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1.
Nat Immunol ; 22(11): 1382-1390, 2021 11.
Artigo em Inglês | MEDLINE | ID: mdl-34663978

RESUMO

Intergenerational inheritance of immune traits linked to epigenetic modifications has been demonstrated in plants and invertebrates. Here we provide evidence for transmission of trained immunity across generations to murine progeny that survived a sublethal systemic infection with Candida albicans or a zymosan challenge. The progeny of trained mice exhibited cellular, developmental, transcriptional and epigenetic changes associated with the bone marrow-resident myeloid effector and progenitor cell compartment. Moreover, the progeny of trained mice showed enhanced responsiveness to endotoxin challenge, alongside improved protection against systemic heterologous Escherichia coli and Listeria monocytogenes infections. Sperm DNA of parental male mice intravenously infected with the fungus C. albicans showed DNA methylation differences linked to immune gene loci. These results provide evidence for inheritance of trained immunity in mammals, enhancing protection against infections.


Assuntos
Candida albicans/imunologia , Candidíase/imunologia , Infecções por Escherichia coli/imunologia , Escherichia coli/imunologia , Hereditariedade , Imunidade Inata/genética , Listeria monocytogenes/imunologia , Listeriose/imunologia , Células Mieloides/imunologia , Animais , Candida albicans/patogenicidade , Candidíase/genética , Candidíase/metabolismo , Candidíase/microbiologia , Células Cultivadas , Metilação de DNA , Modelos Animais de Doenças , Epigênese Genética , Escherichia coli/patogenicidade , Infecções por Escherichia coli/genética , Infecções por Escherichia coli/metabolismo , Infecções por Escherichia coli/microbiologia , Interações Hospedeiro-Patógeno , Listeria monocytogenes/patogenicidade , Listeriose/genética , Listeriose/metabolismo , Listeriose/microbiologia , Masculino , Camundongos Transgênicos , Células Mieloides/metabolismo , Células Mieloides/microbiologia , Espermatozoides/imunologia , Espermatozoides/metabolismo , Transcrição Genética
2.
PLoS Pathog ; 17(9): e1009493, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-34555127

RESUMO

Listeria monocytogenes is an intracellular bacterium that elicits robust CD8+ T-cell responses. Despite the ongoing development of L. monocytogenes-based platforms as cancer vaccines, our understanding of how L. monocytogenes drives robust CD8+ T-cell responses remains incomplete. One overarching hypothesis is that activation of cytosolic innate pathways is critical for immunity, as strains of L. monocytogenes that are unable to access the cytosol fail to elicit robust CD8+ T-cell responses and in fact inhibit optimal T-cell priming. Counterintuitively, however, activation of known cytosolic pathways, such as the inflammasome and type I IFN, lead to impaired immunity. Conversely, production of prostaglandin E2 (PGE2) downstream of cyclooxygenase-2 (COX-2) is essential for optimal L. monocytogenes T-cell priming. Here, we demonstrate that vacuole-constrained L. monocytogenes elicit reduced PGE2 production compared to wild-type strains in macrophages and dendritic cells ex vivo. In vivo, infection with wild-type L. monocytogenes leads to 10-fold increases in PGE2 production early during infection whereas vacuole-constrained strains fail to induce PGE2 over mock-immunized controls. Mice deficient in COX-2 specifically in Lyz2+ or CD11c+ cells produce less PGE2, suggesting these cell subsets contribute to PGE2 levels in vivo, while depletion of phagocytes with clodronate abolishes PGE2 production completely. Taken together, this work demonstrates that optimal PGE2 production by phagocytes depends on L. monocytogenes access to the cytosol, suggesting that one reason cytosolic access is required to prime CD8+ T-cell responses may be to facilitate production of PGE2.


Assuntos
Células Dendríticas/imunologia , Dinoprostona/biossíntese , Dinoprostona/imunologia , Listeriose/imunologia , Macrófagos/imunologia , Animais , Células Dendríticas/metabolismo , Células Dendríticas/microbiologia , Feminino , Listeria monocytogenes/imunologia , Ativação Linfocitária/imunologia , Macrófagos/metabolismo , Macrófagos/microbiologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL
3.
Nat Commun ; 12(1): 4999, 2021 08 17.
Artigo em Inglês | MEDLINE | ID: mdl-34404769

RESUMO

The type I interferon (IFN) signaling pathway has important functions in resistance to viral infection, with the downstream induction of interferon stimulated genes (ISG) protecting the host from virus entry, replication and spread. Listeria monocytogenes (Lm), a facultative intracellular foodborne pathogen, can exploit the type I IFN response as part of their pathogenic strategy, but the molecular mechanisms involved remain unclear. Here we show that type I IFN suppresses the antibacterial activity of phagocytes to promote systemic Lm infection. Mechanistically, type I IFN suppresses phagosome maturation and proteolysis of Lm virulence factors ActA and LLO, thereby promoting phagosome escape and cell-to-cell spread; the antiviral protein, IFN-induced transmembrane protein 3 (IFITM3), is required for this type I IFN-mediated alteration. Ifitm3-/- mice are resistant to systemic infection by Lm, displaying decreased bacterial spread in tissues, and increased immune cell recruitment and pro-inflammatory cytokine signaling. Together, our findings show how an antiviral mechanism in phagocytes can be exploited by bacterial pathogens, and implicate IFITM3 as a potential antimicrobial therapeutic target.


