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1.
PLoS Pathog ; 17(9): e1009804, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-34529726

RESUMO

Prior studies have demonstrated that immunologic dysfunction underpins severe illness in COVID-19 patients, but have lacked an in-depth analysis of the immunologic drivers of death in the most critically ill patients. We performed immunophenotyping of viral antigen-specific and unconventional T cell responses, neutralizing antibodies, and serum proteins in critically ill patients with SARS-CoV-2 infection, using influenza infection, SARS-CoV-2-convalescent health care workers, and healthy adults as controls. We identify mucosal-associated invariant T (MAIT) cell activation as an independent and significant predictor of death in COVID-19 (HR = 5.92, 95% CI = 2.49-14.1). MAIT cell activation correlates with several other mortality-associated immunologic measures including broad activation of CD8+ T cells and non-Vδ2 γδT cells, and elevated levels of cytokines and chemokines, including GM-CSF, CXCL10, CCL2, and IL-6. MAIT cell activation is also a predictor of disease severity in influenza (ECMO/death HR = 4.43, 95% CI = 1.08-18.2). Single-cell RNA-sequencing reveals a shift from focused IFNα-driven signals in COVID-19 ICU patients who survive to broad pro-inflammatory responses in fatal COVID-19 -a feature not observed in severe influenza. We conclude that fatal COVID-19 infection is driven by uncoordinated inflammatory responses that drive a hierarchy of T cell activation, elements of which can serve as prognostic indicators and potential targets for immune intervention.


Assuntos
COVID-19/imunologia , COVID-19/mortalidade , Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/imunologia , Antígenos CD/imunologia , Antígenos de Diferenciação de Linfócitos T/imunologia , Linfócitos B/imunologia , Biomarcadores/sangue , Proteínas Sanguíneas/metabolismo , Estudos de Coortes , Estado Terminal/mortalidade , Feminino , Humanos , Imunofenotipagem , Influenza Humana/imunologia , Lectinas Tipo C/imunologia , Ativação Linfocitária , Masculino , Pessoa de Meia-Idade , Células T Invariantes Associadas à Mucosa/imunologia , Gravidade do Paciente
2.
Front Immunol ; 12: 650808, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34234773

RESUMO

The myeloid inhibitory C-type lectin receptor CLEC12A limits neutrophil activation, pro-inflammatory pathways and disease in mouse models of inflammatory arthritis by a molecular mechanism that remains poorly understood. We addressed how CLEC12A-mediated inhibitory signaling counteracts activating signaling by cross-linking CLEC12A in human neutrophils. CLEC12A cross-linking induced its translocation to flotillin-rich membrane domains where its ITIM was phosphorylated in a Src-dependent manner. Phosphoproteomic analysis identified candidate signaling molecules regulated by CLEC12A that include MAPKs, phosphoinositol kinases and members of the JAK-STAT pathway. Stimulating neutrophils with uric acid crystals, the etiological agent of gout, drove the hyperphosphorylation of p38 and Akt. Ultimately, one of the pathways through which CLEC12A regulates uric acid crystal-stimulated release of IL-8 by neutrophils is through a p38/PI3K-Akt signaling pathway. In summary this work defines early molecular events that underpin CLEC12A signaling in human neutrophils to modulate cytokine synthesis. Targeting this pathway could be useful therapeutically to dampen inflammation.


Assuntos
Lectinas Tipo C/imunologia , Ativação de Neutrófilo/imunologia , Neutrófilos/imunologia , Fosfatidilinositol 3-Quinases/imunologia , Proteínas Proto-Oncogênicas c-akt/imunologia , Receptores Mitogênicos/imunologia , Transdução de Sinais/imunologia , Adulto , Células Cultivadas , Citocinas/imunologia , Citocinas/metabolismo , Células HEK293 , Células HeLa , Humanos , Lectinas Tipo C/genética , Lectinas Tipo C/metabolismo , Microscopia Confocal , Neutrófilos/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Fosforilação , Proteínas Proto-Oncogênicas c-akt/metabolismo , Receptores Mitogênicos/genética , Receptores Mitogênicos/metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno/imunologia , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo
3.
Fish Shellfish Immunol ; 115: 104-111, 2021 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-34062237

RESUMO

C-type lectins (CTLs) are important pathogen pattern recognition receptors that recognize carbohydrate structures. In present study, a C-type lectin domain family 4 member E-like gene from turbot, which tentatively named SmCLEC4E-like (SmCLEC4EL), was identified, and the expressional and functional analyses were performed. In our results, SmCLEC4EL showed conserved synteny with CLEC4E-like genes from several fish species in genome, and possessed a typical type II transmembrane CTL architecture: an N-terminal intracellular region, a transmembrane domain and a C-terminal extracellular region which contained a predicted carbohydrate recognition domain (CRD). In addition, SmCLEC4EL exhibited the highest expression level in spleen in healthy fish, and showed significantly induced expression in mucosal tissues, intestine and skin, under bacteria challenge. Finally, the recombinant SmCLEC4EL protein combined with LPS, PGN, LTA and five different kinds of bacteria in a dose-dependent manner, and agglutinated these bacteria strains in the presence of calcium. These findings collectively demonstrated that SmCLEC4EL, a calcium-dependent CTL, could function as a pattern recognition receptor in pathogen recognition and participate in host anti-bacteria immunity.


