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1.
Mol Pharmacol ; 97(5): 324-335, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-32173651

RESUMO

Skin serves not only as a protective barrier to microbial entry into the body but also as an immune organ. The outer layer, the epidermis, is composed predominantly of keratinocytes, which can be stimulated to produce proinflammatory mediators. Although some inflammation is useful to defend against infection, excessive or persistent inflammation can lead to the development of inflammatory skin diseases, such as psoriasis, a common skin disorder affecting approximately 2% of the US population. We have previously found that phosphatidylglycerol (PG) derived from soy can inhibit inflammation in a contact irritant ear edema mouse model. Here, we investigated the ability of soy PG to inhibit inflammatory mediator expression in response to activators of the pattern recognition receptors, toll-like receptor-2 (TLR2) and -4 (TLR4). We found that in epidermal keratinocytes, soy PG inhibited TLR2 and TLR4 activation and inflammatory mediator expression in response to a synthetic triacylated lipopeptide and lipopolysaccharide, respectively, as well as an endogenous danger-associated molecular pattern. However, at higher concentrations, soy PG alone enhanced the expression of some proinflammatory cytokines, suggesting a narrow therapeutic window for this lipid. Dioleoylphosphatidylglycerol (DOPG), but not dioleoylphosphatidylcholine, exerted a similar inhibitory effect, completely blocking keratinocyte inflammatory mediator expression induced by TLR2 and TLR4 activators as well as NFκB activation in a macrophage cell line (RAW264.7); however, DOPG was not itself proinflammatory even at high concentrations. Furthermore, DOPG had no effect on NFκB activation in response to a TLR7/8 agonist. Our results suggest that DOPG could be used to inhibit excessive skin inflammation. SIGNIFICANCE STATEMENT: Although inflammation is beneficial for clearing an infection, in some cases, the infection can be excessive and/or become chronic, thereby resulting in considerable tissue damage and pathological conditions. We show here that the phospholipid phosphatidylglycerol can inhibit the activation of toll-like receptors 2 and 4 of the innate immune system as well as the downstream inflammatory mediator expression in response to microbial component-mimicking agents in epidermal keratinocytes that form the physical barrier of the skin.


Assuntos
Mediadores da Inflamação/metabolismo , Queratinócitos/metabolismo , Padrões Moleculares Associados a Patógenos/farmacologia , Fosfatidilgliceróis/farmacologia , Receptor 2 Toll-Like/metabolismo , Receptor 4 Toll-Like/metabolismo , Animais , Calgranulina B/farmacologia , Humanos , Imidazóis/farmacologia , Lipopeptídeos/farmacologia , Lipopolissacarídeos/farmacologia , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Camundongos , NF-kappa B/metabolismo , Células RAW 264.7 , Receptores de Reconhecimento de Padrão/metabolismo , Proteínas Recombinantes/farmacologia , Soja/química
2.
Life Sci ; 250: 117543, 2020 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-32169518

RESUMO

AIMS: HMGB1 has been reported to play a crucial role in the physiological and pathophysiological responses during pregnancy. However, it is still unknown whether excessively expressed HMGB1 at the maternal-fetal interface related to Unexplained Recurrent Spontaneous Abortion (URSA). This study was designed to investigate the local capability of HMGB1 in the pathology of URSA, determined the distributions and characteristics of HMGB1, its receptors (RAGE/TLR2/TLR4) and important signaling molecule NF-κB p65 expression at the maternal-fetal interface,as well as compared the differences of HMGB1 expression between the URSA group, control group and aspirin treatment group. MATERIAL AND METHODS: H&E staining, Western blot analysis, immunofluorescence assay and immunohistochemical staining were applied to determine the effect of HMGB1 and its receptors at the maternal-fetal interface. ELISA was utilized to detect the concentration of HMGB1 in plasma. KEY FINDINGS: Our study demonstrated that the activation of the HMGB1-RAGE/TLR2/TLR4-NF-κB pathway at the maternal-fetal interface may have occurred in the URSA group. HMGB1 concentration in plasma was higher in the URSA group than the control group. Furthermore, the levels of HMGB1 of subjects with URSA could be reduced by administrating low doses of aspirin (ASPL). SIGNIFICANCE: This is the first report indicating the roles of HMGB1 at the maternal-fetal interface of URSA patients and broadening the horizons for clinical treatment of URSA. HMGB1-RAGE/TLR2/TLR4-NF-κB signaling pathway may be activated at the maternal-fetal interface in URSA and account for its pathogenesis. HMGB1 have the potential to be promising biomarkers in prevention and therapy of URSA.


Assuntos
Aborto Habitual/metabolismo , Aborto Espontâneo/metabolismo , Proteína HMGB1/metabolismo , Transdução de Sinais , Antígenos de Neoplasias/metabolismo , Aspirina/administração & dosagem , Feminino , Feto , Regulação da Expressão Gênica , Humanos , Antígenos Comuns de Leucócito/metabolismo , Troca Materno-Fetal , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Gravidez , Receptor 2 Toll-Like/metabolismo , Receptor 4 Toll-Like/metabolismo , Fator de Transcrição RelA/metabolismo , Regulação para Cima
3.
PLoS One ; 15(2): e0228958, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32069296

RESUMO

Obstructive sleep apnea (OSA) is a syndrome leading to chronic intermittent hypoxia, and the up-regulation of toll-like receptors (TLR) 2 and 6 on peripheral blood cells has been reported. We hypothesized that DNA methylation in TLR2 and TLR6 genes may play a role in the development of OSA and its excessive daytime sleepiness (EDS) phenotype. DNA methylation over 28 cytosine-phosphate-guanine (CpG) sites of the TLR2 promoter region and 3 CpG sites of the TLR6 gene body, and their protein expressions were measured by using pyrosequencing and ELISA methods in 18 heathy subjects (HS) and 58 patients with severe OSA (divided into 18 non-EDS and 40 EDS group). Patients with severe OSA had higher DNA methylation levels over five CpG sites (#1, #2, #3, #25 and #28) and lower DNA methylation levels over CpG site #18 of the TLR2 promoter region, higher DNA methylation levels over two CpG sites (#1 and #3) of the TLR6 gene body, and higher protein expressions of TLR6 than HS. The CpG site #2 of the TLR6 gene body was hypermethylated in severe OSA patients with EDS. Both DNA methylation levels over CpG site #1 of the TLR6 gene body and protein expressions of TLR6 were reduced after more than 6 months of nasal CPAP treatment in seven selected patients. Aberrant DNA methylation of the TLR2 promoter region and TLR6 gene body are associated with the consequence of severe OSA and its EDS phenotype.