Assuntos
Antibacterianos/farmacologia , Listeria/efeitos dos fármacos , Listeriose/imunologia , Proteínas de Membrana/antagonistas & inibidores , Proteínas de Membrana/metabolismo , Fagócitos/imunologia , Fagócitos/microbiologia , Animais , Modelos Animais de Doenças , Interações Hospedeiro-Patógeno , Interferon Tipo I/metabolismo , Listeria monocytogenes/imunologia , Masculino , Proteínas de Membrana/genética , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fagossomos/imunologia , Células RAW 264.7 , Transcriptoma , Fatores de Virulência , Internalização do Vírus/efeitos dos fármacos
4.
Sci Rep ; 11(1): 16347, 2021 08 11.
Artigo em Inglês | MEDLINE | ID: mdl-34381163

RESUMO

Gamma-delta (γδ) T cells express T cell receptors (TCR) that are preconfigured to recognize signs of pathogen infection. In primates, γδ T cells expressing the Vγ9Vδ2 TCR innately recognize (E)-4-hydroxy-3-methyl-but- 2-enyl pyrophosphate (HMBPP), a product of the 2-C-methyl-D-erythritol 4- phosphate (MEP) pathway in bacteria that is presented in infected cells via interaction with members of the B7 family of costimulatory molecules butyrophilin (BTN) 3A1 and BTN2A1. In humans, Listeria monocytogenes (Lm) vaccine platforms have the potential to generate potent Vγ9Vδ2 T cell recognition. To evaluate the activation of Vγ9Vδ2 T cells by Lm-infected human monocyte-derived dendritic cells (Mo-DC) we engineered Lm strains that lack components of the MEP pathway. Direct infection of Mo-DC with these bacteria were unchanged in their ability to activate CD107a expression in Vγ9Vδ2 T cells despite an inability to synthesize HMBPP. Importantly, functional BTN3A1 was essential for this activation. Unexpectedly, we found that cytoplasmic entry of Lm into human dendritic cells resulted in upregulation of cholesterol metabolism in these cells, and the effect of pathway regulatory drugs suggest this occurs via increased synthesis of the alternative endogenous Vγ9Vδ2 ligand isoprenyl pyrophosphate (IPP) and/or its isomer dimethylallyl pyrophosphate (DMAPP). Thus, following direct infection, host pathways regulated by cytoplasmic entry of Lm can trigger Vγ9Vδ2 T cell recognition of infected cells without production of the unique bacterial ligand HMBPP.


Assuntos
Células Dendríticas/imunologia , Listeria monocytogenes/imunologia , Monócitos/imunologia , Organofosfatos/imunologia , Receptores de Antígenos de Linfócitos T gama-delta/imunologia , Linfócitos T/imunologia , Butirofilinas/imunologia , Células Cultivadas , Hemiterpenos/imunologia , Humanos , Ativação Linfocitária/imunologia , Proteína 1 de Membrana Associada ao Lisossomo/imunologia , Compostos Organofosforados/imunologia , Ligação Proteica/imunologia
5.
Front Immunol ; 12: 650779, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34194428

RESUMO

Listeria monocytogenes (L.m) is efficiently controlled by several cells of the innate immunity, including the Mast Cell (MC). MC is activated by L.m inducing its degranulation, cytokine production and microbicidal mechanisms. TLR2 is required for the optimal control of L.m infection by different cells of the immune system. However, little is known about the MC receptors involved in recognizing this bacterium and whether these interactions mediate MC activation. In this study, we analyzed whether TLR2 is involved in mediating different MC activation responses during L.m infection. We found that despite MC were infected with L.m, they were able to clear the bacterial load. In addition, MC degranulated and produced ROS, TNF-α, IL-1ß, IL-6, IL-13 and MCP-1 in response to bacterial infection. Interestingly, L.m induced the activation of signaling proteins: ERK, p38 and NF-κB. When TLR2 was blocked, L.m endocytosis, bactericidal activity, ROS production and mast cell degranulation were not affected. Interestingly, only IL-6 and IL-13 production were affected when TLR2 was inhibited in response to L.m infection. Furthermore, p38 activation depended on TLR2, but not ERK or NF-κB activation. These results indicate that TLR2 mediates only some MC activation pathways during L.m infection, mainly those related to IL-6 and IL-13 production.


Assuntos
Interleucina-13/imunologia , Interleucina-6/imunologia , Listeria monocytogenes/imunologia , Mastócitos/imunologia , Receptor 2 Toll-Like/imunologia , Animais , Degranulação Celular/imunologia , Degranulação Celular/fisiologia , Células Cultivadas , Citocinas/imunologia , Citocinas/metabolismo , Ativação Enzimática/imunologia , Interações Hospedeiro-Patógeno/imunologia , Interleucina-13/metabolismo , Interleucina-6/metabolismo , Listeria monocytogenes/fisiologia , Mastócitos/microbiologia , Mastócitos/fisiologia , Camundongos Endogâmicos C57BL , NF-kappa B/imunologia , NF-kappa B/metabolismo , Espécies Reativas de Oxigênio/imunologia , Espécies Reativas de Oxigênio/metabolismo , Receptor 2 Toll-Like/metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno/imunologia , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo
6.
PLoS Pathog ; 17(7): e1009697, 2021 07.
Artigo em Inglês | MEDLINE | ID: mdl-34237114

RESUMO

Listeria monocytogenes (L. monocytogenes) is a food-borne bacterial pathogen. Innate immunity to L. monocytogenes is profoundly affected by type I interferons (IFN-I). Here we investigated host metabolism in L. monocytogenes-infected mice and its potential control by IFN-I. Accordingly, we used animals lacking either the IFN-I receptor (IFNAR) or IRF9, a subunit of ISGF3, the master regulator of IFN-I-induced genes. Transcriptomes and metabolite profiles showed that L. monocytogenes infection induces metabolic rewiring of the liver. This affects various metabolic pathways including fatty acid (FA) metabolism and oxidative phosphorylation and is partially dependent on IFN-I signaling. Livers and macrophages from Ifnar1-/- mice employ increased glutaminolysis in an IRF9-independent manner, possibly to readjust TCA metabolite levels due to reduced FA oxidation. Moreover, FA oxidation inhibition provides protection from L. monocytogenes infection, explaining part of the protection of Irf9-/- and Ifnar1-/- mice. Our findings define a role of IFN-I in metabolic regulation during L. monocytogenes infection. Metabolic differences between Irf9-/- and Ifnar1-/- mice may underlie the different susceptibility of these mice against lethal infection with L. monocytogenes.


Assuntos
Interferon Tipo I/metabolismo , Listeria monocytogenes/metabolismo , Listeriose/metabolismo , Fígado/metabolismo , Animais , Ácidos Graxos/metabolismo , Interferon Tipo I/imunologia , Listeria monocytogenes/imunologia , Listeriose/imunologia , Fígado/imunologia , Camundongos , Camundongos Endogâmicos C57BL
7.
Nat Commun ; 12(1): 3392, 2021 06 07.
Artigo em Inglês | MEDLINE | ID: mdl-34099666

RESUMO

Cells infected with pathogens can contribute to clearing infections by releasing signals that instruct neighbouring cells to mount a pro-inflammatory cytokine response, or by other mechanisms that reduce bystander cells' susceptibility to infection. Here, we show the opposite effect: epithelial cells infected with Salmonella Typhimurium secrete host factors that facilitate the infection of bystander cells. We find that the endoplasmic reticulum stress response is activated in both infected and bystander cells, and this leads to activation of JNK pathway, downregulation of transcription factor E2F1, and consequent reprogramming of microRNA expression in a time-dependent manner. These changes are not elicited by infection with other bacterial pathogens, such as Shigella flexneri or Listeria monocytogenes. Remarkably, the protein HMGB1 present in the secretome of Salmonella-infected cells is responsible for the activation of the IRE1 branch of the endoplasmic reticulum stress response in non-infected, neighbouring cells. Furthermore, E2F1 downregulation and the associated microRNA alterations promote Salmonella replication within infected cells and prime bystander cells for more efficient infection.


Assuntos
Efeito Espectador/genética , Fator de Transcrição E2F1/metabolismo , MicroRNAs/metabolismo , Infecções por Salmonella/imunologia , Salmonella typhimurium/imunologia , Animais , Efeito Espectador/imunologia , Modelos Animais de Doenças , Regulação para Baixo/imunologia , Fator de Transcrição E2F1/genética , Estresse do Retículo Endoplasmático/imunologia , Endorribonucleases/metabolismo , Proteína HMGB1/metabolismo , Células HeLa , Interações Hospedeiro-Patógeno/genética , Interações Hospedeiro-Patógeno/imunologia , Humanos , Listeria monocytogenes/imunologia , Sistema de Sinalização das MAP Quinases/genética , Sistema de Sinalização das MAP Quinases/imunologia , Proteínas Serina-Treonina Quinases/metabolismo , RNA-Seq , Infecções por Salmonella/genética , Infecções por Salmonella/microbiologia , Salmonella typhimurium/patogenicidade , Shigella flexneri/imunologia , Suínos
8.
Front Immunol ; 12: 667664, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34135896

RESUMO

The yellow mealworm beetle (Tenebrio molitor) has been exploited as an experimental model to unravel the intricacies of cellular and humoral immunity against pathogenic infections. Studies on this insect model have provided valuable insights into the phenotypic plasticity of immune defenses against parasites and pathogens. It has thus been possible to characterize the hemocoelic defenses of T. molitor that rely on the recognition of non-self-components of pathogens by pattern recognition receptors (PRRs). The subsequent signaling cascade activating pathways such as the NF-κB controlled by Toll and IMD pathways lead to the synthesis of antimicrobial peptides (AMPs), onset of hemocyte-driven phagocytosis, and activation of the prophenoloxidase cascade regulating the process of melanization. Nevertheless, the activation of autophagy-mediated defenses of T. molitor against the facultative intracellular gram-positive bacterium Listeria monocytogenes provides clear evidence of the existence of a cross-talk between autophagy and the IMD pathway. Moreover, the identification of several autophagy-related genes (Atgs) in T. molitor transcriptome and expressed sequence tag (EST) databases has contributed to the understanding of the autophagy-signaling cascade triggered by L. monocytogenes challenge. Providing further evidence of the cross-talk hypothesis, TmRelish has been shown to be required not only for regulating the synthesis of AMPs through the PGRP-LE/IMD pathway activation but also for the expression of Atgs in T. molitor larvae following L. monocytogenes challenge. Notably, L. monocytogenes can stimulate the T. molitor innate immune system by producing molecules recognized by the multifunctional PRR (TmPGRP-LE), which stimulates intracellular activation of the IMD pathway and autophagy. Considering the conservation of autophagy components involved in combating intracellular pathogens, it will be interesting to extrapolate a dynamic cross-talk model of immune activation. This review summarizes the most significant findings on the regulation of autophagy in T. molitor during L. monocytogenes infection and on the role of the innate immunity machinery, including the NF-κB pathway, in the control of pathogenic load.


Assuntos
Proteínas Relacionadas à Autofagia/metabolismo , Imunidade Inata , Proteínas de Insetos/metabolismo , Listeria monocytogenes/patogenicidade , Listeriose/microbiologia , Macroautofagia , Tenebrio/microbiologia , Animais , Proteínas Relacionadas à Autofagia/genética , Carga Bacteriana , Interações Hospedeiro-Patógeno , Proteínas de Insetos/genética , Listeria monocytogenes/imunologia , Listeriose/genética , Listeriose/imunologia , Listeriose/metabolismo , Transdução de Sinais , Tenebrio/genética , Tenebrio/imunologia , Tenebrio/metabolismo
9.
Cell Physiol Biochem ; 55(3): 256-264, 2021 May 14.
Artigo em Inglês | MEDLINE | ID: mdl-33984198

RESUMO

BACKGROUND/AIMS: During an immune response, type I interferon (IFN-I) signaling induces a wide range of changes, including those which are required to overcome viral infection and those which suppress cytotoxic T cells to avoid immunopathology. During certain bacterial infections, IFN-I signaling exerts largely detrimental effects. Although the IFN-I family of proteins all share one common receptor, biologic responses to signaling vary depending on IFN-I subtype. Here, we asked if one IFN-I subtype dominates the pro-bacterial effect of IFN-I signaling and found that control of Listeria monocytogenes (L.m.) infection is more strongly suppressed by IFN-ß than IFN-α. METHODS: To study this, we measured bacterial titers in IFNAR-/-, IFN-ß­/­, Stat2-/-, Usp18fl/fl and Usp18fl/fl x CD11c-Cre mice models in addition to IFN-I blocking antibodies. Moreover, we measured interferon stimulated genes in bone marrow derived dendritic cells after treatment with IFN-α4 and IFN-ß. RESULTS: Specifically, we show that genetic deletion of IFN-ß or antibody-mediated IFN-ß neutralization was sufficient to reduce bacterial titers to levels similar to those observed in mice that completely lack IFN-I signaling (IFNAR-/- mice). However, IFN-α blockade failed to significantly reduce L.m. titers, suggesting that IFN-ß is the dominant IFN-I subtype responsible for the pro-bacterial effect of IFN-I. Mechanistically, when focusing on IFN-I signals to dendritic cells, we found that IFN-ß induces ISGs more robustly than IFN-α, including USP18, the protein we previously identified as driving the pro-bacterial effects of IFN-I. Further, we found that this induction was STAT1/STAT2 heterodimer- or STAT2/STAT2 homodimer-dependent, as STAT2-deficient mice were more resistant to L.m. infection. CONCLUSION: In conclusion, IFN-Β is the principal member of the IFN-I family responsible for driving the pro-bacterial effect of IFN-I.


Assuntos
Interferon-alfa/imunologia , Interferon beta/imunologia , Listeria monocytogenes/imunologia , Listeriose/imunologia , Animais , Feminino , Interferon-alfa/genética , Interferon beta/genética , Listeriose/genética , Masculino , Camundongos , Camundongos Knockout , Receptor de Interferon alfa e beta/deficiência , Receptor de Interferon alfa e beta/imunologia
10.
J Immunol ; 206(11): 2638-2651, 2021 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-34031148

RESUMO

HIV reservoirs persist in gut-homing CD4+ T cells of people living with HIV and receiving antiretroviral therapy, but the antigenic specificity of such reservoirs remains poorly documented. The imprinting for gut homing is mediated by retinoic acid (RA), a vitamin A-derived metabolite produced by dendritic cells (DCs) exhibiting RA-synthesizing (RALDH) activity. RALDH activity in DCs can be induced by TLR2 ligands, such as bacterial peptidoglycans and fungal zymosan. Thus, we hypothesized that bacterial/fungal pathogens triggering RALDH activity in DCs fuel HIV reservoir establishment/outgrowth in pathogen-reactive CD4+ T cells. Our results demonstrate that DCs derived from intermediate/nonclassical CD16+ compared with classical CD16- monocytes exhibited superior RALDH activity and higher capacity to transmit HIV infection to autologous Staphylococcus aureus-reactive T cells. Exposure of total monocyte-derived DCs (MDDCs) to S. aureus lysates as well as TLR2 (zymosan and heat-killed preparation of Listeria monocytogenes) and TLR4 (LPS) agonists but not CMV lysates resulted in a robust upregulation of RALDH activity. MDDCs loaded with S. aureus or zymosan induced the proliferation of T cells with a CCR5+integrin ß7+CCR6+ phenotype and efficiently transmitted HIV infection to these T cells via RALDH/RA-dependent mechanisms. Finally, S. aureus- and zymosan-reactive CD4+ T cells of antiretroviral therapy-treated people living with HIV carried replication-competent integrated HIV-DNA, as demonstrated by an MDDC-based viral outgrowth assay. Together, these results support a model in which bacterial/fungal pathogens in the gut promote RALDH activity in MDDCs, especially in CD16+ MDDCs, and subsequently imprint CD4+ T cells with gut-homing potential and HIV permissiveness. Thus, nonviral pathogens play key roles in fueling HIV reservoir establishment/outgrowth via RALDH/RA-dependent mechanisms that may be therapeutically targeted.