Assuntos
Doenças dos Peixes/imunologia , Linguados/genética , Linguados/imunologia , Regulação da Expressão Gênica/imunologia , Imunidade Inata/genética , Lectinas Tipo C/genética , Lectinas Tipo C/imunologia , Sequência de Aminoácidos , Animais , Infecções Bacterianas/imunologia , Infecções Bacterianas/veterinária , Proteínas de Peixes/química , Proteínas de Peixes/genética , Proteínas de Peixes/imunologia , Perfilação da Expressão Gênica/veterinária , Lectinas Tipo C/química , Lipopolissacarídeos/farmacologia , Peptidoglicano/farmacologia , Filogenia , Alinhamento de Sequência/veterinária , Ácidos Teicoicos/farmacologia
4.
Fish Shellfish Immunol ; 115: 160-170, 2021 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-34147614

RESUMO

C-type lectins (CTL) are a large group of pattern-recognition proteins and to play important roles in glycoprotein metabolism, multicellular integration, and immunity. Based on their overall domain structure, they can be classified as different groups that possess different physiological functions. A typical C-type lectin (named as OmLec1) was identified from the fish, Onychostoma macrolepis, an important cultured fish in China. Open reading frame of OmLec1 contains a 570 bp, encoding a protein of 189 amino acids that includes a signal peptide and a single carbohydrate-recognition domain. The phylogenetic analysis showed that OmLec1 could be grouped with C-type lectin from other fish. OmLec1 was expressed in all the tissues in our study, and the expression level was highest in liver. And its relative expression levels were significantly upregulated following infection with Aeromonas hydrophila. The recombinant OmLec1 protein (rOmLec1) could agglutinate some Gram-negative bacteria and Gram-positive bacteria in vitro in the presence of Ca2+, showing a typical Ca2+-dependent carbohydrate-binding protein. Furthermore, rOmLec1 purified from E. coli BL21 (DE3), strongly bound to LPS and PGN, as well as all tested bacteria in a Ca2+-dependent manner. These results indicate that OmLec1 plays a central role in the innate immune response and as a pattern recognition receptor that recognizes diverse pathogens among O. macrolepis.


Assuntos
Cyprinidae/genética , Cyprinidae/imunologia , Doenças dos Peixes/imunologia , Regulação da Expressão Gênica/imunologia , Imunidade Inata/genética , Lectinas Tipo C/genética , Lectinas Tipo C/imunologia , Aeromonas hydrophila/fisiologia , Sequência de Aminoácidos , Animais , Sequência de Bases , Proteínas de Peixes/química , Proteínas de Peixes/genética , Proteínas de Peixes/imunologia , Perfilação da Expressão Gênica/veterinária , Infecções por Bactérias Gram-Negativas/imunologia , Infecções por Bactérias Gram-Negativas/veterinária , Lectinas Tipo C/química , Filogenia , Alinhamento de Sequência/veterinária
5.
Theranostics ; 11(15): 7308-7321, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34158852

RESUMO

Dendritic cells (DCs) can process the antigens of cancer vaccine and thus stimulate the CD8+ T cells to recognize and kill the tumor cells that express these antigens. However, lack of promising carriers for presenting the antigens to DCs is one of the main barriers to the development of clinically effective cancer vaccines. Another limitation is the risk of inflammatory side effects induced by the adjuvants. It is still unclear how we can develop ideal adjuvant-free DC vaccine carriers without adjuvants. Methods: A 12-mer peptide carrier (CBP-12) with high affinity for Clec9a expressed on DCs was developed using an in silico rational optimization method. The therapeutic effects of the adjuvant-free vaccine comprising CBP-12 and exogenous or endogenous antigenic peptides were investigated in terms of antigen cross-presentation efficacy, specific cytotoxic T lymphocyte response, and antitumor activity. We also explored the mechanism involved in the antitumor effects of the adjuvant-free CBP-12 vaccine. Finally, we assessed the effects of the CBP-12 conjugated peptide vaccine combined with radiotherapy. Results: Here, we developed CBP-12 as a vaccine carrier that enhanced the uptake and cross-presentation of the antigens, thus inducing strong CD8+ T cell responses and antitumor effects in both anti-PD-1-responsive (B16-OVA) and -resistant (B16) models, even in adjuvant-free conditions. CBP-12 bound to and activated Clec9a, thereby stimulating Clec9a+ DC to product IL-21, but not IL-12 by activating of Syk. The antitumor effects of the CBP-12 conjugated peptide vaccines could be blocked by an IL-21 neutralizing antibody. We also observed the synergistic antitumor effects of the CBP-12 conjugated peptide vaccine combined with radiotherapy. Conclusions: CBP-12 could serve as an adjuvant-free peptide vaccine carrier for cancer immunotherapy.


Assuntos
Vacinas Anticâncer , Células Dendríticas/imunologia , Sistemas de Liberação de Medicamentos , Interleucinas/imunologia , Lectinas Tipo C/imunologia , Melanoma Experimental/imunologia , Peptídeos , Receptores Imunológicos/imunologia , Transdução de Sinais/efeitos dos fármacos , Quinase Syk/imunologia , Animais , Vacinas Anticâncer/imunologia , Vacinas Anticâncer/farmacologia , Feminino , Interleucinas/genética , Lectinas Tipo C/genética , Melanoma Experimental/genética , Melanoma Experimental/terapia , Camundongos , Camundongos Knockout , Peptídeos/imunologia , Peptídeos/farmacologia , Receptores Imunológicos/genética , Transdução de Sinais/genética , Transdução de Sinais/imunologia , Quinase Syk/genética , Vacinas de Subunidades/imunologia , Vacinas de Subunidades/farmacologia
6.
J Biomed Sci ; 28(1): 46, 2021 Jun 11.
Artigo em Inglês | MEDLINE | ID: mdl-34116654

RESUMO

Dysregulated formation of neutrophil extracellular traps (NETs) is observed in acute viral infections. Moreover, NETs contribute to the pathogenesis of acute viral infections, including those caused by the dengue virus (DV) and severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2). Furthermore, excessive NET formation (NETosis) is associated with disease severity in patients suffering from SARS-CoV-2-induced multiple organ injuries. Dendritic cell-specific intercellular adhesion molecule-3-grabbing non-integrin (DC-SIGN) and other members of C-type lectin family (L-SIGN, LSECtin, CLEC10A) have been reported to interact with viral glycans to facilitate virus spreading and exacerbates inflammatory reactions. Moreover, spleen tyrosine kinase (Syk)-coupled C-type lectin member 5A (CLEC5A) has been shown as the pattern recognition receptor for members of flaviviruses, and is responsible for DV-induced cytokine storm and Japanese encephalomyelitis virus (JEV)-induced neuronal inflammation. Moreover, DV activates platelets via CLEC2 to release extracellular vesicles (EVs), including microvesicles (MVs) and exosomes (EXOs). The DV-activated EXOs (DV-EXOs) and MVs (DV-MVs) stimulate CLEC5A and Toll-like receptor 2 (TLR2), respectively, to enhance NET formation and inflammatory reactions. Thus, EVs from virus-activated platelets (PLT-EVs) are potent endogenous danger signals, and blockade of C-type lectins is a promising strategy to attenuate virus-induced NETosis and intravascular coagulopathy.


Assuntos
COVID-19/imunologia , Vírus da Encefalite Japonesa (Espécie)/imunologia , Encefalite Japonesa/imunologia , Armadilhas Extracelulares/imunologia , Lectinas Tipo C/imunologia , SARS-CoV-2/imunologia , Plaquetas/imunologia , Plaquetas/patologia , COVID-19/patologia , Síndrome da Liberação de Citocina/imunologia , Síndrome da Liberação de Citocina/patologia , Encefalite Japonesa/patologia , Humanos , Ativação Plaquetária/imunologia , Transdução de Sinais/imunologia
7.
Biochem Biophys Res Commun ; 561: 101-105, 2021 07 05.
Artigo em Inglês | MEDLINE | ID: mdl-34020140

RESUMO

Mast cells (MCs) are present in various organs including the skin, peritoneal cavity, lung, and intestine and involved in the development of allergic diseases and host defense against infection. However, the regulatory mechanism of mast cell activation remains incompletely understood. We found in a database that Clec12b encoding a C-type lectin receptor Clec12b is preferentially expressed in skin MCs in mice. However, neither MCs in other tissues such as trachea, tongue, esophagus, or peritoneal cavity nor most lymphocytes and myeloid cells express Clec12b. To analyze the protein expression of Clec12b, we newly generated a monoclonal antibody (named TX109), which recognizes both mouse and human Clec12b. Consistent with the gene expression profile, flow cytometry analysis demonstrated that Clec12b is expressed only on MCs in the skin, but not on any other immune cell types in various tissues, in mice. Similarly, Clec12b is also expressed on skin MCs, but not on circulating lymphocytes and myeloid cells, in humans. Our results suggest that Clec12b plays an important role in the regulation of MCs activation in the skin.


Assuntos
Anticorpos Monoclonais/imunologia , Lectinas Tipo C/metabolismo , Mastócitos/metabolismo , Receptores Mitogênicos/metabolismo , Pele/metabolismo , Animais , Diferenciação Celular , Células Cultivadas , Citometria de Fluxo/métodos , Humanos , Lectinas Tipo C/imunologia , Mastócitos/citologia , Mastócitos/imunologia , Camundongos , Receptores Mitogênicos/imunologia , Pele/citologia , Pele/imunologia
8.
Front Immunol ; 12: 635411, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33995354

RESUMO

More than fifty c-type lectin receptors (CLR) are known and have been identified so far. Moreover, we know the group of galectins and sialic acid-binding immunoglobulin-type lectins that also belong to the carbohydrate-binding receptors of the immune system. Thus, the lectin receptors form the largest receptor family among the pathogen recognition receptors. Similar to the toll-like receptors (TLRs), the CLR do not only recognize foreign but also endogenous molecules. In contrast to TLRs, which have a predominantly activating effect on the immune system, lectin receptors also mediate inhibitory signals. They play an important role in innate and adaptive immunity for the induction, regulation and shaping of the immune response. The hygiene hypothesis links enhanced infection to protection from allergic disease. Yet, the microbial substances that are responsible for mediating this allergy-protective activity still have to be identified. Microbes contain both ligands binding to TLRs and carbohydrates that are recognized by CLR and other lectin receptors. In the current literature, the CLR are often recognized as the 'bad guys' in allergic inflammation, because some glycoepitopes of allergens have been shown to bind to CLR, facilitating their uptake and presentation. On the other hand, there are many reports revealing that sugar moieties are involved in immune regulation. In this review, we will summarize what is known about the role of carbohydrate interaction with c-type lectins and other sugar-recognizing receptors in anti-inflammation, with a special focus on the regulation of the allergic immune response.