Assuntos
Apneia Obstrutiva do Sono/genética , Receptor 2 Toll-Like/genética , Receptor 6 Toll-Like/genética , Adulto , Estudos de Casos e Controles , Pressão Positiva Contínua nas Vias Aéreas/métodos , Ilhas de CpG/genética , Metilação de DNA/genética , Epigênese Genética/genética , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Fenótipo , Polissonografia/métodos , Regiões Promotoras Genéticas/genética , Apneia Obstrutiva do Sono/metabolismo , Apneia Obstrutiva do Sono/fisiopatologia , Taiwan , Receptor 2 Toll-Like/metabolismo , Receptor 6 Toll-Like/metabolismo , Receptores Toll-Like/genética
4.
Chemosphere ; 249: 126200, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32086066

RESUMO

Perfluorooctanoic acid (PFOA) has been identified as a new persistent organic pollutant. This pollutant is ubiquitous in water and environments. Although PFOA is toxic to fishes, the precise immunotoxicological mechanism remains unclear. In this study, HPLC-MS analysis proved that PFOA can accumulate in the spleen of zebrafish. As comparison of 7-day and 14-day data, the cumulative content in the spleen significantly increased by 26% even in the 0.1 mg/L PFOA-treated group. Morphological observations revealed that PFOA can damage immune cells in zebrafish spleen by inducing vacuolization, lipofuscin granule production, and mitochondrial swelling. The Toll-like receptor 2 (TLR2)/myeloid differentiation factor 88 (myd88)/NF-κB (P65) pathway can mediate the mRNA expression levels of interferon (IFN) and B cell-activating factor (BAFF); immunoglobulin (Ig) secretion is further regulated. RT-PCR results indicated that the expression levels of P65 and IFN in the 1 mg/L group after PFOA exposure for 7 d increased by 4.03- and 3.28-fold, respectively, in a dose-dependent manner compared with those of the control group. The linear correlation coefficient (r2) was analyzed, and the results indicated that the Ig-mediated pathway can be affected by PFOA. For example, the r2 between IgD and P65 decreased from 0.641 (7 d) to 0.295 (14 d) after the cells were exposed to PFOA for a prolonged time; the r2 between IgD and IFN increased from 0.562 (7 d) to 0.808 (14 d). The triangle plot method strongly demonstrated that increased PFOA concentration and prolonged exposure to PFOA can inhibit Ig secretion. Therefore, immune organs, particularly the spleen, of zebrafish are vulnerable to PFOA. These results can help to improve the understanding of the possible noncarcinogenic risk mechanisms induced by PFOA.


Assuntos
Caprilatos/toxicidade , Fluorcarbonetos/toxicidade , Baço/imunologia , Poluentes Químicos da Água/toxicidade , Peixe-Zebra/imunologia , Animais , Poluentes Ambientais/metabolismo , Imunossupressão , Baço/metabolismo , Receptor 2 Toll-Like , Fator de Transcrição RelA/metabolismo , Peixe-Zebra/metabolismo
5.
Braz Oral Res ; 34: e012, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32049112

RESUMO

Lipoproteins are important bacterial immunostimulating molecules capable of inducing receptor activator of nuclear factor-κB (RANKL) and osteoclast formation in vitro and in vivo . Although these molecules are present in periodontopathogenic bacteria, their role in periodontitis is not known. In this study, we used Pam2CSK4 (PAM2), a synthetic molecule that mimics bacterial lipoprotein, to investigate the effects of lipoproteins on periodontitis in mice. C57BL/6 male mice were randomly divided into three experimental groups: 1) Negative control group: animals received vehicle injection; 2) Positive control group: animals received injection of Escherichia coli lipopolysaccharide (LPS); 3) PAM2 group: animals received PAM2 injection. All the injections were performed bilaterally every other day into the palatal mucosa between first and second molars. After twenty-four days, the animals were euthanized to assess alveolar bone volume (micro-CT), cellular and extracellular composition in the gingiva (stereometric analysis), and osteoclast numbers (TRAP staining). Treatment with either PAM2 or LPS induced gingival inflammation, as demonstrated by increased infiltration of inflammatory cells and enhanced angiogenesis, associated with a smaller number of fibroblasts and decreased extracellular matrix. Importantly, treatment not only with LPS but also with PAM2 resulted in a larger number of TRAP+ multinucleated osteoclasts and significant loss of alveolar bone. Collectively, our data demonstrate that PAM2 can induce gingival inflammation and bone loss in mice, broadening the avenues of investigation into the role of lipoproteins in the pathogenesis of periodontal disease.