Assuntos
Linfócitos T CD4-Positivos/imunologia , Células Dendríticas/imunologia , Infecções por HIV/imunologia , Monócitos/imunologia , Receptores de IgG/imunologia , Tretinoína/imunologia , Adulto , Feminino , Fungos/imunologia , Proteínas Ligadas por GPI/imunologia , Humanos , Listeria monocytogenes/imunologia , Masculino , Pessoa de Meia-Idade , Staphylococcus aureus/imunologia , Adulto Jovem
11.
PLoS Pathog ; 17(4): e1009531, 2021 04.
Artigo em Inglês | MEDLINE | ID: mdl-33878120

RESUMO

Most individuals who consume foods contaminated with the bacterial pathogen Listeria monocytogenes (Lm) develop mild symptoms, while others are susceptible to life-threatening systemic infections (listeriosis). Although it is known that the risk of severe disease is increased in certain human populations, including the elderly, it remains unclear why others who consume contaminated food develop listeriosis. Here, we used a murine model to discover that pulmonary coinfections can impair the host's ability to adequately control and eradicate systemic Lm that cross from the intestines to the bloodstream. We found that the resistance of mice to oral Lm infection was dramatically reduced by coinfection with Streptococcus pneumoniae (Spn), a bacterium that colonizes the respiratory tract and can also cause severe infections in the elderly. Exposure to Spn or microbial products, including a recombinant Lm protein (L1S) and lipopolysaccharide (LPS), rendered otherwise resistant hosts susceptible to severe systemic Lm infection. In addition, we show that this increase in susceptibility was dependent on an increase in the production of interleukin-10 (IL-10) from Ncr1+ cells, including natural killer (NK) cells. Lastly, the ability of Ncr1+ cell derived IL-10 to increase disease susceptibility correlated with a dampening of both myeloid cell accumulation and myeloid cell phagocytic capacity in infected tissues. These data suggest that efforts to minimize inflammation in response to an insult at the respiratory mucosa render the host more susceptible to infections by Lm and possibly other pathogens that access the oral mucosa.


Assuntos
Listeria monocytogenes/imunologia , Listeriose/imunologia , Pneumonia/imunologia , Animais , Progressão da Doença , Suscetibilidade a Doenças , Feminino , Interleucina-10/metabolismo , Células Matadoras Naturais/metabolismo , Células Matadoras Naturais/fisiologia , Lipopolissacarídeos , Listeria monocytogenes/patogenicidade , Listeriose/complicações , Listeriose/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Doenças da Boca/complicações , Doenças da Boca/imunologia , Doenças da Boca/microbiologia , Doenças da Boca/patologia , Pneumonia/complicações , Pneumonia/etiologia , Pneumonia/patologia
12.
Genes (Basel) ; 12(3)2021 03 13.
Artigo em Inglês | MEDLINE | ID: mdl-33805671

RESUMO

Listeriosis is a food-borne illness caused by Listeria monocytogenes. Ampicillin (AMP) alone or in combination with gentamicin (GEN) is the first-line treatment option. Membrane vesicle (MV) production in L. monocytogenes under antibiotic stress conditions and pathologic roles of these MVs in hosts have not been reported yet. Thus, the aim of this study was to investigate the production of MVs in L. monocytogenes cultured with sub-minimum inhibitory concentrations (MICs) of AMP, GEN, or trimethoprim/sulfamethoxazole (SXT) and determine pathologic effects of these MVs in colon epithelial Caco-2 cells. L. monocytogenes cultured in tryptic soy broth with 1/2 MIC of AMP, GEN, or SXT produced 6.0, 2.9, or 1.5 times more MV particles, respectively, than bacteria cultured without antibiotics. MVs from L. monocytogenes cultured with AMP (MVAMP), GEN (MVGEN), or SXT (MVSXT) were more cytotoxic to Caco-2 cell than MVs obtained from cultivation without antibiotics (MVTSB). MVAMP induced more expression of tumor necrosis factor (TNF)-α gene than MVTSB, MVGEN and MVSXT, whereas MVTSB induced more expression of interleukin (IL)-1ß and IL-8 genes than other MVs. Expression of pro-inflammatory cytokine genes by L. monocytogenes MVs was significantly inhibited by proteinase K treatment of MVs. In conclusion, antibiotic stress can trigger the biogenesis of MVs in L. monocytogenes and MVs produced by L. monocytogenes exposed to sub-MIC of AMP can induce strong pro-inflammatory responses by expressing TNF-α gene in host cells, which may contribute to the pathology of listeriosis.


Assuntos
Antibacterianos/farmacologia , Imunidade Inata/efeitos dos fármacos , Listeria monocytogenes/efeitos dos fármacos , Listeria monocytogenes/imunologia , Proteínas de Bactérias/imunologia , Células CACO-2 , Linhagem Celular Tumoral , Citocinas/imunologia , Humanos , Listeriose/tratamento farmacológico , Listeriose/imunologia , Testes de Sensibilidade Microbiana/métodos , Fatores de Virulência/imunologia
13.
J Microbiol ; 59(7): 681-692, 2021 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-33877580