Assuntos
Hipersensibilidade/metabolismo , Inflamação/metabolismo , Lectinas Tipo C/metabolismo , Animais , Humanos , Hipersensibilidade/imunologia , Inflamação/imunologia , Lectinas Tipo C/imunologia , Ligantes , Transdução de Sinais
9.
J Exp Med ; 218(7)2021 07 05.
Artigo em Inglês | MEDLINE | ID: mdl-33944900

RESUMO

Food allergies are a leading cause of anaphylaxis, and cellular mechanisms involving antigen presentation likely play key roles in their pathogenesis. However, little is known about the response of specific antigen-presenting cell (APC) subsets to food allergens in the setting of food allergies. Here, we show that in peanut-allergic humans, peanut allergen drives the differentiation of CD209+ monocyte-derived dendritic cells (DCs) and CD23+ (FcєRII) myeloid dendritic cells through the action of allergen-specific CD4+ T cells. CD209+ DCs act reciprocally on the same peanut-specific CD4+ T cell population to reinforce Th2 cytokine expression in a positive feedback loop, which may explain the persistence of established food allergy. In support of this novel model, we show clinically that the initiation of oral immunotherapy (OIT) in peanut-allergic patients is associated with a decrease in CD209+ DCs, suggesting that breaking the cycle of positive feedback is associated with therapeutic effect.


Assuntos
Alérgenos/imunologia , Arachis/imunologia , Imunidade/imunologia , Hipersensibilidade a Amendoim/imunologia , Animais , Células Apresentadoras de Antígenos/imunologia , Moléculas de Adesão Celular/imunologia , Citocinas/imunologia , Células Dendríticas/imunologia , Retroalimentação , Humanos , Imunoterapia/métodos , Lectinas Tipo C/imunologia , Camundongos , Monócitos/imunologia , Receptores de Superfície Celular/imunologia , Receptores de IgE/imunologia , Linfócitos T Reguladores/imunologia , Células Th2/imunologia
10.
Int J Mol Sci ; 22(9)2021 Apr 30.
Artigo em Inglês | MEDLINE | ID: mdl-33946381

RESUMO

Understanding the host anti-fungal immunity induced by beta-glucan has been one of the most challenging conundrums in the field of biomedical research. During the last couple of decades, insights on the role of beta-glucan in fungal disease progression, susceptibility, and resistance have been greatly augmented through the utility of various beta-glucan cognate receptor-deficient mouse models. Analysis of dectin-1 knockout mice has clarified the downstream signaling pathways and adaptive effector responses triggered by beta-glucan in anti-fungal immunity. On the other hand, assessment of CR3-deficient mice has elucidated the compelling action of beta-glucans in neutrophil-mediated fungal clearance, and the investigation of EphA2-deficient mice has highlighted its novel involvement in host sensing and defense to oral mucosal fungal infection. Based on these accounts, this review focuses on the recent discoveries made by these gene-targeted mice in beta-glucan research with particular emphasis on the multifaceted aspects of fungal immunity.


Assuntos
Fungos/imunologia , Micoses/imunologia , beta-Glucanas/imunologia , Imunidade Adaptativa , Animais , Modelos Animais de Doenças , Deleção de Genes , Humanos , Imunidade , Lectinas Tipo C/genética , Lectinas Tipo C/imunologia , Antígeno de Macrófago 1/genética , Antígeno de Macrófago 1/imunologia , Camundongos , Camundongos Knockout , Micoses/genética , Micoses/microbiologia , Receptor EphA2/genética , Receptor EphA2/imunologia
11.
Nat Commun ; 12(1): 2715, 2021 05 11.
Artigo em Inglês | MEDLINE | ID: mdl-33976157

RESUMO

Efficient immune responses rely on heterogeneity, which in CD8+ T cells, amongst other mechanisms, is achieved by asymmetric cell division (ACD). Here we find that ageing, known to negatively impact immune responses, impairs ACD in murine CD8+ T cells, and that this phenotype can be rescued by transient mTOR inhibition. Increased ACD rates in mitotic cells from aged mice restore the expansion and memory potential of their cellular progenies. Further characterization of the composition of CD8+ T cells reveals that virtual memory cells (TVM cells), which accumulate during ageing, have a unique proliferation and metabolic profile, and retain their ability to divide asymmetrically, which correlates with increased memory potential. The opposite is observed for naive CD8+ T cells from aged mice. Our data provide evidence on how ACD modulation contributes to long-term survival and function of T cells during ageing, offering new insights into how the immune system adapts to ageing.


