural killer (NK) cell functions in mice with pulmonary Chlamydia muridarum infection and explore the possible mechanism. METHODS: Male BALB/c mice were randomly divided into infection group (Cm group), anti-CD96 treatment group (anti-CD96 group) and control group (n=5). In the former two groups, C. muridarum was inoculated via intranasal administration to establish mouse models of pulmonary C. muridarum infection, and the mice in the control group received intranasal administration of the inhalation buffer. In anti-CD96 group, the mice were injected with anti-CD96 antibody intraperitoneally at the dose of 250 µg every 3 days after the infection; the mice in Cm group received intraperitoneal injections of saline. The body weight of the mice was recorded daily. The mice were sacrificed 5 days after C. muridarum infection, and CD96 expression was detected by quantitative real-time PCR and Western blotting. HE staining and pathological scores were used to evaluate pneumonia of the mice. The inclusion body forming units (IFUs) were detected in the lung tissue homogenates to assess lung tissue chlamydia load. Flow cytometry and ELISA were used to assess the capacity of the lung NK cells to produce interferon-γ (IFN-γ) and regulate macrophages and Th1 cells. RESULTS: C. muridarum infection inhibited CD96 expression in NK cells of the mice. Compared with those in Cm group, the mice in antiCD96 mice showed significantly milder lung inflammation (P < 0.05) and reduced chlamydia load in the lung tissue (P < 0.05). Neutralizing CD96 with anti-CD96 significantly enhanced IFN-γ secretion by the NK cells (P < 0.05) and augmented the immunoregulatory effect of the NK cells shown by enhanced responses of the lung macrophages (P < 0.05) and Th1 cells (P < 0.05). CONCLUSIONS: Inhibition of CD96 alleviates pneumonia in C. muridarum-infected mice possibly by enhancing IFN-γ secretion by NK cells and augmenting the immunoregulatory effect of the NK cells on innate and adaptive immunity.


Assuntos
Infecções por Chlamydia , Chlamydia muridarum , Interferon gama , Lesão Pulmonar , Animais , Antígenos CD/metabolismo , Infecções por Chlamydia/complicações , Infecções por Chlamydia/imunologia , Infecções por Chlamydia/fisiopatologia , Interferon gama/genética , Interferon gama/metabolismo , Células Matadoras Naturais/metabolismo , Lesão Pulmonar/etiologia , Lesão Pulmonar/genética , Lesão Pulmonar/prevenção & controle , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL
18.
Mol Cell ; 78(6): 1207-1223.e8, 2020 06 18.
Artigo em Inglês | MEDLINE | ID: mdl-32504554

RESUMO

Tumor interferon (IFN) signaling promotes PD-L1 expression to suppress T cell-mediated immunosurveillance. We identify the IFN-stimulated non-coding RNA 1 (INCR1) as a long noncoding RNA (lncRNA) transcribed from the PD-L1 locus and show that INCR1 controls IFNγ signaling in multiple tumor types. Silencing INCR1 decreases the expression of PD-L1, JAK2, and several other IFNγ-stimulated genes. INCR1 knockdown sensitizes tumor cells to cytotoxic T cell-mediated killing, improving CAR T cell therapy. We discover that PD-L1 and JAK2 transcripts are negatively regulated by binding to HNRNPH1, a nuclear ribonucleoprotein. The primary transcript of INCR1 binds HNRNPH1 to block its inhibitory effects on the neighboring genes PD-L1 and JAK2, enabling their expression. These findings introduce a mechanism of tumor IFNγ signaling regulation mediated by the lncRNA INCR1 and suggest a therapeutic target for cancer immunotherapy.


Assuntos
Antígeno B7-H1/genética , Interferon gama/metabolismo , RNA Longo não Codificante/genética , Idoso , Animais , Linhagem Celular Tumoral , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Imunoterapia , Imunoterapia Adotiva/métodos , Interferon gama/genética , Interferons/genética , Interferons/metabolismo , Janus Quinase 2/genética , Janus Quinase 2/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos NOD , Pessoa de Meia-Idade , Proteína 2 Ligante de Morte Celular Programada 1/genética , Fator de Transcrição STAT1/metabolismo , Transdução de Sinais/efeitos dos fármacos , Linfócitos T Citotóxicos
20.
Proc Natl Acad Sci U S A ; 117(27): 15935-15946, 2020 07 07.
Artigo em Inglês | MEDLINE | ID: mdl-32571912

RESUMO

Excessive tumor necrosis factor (TNF) is known to cause significant pathology. Paradoxically, deficiency in TNF (TNF-/-) also caused substantial pathology during respiratory ectromelia virus (ECTV) infection, a surrogate model for smallpox. TNF-/- mice succumbed to fulminant disease whereas wild-type mice, and those engineered to express only transmembrane TNF (mTNF), fully recovered. TNF deficiency did not affect viral load or leukocyte recruitment but caused severe lung pathology and excessive production of the cytokines interleukin (IL)-6, IL-10, transforming growth factor beta (TGF-ß), and interferon gamma (IFN-γ). Short-term blockade of these cytokines significantly reduced lung pathology in TNF-/- mice concomitant with induction of protein inhibitor of activated STAT3 (PIAS3) and/or suppressor of cytokine signaling 3 (SOCS3), factors that inhibit STAT3 activation. Consequently, inhibition of STAT3 activation with an inhibitor reduced lung pathology. Long-term neutralization of IL-6 or TGF-ß protected TNF-/- mice from an otherwise lethal infection. Thus, mTNF alone is necessary and sufficient to regulate lung inflammation but it has no direct antiviral activity against ECTV. The data indicate that targeting specific cytokines or cytokine-signaling pathways to reduce or ameliorate lung inflammation during respiratory viral infections is possible but that the timing and duration of the interventive measure are critical.


Assuntos
Citocinas/metabolismo , Infecções por Poxviridae/virologia , Poxviridae/metabolismo , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/metabolismo , Animais , Linhagem Celular Tumoral , Feminino , Interferon gama/metabolismo , Interleucina-10/metabolismo , Interleucina-6/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Poxviridae/imunologia , Infecções por Poxviridae/imunologia , Infecções por Poxviridae/patologia , Infecções Respiratórias/imunologia , Infecções Respiratórias/virologia , Fator de Transcrição STAT3/genética , Fator de Transcrição STAT3/metabolismo , Proteína 3 Supressora da Sinalização de Citocinas/metabolismo , Fator de Crescimento Transformador beta/metabolismo
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA