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1.
Adv Exp Med Biol ; 1197: 55-67, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31732934

RESUMO

Gingival epithelium plays a pivotal role in protecting the underlying periodontium from the microbial colonization found in the gingival sulcus. Having an appropriate phenotype displayed by gingival epithelial cells is a critical host component required for protection against bacterial invasion into gingival tissues. In the present study, gingival epithelial homeostasis associated with the CXCL-8/IL-8 chemokine response was investigated in vitro to determine the mechanisms that gingival epithelial cells utilize for sensing gram-positive and gram-negative microorganisms. The findings of this study have demonstrated, by using Fusobacterium nucleatum, a heterogeneity of gingival epithelial cell response by Toll-like receptor (TLR) 2, a lipoprotein sensor. Notably, however, lipopolysaccharide (LPS), a major virulence factor of gram-negative bacteria, is not recognized by gingival epithelial cells unless the LPS is internalized into the cells. Activation of TLR4 in gingival epithelial cells occurs in the endosome, an intracellular event that requires a vesicular acidification to turn on TLR4 signaling, indicating their stringency for fine-tuning a local LPS response. This study has identified a unique LPS sensing mechanism of the oral epithelium to overcome a periodontal infection associated with LPS derived from gram-negative microbes that arises during dysbiosis.


Assuntos
Gengiva , Lipopolissacarídeos , Periodontite , Células Epiteliais/imunologia , Células Epiteliais/microbiologia , Gengiva/citologia , Gengiva/imunologia , Gengiva/microbiologia , Humanos , Interleucina-8/imunologia , Lipopolissacarídeos/metabolismo , Periodontite/imunologia , Periodontite/microbiologia
2.
J Agric Food Chem ; 67(32): 9009-9021, 2019 Aug 14.
Artigo em Inglês | MEDLINE | ID: mdl-31319030

RESUMO

Soybean allergy is a serious health risk to humans and animals; ß-conglycinin is the primary antigenic protein in soybean. Intestinal porcine epithelial (IPEC-J2) cells were used as an in vitro physiological model of the intestinal epithelium to study the effects of different concentrations of soybean antigen protein ß-conglycinin to identify the involved signaling pathways. The cells were divided into eight groups and either untreated or treated with different concentrations of ß-conglycinin, pyrrolidine dithiocarbamate (PDTC), Nω-nitro-l-arginine methyl ester hydrochloride (l-NAME), SP600125, and SB202190 either alone or in combination. The cells were incubated with 1, 5, and 10 mg·mL-1 ß-conglycinin or 5 mg·mL-1 ß-conglycinin and 1 µmol·L-1 nuclear factor κB (NF-κB) inhibitor (PDTC), inducible nitric oxide synthase inhibitor (l-NAME), c-Jun N-terminal kinase (JNK) inhibitor (SP600125), and p38 inhibitor (SB202190) for 24 h, separately; controls were left untreated. The mRNA, protein, and phosphorylation levels of NF-κB, p38, and JNK were higher in the treated groups than in the control group. ß-Conglycinin decreased tight junction distribution, destroyed the cytoskeleton of IPEC-J2 cells, and caused cell death. After the addition of the inhibitors, ß-conglycinin-induced IPEC-J2 cell damage was significantly reduced. ß-Conglycinin caused damage to IPEC-J2 cells via the mitogen-activated protein kinase/NF-κB signaling pathway. The results of this study are crucial for exploring the mechanisms underlying allergic reactions caused by soybean antigen proteins.


Assuntos
Antígenos de Plantas/imunologia , Células Epiteliais/imunologia , Hipersensibilidade Alimentar/imunologia , Globulinas/imunologia , Proteínas Quinases Ativadas por Mitógeno/imunologia , NF-kappa B/imunologia , Proteínas de Armazenamento de Sementes/imunologia , Proteínas de Soja/imunologia , Soja/imunologia , Animais , Humanos , Mucosa Intestinal/citologia , Mucosa Intestinal/imunologia , Proteínas Quinases JNK Ativadas por Mitógeno/genética , Proteínas Quinases JNK Ativadas por Mitógeno/imunologia , Proteínas Quinases Ativadas por Mitógeno/genética , NF-kappa B/genética , Óxido Nítrico Sintase Tipo II/genética , Óxido Nítrico Sintase Tipo II/imunologia , Fosforilação , Transdução de Sinais , Suínos , Junções Íntimas/genética , Junções Íntimas/imunologia
3.
Artigo em Chinês | MEDLINE | ID: mdl-31315358