Assuntos
Envelhecimento/genética , Divisão Celular Assimétrica/genética , Linfócitos T CD8-Positivos/imunologia , Memória Imunológica/genética , Serina-Treonina Quinases TOR/genética , Envelhecimento/imunologia , Animais , Divisão Celular Assimétrica/imunologia , Linfócitos T CD8-Positivos/citologia , Linfócitos T CD8-Positivos/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Regulação da Expressão Gênica , Imunidade Inata , Interferon gama/genética , Interferon gama/imunologia , Interleucina-1beta/genética , Interleucina-1beta/imunologia , Subunidade beta de Receptor de Interleucina-2/genética , Subunidade beta de Receptor de Interleucina-2/imunologia , Lectinas Tipo C/genética , Lectinas Tipo C/imunologia , Ativação Linfocitária , Camundongos , Subfamília K de Receptores Semelhantes a Lectina de Células NK/genética , Subfamília K de Receptores Semelhantes a Lectina de Células NK/imunologia , Receptores CXCR3/genética , Receptores CXCR3/imunologia , Receptores Imunológicos/genética , Receptores Imunológicos/imunologia , Receptores de Interleucina-7/genética , Receptores de Interleucina-7/imunologia , Transdução de Sinais , Sirolimo/farmacologia , Serina-Treonina Quinases TOR/antagonistas & inibidores , Serina-Treonina Quinases TOR/imunologia
12.
Eur J Immunol ; 51(7): 1715-1731, 2021 07.
Artigo em Inglês | MEDLINE | ID: mdl-33891704

RESUMO

Hemocyanins are used as immunomodulators in clinical applications because they induce a strong Th1-biased cell-mediated immunity, which has beneficial effects. They are multiligand glycosylated molecules with abundant and complex mannose-rich structures. It remains unclear whether these structures influence hemocyanin-induced immunostimulatory processes in human APCs. We have previously shown that hemocyanin glycans from Concholepas concholepas (CCH), Fissurella latimarginata (FLH), and Megathura crenulata (KLH), participate in their immune recognition and immunogenicity in mice, interacting with murine C-type lectin receptors (CLRs). Here, we studied the interactions of these hemocyanins with two major mannose-binding CLRs on monocyte-derived human DCs: MR (mannose receptor) and DC-SIGN (DC-specific ICAM-3-grabbing nonintegrin). Diverse analyses showed that hemocyanins are internalized by a mannose-sensitive mechanism. This process was calcium dependent. Moreover, hemocyanins colocalized with MR and DC-SIGN, and were partly internalized through clathrin-mediated endocytosis. The hemocyanin-mediated proinflammatory cytokine response was impaired when using deglycosylated FLH and KLH compared to CCH. We further showed that hemocyanins bind to human MR and DC-SIGN in a carbohydrate-dependent manner with affinity constants in the physiological concentration range. Overall, we showed that these three clinically valuable hemocyanins interact with human mannose-sensitive CLRs, initiating an immune response and promoting a Th1 cell-driving potential.


Assuntos
Moléculas de Adesão Celular/imunologia , Células Dendríticas/imunologia , Hemocianinas/imunologia , Fatores Imunológicos/imunologia , Lectinas Tipo C/imunologia , Lectinas de Ligação a Manose/imunologia , Receptores de Superfície Celular/imunologia , Animais , Células CHO , Linhagem Celular Tumoral , Células Cultivadas , Cricetulus , Humanos , Imunidade Celular/imunologia , Imunização/métodos , Monócitos/imunologia , Células U937
13.
Front Immunol ; 12: 661823, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33897708

RESUMO

C-type lectin (CTL), a well-known immune-related molecule, has received more and more attention due to its diverse functions, especially its important role in development and host defense of vertebrate and invertebrate. Since the research on crab CTLs is still lack, we screened a new CTL homolog, named SpCTL6 from mud crab Scylla paramamosain. The full-length cDNA sequence of SpCTL6 was 738 bp with a 486 bp of ORF, and the deduced amino acids were 161 aa. SpCTL6 was predicted to have a 17 aa signal peptide and its mature peptide was 144 aa (MW 16.7 kDa) with pI value of 5.22. It had typical CTL structural characteristics, such as a single C-type lectin-like domain, 4 conserved cysteines, similar tertiary structure to that of vertebrate CTLs and a mutated Ca2+ binding motif Gln-Pro-Thr (QPT), clustering into the same branch as the crustacean CTLs. SpCTL6 was highly expressed in the entire zoeal larval stages and widely distributed in adult crab tissues with the highest transcription level in testis. During the molting process of juvenile crabs, the expression level of SpCTL6 was remarkably increased after molting. SpCTL6 could be significantly upregulated in two larval stages (Z1 and megalopa) and adult crab testis under immune challenges. Recombinant SpCTL6 (rSpCTL6) was successfully obtained from eukaryotic expression system. rSpCTL6 exhibited binding activity with PAMPs (LPS, lipoteichoic acid, peptidoglycan, and glucan) and had a broad spectrum bacterial agglutination activity in a Ca2+-dependent manner. In addition, rSpCTL6 could enhance the encapsulation activity of hemocytes and has no cytotoxic effect on hemocytes. Although rSpCTL6 had no bactericidal activity on Vibrio alginolyticus, rSpCTL6 treatment could significantly reduce the bacterial endotoxin level in vitro and greatly improved the survival of S. paramamosain under V. alginolyticus infection in vivo. The immunoprotective effect of rSpCTL6 might be due to the regulatory role of rSpCTL6 in immune-related genes and immunological parameters. Our study provides new information for understanding the immune defense of mud crabs and would facilitate the development of effective strategies for mud crab aquaculture disease control.


Assuntos
Proteínas de Artrópodes/genética , Proteínas de Artrópodes/metabolismo , Braquiúros/imunologia , Regulação da Expressão Gênica/imunologia , Lectinas Tipo C/genética , Lectinas Tipo C/imunologia , Vibrioses/veterinária , Animais , Braquiúros/efeitos dos fármacos , Braquiúros/genética , Braquiúros/microbiologia , Clonagem Molecular , DNA Complementar/genética , Feminino , Perfilação da Expressão Gênica , Hemócitos/imunologia , Imunidade Inata , Masculino , Alinhamento de Sequência , Vibrioses/imunologia , Vibrioses/prevenção & controle , Vibrio alginolyticus/imunologia
14.
J Exp Med ; 218(6)2021 06 07.
Artigo em Inglês | MEDLINE | ID: mdl-33914023

RESUMO

Tissue-resident memory T cells (TRM cells) are key elements of tissue immunity. Here, we investigated the role of the regulator of T cell receptor and cytokine signaling, Ptpn2, in the formation and function of TRM cells in skin. Ptpn2-deficient CD8+ T cells displayed a marked defect in generating CD69+ CD103+ TRM cells in response to herpes simplex virus type 1 (HSV-1) skin infection. This was accompanied by a reduction in the proportion of KLRG1- memory precursor cells and a transcriptional bias toward terminal differentiation. Of note, forced expression of KLRG1 was sufficient to impede TRM cell formation. Normalizing memory precursor frequencies by transferring equal numbers of KLRG1- cells restored TRM generation, demonstrating that Ptpn2 impacted skin seeding with precursors rather than downstream TRM cell differentiation. Importantly, Ptpn2-deficient TRM cells augmented skin autoimmunity but also afforded superior protection from HSV-1 infection. Our results emphasize that KLRG1 repression is required for optimal TRM cell formation in skin and reveal an important role of Ptpn2 in regulating TRM cell functionality.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Memória Imunológica/imunologia , Lectinas Tipo C/imunologia , Proteína Tirosina Fosfatase não Receptora Tipo 2/imunologia , Receptores Imunológicos/imunologia , Animais , Autoimunidade/imunologia , Feminino , Herpes Simples/imunologia , Herpesvirus Humano 1/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Pele/imunologia
15.
Arch Insect Biochem Physiol ; 107(2): e21786, 2021 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-33818830

RESUMO

Cotesia kariyai (Ck) larvae implanted into the body cavity of the Mythimna separata (armyworm) larvae get melanized and encapsulated after adhesion by hemocytes called hyperspread cells (HSCs). The present study showed that HSCs could not adhere to the implanted Ck larvae in armyworm larvae after injection of Ck polydnavirus (CkPDV) + venom (V), thus melanization and encapsulation could not occur. A C-type lectin called Mys-IML of the host armyworm larvae was considered to be involved in the recognition of foreign substances which always expressed in hemocytes. The CkPDV DNA encodes a C-type lectin called Cky811 that has high amino acid homology to Mys-IML. HSCs did not adhere when CkPDV + V was mixed with the hemolymph of armyworm larvae on glass slides and incubated in vitro, but the addition of anti-Cky811 antibody enabled HSCs to adhere. The messenger RNA (mRNA) expression of Mys-IML in armyworm larvae injected with CkPDV + V became undetectable by 6 h. On the contrary, Cky811 mRNA was well expressed in the hemocytes of armyworm larvae injected with CkPDV + V from 0.5 to 6 h. Cky811 protein was also detected in the crude extracts from Ck venom gland + Ck venom reservoir, suggesting that these proteins regulate foreign substance recognition by the armyworm within 0.5 h. These results suggest that CkPDV + V suppresses mRNA expression of Mys-IML, and that Cky811 protein expressed in hemocytes regulates foreign substance recognition of Mys-IML, resulting in inhibition of the downstream reaction steps: HSCs adhesion, melanization, and encapsulation.


Assuntos
Lectinas Tipo C/imunologia , Mariposas/parasitologia , Polydnaviridae , Vespas , Animais , Anticorpos Antivirais/metabolismo , Hemócitos/imunologia , Hemócitos/metabolismo , Interações Hospedeiro-Parasita/imunologia , Imunidade , Larva/imunologia , Larva/metabolismo , Larva/parasitologia , Lectinas Tipo C/metabolismo , Mariposas/imunologia , Polydnaviridae/metabolismo , Proteínas Virais/imunologia , Proteínas Virais/metabolismo , Vespas/patogenicidade , Vespas/virologia
16.
Fish Shellfish Immunol ; 113: 51-60, 2021 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-33798718

RESUMO

Killer cell lectin-like receptor G subfamily 1 (KLRG1) is a receptor generally expressed on effector CD8+ T cells or NK cells at terminal differentiation stage, and it will be highly induced for lymphocyte cytotoxicity upon pathogen infection or lymphocyte activation. However, little is known about the character or function of KLRG1 in lower vertebrates. In present study, we reappraised a molecule that previously defined as KLRG1 in the genomic sequence of Nile tilapia Oreochromis niloticus, and identified it as an atypical KLRG1-like molecule (defined as On-KLRG1-L), and illustrated its potential function serving as a marker representing effector T lymphocytes of fish species. On-KLRG1-L consists of two C-type lectin-like domains (CTLDs) without transmembrane region, and the tertiary structure of the CTLD is highly alike to that in mouse KLRG1. As a CTLD-containing protein, the recombinant On-KLRG1-L could bind PGN and several microbes in vitro. On-KLRG1-L was widely expressed in immune-associated tissues, with the highest expression level in the gill. Once Nile tilapia is infected by Aeromonas hydrophila, mRNA level of On-KLRG1-L in spleen lymphocytes were significantly up-regulated on 5 days after infection. Meanwhile, On-KLRG1-L protein was also induced on 5 or 8 days after A. hydrophila infection. Furthermore, we found both mRNA and protein levels of On-KLRG1-L were dramatically enhanced within several hours after spleen lymphocytes were activated by T cell-specific mitogen PHA in vitro. More importantly, the ratio of On-KLRG1-L+ T cells was also augmented after PHA stimulation. The observations suggested that the KLRG1-like molecule from Nile tilapia participated in lymphocyte activation and anti-bacterial adaptive immune response, and could serve as an activation marker of T lymphocytes. Our study thus provided new evidences to understand lymphocyte-mediated adaptive immunity of teleost.