RESUMO

Listeria monocytogenes (L. monocytogenes) is a Gram-positive intracellular foodborne pathogen that causes severe diseases, such as meningitis and sepsis. The NLR family pyrin domain-containing 3 (NLRP3) inflammasome has been reported to participate in host defense against pathogen infection. However, the exact molecular mechanisms underlying NLRP3 inflammasome activation remain to be fully elucidated. In the present study, the roles of mammalian Ste20-like kinases 1/2 (Mst1/2) and Anaplastic Lymphoma Kinase (ALK) in the activation of the NLRP3 inflammasome induced by L. monocytogenes infection were investigated. The expression levels of Mst1/2, phospho (p)-ALK, p-JNK, Nek7, and NLRP3 downstream molecules including activated cas-pase-1 (p20) and mature interleukin (IL)-1ß (p17), were up-regulated in L. monocytogenes-infected macrophages. The ALK inhibitor significantly decreased the expression of p-JNK, Nek7, and NLRP3 downstream molecules in macrophages infected with L. monocytogenes. Furthermore, the Mst1/2 inhibitor markedly inhibited the L. monocytogenes-induced activation of ALK, subsequently downregulating the expression of p-JNK, Nek7, and NLRP3 downstream molecules. Therefore, our study demonstrated that Mst1/2-ALK mediated the activation of the NLRP3 inflammasome by promoting the interaction between Nek7 and NLRP3 via JNK during L. monocytogenes infection, which subsequently increased the maturation and release of proinflammatory cytokine to resist pathogen infection. Moreover, Listeriolysin O played a key role in the process. In addition, we also found that the L. monocytogenes-induced apoptosis of J774A.1 cells was reduced by the Mst1/2 or ALK inhibitor. The present study reported, for the first time, that the Mst1/2-ALK-JNK-NLRP3 signaling pathway plays a vital proinflammatory role during L. monocytogenes infection.


Assuntos
Quinase do Linfoma Anaplásico/metabolismo , Apoptose , Inflamassomos/imunologia , Listeria monocytogenes/imunologia , Macrófagos/imunologia , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Animais , Linhagem Celular Tumoral , Citocinas/metabolismo , Regulação para Baixo , Feminino , Inflamassomos/metabolismo , Proteínas Quinases JNK Ativadas por Mitógeno/genética , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Macrófagos/metabolismo , Macrófagos/microbiologia , Macrófagos Peritoneais/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Quinases Relacionadas a NIMA/genética , Quinases Relacionadas a NIMA/metabolismo , Transdução de Sinais , Regulação para Cima
14.
Front Immunol ; 12: 640644, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33717195

RESUMO

Infection with SARS-CoV-2 can lead to Coronavirus disease-2019 (COVID-19) and result in severe acute respiratory distress syndrome (ARDS). Recent reports indicate an increased rate of fungal coinfections during COVID-19. With incomplete understanding of the pathogenesis and without any causative therapy available, secondary infections may be detrimental to the prognosis. We monitored 11 COVID-19 patients with ARDS for their immune phenotype, plasma cytokines, and clinical parameters on the day of ICU admission and on day 4 and day 7 of their ICU stay. Whole blood stimulation assays with lipopolysaccharide (LPS), heat-killed Listeria monocytogenes (HKLM), Aspergillus fumigatus, and Candida albicans were used to mimic secondary infections, and changes in immune phenotype and cytokine release were assessed. COVID-19 patients displayed an immune phenotype characterized by increased HLA-DR+CD38+ and PD-1+ CD4+ and CD8+ T cells, and elevated CD8+CD244+ lymphocytes, compared to healthy controls. Monocyte activation markers and cytokines IL-6, IL-8, TNF, IL-10, and sIL2Rα were elevated, corresponding to monocyte activation syndrome, while IL-1ß levels were low. LPS, HKLM and Aspergillus fumigatus antigen stimulation provoked an immune response that did not differ between COVID-19 patients and healthy controls, while COVID-19 patients showed an attenuated monocyte CD80 upregulation and abrogated release of IL-6, TNF, IL-1α, and IL-1ß toward Candida albicans. This study adds further detail to the characterization of the immune response in critically ill COVID-19 patients and hints at an increased susceptibility for Candida albicans infection.


Assuntos
Aspergillus fumigatus/imunologia , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , COVID-19/imunologia , Candida albicans/imunologia , Listeria monocytogenes/imunologia , SARS-CoV-2/fisiologia , Idoso , Células Cultivadas , Citocinas/metabolismo , Suscetibilidade a Doenças , Feminino , Humanos , Tolerância Imunológica , Masculino , Pessoa de Meia-Idade , Receptor de Morte Celular Programada 1/metabolismo , Síndrome do Desconforto Respiratório
15.
Immunity ; 54(4): 829-844.e5, 2021 04 13.
Artigo em Inglês | MEDLINE | ID: mdl-33705706

RESUMO

Memory T cells are thought to rely on oxidative phosphorylation and short-lived effector T cells on glycolysis. Here, we investigated how T cells arrive at these states during an immune response. To understand the metabolic state of rare, early-activated T cells, we adapted mass cytometry to quantify metabolic regulators at single-cell resolution in parallel with cell signaling, proliferation, and effector function. We interrogated CD8+ T cell activation in vitro and in response to Listeria monocytogenes infection in vivo. This approach revealed a distinct metabolic state in early-activated T cells characterized by maximal expression of glycolytic and oxidative metabolic proteins. Cells in this transient state were most abundant 5 days post-infection before rapidly decreasing metabolic protein expression. Analogous findings were observed in chimeric antigen receptor (CAR) T cells interrogated longitudinally in advanced lymphoma patients. Our study demonstrates the utility of single-cell metabolic analysis by mass cytometry to identify metabolic adaptations of immune cell populations in vivo and provides a resource for investigations of metabolic regulation of immune responses across a variety of applications.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Ativação Linfocitária/imunologia , Transdução de Sinais/imunologia , Animais , Proliferação de Células/fisiologia , Feminino , Glicólise/imunologia , Memória Imunológica/imunologia , Listeria monocytogenes/imunologia , Listeriose/imunologia , Listeriose/microbiologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Fosforilação Oxidativa , Receptores de Antígenos Quiméricos/imunologia , Análise de Célula Única/métodos
16.
Int J Biol Macromol ; 176: 567-577, 2021 Apr 15.
Artigo em Inglês | MEDLINE | ID: mdl-33581203