RESUMO

Objective: To explore the role of autophagy in PM2.5-induced inflammation in human nasal epithelial cells and related mechanism. Methods: Human nasal epithelial cells were exposed to different concentration of PM2.5 for different times, and the expression levels of microtubule-associated protein-1 light chain-3 Ⅱ (LC3 Ⅱ) and Beclin1 proteins were measured by Western blot. The typical autophagosome and autolysosome were observed by using transmission electron microscopy (TEM). To observe autophagic flux, mRFP-GFP-LC3 plasmid was transfected to nasal epithelial cells and the punctate staining of mRFP-GFP-LC3 were determined by confocal laser scanning microscope. The expression of inflammatory cytokines interleukin 6 (IL-6) and tumor necrosis factor-α (TNF-α) in cell culture supernatant were assessed by enzyme-linked immunosorbent assay (ELISA). To assess the role of autophagy in PM2.5-mediated inflammation, autophagy related gene Atg5 and Beclin-1 were silenced by siRNA knockdown, and inflammatory cytokines were analyzed.GraphPad Prism 6.0 was used for statistical analysis. Results: PM2.5 exposure increased the expression of LC3 Ⅱ and Beclin-1 proteins in a dose- (in PM2.5 group with concentration of 0, 15, 30, 60, 120 µg/ml, the expression of LC3 Ⅱ was 0.021±0.001(x±s), 0.037±0.002, 0.058±0.005, 0.075±0.006, 0.085±0.004, respectively, F=126.8, P<0.05; the expression of Beclin-1 was 0.002±0.000, 0.003±0.000, 0.005±0.000, 0.007±0.001, 0.008±0.001, respectively, F=137.3, P<0.05) and time-dependent manner (in PM2.5 group with exposure time of 0, 3, 6, 12, 24 h, the expression of LC3Ⅱ was 0.160±0.007, 0.222±0.003, 0.251±0.015, 0.483±0.029, 0.585±0.035, respectively, F=215.3, P<0.05; the expression of Beclin-1 was 0.059±0.002, 0.080±0.002, 0.087±0.002, 0.183±0.007, 0.228±0.005, respectively, F=137.3, P<0.05) in human nasal epithelial cells. TEM analysis showed typical autophagosome and autolysosome in cells after PM2.5 exposure for 24 h. PM2.5 significantly increased the number of yellow and red dots representing autophagosomes and autolysosomes respectively, indicating autophagic flux was elevated. Moreover, PM2.5 enhanced the secretion of inflammatory cytokines such as IL-6 and TNF-α, which was dramatically prevented by Atg5-siRNA and Beclin-1-siRNA. Conclusion: Autophagy plays an important role in PM2.5-caused inflammation response in nasal epithelial cells, which can induce release of inflammatory factors such as IL-6 and TNF-α and advance the inflammatory reaction.


Assuntos
Autofagia/imunologia , Células Epiteliais/imunologia , Inflamação/imunologia , Mucosa Nasal/imunologia , Material Particulado/imunologia , Proteína Beclina-1/biossíntese , Humanos , Interleucina-6/biossíntese , Proteínas Associadas aos Microtúbulos/biossíntese , Material Particulado/efeitos adversos , Fator de Necrose Tumoral alfa/biossíntese
4.
Nat Commun ; 10(1): 3060, 2019 07 16.
Artigo em Inglês | MEDLINE | ID: mdl-31311921

RESUMO

Control of Streptococcus pneumoniae colonisation at human mucosal surfaces is critical to reducing the burden of pneumonia and invasive pneumococcal disease, interrupting transmission, and achieving herd protection. Here, we use an experimental human pneumococcal carriage model (EHPC) to show that S. pneumoniae colonisation is associated with epithelial surface adherence, micro-colony formation and invasion, without overt disease. Interactions between different strains and the epithelium shaped the host transcriptomic response in vitro. Using epithelial modules from a human epithelial cell model that recapitulates our in vivo findings, comprising of innate signalling and regulatory pathways, inflammatory mediators, cellular metabolism and stress response genes, we find that inflammation in the EHPC model is most prominent around the time of bacterial clearance. Our results indicate that, rather than being confined to the epithelial surface and the overlying mucus layer, the pneumococcus undergoes micro-invasion of the epithelium that enhances inflammatory and innate immune responses associated with clearance.


Assuntos
Portador Sadio/imunologia , Nasofaringe/imunologia , Infecções Pneumocócicas/imunologia , Mucosa Respiratória/imunologia , Streptococcus pneumoniae/imunologia , Adulto , Portador Sadio/microbiologia , Células Epiteliais/imunologia , Células Epiteliais/microbiologia , Feminino , Voluntários Saudáveis , Humanos , Imunidade Inata , Masculino , Pessoa de Meia-Idade , Nasofaringe/microbiologia , Infecções Pneumocócicas/microbiologia , Mucosa Respiratória/microbiologia , Streptococcus pneumoniae/patogenicidade , Adulto Jovem
5.
Korean J Parasitol ; 57(3): 217-223, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31284343

RESUMO

Acanthamoeba castellanii has ubiquitous distribution and causes primary acanthamoebic keratitis (AK). AK is a common disease in contact lens wearers and results in permanent visual impairment or blindness. In this study, we observed the cytopathic effect, in vitro cytotoxicity, and secretion pattern of cytokines in human corneal epithelial cells (HCECs) induced by A. castellanii trophozoites and/or cysts. Morphological observation revealed that panked dendritic HCECs co-cultured with amoeba cysts had changed into round shape and gradually died. Such changes were more severe in co-culture with cyst than those of co-cultivation with trophozoites. In vitro cytotoxicity assay revealed the highest cytotoxicity to HCECs in the co-culture system with amoeba cysts. A. castellanii induced the expression of IL-1α, IL-6, IL-8, and CXCL1 in HCECs. Secreted levels of IL-1α, IL-6, and IL-8 in HCECs co-cultured with both trophozoites and cysts were increased at an early incubation time (3 and 6 hr). These results suggested that cytopathic changes and pro-inflammatory cytokines release of HCECs in response to A. castellanii, especially amoebic cysts, are an important mechanism for AK development.


Assuntos
Ceratite por Acanthamoeba/imunologia , Acanthamoeba castellanii/fisiologia , Córnea/citologia , Células Epiteliais/imunologia , Trofozoítos/fisiologia , Ceratite por Acanthamoeba/parasitologia , Acanthamoeba castellanii/crescimento & desenvolvimento , Células Cultivadas , Córnea/imunologia , Córnea/parasitologia , Células Epiteliais/parasitologia , Humanos , Interleucina-1/genética , Interleucina-1/imunologia , Interleucina-6/genética , Interleucina-6/imunologia , Interleucina-8/genética , Interleucina-8/imunologia , Trofozoítos/crescimento & desenvolvimento
6.
PLoS Negl Trop Dis ; 13(6): e0007537, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-31251739

RESUMO

Infection by Zika virus (ZIKV) is linked to microcephaly and other neurological disorders, posing a significant health threat. Innate immunity is the first line of defense against invading pathogens, but relatively little is understood regarding host intrinsic mechanisms that guard against ZIKV. Here, we show that host tripartite motif-containing protein 56 (TRIM56) poses a barrier to ZIKV infection in cells of neural, epithelial and fibroblast origins. Overexpression of TRIM56, but not an E3 ligase-dead mutant or one lacking a short C-terminal portion, inhibited ZIKV RNA replication. Conversely, depletion of TRIM56 increased viral RNA levels. Although the C-terminal region of TRIM56 bears sequence homology to NHL repeat of TRIM-NHL proteins that regulate miRNA activity, knockout of Dicer, which abolishes production of miRNAs, had no demonstrable effect on ZIKV restriction imposed by TRIM56. Rather, we found that TRIM56 is an RNA-binding protein that associates with ZIKV RNA in infected cells. Moreover, a recombinant TRIM56 fragment comprising the C-terminal 392 residues captured ZIKV RNA in cell-free reactions, indicative of direct interaction. Remarkably, deletion of a short C-terminal tail portion abrogated the TRIM56-ZIKV RNA interaction, concomitant with a loss in antiviral activity. Altogether, our study reveals TRIM56 is an RNA binding protein that acts as a ZIKV restriction factor and provides new insights into the antiviral mechanism by which this E3 ligase tackles flavivirus infections.


Assuntos
Fatores Imunológicos/metabolismo , MicroRNAs/metabolismo , RNA Viral/metabolismo , Proteínas de Ligação a RNA/metabolismo , Proteínas com Motivo Tripartido/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Zika virus/imunologia , Células Epiteliais/imunologia , Células Epiteliais/virologia , Fibroblastos/imunologia , Fibroblastos/virologia , Humanos , Neurônios/imunologia , Neurônios/virologia , Ligação Proteica , Replicação Viral
7.
Vet Res ; 50(1): 49, 2019 Jun 20.
Artigo em Inglês | MEDLINE | ID: mdl-31221210

RESUMO

An ethanolic extract from Rhodomyrtus tomentosa leaves (RTL) was studied as a natural alternative to control Staphylococcus aureus, which is an important pathogen responsible for bovine mastitis. The minimal inhibitory concentrations (MICs) of the RTL extract and of rhodomyrtone, a pure compound isolated from the plant, were determined by a microdilution method. Rhodomyrtone and the RTL extract exhibited antibacterial activity against S. aureus, including its persistent phenotype (SCV: small-colony variant) and a biofilm hyperproducer strain, with MICs of 0.25-0.5 and 8-16 µg/mL, respectively. Time-kill kinetics showed a strong bactericidal activity for both the RTL extract- and rhodomyrtone-treated bacteria at 2 × MIC as early as 4 h post-exposure. An additive effect of the extract at 0.5 × MIC was observed in a combination with oxytetracycline or pirlimycin against S. aureus by showing a 64- to 128-fold reduction in antibiotic MICs. Moreover, the RTL extract significantly decreased the number of intracellular SCVs inside bovine mammary epithelial cells. However, the extract or its combination with pirlimycin only slightly improved the activity of pirlimycin against the bacterial colonization of mouse mammary glands. In vitro MICs determined in the presence of casein indicated that the limited activity of the RTL extract in the murine model of mastitis could be linked to neutralization of active components by milk proteins. While the RTL extract showed interesting antibacterial properties in vitro, to be considered as an alternative to antibiotics in dairy farms, formulation studies are needed to cope with the observed reduction of activity in vivo.


Assuntos
Antibacterianos/farmacologia , Mastite Bovina/tratamento farmacológico , Myrtaceae/química , Extratos Vegetais/farmacologia , Infecções Estafilocócicas/veterinária , Staphylococcus aureus/efeitos dos fármacos , Animais , Bovinos , Modelos Animais de Doenças , Células Epiteliais/imunologia , Células Epiteliais/microbiologia , Feminino , Mastite Bovina/microbiologia , Camundongos , Testes de Sensibilidade Microbiana , Infecções Estafilocócicas/tratamento farmacológico , Infecções Estafilocócicas/microbiologia , Xantonas/farmacologia
8.
Nat Med ; 25(7): 1153-1163, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-31209336

RESUMO

Human lungs enable efficient gas exchange and form an interface with the environment, which depends on mucosal immunity for protection against infectious agents. Tightly controlled interactions between structural and immune cells are required to maintain lung homeostasis. Here, we use single-cell transcriptomics to chart the cellular landscape of upper and lower airways and lung parenchyma in healthy lungs, and lower airways in asthmatic lungs. We report location-dependent airway epithelial cell states and a novel subset of tissue-resident memory T cells. In the lower airways of patients with asthma, mucous cell hyperplasia is shown to stem from a novel mucous ciliated cell state, as well as goblet cell hyperplasia. We report the presence of pathogenic effector type 2 helper T cells (TH2) in asthmatic lungs and find evidence for type 2 cytokines in maintaining the altered epithelial cell states. Unbiased analysis of cell-cell interactions identifies a shift from airway structural cell communication in healthy lungs to a TH2-dominated interactome in asthmatic lungs.


Assuntos
Asma/patologia , Pulmão/citologia , Adulto , Idoso , Linfócitos T CD4-Positivos/fisiologia , Comunicação Celular , Células Epiteliais/imunologia , Células Epiteliais/fisiologia , Feminino , Estudo de Associação Genômica Ampla , Células Caliciformes/metabolismo , Humanos , Pulmão/imunologia , Pulmão/patologia , Masculino , Metaplasia , Pessoa de Meia-Idade , Células Th2/fisiologia , Transcriptoma
9.
Nat Commun ; 10(1): 2423, 2019 06 03.
Artigo em Inglês | MEDLINE | ID: mdl-31160559

RESUMO

The germinal center (GC) reaction in Peyer's patches (PP) requires continuous access to antigens, but how this is achieved is not known. Here we show that activated antigen-specific CCR6+CCR1+GL7- B cells make close contact with M cells in the subepithelial dome (SED). Using in situ photoactivation analysis of antigen-specific SED B cells, we find migration of cells towards the GC. Following antigen injection into ligated intestinal loops containing PPs, 40% of antigen-specific SED B cells bind antigen within 2 h, whereas unspecifc cells do not, indicating B cell-receptor involvment. Antigen-loading is not observed in M cell-deficient mice, but is unperturbed in mice depleted of classical dendritic cells (DC). Thus, we report a M cell-B cell antigen-specific transporting pathway in PP that is independent of DC. We propose that this antigen transporting pathway has a critical role in gut IgA responses, and should be taken into account when developing mucosal vaccines.


Assuntos
Células Apresentadoras de Antígenos/imunologia , Antígenos/imunologia , Linfócitos B/imunologia , Células Epiteliais/imunologia , Nódulos Linfáticos Agregados/imunologia , Receptores de Antígenos de Linfócitos B/imunologia , Animais , Células Dendríticas/imunologia , Centro Germinativo/imunologia , Imunoglobulina A/imunologia , Ativação Linfocitária , Camundongos
10.
Nat Commun ; 10(1): 2699, 2019 06 20.
Artigo em Inglês | MEDLINE | ID: mdl-31221976

RESUMO

Human cytomegalovirus (CMV) causes a wide array of disease to diverse populations of immune-compromised individuals. Thus, a more comprehensive understanding of how CMV enters numerous host cell types is necessary to further delineate the complex nature of CMV pathogenesis and to develop targeted therapeutics. To that end, we establish a vaccination strategy utilizing membrane vesicles derived from epithelial cells to generate a library of monoclonal antibodies (mAbs) targeting cell surface proteins in their native conformation. A high-throughput inhibition assay is employed to screen these antibodies for their ability to limit infection, and mAbs targeting CD46 are identified. In addition, a significant reduction of viral proliferation in CD46-KO epithelial cells confirms a role for CD46 function in viral dissemination. Further, we demonstrate a CD46-dependent entry pathway of virus infection in trophoblasts, but not in fibroblasts, highlighting the complexity of CMV entry and identifying CD46 as an entry factor in congenital infection.


Assuntos
Infecções por Citomegalovirus/imunologia , Citomegalovirus/imunologia , Interações Hospedeiro-Patógeno/imunologia , Proteína Cofatora de Membrana/imunologia , Internalização do Vírus , Anticorpos Monoclonais/administração & dosagem , Anticorpos Monoclonais/imunologia , Anticorpos Neutralizantes/administração & dosagem , Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/administração & dosagem , Anticorpos Antivirais/imunologia , Linhagem Celular , Infecções por Citomegalovirus/prevenção & controle , Infecções por Citomegalovirus/virologia , Células Epiteliais/imunologia , Células Epiteliais/virologia , Fibroblastos/imunologia , Fibroblastos/virologia , Técnicas de Inativação de Genes , Humanos , Proteína Cofatora de Membrana/genética , RNA Interferente Pequeno/metabolismo , Trofoblastos/imunologia , Trofoblastos/virologia , Vacinas Virais/administração & dosagem , Vacinas Virais/imunologia
11.
Gene ; 710: 114-121, 2019 Aug 20.
Artigo em Inglês | MEDLINE | ID: mdl-31153885

RESUMO

Mastitis impairs animal health and results in economic loss. Lipopolysaccharide (LPS) may cause immune response and inflammation in the bovine mammary gland. Hydrogen sulfide (H2S) is the third gasotransmitter that acts as an anti-inflammation regulator in many cells. Despite the importance of H2S in regulating inflammation, the effect and mechanism of exogenous H2S on LPS-induced inflammation in bovine mammary epithelial cells are unknown. In the present study, with NaHS as a donor of H2S, the bovine mammary epithelial cell line (MAC-T) was applied as an in vitro model to study the role of H2S on LPS-induced MAC-T cells. The results verified that the cell viability was diminished by LPS but restored by exogenous H2S at a physiologically relevant concentration (10 µM). Additionally, the production of H2S was mitigated in the LPS-induced MAC-T cells. Meanwhile, exogenous H2S decreased the intracellular ROS production and mRNA expression levels of the pro-inflammatory cytokines, TNF-α, IL-1ß, IL-8, and IL-6. Furthermore, exogenous H2S inhibited the mRNA expression of TLR4 and activation of NF-κB signaling pathway. In summary, exogenous H2S exerts anti-inflammatory effects through attenuating oxidative stress and blocking the TLR4/NF-κB pathway in the LPS-induced bovine mammary epithelial cells. Our findings might clarify new prophylactic approaches for mastitis.


Assuntos
Anti-Inflamatórios/farmacologia , Sulfeto de Hidrogênio/farmacologia , Lipopolissacarídeos/efeitos adversos , Glândulas Mamárias Animais/citologia , Transdução de Sinais/efeitos dos fármacos , Animais , Apoptose/efeitos dos fármacos , Bovinos , Linhagem Celular , Citocinas/genética , Regulação para Baixo , Células Epiteliais/citologia , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/imunologia , Feminino , Regulação da Expressão Gênica/efeitos dos fármacos , Glândulas Mamárias Animais/efeitos dos fármacos , Glândulas Mamárias Animais/imunologia , NF-kappa B/genética , NF-kappa B/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Receptor 4 Toll-Like/genética , Receptor 4 Toll-Like/metabolismo
12.
Expert Opin Investig Drugs ; 28(7): 605-616, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-31203680

RESUMO

INTRODUCTION: The salivary gland (SG) in primary Sjögren's syndrome (pSS) is characterized by its lack of function (hyposalivation) and lymphocytic invasion. Small-molecule inhibitors (SMIs) are a new class of drugs, whose diminutive size permits diffusion into cells. SMIs targeting components of the immune system are eagerly being trialed for their potential therapeutic utility in pSS. Neglected until now, however, is a discussion of the potential effects of SMIs on the SG epithelium. AREAS COVERED: We begin by reminding the reader of the SG epithelial compartment, its complicity in inflammatory milieu formation in pSS, and categories of SMIs which merit attention. We discuss each SMI category, including pre-clinical data concerning pSS and likely consequences of their application on the SG epithelium. EXPERT OPINION: Recovery of saliva production in pSS requires restoring the function of the SG epithelium, not solely on inflammation resolution. Many SMIs, for example, those blocking JAK-STAT signaling, interfere with critical epithelial cell pathways, most notably EGF signaling. If the effect of SMIs on SG epithelium is ignored, recovery of SG function will be challenging. We predict that NFκB signaling blockade will impart the least SG epithelium damage whilst reducing inflammation and facilitating recovery from hyposalivation in pSS.


Assuntos
Inflamação/tratamento farmacológico , Glândulas Salivares/efeitos dos fármacos , Síndrome de Sjogren/tratamento farmacológico , Animais , Desenho de Drogas , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/imunologia , Humanos , Inflamação/imunologia , NF-kappa B/metabolismo , Saliva/metabolismo , Glândulas Salivares/imunologia , Transdução de Sinais/efeitos dos fármacos , Síndrome de Sjogren/imunologia
13.
Hum Cell ; 32(3): 306-315, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31104300

RESUMO

Allergic rhinitis (AR) is a common hypersensitive disease that troubles patients a lot. Nasal epithelial cells (NECs), as the outmost protection of inhalation, play an important role in AR allergic response. Adrenoceptor beta 2 (ADRB2) is an important gene in inflammatory response, which has become the hot spot for AR development and treatment in recent years. MiR-15a-5p has been proved to be involved in AR immune response as the upstream regulator of ADRB2. Human primary NECs were isolated and stimulated by IL-13. qRT-PCR assay was used to detect the RNA level of target genes. ELISA and Western blotting were applied to detect target protein levels. Luciferase reporter assay and biotin pull-down assay were performed to test molecules interaction. ADRB2 was highly expressed in nasal mucosa of AR patients and was positively correlated with IL-13 stimulation, and knockdown of ADRB2 inhibited IL-13-induced expression of GM-CSF, eotaxin, and MUC5AC in NECs. ADRB2 was directly targeted by miR-15a-5p, and miR-15a-5p inhibited IL-13-induced expression of GM-CSF, eotaxin, and MUC5AC in NECs. ADRB2 mediated the effect of miR-15a-5p on the regulation of nasal epithelial immune responses. ADRB2 is negatively regulated by miR-15a-5p, which inhibits IL-13-induced nasal epithelial inflammatory responses.


Assuntos
Células Epiteliais/imunologia , Interleucina-13/imunologia , MicroRNAs/imunologia , Mucosa Nasal/citologia , Receptores Adrenérgicos beta 2/fisiologia , Rinite Alérgica/genética , Rinite Alérgica/imunologia , Células Cultivadas , Expressão Gênica , Regulação da Expressão Gênica , Humanos , Receptores Adrenérgicos beta 2/genética , Receptores Adrenérgicos beta 2/metabolismo
14.
Microb Pathog ; 133: 103554, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31121271

RESUMO

We have previously shown that Listeria monocytogenes, a causative agent of listeriosis, can produce membrane vesicles (MVs) during in vitro culture. The aim of this study was to investigate the ability of MVs from L. monocytogenes cultured with or without salt stress to induce cytotoxicity and pro-inflammatory responses in colon epithelial Caco-2 cells. MVs were purified from wild-type L. monocytogenes 10403S strain and an isogenic ΔsigB mutant strain. MVs from both wild-type and ΔsigB mutant strains increased viability of Caco-2 cells regardless of salt stress. Both MVs from wild-type and ΔsigB mutant strains stimulated expression of pro-inflammatory cytokine and chemokine genes in Caco-2 cells. Expression levels of pro-inflammatory cytokine genes in cells treated with MVs from bacteria cultured without salt stress were significantly higher than those in cells treated with MVs from bacteria cultured with salt stress. However, expression levels of chemokine genes in cells treated with MVs from bacteria cultured with salt stress were significantly higher than those in cells treated with MVs from bacteria cultured without salt stress. In addition, expression levels of interleukin (IL)-1ß and IL-8 genes were partially inhibited by either lysozyme-treated MVs or ethylenediaminetetraacetic acid-treated MVs compared to those after treatment with intact MVs. Our results suggest that salt stress can affect the production of L. monocytogenes MVs, thus causing different pro-inflammatory responses in host cells.


Assuntos
Proteínas de Bactérias/imunologia , Células CACO-2/imunologia , Células Epiteliais/imunologia , Listeria monocytogenes/metabolismo , Estresse Salino/fisiologia , Proteínas de Bactérias/genética , Sobrevivência Celular , Quimiocinas/genética , Quimiocinas/metabolismo , Citocinas/genética , Citocinas/metabolismo , Perfilação da Expressão Gênica , Regulação Bacteriana da Expressão Gênica , Humanos , Interleucina-1beta/genética , Interleucina-1beta/metabolismo , Interleucina-8/genética , Interleucina-8/metabolismo , Fator sigma/genética , Fator sigma/imunologia , Estresse Fisiológico/fisiologia
15.
Cell Host Microbe ; 25(5): 706-718.e7, 2019 05 08.
Artigo em Inglês | MEDLINE | ID: mdl-31053533

RESUMO

Metabolic programs and host defense are highly integrated to ensure proper immune responses during stress. Central to these responses, mTOR regulates immune functions by sensing and integrating environmental cues, yet how these systems are coordinated at the intestinal surface remains undefined. We show that the antimicrobial peptide α-defensin is functionally sustained during nutrient deprivation because of regulation of the defensin-processing enzyme MMP7 by microbiota- and host-derived factors. Unlike other antimicrobial peptides, the MMP7-α-defensin axis remains active during nutrient fluctuations, providing essential protection against enteric pathogens. Sustained Mmp7 expression requires the microbiota and is mediated by de-repression of the transcription activator Atoh1 upon attenuation of the transcriptional repressor Hes1 in intestinal epithelial cells. Hes1 levels are regulated via mTOR and controlled translationally, constituting a metabolism-translation-transcription loop. Disrupting this loop by supplying nutrients paradoxically compromises antibacterial defense. Together, these results uncover a regulatory circuit that couples host nutrient status to epithelial antimicrobial immunity.


Assuntos
Células Epiteliais/imunologia , Regulação da Expressão Gênica , Imunidade nas Mucosas , Metaloproteinase 7 da Matriz/biossíntese , Nutrientes/metabolismo , Fatores de Transcrição HES-1/metabolismo , alfa-Defensinas/biossíntese , Animais , Linhagem Celular , Células Epiteliais/efeitos dos fármacos , Mucosa Intestinal/efeitos dos fármacos , Mucosa Intestinal/imunologia , Camundongos Endogâmicos C57BL
16.
Food Funct ; 10(5): 2906-2913, 2019 May 22.
Artigo em Inglês | MEDLINE | ID: mdl-31070650

RESUMO

Dysfunction of the intestinal epithelial barrier plays an important role in the pathogenesis of several intestinal diseases, including celiac disease, inflammatory bowel disease, and irritable bowel syndrome. The present research was carried out to investigate the protective effect of total polysaccharides of adlay bran (TPA) on TNF-α-evoked epithelial barrier dysfunction in Caco-2 cells. Caco-2 cells were treated with or without TPA in the absence or presence of TNF-α, and transepithelial electrical resistance (TEER) and Phenol Red flux were assayed to evaluate the intestinal epithelial barrier function. The results indicated that TPA suppressed the TNF-α-induced release of pro-inflammatory factors. Furthermore, TPA obviously assuaged both the increased paracellular permeability and the decrease of TEER in TNF-α-challenged Caco-2 cells. Furthermore, TPA obviously assuaged TNF-α-evoked up-regulation of IL-8 and IL-6 expression, down-regulation of occludin and ZO-3 expression, and markedly suppressed the activation and protein expression of NF-κB p65. Our results indicated that TPA assuages the TNF-α-evoked dysfunction of the intestinal epithelial barrier by inhibiting the NF-κB p65-mediated inflammatory response.


Assuntos
Coix/química , Mucosa Intestinal/imunologia , Extratos Vegetais/farmacologia , Polissacarídeos/farmacologia , Fator de Necrose Tumoral alfa/imunologia , Células CACO-2 , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/imunologia , Humanos , Interleucina-6/genética , Interleucina-6/imunologia , Interleucina-8/genética , Interleucina-8/imunologia , Mucosa Intestinal/efeitos dos fármacos , Ocludina/genética , Ocludina/imunologia , Fosforilação , Fator de Transcrição RelA/genética , Fator de Transcrição RelA/imunologia , Fator de Necrose Tumoral alfa/genética
17.
Infect Immun ; 87(7)2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-31061143

RESUMO

Interleukin 1 receptor-like 1 (IL1RL1), also known as suppression of tumorigenicity 2 (ST2), is the receptor for interleukin 33 (IL-33) and has been increasingly studied in type 2 inflammation. An increase in airway IL-33/ST2 signaling in asthma has been associated with eosinophilic inflammation, but little is known about the role of ST2 in neutrophilic inflammation. Airway Mycoplasma pneumoniae and human rhinovirus (HRV) infections are linked to neutrophilic inflammation during acute exacerbations of asthma. However, whether ST2 contributes to M. pneumoniae- and HRV-mediated airway inflammation is poorly understood. The current study sought to determine the functions of ST2 during airway M. pneumoniae or HRV infection. In cultured normal human primary airway epithelial cells, ST2 overexpression (OE) increased the production of neutrophilic chemoattractant IL-8 in the absence or presence of M. pneumoniae or HRV1B infection. ST2 OE also enhanced HRV1B-induced IP-10, a chemokine involved in asthma exacerbations. In the M. pneumoniae-infected mouse model, ST2 deficiency, in contrast to sufficiency, significantly reduced the levels of neutrophils following acute (≤24 h) infection, while in the HRV1B-infected mouse model, ST2 deficiency significantly reduced the levels of proinflammatory cytokines KC, IP-10, and IL-33 in bronchoalveolar lavage (BAL) fluid. Overall, ST2 overexpression in human epithelial cells and ST2 sufficiency in mice increased the M. pneumoniae and HRV loads in cell supernatants and BAL fluid. After pathogen infection, ST2-deficient mice showed a higher level of the host defense protein lactotransferrin in BAL fluid. Our data suggest that ST2 promotes proinflammatory responses (e.g., neutrophils) to airway bacterial and viral infection and that blocking ST2 signaling may broadly attenuate airway infection and inflammation.


Assuntos
Infecções por Enterovirus/imunologia , Enterovirus/fisiologia , Proteína 1 Semelhante a Receptor de Interleucina-1/imunologia , Mycoplasma pneumoniae/fisiologia , Pneumonia por Mycoplasma/microbiologia , Sistema Respiratório/imunologia , Animais , Líquido da Lavagem Broncoalveolar/imunologia , Líquido da Lavagem Broncoalveolar/microbiologia , Líquido da Lavagem Broncoalveolar/virologia , Enterovirus/genética , Infecções por Enterovirus/genética , Infecções por Enterovirus/virologia , Células Epiteliais/imunologia , Células Epiteliais/microbiologia , Células Epiteliais/virologia , Humanos , Proteína 1 Semelhante a Receptor de Interleucina-1/genética , Interleucina-33/genética , Interleucina-33/imunologia , Interleucina-8/genética , Interleucina-8/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Mycoplasma pneumoniae/genética , Pneumonia por Mycoplasma/genética , Pneumonia por Mycoplasma/imunologia , Sistema Respiratório/microbiologia , Sistema Respiratório/virologia
18.
Mol Immunol ; 112: 140-150, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31102986

RESUMO

The prevalence of fish allergy among fish-processing workers is higher than in the general population, possibly due to sensitization via inhalation and higher exposure. However, the response of the bronchial epithelium to fish allergens has never been explored. Parvalbumins (PVs) from bony fish are major sensitizers in fish allergy, while cartilaginous fish and their PVs are considered less allergenic. Increasing evidence demonstrates that components other than proteins from the allergen source, such as low molecular weight components smaller than 3 kDa (LMC) from pollen, may act as adjuvants during allergic sensitization. We investigated the response of bronchial epithelial cells to PVs and to LMC from Atlantic cod, a bony fish, and gummy shark, a cartilaginous fish. Polarized monolayers of the bronchial epithelial cell line 16HBE14o- were stimulated apically with fish PVs and/-or the corresponding fish LMC. Barrier integrity, transport of PVs across the monolayers and release of mediators were monitored. Intact PVs from both the bony and the cartilaginous fish were rapidly internalized by the cells and transported to the basolateral side of the monolayers. The PVs did not disrupt the epithelial barrier integrity nor did they modify the release of proinflammatory cytokines. In contrast, LMC from both fish species modified the physical and immunological properties of the epithelial barrier and the responses differed between bony and cartilaginous fish. While the barrier integrity was lowered by cod LMC 24 h after cell stimulation, it was increased by up to 2.3-fold by shark LMC. Furthermore, LMC from both fish species increased basolateral and apical release of IL-6 and IL-8, while CCL2 release was increased by cod but not by shark LMC. In summary, our study demonstrated the rapid transport of PVs across the epithelium which may result in their availability to antigen presenting cells required for allergic sensitization. Moreover, different cell responses to LMC derived from bony versus cartilaginous fish were observed, which may play a role in different allergenic potentials of these two fish classes.