Assuntos
Imunidade Adaptativa/genética , Ciclídeos/imunologia , Doenças dos Peixes/imunologia , Lectinas Tipo C/imunologia , Ativação Linfocitária/imunologia , Receptores Imunológicos/imunologia , Aeromonas hydrophila/fisiologia , Sequência de Aminoácidos , Animais , Biomarcadores/metabolismo , Doenças dos Peixes/microbiologia , Proteínas de Peixes/genética , Proteínas de Peixes/imunologia , Infecções por Bactérias Gram-Negativas/imunologia , Infecções por Bactérias Gram-Negativas/microbiologia , Infecções por Bactérias Gram-Negativas/veterinária , Lectinas Tipo C/genética , Estrutura Terciária de Proteína , Receptores Imunológicos/genética , Alinhamento de Sequência/veterinária
17.
Int J Mol Sci ; 22(5)2021 Mar 05.
Artigo em Inglês | MEDLINE | ID: mdl-33808005

RESUMO

Heat-inactivation of sera is used to reduce possible disturbing effects of complement factors in cell-culture experiments, but it is controversially discussed whether this procedure is appropriate or could be neglected. Here, we report a strong impact of heat-inactivation of human sera on the activation and effector functions of human CD4+ T cells. While T cells cultured with native sera were characterized by a higher proliferation rate and higher expression of CD28, heat-inactivated sera shaped T cells towards on-blast formation, higher cytokine secretion (interferon γ, tumor necrosis factor, and interleukin-17), stronger CD69 and PD-1 expression, and increased metabolic activity. Heat-inactivated sera contained reduced amounts of complement factors and regulators like C1 inhibitor, but increased concentrations of circulating immune complexes. Substitution of C1 inhibitor reduced the beneficial effect of heat-inactivation in terms of cytokine release, whereas surface-molecule expression was affected by the addition of complex forming anti-C1q antibody. Our data clearly demonstrate a beneficial effect of heat-inactivation of human sera for T cell experiments but indicate that beside complement regulators and immune complexes other components might be relevant. Beyond that, this study further underpins the strong impact of the complement system on T cell function.


Assuntos
Complexo Antígeno-Anticorpo/imunologia , Linfócitos T CD4-Positivos/imunologia , Proteína Inibidora do Complemento C1/imunologia , Complexo Antígeno-Anticorpo/sangue , Antígenos CD/sangue , Antígenos CD/imunologia , Antígenos de Diferenciação de Linfócitos T/sangue , Antígenos de Diferenciação de Linfócitos T/imunologia , Antígenos CD28/sangue , Antígenos CD28/imunologia , Linfócitos T CD4-Positivos/metabolismo , Proteína Inibidora do Complemento C1/metabolismo , Citocinas/sangue , Citocinas/imunologia , Temperatura Alta , Humanos , Lectinas Tipo C/sangue , Lectinas Tipo C/imunologia , Receptor de Morte Celular Programada 1/sangue , Receptor de Morte Celular Programada 1/imunologia
18.
PLoS One ; 16(3): e0247658, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33667240

RESUMO

Microsporidia are recognized as opportunistic pathogens in individuals with immunodeficiencies, especially related to T cells. Although the activity of CD8+ T lymphocytes is essential to eliminate these pathogens, earlier studies have shown significant participation of macrophages at the beginning of the infection. Macrophages and other innate immunity cells play a critical role in activating the acquired immunity. After programmed cell death, the cell fragments or apoptotic bodies are cleared by phagocytic cells, a phenomenon known as efferocytosis. This process has been recognized as a way of evading immunity by intracellular pathogens. The present study evaluated the impact of efferocytosis of apoptotic cells either infected or not on macrophages and subsequently challenged with Encephalitozoon cuniculi microsporidia. Macrophages were obtained from the bone marrow monocytes from C57BL mice, pre-incubated with apoptotic Jurkat cells (ACs), and were further challenged with E. cuniculi spores. The same procedures were performed using the previously infected Jurkat cells (IACs) and challenged with E. cuniculi spores before macrophage pre-incubation. The average number of spores internalized by macrophages in phagocytosis was counted. Macrophage expression of CD40, CD206, CD80, CD86, and MHCII, as well as the cytokines released in the culture supernatants, was measured by flow cytometry. The ultrastructural study was performed to analyze the multiplication types of pathogens. Macrophages pre-incubated with ACs and challenged with E. cuniculi showed a higher percentage of phagocytosis and an average number of internalized spores. Moreover, the presence of stages of multiplication of the pathogen inside the macrophages, particularly after efferocytosis of infected apoptotic bodies, was observed. In addition, pre-incubation with ACs or IACs and/or challenge with the pathogen decreased the viability of macrophages, reflected as high percentages of apoptosis. The marked expression of CD206 and the release of large amounts of IL-10 and IL-6 indicated the polarization of macrophages to an M2 profile, compatible with efferocytosis and favorable for pathogen development. We concluded that the pathogen favored efferocytosis and polarized the macrophages to an M2 profile, allowing the survival and multiplication of E. cuniculi inside the macrophages and explaining the possibility of macrophages acting as Trojan horses in microsporidiosis.