Assuntos
Lipopeptídeos/farmacologia , Periodontite/etiologia , Periodontite/patologia , Receptor 2 Toll-Like/antagonistas & inibidores , Perda do Osso Alveolar/etiologia , Perda do Osso Alveolar/patologia , Processo Alveolar/efeitos dos fármacos , Processo Alveolar/patologia , Animais , Modelos Animais de Doenças , Gengiva/efeitos dos fármacos , Gengiva/patologia , Gengivite/etiologia , Gengivite/patologia , Masculino , Camundongos Endogâmicos C57BL , Osteoclastos/efeitos dos fármacos , Osteoclastos/fisiologia , Periodontite/microbiologia , Distribuição Aleatória , Fosfatase Ácida Resistente a Tartarato , Fatores de Tempo , Microtomografia por Raio-X
6.
Infect Immun ; 88(3)2020 02 20.
Artigo em Inglês | MEDLINE | ID: mdl-31907194

RESUMO

Yersinia pestis is the causative agent of bubonic, pneumonic, and septicemic plague. We demonstrate that Toll-like receptor 2-deficient (TLR2-/-) mice are resistant to septicemic infection by the KIM5 strain of Y. pestis but not to infection by the CO92 Δpgm strain. This resistance is dependent on TLR2, the route of infection, and the isoform of YopJ. Elevated bacterial burdens were found in the spleens of CO92 Δpgm-infected animals by 24 h postinfection and in the livers by 4 days. The YopJ isoform present contributed directly to cytotoxicity and inflammatory cytokine production of bone marrow-derived macrophages from TLR2-/- mice. Immune cell trafficking is altered in CO92 Δpgm infections, with an increased neutrophil infiltration to the spleen 5 days postinfection. Immune cell infiltration to the liver was greater and earlier in KIM5-infected TLR2-/- mice. The functionality of the immune cells was assessed by the ability to develop reactive oxygen and nitrogen species. Our data suggest an inhibition of granulocytes in forming these species in CO92 Δpgm-infected TLR2-/- mice. These findings suggest that resistance to KIM5 in TLR2-/- mice is dependent on early immune cell trafficking and functionality.


Assuntos
Peste/imunologia , Receptor 2 Toll-Like/deficiência , Yersinia pestis/patogenicidade , Animais , Carga Bacteriana , Proteínas de Bactérias/genética , Citocinas/metabolismo , Modelos Animais de Doenças , Feminino , Granulócitos/metabolismo , Fígado/imunologia , Macrófagos/imunologia , Macrófagos/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Neutrófilos/imunologia , Neutrófilos/metabolismo , Peste/metabolismo , Peste/microbiologia , Espécies Reativas de Nitrogênio/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Baço/imunologia , Baço/microbiologia , Receptor 2 Toll-Like/imunologia , Virulência/genética , Yersinia pestis/genética
7.
Life Sci ; 242: 117189, 2020 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-31891724

RESUMO

AIMS: Neointimal hyperplasia contributes to arterial restenosis after percutaneous transluminal coronary angioplasty or vascular surgery. Neointimal thickening after arterial injury is determined by inflammatory processes. We investigated the role of the innate immune receptor toll-like receptor 2 (TLR2) in neointima formation after arterial injury in mice. MATERIALS AND METHODS: Carotid artery injury was induced by 10% ferric chloride in C57Bl/6J wild type (WT), TLR2 deficient (B6.129-Tlr2tm1Kir/J, TLR2-/-) and WT mice treated with a TLR2 blocking antibody. 21 days after injury, carotid arteries were assessed histomorphometrically and for smooth muscle cell (SMC) content. To identify the contribution of circulating cells in mediating the effects of TLR2-deficiency, arterial injury was induced in WT/TLR2-/--chimeric mice and the paracrine modulation of bone marrow-derived cells from WT and TLR2-/- on SMC migration compared in vitro. KEY FINDINGS: TLR2-/- mice and WT mice treated with TLR2 blocking antibodies exhibited reduced neointimal thickening (23.7 ± 4.2 and 6.5 ± 3.0 vs. 43.1 ± 5.9 µm, P < 0.05 and P < 0.01), neointimal area (5491 ± 1152 and 315 ± 76.7 vs. 13,756 ± 2627 µm2, P < 0.05 and P < 0.01) and less luminal stenosis compared to WT mice (8.5 ± 1.6 and 5.0 ± 1.3 vs. 22.4 ± 2.2%, both P < 0.001n = 4-8 mice/group). The phenotypes of TLR2-/- vs. WT mice were completely reverted in WT/TLR2-/- bone marrow chimeric mice (5.9 ± 1.5 µm neointimal thickness, 874.2 ± 290.2 µm2 neointima area and 2.7 ± 0.6% luminal stenoses in WT mice transplanted with TLR2-/- bone marrow vs. 23.6 ± 5.1 µm, 3555 ± 511 µm2 and 12.0 ± 1.3% in WT mice receiving WT bone marrow, all P < 0.05, n = 6/group). Neointimal lesions of WT and WT mice transplanted with TLR2-/- bone marrow chimeric mice showed increased numbers of SMC (10.8 ± 1.4 and 12.6 ± 1.4 vs. 3.8 ± 0.9 in TLR2-/- and 3.5 ± 1.1 cells in WT mice transplanted with TLR2-/- bone marrow, all P < 0.05, n = 6). WT bone marrow cells stimulated SMC migration more than TLR2-deficient bone marrow cells (1.7 ± 0.05 vs. 1.3 ± 0.06-fold, P < 0.05, n = 7) and this effect was aggravated by TLR2 stimulation and diminished by TLR2 blockade (1.1 ± 0.03-fold after stimulation with TLR2 agonists and 0.8 ± 0.02-fold after TLR2 blockade vs. control treated cells defined as 1.0, P < 0.05, n = 7). SIGNIFICANCE: TLR2-deficiency on hematopoietic but not vessel wall resident cells augments vascular healing after arterial injury. Pharmacological blockade of TLR2 may thus be a promising therapeutic option to improve vessel patency after iatrogenic arterial injury.


Assuntos
Células da Medula Óssea/metabolismo , Receptor 2 Toll-Like/deficiência , Cicatrização , Animais , Artérias/lesões , Transplante de Medula Óssea , Lesões das Artérias Carótidas/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Músculo Liso Vascular/lesões , Músculo Liso Vascular/metabolismo , Músculo Liso Vascular/patologia , Neointima/metabolismo , Neointima/patologia , Receptor 2 Toll-Like/metabolismo
8.
Toxicol Lett ; 321: 146-154, 2020 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-31836503

RESUMO

BACKGROUND: Exposure to particulate matters (PMs) can lead to an acute exacerbation of allergic airway diseases, increasing the severity of symptoms and mortality. However, little is known about the underlying molecular mechanism. This study aimed to investigate the effects of PMs on acute exacerbation of allergic airway inflammation and seek potential therapeutic targets. METHODS: Non-allergic control and ovalbumin (OVA)-allergic wide-type (WT) and Toll-like receptor 2 knockout (Tlr2-/-) mice were exposed to 100 µg of PM (diameter 5.85 µm) or saline by the oropharyngeal instillation. The responses were examined three days after exposure. In the RAW264.7 macrophage cell line, Tlr2 was knocked down by small-interfering RNA or the NF-κB inhibitor JSH-23 was used, and then the cells were stimulated with PMs for 12 h before comparison of the inflammatory responses. RESULTS: PM exposure led to increased inflammatory cell recruitment and airway intensity of PAS + staining in OVA-allergic WT mice, accompanied with an accumulation of inflammatory cells and elevated inflammatory cytokines, such as IL-6 and IL-18, in the bronchoalveolar lavage fluid (BALF). Furthermore, the protein levels of TLR2 and the NLRP3 inflammasome were elevated concomitantly with the airway inflammation post-OVA/PMs challenge. Tlr2 deficiency effectively inhibited the airway inflammation, including pulmonary inflammatory cell recruitment, mucus secretion, serum OVA-specific immunoglobulin E (IgE), and BALF inflammatory cytokine production. Additionally, the P-induced NLRP3 activation in the RAW 264.7 cell line was diminished by the knockdown of Tlr2 or JSH-23 treatment in vitro. CONCLUSION: Our results indicated that PMs exacerbate the allergic airway inflammation mediated by the TLR2/ NF-κB/NLRP3 signaling pathway. Inhibition of NF-κB seems to be a possible treatment.


Assuntos
Pulmão/efeitos dos fármacos , Macrófagos/efeitos dos fármacos , NF-kappa B/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Material Particulado/toxicidade , Hipersensibilidade Respiratória/induzido quimicamente , Receptor 2 Toll-Like/metabolismo , Alérgenos , Animais , Citocinas/metabolismo , Modelos Animais de Doenças , Progressão da Doença , Feminino , Pulmão/metabolismo , Macrófagos/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Ovalbumina , Tamanho da Partícula , Células RAW 264.7 , Hipersensibilidade Respiratória/genética , Hipersensibilidade Respiratória/metabolismo , Transdução de Sinais , Receptor 2 Toll-Like/deficiência , Receptor 2 Toll-Like/genética
9.
J Agric Food Chem ; 68(2): 512-522, 2020 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-31870150

RESUMO

Host defense peptides (HDPs) are vital mucosal defense effectors of the innate immune response. The expression of HDPs is inducible in epithelial cells by potent endogenous inducers. Herein, our results demonstrate that sodium butyrate (NaB) induces the expression of porcine ß-defensin-3 (pBD3) and porcine epididymis protein 2 splicing variant C (pEP2C) in a dose- and time-dependent manner, without modifying the production of proinflammatory cytokines, in porcine intestinal epithelial cells (IPEC J2). Moreover, NaB promotes toll-like receptor 2 (TLR2) expression. TLR2 silencing inhibits the pBD3 and pEP2C expression induced by NaB but does not abolish the histone deacetylase (HDAC) inhibitory activity of NaB. We found that NaB activated the nuclear factor-κB (NF-κB) pathway. Importantly, the degree of cell confluence governs the regulatory responses but does not affect the HDAC activity of NaB. Furthermore, epidermal growth factor receptor (EGFR), but not the mitogen-activated protein kinase (MAPK) pathway, is vital during the NaB-induced pBD3 and pEP2C regulation process. We also demonstrated that pBD3 overexpression increases interleukin-18 levels. This study showed that NaB simultaneously induces pBD3 and pEP2C via TLR2 and EGFR in IPEC J2 cells without increasing the risk of a harmful inflammatory response.


Assuntos
Ácido Butírico/farmacologia , Receptores ErbB/metabolismo , Inibidores de Histona Desacetilases/farmacologia , Histona Desacetilases/metabolismo , Receptor 2 Toll-Like/metabolismo , beta-Defensinas/metabolismo , Animais , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/metabolismo , Receptores ErbB/genética , Regulação da Expressão Gênica/efeitos dos fármacos , Histona Desacetilases/genética , Interleucina-18/genética , Interleucina-18/metabolismo , NF-kappa B/genética , NF-kappa B/metabolismo , Suínos , Receptor 2 Toll-Like/genética , beta-Defensinas/genética
10.
Autoimmun Rev ; 19(1): 102430, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31734400

RESUMO

The pathophysiology of autoimmune diseases such as Multiple Sclerosis (MS) involves a complex interaction between genetic and environmental factors. Studies of monozygotic twins suggest a significant role for environmental factors in susceptibility to MS. Numerous studies, driven by the "Hygiene Hypothesis," have focused on the role of environmental factors in allergic and autoimmune diseases. The hygiene hypothesis postulates that individuals living in environments that are too "clean" lack the requisite exposure to "immune-tolerizing" microbial products, resulting in poorly regulated immune systems and increased immune-mediated diseases. Interestingly, few studies have linked MS with the hygiene hypothesis. Similarly, although numerous studies have examined the role of the microbiome in autoimmune diseases, there has been no consistent documentation of disease-specific alterations in the MS microbiome. In this review, we present evidence that integrating the hygiene hypothesis and the microbiome allows for the identification of novel pathophysiologic mechanisms in MS. Our central hypothesis is that the microbiome in MS represents a "defective environment" that fails to provide normal levels of "TLR2-tolerizing" bacterial products to the systemic immune system. Consistent with the hygiene hypothesis, we posit that this defective microbiome function results in abnormally regulated systemic innate immune TLR2 responses that play a critical role in both the inflammatory and defective remyelinative aspects of MS. We have completed proof of concept studies that support the inflammatory, remyelinating, and human immune response components of this paradigm. Our studies suggest that induction of TLR2 tolerance may represent a novel approach to treating MS, inhibiting autoimmune inflammation while simultaneously facilitating remyelination.


Assuntos
Hipótese da Higiene , Microbiota , Esclerose Múltipla/imunologia , Esclerose Múltipla/microbiologia , Receptor 2 Toll-Like/imunologia , Humanos
11.
J Immunol Res ; 2019: 1790908, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31886295

RESUMO

The glycoprotein gp43 is the major antigenic/diagnostic component of Paracoccidioides brasiliensis, one of the etiologic agents of paracoccidioidomycosis (PCM). Gp43 has protective roles in mice, but due to adhesive properties, this glycoprotein has also been associated with immune evasion mechanisms. The present study evaluated gp43 interaction in vitro with Toll-like receptors 2 and 4 (TLR2 and TLR4) present in polymorphonuclear neutrophils (PMNs) from healthy human individuals and the consequent modulation of the immune response through the expression and release of cytokines and eicosanoids. PMNs were incubated in the absence or presence of monoclonal antibodies anti-TLR2 and anti-TLR4 (individually or in combination) before gp43 stimulation. Then, PMNs were analyzed for the expression of both surface receptors and the detection of intracytoplasmic IL-17A and IL-4 using flow cytometry, while the production of PGE2, LTB4, IL-6, IL-10, IL-12, IFN-γ, and TNF-α was evaluated in the supernatants by enzyme-linked immunosorbent assay (ELISA). Our results showed that gp43 increased TLR2 and TLR4 expression by PMNs and induced PGE2 and IL-17A via TLR4 and TLR2, respectively. Thus, our data suggest that gp43 from P. brasiliensis might modulate host susceptibility to the fungal infection by affecting PGE2 and IL-17A production.


Assuntos
Antígenos de Fungos/imunologia , Dinoprostona/metabolismo , Proteínas Fúngicas/imunologia , Interleucina-17/metabolismo , Neutrófilos/imunologia , Neutrófilos/metabolismo , Paracoccidioides/imunologia , Paracoccidioidomicose/imunologia , Paracoccidioidomicose/metabolismo , Adulto , Biomarcadores , Citocinas/biossíntese , Feminino , Expressão Gênica , Humanos , Mediadores da Inflamação/metabolismo , Masculino , Paracoccidioidomicose/genética , Paracoccidioidomicose/microbiologia , Receptor 2 Toll-Like/genética , Receptor 2 Toll-Like/metabolismo , Receptor 4 Toll-Like/genética , Receptor 4 Toll-Like/metabolismo , Adulto Jovem
12.
BMC Immunol ; 20(1): 48, 2019 12 16.
Artigo em Inglês | MEDLINE | ID: mdl-31842739

RESUMO

BACKGROUND: Yersinia pestis, the etiological pathogen of plague, is capable of repressing the immune response of white blood cells to evade phagocytosis. The V-antigen (LcrV) was found to be involved in this process by binding to human Toll-like Receptor 2 (TLR2). The detailed mechanism behind this LcrV and TLR2 mediated immune response repression, however, is yet to be fully elucidated due to the lack of structural information. RESULTS: In this work, with protein structure modelling, we were able to construct a structure model of the heterotetramer of Y. pestis LcrV and human TLR2. Molecular dynamics simulation suggests the stability of this structure in aquatic environment. The LcrV model has a dumbbell-like structure with two globule domains (G1 at N-terminus and G2 away from membrane) connected with a coiled-coil linker (CCL) domain. The two horseshoe-shape TLR2 subunits form a V-shape structure, are not in direct contact with each other, and are held together by the LcrV homodimer. In this structure model, both the G1 and CCL domains are involved in the formation of LcrV homodimer, while all three domains are involved in LcrV-TLR2 binding. A mechanistic model was proposed based on this heterotetrameric structure model: The LcrV homodimer separates the TLR2 subunits to inhibit the dimerization of TLR2 and subsequent signal transfer for immune response; while LcrV could also inhibit the formation of heterodimers of TLR2 with other TLRs, and leads to immune response repression. CONCLUSIONS: A heterotetrameric structure of Y. pestis LcrV and human TLR2 was modelled in this work. Analysis of this modelled structure showed its stability in aquatic environments and the role of LcrV domains and residues in protein-protein interaction. A mechanistic model for the role of LcrV in Y. pestis pathogenesis is raised based on this heterotetrameric structure model. This work provides a hypothesis of LcrV function, with which further experimental validation may elucidate the role of LcrV in human immune response repression.


Assuntos
Antígenos de Bactérias/química , Antígenos de Bactérias/imunologia , Antígenos de Bactérias/metabolismo , Complexos Multiproteicos/química , Proteínas Citotóxicas Formadoras de Poros/química , Proteínas Citotóxicas Formadoras de Poros/imunologia , Proteínas Citotóxicas Formadoras de Poros/metabolismo , Receptor 2 Toll-Like/química , Receptor 2 Toll-Like/metabolismo , Domínio Catalítico , Interações Hospedeiro-Patógeno/imunologia , Humanos , Ligação de Hidrogênio , Modelos Moleculares , Conformação Molecular , Complexos Multiproteicos/imunologia , Complexos Multiproteicos/metabolismo , Ligação Proteica , Conformação Proteica , Multimerização Proteica , Relação Estrutura-Atividade
13.
Curr Med Sci ; 39(6): 906-912, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31845221

RESUMO

This study aimed to assess whether genetic variants of dendritic cell-associated C-type lectine-1 (Dectin-1), Toll-like receptor 2 (TLR2), Toll-like receptor 4 (TLR4), and myeloid differentiation primary response 88 (MyD88) influence the susceptibility to pulmonary invasive fungal disease (IFD) in patients with acute myeloid leukemia (AML) from a Chinese Han population. Eight single nucleotide polymorphisms (SNPs) of Dectin-1 (rs16910526, rs3901533, and rs7309123), TLR2 (rs5743708), TLR4 (rs4986790 and rs4986791) and MyD88 (rs4988453 and rs4988457) in the genomic DNA of 172 adult AML patients were genotyped. Pulmonary IFD was diagnosed as proven or probable according to the 2008 European Organization for Research and Treatment of Cancer/Invasive Fungal Infections Cooperative Group and the National Institute of Allergy and Infectious Diseases Mycoses Study Group (EORTC/MSG) consensus guidelines. SNPs that were significant in the univariate analysis were further analyzed using the multiple logistic regression analysis to determine their association with the occurrence of pulmonary IFD. The mRNA expression of Dectin-1 was detected according to the genotype by quantitative realtime PCR (qRT-PCR), and the correlation of this expression with the occurrence of pulmonary IFD in AML patients was analyzed. Two Dectin-1 intron SNPs (rs3901533 and rs7309123) were found to be significantly associated with the susceptibility to pulmonary IFD in AML patients in a Chinese Han population. Significant associations were noted between pulmonary IFD and Dectin-1 rs3901533 dominant model (G/T+G/G vs. T/T, OR: 2.158; 95% CI: 1.109-4.2, P=0.02), Dectin-1 rs3901533 G allele (OR: 2.201; 95% CI: 1.206-4.019, P=0.01), or Dectin-1 rs7309123 C allele (OR: 1.919; 95% CI: 1.047-3.518, P=0.03). There were no significant associations between pulmonary IFD and the remaining Dectin-1 SNPs (rs16910526), TLR2 (rs5743708), TLR4 (rs4986790 and rs4986791) or MyD88 (rs4988453 and rs4988457). In conclusion, two Dectin-1 SNPs (rs3901533 and rs7309123) are associated with increased susceptibility to pulmonary IFD in AML patients in a Chinese Han population.


Assuntos
Grupo com Ancestrais do Continente Asiático/etnologia , Infecções Fúngicas Invasivas/genética , Lectinas Tipo C/genética , Leucemia Mieloide Aguda/microbiologia , Pneumopatias Fúngicas/genética , Grupo com Ancestrais do Continente Asiático/genética , China/etnologia , Feminino , Regulação da Expressão Gênica , Predisposição Genética para Doença , Humanos , Infecções Fúngicas Invasivas/etnologia , Leucemia Mieloide Aguda/etnologia , Leucemia Mieloide Aguda/genética , Pneumopatias Fúngicas/etnologia , Masculino , Fator 88 de Diferenciação Mieloide/genética , Polimorfismo de Nucleotídeo Único , Receptor 2 Toll-Like/genética , Receptor 4 Toll-Like/genética
14.
BMC Complement Altern Med ; 19(1): 326, 2019 Nov 21.
Artigo em Inglês | MEDLINE | ID: mdl-31752797

RESUMO

BACKGROUND: It has been testified that Diabetes mellitus (DM) has a close association with chronic inflammation and Toll-like Receptors (TLRs), and DM could be prevented by mulberry leaf. Therefore, a hypothesis came into being that mulberry leaf could ameliorate proinflammation and insulin resistance (IR) through TLRs and insulin signalling pathways. METHODS: Water extracts of mulberry leaf (WEM) was given to diabetic mice by gavage for 10 weeks, and the diabetic mice was injected with low-dose streptozocin, fed with high-fat and high-sugar diet. Oral glucose tolerance tests (OGTTs) were conducted. At the same time, homeostasis model assessment of insulin (HOMA-IR) and the level of the inflammatory factor, tumour necrosis factor-α (TNF-α) was measured. The expressions of critical nodes of TLRs and insulin signalling pathway were also examined. RESULTS: WEM contributed to a significant decrease in fasting blood glucose, AUC from the investigation of OGTTs and HOMA-IR. The levels of the inflammatory factor, tumour necrosis factor-α (TNF-α) also declined. Moreover, WEM suppressed the expression of TLR2, myeloid differentiation primary-response protein 88 (MyD88), tumour-necrosis-factor receptor-associated factor 6 (TRAF6), nuclear factor kappa B (NF-κB) in the skeletal muscle. WEM could up-regulate the expression of insulin receptor (InsR) and insulin receptor substrate 1 (IRS1), and down-regulate the phosphorylation of IRS1 in adipose tissue. CONCLUSION: Through this study, a conclusion could be made that WEM mitigates hyperglycemia, IR, and inflammation through the interactions among TLR2 signalling pathway, insulin signalling pathway and TNF-α.


Assuntos
Diabetes Mellitus Experimental/metabolismo , Inflamação/metabolismo , Resistência à Insulina/fisiologia , Morus , Extratos Vegetais/farmacologia , Animais , Glicemia/efeitos dos fármacos , Peso Corporal/efeitos dos fármacos , Diabetes Mellitus Tipo 2/metabolismo , Insulina/sangue , Insulina/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos ICR , Folhas de Planta/química , Transdução de Sinais/efeitos dos fármacos , Receptor 2 Toll-Like/sangue , Receptor 2 Toll-Like/metabolismo , Fator de Necrose Tumoral alfa/sangue , Fator de Necrose Tumoral alfa/metabolismo
15.
PLoS Negl Trop Dis ; 13(11): e0007789, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31675378

RESUMO

Leptospirosis, caused by pathogenic Leptospira species, has emerged as an important neglected zoonotic disease. Few studies have reported the preventable effects of immunoregulators, except for antibiotics, against leptospirosis. Generally, immunostimulatory agents are considered effective for enhancing innate immune responses. Many studies have found that beta-glucan (ß-glucan) could be a potent and valuable immunostimulant for improving immune responses and controlling diseases. In this study, we investigated the preventable role of ß-glucan against Leptospira infection in hamsters. First, ß-glucan was administered 24 h prior to, during and after infection. The results showed that ß-glucan increased the survival rate to 100%, alleviated tissue injury, and decreased leptospire loads in target organs. Additionally, we found using quantitative real-time PCR that application of ß-glucan significantly enhanced the expression of Toll-like receptor (TLR) 2, interleukin (IL)-1ß and iNOS at 2 dpi (days post infection) and reduced the increase of TLR2, IL-1ß and iNOS induced by Leptospira at 5 dpi. Furthermore, to induce memory immunity, ß-glucan was administered 5 days prior to infection. ß-Glucan also significantly increased the survival rates and ameliorated pathological damage to organs. Moreover, we demonstrated that ß-glucan-trained macrophages exhibited elevated expression of proinflammatory cytokines (IL-1ß and IL-6) in vitro, indicating that ß-glucan induces an enhanced inflammatory response against Leptospira infection. These results indicate that administration of ß-glucan and other immunostimulants could be potential valuable options for the control of Leptospira infection.


Assuntos
Adjuvantes Imunológicos/uso terapêutico , Leptospirose/imunologia , Leptospirose/prevenção & controle , beta-Glucanas/uso terapêutico , Adjuvantes Imunológicos/administração & dosagem , Animais , Cricetinae , Citocinas/metabolismo , Modelos Animais de Doenças , Imunidade Inata/efeitos dos fármacos , Interleucina-1beta/metabolismo , Leptospira/crescimento & desenvolvimento , Leptospira/imunologia , Leptospira/patogenicidade , Leptospira interrogans/crescimento & desenvolvimento , Leptospira interrogans/imunologia , Leptospirose/patologia , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Óxido Nítrico Sintase Tipo II/metabolismo , Receptor 2 Toll-Like/metabolismo , beta-Glucanas/administração & dosagem
16.
BMC Res Notes ; 12(1): 648, 2019 Oct 07.
Artigo em Inglês | MEDLINE | ID: mdl-31590685

RESUMO

OBJECTIVES: Mycobacterium indicus pranii (MIP) is an atypical mycobacterium species with potent antitumor efficacy. Macrophages and dendritic cells (DCs) are antigen-presenting cells, playing key roles in the activation of antitumor immunity. We have previously shown the potent activation of macrophages and DCs by MIP, which is mediated by MyD88-TLR2 signaling axis. In the present study, we further examined the role of MyD88 and TLR2 in MIP-mediated tumor regression. RESULTS: Wild-type and MyD88-/- mice were implanted with B16F10 tumor cells, treated with MIP or phosphate-buffered saline (PBS) and monitored for tumor growth. As expected, MIP therapy led to significant tumor regression in wild-type mice. However, antitumor efficacy of MIP was lost in MyD88-/- animals. Both PBS-treated (control) and MIP-treated MyD88-/- mice developed tumors with comparable volume. Since MyD88 relays TLR engagement signals, we analyzed the antitumor efficacy of MIP in TLR2-/- and TLR4-/- mice. It was observed that MIP therapy reduced tumor burden in wild-type and TLR4-/- mice but not in TLR2-/- mice. Tumor volume in MIP-treated TLR2-/- mice were comparable with those in PBS-treated wild-type animals. These results implicated the MyD88-TLR2 signaling axis in the antitumor efficacy of MIP.


Assuntos
Melanoma Experimental/imunologia , Fator 88 de Diferenciação Mieloide/imunologia , Micobactérias não Tuberculosas/imunologia , Receptor 2 Toll-Like/imunologia , Carga Tumoral/imunologia , Animais , Linhagem Celular Tumoral , Células Dendríticas/imunologia , Células Dendríticas/metabolismo , Humanos , Macrófagos/imunologia , Macrófagos/metabolismo , Melanoma Experimental/genética , Melanoma Experimental/microbiologia , Melanoma Experimental/terapia , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fator 88 de Diferenciação Mieloide/genética , Fator 88 de Diferenciação Mieloide/metabolismo , Micobactérias não Tuberculosas/fisiologia , Transdução de Sinais/genética , Transdução de Sinais/imunologia , Receptor 2 Toll-Like/genética , Receptor 2 Toll-Like/metabolismo , Receptor 4 Toll-Like/genética , Receptor 4 Toll-Like/imunologia , Receptor 4 Toll-Like/metabolismo , Carga Tumoral/genética
17.
Molecules ; 24(19)2019 Sep 27.
Artigo em Inglês | MEDLINE | ID: mdl-31569697

RESUMO

Cancer vaccine is a promising immunotherapeutic approach to train the immune system with vaccines to recognize and eliminate tumors. Adjuvants are compounds that are necessary in cancer vaccines to mimic an infection process and amplify immune responses. The Toll-like receptor 2 and 6 (TLR2/TLR6) agonist dipalmitoyl-S-glyceryl cysteine (Pam2Cys) was demonstrated as an ideal candidate for synthetic vaccine adjuvants. However, the synthesis of Pam2Cys requires expensive N-protected cysteine as a key reactant, which greatly limits its application as a synthetic vaccine adjuvant in large-scaled studies. Here, we report the development of N-acetylated Pam2Cys analogs as TLR2/TLR6 agonists. Instead of N-protected cysteine, the synthesis utilizes N-acetylcysteine to bring down the synthetic costs. The N-acetylated Pam2Cys analogs were demonstrated to activate TLR2/TLR6 in vitro. Moreover, molecular docking studies were performed to provide insights into the molecular mechanism of how N-acetylated Pam2Cys analogs bind to TLR2/TLR6. Together, these results suggest N-acetylated Pam2Cys analogs as inexpensive and promising synthetic vaccine adjuvants to accelerate the development of cancer vaccines in the future.


Assuntos
Lipopeptídeos/química , Lipopeptídeos/farmacologia , Receptor 2 Toll-Like/agonistas , Receptor 2 Toll-Like/química , Receptor 6 Toll-Like/agonistas , Receptor 6 Toll-Like/química , Humanos , Lipopeptídeos/síntese química , Conformação Molecular , Simulação de Acoplamento Molecular , Simulação de Dinâmica Molecular , Estrutura Molecular
18.
Carbohydr Polym ; 226: 115295, 2019 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-31582086

RESUMO

Tumor-associated macrophages (TAMs) with an M2-like phenotype have been linked to the proliferation, invasion and metastasis of tumor cells. Resetting tumor-associated macrophages represents an attractive target for an effective cancer immunotherapy. WCCP-N-b, a novel linear 3-O-methylated galactan, isolated from Cantharellus cibarius, can convert tumor-promoting M2-like macrophages to tumor-inhibiting M1-like phenotype. On a cellular mechanistic level, WCCP-N-b inhibited M2-like macrophages polarization through suppression of STAT6 activation. Furthermore, WCCP-N-b increased the phosphorylation of mitogen-activated protein kinases (MAPKs) and degradation of IκB-α through targeting Toll-like receptor 2 (TLR2). The activation of MAPKs and degradation of IκB-α were responsible for converting M2-like macrophages to M1-like macrophages. Importantly, cell culture supernatants of WCCP-N-b-treated M2-like macrophages could inhibit the cell viability of B16F1 and B16F10. Our findings provide a potential natural and harmless polysaccharide for macrophage-based tumor immunotherapy.


Assuntos
Antineoplásicos/farmacologia , Galactanos/farmacologia , Ativação de Macrófagos/efeitos dos fármacos , Macrófagos/efeitos dos fármacos , Animais , Basidiomycota/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Proteínas Quinases Ativadas por Mitógeno/metabolismo , NF-kappa B/metabolismo , Células RAW 264.7 , Fator de Transcrição STAT6/metabolismo , Receptor 2 Toll-Like/metabolismo
19.
Infect Immun ; 87(12)2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31570556

RESUMO

The Porphyromonas gingivalis strain ATCC 33277 (33277) and 381 genomes are nearly identical. However, strain 33277 displays a significantly diminished capacity to stimulate host cell Toll-like receptor 2 (TLR2)-dependent signaling and interleukin-1ß (IL-1ß) production relative to 381, suggesting that there are strain-specific differences in one or more bacterial immune-modulatory factors. Genomic sequencing identified a single nucleotide polymorphism in the 33277 fimB allele (A→T), creating a premature stop codon in the 33277 fimB open reading frame relative to the 381 fimB allele. Gene exchange experiments established that the 33277 fimB allele reduces the immune-stimulatory capacity of this strain. Transcriptome comparisons revealed that multiple genes related to carboxy-terminal domain (CTD) family proteins, including the gingipains, were upregulated in 33277 relative to 381. A gingipain substrate degradation assay demonstrated that cell surface gingipain activity is higher in 33277, and an isogenic mutant strain deficient for the gingipains exhibited an increased ability to induce TLR2 signaling and IL-1ß production. Furthermore, 33277 and 381 mutant strains lacking CTD cell surface proteins were more immune-stimulatory than the parental wild-type strains, consistent with an immune-suppressive role for the gingipains. Our data show that the combination of an intact fimB allele and limited cell surface gingipain activity in P. gingivalis 381 renders this strain more immune-stimulatory. Conversely, a defective fimB allele and high-level cell surface gingipain activity reduce the capacity of P. gingivalis 33277 to stimulate host cell innate immune responses. In summary, genomic and transcriptomic comparisons identified key virulence characteristics that confer divergent host cell innate immune responses to these highly related P. gingivalis strains.


Assuntos
Proteínas de Fímbrias/genética , Proteínas de Fímbrias/imunologia , Porphyromonas gingivalis/genética , Porphyromonas gingivalis/imunologia , Infecções por Bacteroidaceae/imunologia , Infecções por Bacteroidaceae/microbiologia , Linhagem Celular Tumoral , Células HEK293 , Humanos , Imunidade Inata/genética , Imunidade Inata/imunologia , Interleucina-1beta/metabolismo , Polimorfismo de Nucleotídeo Único/genética , Transdução de Sinais/imunologia , Células THP-1 , Receptor 2 Toll-Like/metabolismo
20.
Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi ; 35(8): 689-694, 2019 Aug.
Artigo em Chinês | MEDLINE | ID: mdl-31638565

RESUMO

Objective To study the effect of deletion of Toll-like receptor 2 (TLR2) gene on insulin resistance and polarization of macrophages in mice. Methods The wild-type (WT) and TLR2 knockout (TLR2-/-) C57BL/6 male mice, aged 28 days, were selected, with 12 mice in each group. The genotype of each mouse was identified by PCR. After mice were fed with basic diet for 3 months, the glucose tolerance test (GTT) and insulin resistance test (ITT) were performed. The mononuclear cells isolated from peripheral blood were stimulated with GM-CSF/IFN-γ and M-CSF/IL-4/IL-13, respectively, to induce differentiation to M1-like and M2-like macrophages. The CD11b, F4/80, CD11c, CD206 and early growth response 2 (EGR2) were detected by flow cytometry to determine the phenotype of macrophages. The levels of TNF-α, IL-6 and IL-10 in the culture supernatant of macrophages were detected using ELISA. Results The result of PCR identification was consistent with the genotype of mice. Compared to WT mice, TLR2-/- mice exhibited the significantly improved glycemic control at 30 min during GTT and the significantly increased insulin sensitivity at 15 minutes during ITT. The flow cytometry showed that M1 markers decreased and M2 macrophages increased in the TLR2-/- mice. ELISA showed that the levels of IL-6 and TNF-α significantly decreased in the culture supernatant of M1 macrophages, while the level of IL-10 significantly increased in the culture supernatant of M2 macrophages in TLR2-/- mice compared with WT mice. Conclusion TLR2 signal has an effect on the polarization of macrophages and makes macrophages tend to switch to M1 phenotype. A higher number of pro-inflammatory factors secreted by M1 macrophages contribute to a low-grade inflammation state in the body, which leads to a decrease in glucose tolerance and insulin sensitivity.


Assuntos
Resistência à Insulina , Macrófagos , Receptor 2 Toll-Like , Animais , Resistência à Insulina/genética , Macrófagos/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Receptor 2 Toll-Like/genética
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