RESUMO

Listeria monocytogenes is a cause of infectious food-borne disease in humans, characterized by neurological manifestations, abortion, and neonatal septicemia. It is intracellular bacterium, which limits the development of protective inactivated vacines. Adjuvants capable of stimulating cellular immune response are important tools for developing novel vaccines against intracellular bacteria. The aim of this study was to evaluate the vaccine potential of L. monocytogenes inactivated by gamma irradiation (KLM-γ) encapsulated in alginate microcapsules associated or not with chitosan against listeriosis in the murine model. At the fourth day after challenge there was a reduction in bacterial recovery in mice vaccinated with KLM-γ encapsulated with alginate or alginate-chitosan, with lower bacterial loads in the spleen (10 fold) and liver (100 fold) when compared to non-vaccinated mice. In vitro stimulation of splenocytes from mice vaccinated with alginate-chitosan-encapsulated KLM-γ resulted in lymphocyte proliferation, increase of proportion of memory CD4+ and CD8+ T cell and production of IL-10 and IFN-γ. Interestingly, the group vaccinated with alginate-chitosan-encapsulated KLM-γ had increased survival to lethal infection with lower L. monocytogenes-induced hepatic inflammation and necrosis. Therefore, KLM-γ encapsulation with alginate-chitosan proved to have potential for development of novel and safe inactivated vaccine formulations against listeriosis.


Assuntos
Alginatos , Vacinas Bacterianas , Quitosana , Raios gama , Listeria monocytogenes , Listeriose , Alginatos/química , Alginatos/farmacologia , Animais , Vacinas Bacterianas/química , Vacinas Bacterianas/imunologia , Vacinas Bacterianas/farmacologia , Quitosana/química , Quitosana/farmacologia , Modelos Animais de Doenças , Feminino , Listeria monocytogenes/química , Listeria monocytogenes/imunologia , Listeriose/imunologia , Listeriose/prevenção & controle , Camundongos , Camundongos Endogâmicos BALB C , Vacinas de Produtos Inativados/química , Vacinas de Produtos Inativados/imunologia , Vacinas de Produtos Inativados/farmacologia
17.
Toxicol Appl Pharmacol ; 415: 115441, 2021 03 15.
Artigo em Inglês | MEDLINE | ID: mdl-33556388

RESUMO

The immunotoxicity of zearalenone (ZEA) and deoxynivalenol (DON), two of the most common environmental mycotoxins, has been well investigated. However, due to the complexity of the immune system, especially during bacterial infection, many types of immune cells are involved in invasion resistance and bacterial clearance. Of these, T helper 2 (Th2) cells, which are members of the helper T cell family, assist B cells to activate and differentiate into antibody-secreting cells, participate in humoral immune response, and, ultimately, eliminate pathogens. Thus, it is important to identify the stage at which these toxins affect the immune function, and to clarity the underlying mechanisms. In this study, mice infected with Listeria monocytogenes (Listeria) were used to study the effects of ZEA, DON, and ZEA + DON on Th2 differentiation, Interleukin-4 Receptor (IL-4R) expression, costimulatory molecules expression and cytokine secretion after Listeria infection. Naive CD4+ T cells, isolated from mice, were used to verify the in vivo effects and the associated mechanisms. In vivo experiments showed that these toxins aggravated spleen damage after Listeria infection and reduced the differentiation of Th2 cells by affecting the synthesis of IL-4R of CD4+ T cells. In addition, the level of the costimulatory molecule CD154 decreased. Consistent with this, in vitro studies showed that these toxins inhibited the differentiation of mouse naive CD4+ T cell into Th2 subtype and decreased IL-4R levels. In addition, the levels of costimulatory molecules CD154, CD278 and the Th2 cells secrete cytokines IL-4, IL-6, and IL-10 decreased. Based on our in vivo and in vitro experiments, we suggest that ZEA, DON, and ZEA + DON inhibit the expression of costimulatory molecules on CD4+ T cell, and inhibit the IL-4R-mediated Th2 cell differentiation. This may indicate that the body cannot normally resist or clear the pathogen after mycotoxin poisoning.


Assuntos
Diferenciação Celular/efeitos dos fármacos , Listeria monocytogenes/patogenicidade , Listeriose/induzido quimicamente , Ativação Linfocitária/efeitos dos fármacos , Receptores de Interleucina-4/metabolismo , Baço/efeitos dos fármacos , Células Th2/efeitos dos fármacos , Tricotecenos/toxicidade , Zearalenona/toxicidade , Animais , Ligante de CD40/metabolismo , Células Cultivadas , Citocinas/metabolismo , Modelos Animais de Doenças , Feminino , Interações Hospedeiro-Patógeno , Proteína Coestimuladora de Linfócitos T Induzíveis/metabolismo , Listeria monocytogenes/imunologia , Listeriose/imunologia , Listeriose/metabolismo , Listeriose/microbiologia , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Transdução de Sinais , Baço/imunologia , Baço/metabolismo , Baço/microbiologia , Células Th2/imunologia , Células Th2/metabolismo , Células Th2/microbiologia
18.
Infect Immun ; 89(4)2021 03 17.
Artigo em Inglês | MEDLINE | ID: mdl-33431704