Assuntos
Alérgenos/imunologia , Brônquios/imunologia , Citocinas/imunologia , Células Epiteliais/imunologia , Peixes/imunologia , Hipersensibilidade Alimentar/imunologia , Inflamação/imunologia , Animais , Linhagem Celular , Quimiocina CCL2/imunologia , Humanos , Interleucina-6/imunologia , Interleucina-8/imunologia , Peso Molecular , Parvalbuminas/imunologia , Alimentos Marinhos
19.
Nat Commun ; 10(1): 2297, 2019 05 24.
Artigo em Inglês | MEDLINE | ID: mdl-31127085

RESUMO

Candida albicans is a fungal pathobiont, able to cause epithelial cell damage and immune activation. These functions have been attributed to its secreted toxin, candidalysin, though the molecular mechanisms are poorly understood. Here, we identify epidermal growth factor receptor (EGFR) as a critical component of candidalysin-triggered immune responses. We find that both C. albicans and candidalysin activate human epithelial EGFR receptors and candidalysin-deficient fungal mutants poorly induce EGFR phosphorylation during murine oropharyngeal candidiasis. Furthermore, inhibition of EGFR impairs candidalysin-triggered MAPK signalling and release of neutrophil activating chemokines in vitro, and diminishes neutrophil recruitment, causing significant mortality in an EGFR-inhibited zebrafish swimbladder model of infection. Investigation into the mechanism of EGFR activation revealed the requirement of matrix metalloproteinases (MMPs), EGFR ligands and calcium. We thus identify a PAMP-independent mechanism of immune stimulation and highlight candidalysin and EGFR signalling components as potential targets for prophylactic and therapeutic intervention of mucosal candidiasis.


Assuntos
Candida albicans/imunologia , Proteínas Fúngicas/imunologia , Interações Hospedeiro-Patógeno/imunologia , Sacos Aéreos/microbiologia , Animais , Candida albicans/genética , Candida albicans/metabolismo , Candidíase/imunologia , Candidíase/microbiologia , Linhagem Celular Tumoral , Modelos Animais de Doenças , Células Epiteliais/imunologia , Células Epiteliais/metabolismo , Células Epiteliais/microbiologia , Receptores ErbB/genética , Receptores ErbB/imunologia , Receptores ErbB/metabolismo , Feminino , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , Humanos , Sistema de Sinalização das MAP Quinases/imunologia , Metaloproteinases da Matriz/imunologia , Metaloproteinases da Matriz/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Membrana Mucosa/imunologia , Membrana Mucosa/microbiologia , Faringite/imunologia , Faringite/microbiologia , Fosforilação , Peixe-Zebra
20.
Nat Commun ; 10(1): 2220, 2019 05 17.
Artigo em Inglês | MEDLINE | ID: mdl-31101805

RESUMO

Both medullary thymic epithelial cells (mTEC) and dendritic cells (DC) present tissue-restricted antigens (TRA) to thymocytes to induce central tolerance, but the relative contributions of these antigen-presenting cell (APC) subsets remain unresolved. Here we developed a two-photon microscopy approach to observe thymocytes interacting with intact APCs presenting TRAs. We find that mTECs and DCs cooperate extensively to induce tolerance, with their relative contributions regulated by the cellular form of the TRA and the class of major histocompatibility complex (MHC) on which antigen is presented. Even when TRA expression is restricted to mTECs, DCs still present self-antigens at least as frequently as mTECs. Notably, the DC subset cDC2 efficiently acquires secreted mTEC-derived TRAs for cross-presentation on MHC-I. By directly imaging interactions between thymocytes and APCs, while monitoring intracellular signaling, this study reveals that distinct DC subsets and AIRE+ mTECs contribute substantially to presentation of diverse self-antigens for establishing central tolerance.


Assuntos
Tolerância Central/imunologia , Células Dendríticas/imunologia , Timócitos/imunologia , Timo/imunologia , Animais , Apresentação do Antígeno/imunologia , Autoantígenos/imunologia , Autoantígenos/metabolismo , Transplante de Medula Óssea , Separação Celular/métodos , Células Dendríticas/metabolismo , Células Epiteliais/imunologia , Feminino , Citometria de Fluxo/métodos , Microscopia Intravital/métodos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Microscopia de Fluorescência por Excitação Multifotônica/métodos , Linfócitos T Reguladores/imunologia , Timócitos/metabolismo , Timo/citologia , Fatores de Transcrição/imunologia , Fatores de Transcrição/metabolismo , Quimeras de Transplante/imunologia
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