Assuntos
Apoptose/genética , Encephalitozoon cuniculi/imunologia , Evasão da Resposta Imune , Macrófagos/microbiologia , Esporos Fúngicos/imunologia , Animais , Medula Óssea/imunologia , Medula Óssea/microbiologia , Diferenciação Celular , Técnicas de Cocultura , Encephalitozoon cuniculi/genética , Encephalitozoon cuniculi/crescimento & desenvolvimento , Feminino , Expressão Gênica , Humanos , Interleucina-10/genética , Interleucina-10/imunologia , Interleucina-6/genética , Interleucina-6/imunologia , Células Jurkat , Lectinas Tipo C/genética , Lectinas Tipo C/imunologia , Macrófagos/imunologia , Lectinas de Ligação a Manose/genética , Lectinas de Ligação a Manose/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Fagocitose , Cultura Primária de Células , Receptores de Superfície Celular/genética , Receptores de Superfície Celular/imunologia , Esporos Fúngicos/genética , Esporos Fúngicos/crescimento & desenvolvimento
19.
J Biol Chem ; 296: 100487, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33676898

RESUMO

Numerous observations indicate that red blood cells (RBCs) affect T-cell activation and proliferation. We have studied effects of packed RBCs (PRBCs) on T-cell receptor (TCR) signaling and the molecular mechanisms whereby (P)RBCs modulate T-cell activation. In line with previous reports, PRBCs attenuated the expression of T-cell activation markers CD25 and CD69 upon costimulation via CD3/CD28. In addition, T-cell proliferation and cytokine expression were markedly reduced when T-cells were stimulated in the presence of PRBCs. Inhibitory activity of PRBCs required direct cell-cell contact and intact PRBCs. The production of activation-induced cellular reactive oxygen species, which act as second messengers in T-cells, was completely abrogated to levels of unstimulated T-cells in the presence of PRBCs. Phosphorylation of the TCR-related zeta chain and thus proximal TCR signal transduction was unaffected by PRBCs, ruling out mechanisms based on secreted factors and steric interaction restrictions. In large part, downstream signaling events requiring reactive oxygen species for full functionality were affected, as confirmed by an untargeted MS-based phosphoproteomics approach. PRBCs inhibited T-cell activation more efficiently than treatment with 1 mM of the antioxidant N-acetyl cysteine. Taken together, our data imply that inflammation-related radical reactions are modulated by PRBCs. These immunomodulating effects may be responsible for clinical observations associated with transfusion of PRBCs.


Assuntos
Antígenos CD/imunologia , Antígenos de Diferenciação de Linfócitos T/imunologia , Eritrócitos/imunologia , Subunidade alfa de Receptor de Interleucina-2/imunologia , Lectinas Tipo C/imunologia , Espécies Reativas de Oxigênio/metabolismo , Linfócitos T/imunologia , Antígenos CD/metabolismo , Antígenos de Diferenciação de Linfócitos T/metabolismo , Proliferação de Células/fisiologia , Células Cultivadas , Eritrócitos/metabolismo , Humanos , Imunomodulação , Subunidade alfa de Receptor de Interleucina-2/metabolismo , Lectinas Tipo C/metabolismo , Leucócitos Mononucleares , Ativação Linfocitária , Fosforilação , Transdução de Sinais , Linfócitos T/metabolismo
20.
Front Immunol ; 12: 640367, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33767710

RESUMO

The C-type lectins, one family of lectins featuring carbohydrate binding domains which participate in a variety of bioprocesses in both humans and mosquitoes, including immune response, are known to target DENV. A human C-type lectin protein CLEC18A in particular shows extensive glycan binding abilities and correlates with type-I interferon expression, making CLEC18A a potential player in innate immune responses to DENV infection; this potential may provide additional regulatory point in improving mosquito immunity. Here, we established for the first time a transgenic Aedes aegypti line that expresses human CLEC18A. This expression enhanced the Toll immune pathway responses to DENV infection. Furthermore, viral genome and virus titers were reduced by 70% in the midgut of transgenic mosquitoes. We found significant changes in the composition of the midgut microbiome in CLEC18A expressing mosquitoes, which may result from the Toll pathway enhancement and contribute to DENV inhibition. Transgenic mosquito lines offer a compelling option for studying DENV pathogenesis, and our analyses indicate that modifying the mosquito immune system via expression of a human immune gene can significantly reduce DENV infection.


Assuntos
Aedes/imunologia , Aedes/virologia , Animais Geneticamente Modificados , Dengue/imunologia , Lectinas Tipo C/imunologia , Aedes/genética , Animais , Animais Geneticamente Modificados/genética , Animais Geneticamente Modificados/imunologia , Vírus da Dengue , Modelos Animais de Doenças , Humanos , Mosquitos Vetores/genética , Mosquitos Vetores/imunologia , Mosquitos Vetores/virologia
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