RESUMO

The mucin Muc2 is a major constituent of the mucus layer that covers the intestinal epithelium and creates a barrier between epithelial cells and luminal commensal or pathogenic microorganisms. The Gram-positive foodborne pathogen Listeria monocytogenes can cause enteritis and also disseminate from the intestine to give rise to systemic disease. L. monocytogenes can bind to intestinal Muc2, but the influence of the Muc2 mucin barrier on L. monocytogenes intestinal colonization and systemic dissemination has not been explored. Here, we used an orogastric L. monocytogenes infection model to investigate the role of Muc2 in host defense against L. monocytogenes Compared to wild-type mice, we found that Muc2-/- mice exhibited heightened susceptibility to orogastric challenge with L. monocytogenes, with higher mortality, elevated colonic pathology, and increased pathogen burdens in both the intestinal tract and distal organs. In contrast, L. monocytogenes burdens were equivalent in wild-type and Muc2-/- animals when the pathogen was administered intraperitoneally, suggesting that systemic immune defects related to Muc2 deficiency do not explain the heightened pathogen dissemination observed in oral infections. Using a barcoded L. monocytogenes library to measure intrahost pathogen population dynamics, we found that Muc2-/- animals had larger pathogen founding population sizes in the intestine and distal sites than observed in wild-type animals. Comparisons of barcode frequencies suggested that the colon becomes the major source for seeding the internal organs in Muc2-/- animals. Together, our findings reveal that Muc2 mucin plays a key role in controlling L. monocytogenes colonization, dissemination, and population dynamics.


Assuntos
Listeria monocytogenes , Listeriose/microbiologia , Mucina-2/deficiência , Animais , Carga Bacteriana , Modelos Animais de Doenças , Suscetibilidade a Doenças , Genótipo , Mucosa Intestinal/microbiologia , Mucosa Intestinal/patologia , Listeria monocytogenes/imunologia , Listeriose/genética , Listeriose/mortalidade , Camundongos , Camundongos Knockout , Mortalidade , Especificidade de Órgãos
19.
Foodborne Pathog Dis ; 18(4): 267-275, 2021 04.
Artigo em Inglês | MEDLINE | ID: mdl-33493413

RESUMO

Consumption of raw food, especially smoked fish, meat, soft cheeses, and vegetables, contaminated with Listeria monocytogenes can cause listeriosis, which can be invasive in pregnant women, elderly, and immunocompromised and diabetic patients. Through June to November of 2017, 11 patients developed invasive listeriosis in a small area of northern Italy. In the same period, 15 food samples (ready-to-eat seafood, raw vegetables, cheese samples, and salami) collected during the routine screening programs in the same area were found to be contaminated with L. monocytogenes. We characterized the isolates to determine the relatedness of L. monocytogenes strains isolated from patients and isolates from food samples and food-processing plants. Whole genome sequencing analysis showed that multiple L. monocytogenes strains were circulating in the area and no association was found between clinical and food isolates.


Assuntos
Manipulação de Alimentos/estatística & dados numéricos , Microbiologia de Alimentos/estatística & dados numéricos , Listeria monocytogenes/isolamento & purificação , Listeriose/epidemiologia , Adulto , Idoso , Queijo/microbiologia , Feminino , Humanos , Hospedeiro Imunocomprometido/imunologia , Itália/epidemiologia , Listeria monocytogenes/genética , Listeria monocytogenes/imunologia , Listeriose/imunologia , Masculino , Carne/microbiologia , Pessoa de Meia-Idade , Alimentos Marinhos/microbiologia , Verduras/microbiologia , Sequenciamento Completo do Genoma
20.
Scand J Immunol ; 93(3): e12996, 2021 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-33205443

RESUMO

Intestinal T cells form a central part of the front-line defence against foreign organisms and need to be situated in the mucosa where infection occurs. It is well accepted that immunization by a mucosal route favours localization of antigen-specific effector T cells in the mucosal epithelium, while systemic immunization does not. The aim of the study is to determine how homing receptors are specifically involved in retaining effector T cells in the small intestine after oral immunization. We here demonstrate that the chemokine receptor CXCR6, integrins ß7 and CD29 contribute differentially to the epithelial retention phenotype of CD8+ T cells in the small intestine of mice. CD8+ intraepithelial lymphocytes (IELs) of unvaccinated mice are predominantly ß7 single positives, and subcutaneous immunization-induced antigen-specific CD8+ effector IELs are mainly composed of CXCR6+ , CD29+ and CXCR6+ CD29+ cells. Strikingly, the majority of oral immunization-induced antigen-specific CD8+ effector IELs exhibit a distinct, tissue-specific CXCR6+ ß7+ double-positive phenotype, cytotoxic potential and enhanced intraepithelial localization. Transfer of antigen-specific CD8+ T cells preactivated with certain immuno-stimuli (such as monophosphoryl lipid A) results in increased accumulation of donor IELs with the CXCR6+ ß7+ phenotype. As ß7 exclusively paired with αE on IELs, our results strongly suggest that CXCR6 may cooperate with the heterodimer αEß7 to preferentially retain intestinally induced effector IELs in the epithelium. The identification of this novel IEL phenotype has significant implications for the development of vaccines and therapeutic strategies to enhance gut immunity.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Cadeias beta de Integrinas/metabolismo , Intestino Delgado/imunologia , Linfócitos Intraepiteliais/imunologia , Receptores CXCR6/metabolismo , Transferência Adotiva , Animais , Linfócitos T CD8-Positivos/transplante , Integrina beta1/metabolismo , Mucosa Intestinal/citologia , Mucosa Intestinal/imunologia , Intestino Delgado/citologia , Linfócitos Intraepiteliais/transplante , Listeria monocytogenes/imunologia , Ativação Linfocitária/imunologia , Subpopulações de Linfócitos/imunologia , Subpopulações de Linfócitos/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Vacinação
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