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1.
Mol Med Rep ; 30(3)2024 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-38963029

RESUMO

Viral infections in the respiratory tract are common, and, in recent years, severe acute respiratory syndrome coronavirus 2 outbreaks have highlighted the effect of viral infections on antiviral innate immune and inflammatory reactions. Specific treatments for numerous viral respiratory infections have not yet been established and they are mainly treated symptomatically. Therefore, understanding the details of the innate immune system underlying the airway epithelium is crucial for the development of new therapies. The present study aimed to investigate the function and expression of interferon (IFN)­stimulated gene (ISG)60 in non­cancerous bronchial epithelial BEAS­2B cells exposed to a Toll­like receptor 3 agonist. BEAS­2B cells were treated with a synthetic TLR3 ligand, polyinosinic­polycytidylic acid (poly IC). The mRNA and protein expression levels of ISG60 were analyzed using reverse transcription­quantitative PCR and western blotting, respectively. The levels of C­X­C motif chemokine ligand 10 (CXCL10) were examined using an enzyme­linked immunosorbent assay, and the effects of knockdown of IFN­ß, ISG60 and ISG56 were examined using specific small interfering RNAs. Notably, ISG60 expression was increased in proportion to poly IC concentration, and recombinant human IFN­ß also induced ISG60 expression. By contrast, knockdown of IFN­ß and ISG56 decreased ISG60 expression, and ISG60 knockdown reduced CXCL10 and ISG56 expression. These findings suggested that ISG60 is partly implicated in CXCL10 expression and that ISG60 may serve a role in the innate immune response of bronchial epithelial cells. The present study highlights ISG60 as a potential target for new therapeutic strategies against viral infections in the airway.


Assuntos
Brônquios , Quimiocina CXCL10 , Células Epiteliais , Poli I-C , Transdução de Sinais , Receptor 3 Toll-Like , Humanos , Receptor 3 Toll-Like/metabolismo , Receptor 3 Toll-Like/genética , Quimiocina CXCL10/metabolismo , Quimiocina CXCL10/genética , Células Epiteliais/metabolismo , Células Epiteliais/efeitos dos fármacos , Brônquios/citologia , Brônquios/metabolismo , Poli I-C/farmacologia , Transdução de Sinais/efeitos dos fármacos , Linhagem Celular , Imunidade Inata , Interferon beta/metabolismo , Interferon beta/genética , Regulação da Expressão Gênica/efeitos dos fármacos , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/genética , Proteínas de Ligação a RNA , Proteínas Adaptadoras de Transdução de Sinal , Proteínas Reguladoras de Apoptose
2.
Expert Rev Mol Diagn ; 24(6): 525-531, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38864429

RESUMO

BACKGROUND: A distinct phenotype in Coronavirus disease 2019 (Covid-19) was observed in severe patients, consisting of a highly impaired interferon (IFN) type I response, an exacerbated inflammatory response. OBJECTIVE: The aim of the present study was to investigate a possible association of single nucleotide polymorphisms (SNPs), in five genes related to the immune response, rs3775291 in TLR3; rs2292151 in TICAM1; rs1758566 in IFNA1; rs1800629 in TNF, and rs1800795 in IL6 with the severity of Covid-19. METHODS: A cross-sectional study was performed, with non-severe and severe/critical patients diagnosed with Covid-19, by two public hospitals in Brazil. In total, 300 patients were genotyped for the SNPs, 150 with the non-severe form of the disease and 150 with severe/critical form. RESULTS: The T/T genotype of TLR3 in recessive model shows 58% of protection against severe/critical Covid-19; as well as the genotypes G/A+A/A of TICAM1 in dominant model with 60% of protection, and in a codominant model G/A with 57% and A/A with 71% of protection against severe/critical Covid-19. Comparing severe and critical cases, the T/C genotype of IFNA1 in the codominant model and TC+C/C in the dominant model showed twice the risk of critical Covid-19. CONCLUSION: We can conclude that rs3775291, rs2292151 and rs1758566 can influence the COVID-19 severity.


Assuntos
COVID-19 , Predisposição Genética para Doença , Polimorfismo de Nucleotídeo Único , SARS-CoV-2 , Índice de Gravidade de Doença , Receptor 3 Toll-Like , Humanos , COVID-19/genética , COVID-19/virologia , Brasil/epidemiologia , Receptor 3 Toll-Like/genética , Masculino , Feminino , Pessoa de Meia-Idade , SARS-CoV-2/genética , Estudos Transversais , Adulto , Genótipo , Idoso , Interferon Tipo I/genética , Interferon-alfa
3.
BMC Infect Dis ; 24(1): 616, 2024 Jun 21.
Artigo em Inglês | MEDLINE | ID: mdl-38907187

RESUMO

BACKGROUND: Toll-Like receptors (TLRs) play an important role in the immune response during hepatitis B virus (HBV) infection. In this study, we evaluated the association between two SNP variants (TLR3 rs3775290 and TLR4 rs4986790) and susceptibility to chronic HBV infection in Mauritania. SUBJECTS AND METHODS: A total of 188 subjects were recruited for this study: 102 chronically infected patients and 86 individuals with spontaneously resolved HBV infection who were considered controls. Targeted PCR products were sequenced using Sanger sequencing. RESULTS: We found that TLR3 rs3775290 was significantly more frequent in patients with chronic HBV than in the control population (p = 0.03). However, no association was found between the TLR4 rs3775290 polymorphism and chronic infection. CONCLUSION: Our results suggest that the TLR3 rs3775290 polymorphism may be a risk factor for susceptibility to chronic HBV infection in the Mauritanian population.


Assuntos
Predisposição Genética para Doença , Hepatite B Crônica , Polimorfismo de Nucleotídeo Único , Receptor 3 Toll-Like , Humanos , Receptor 3 Toll-Like/genética , Masculino , Feminino , Estudos de Casos e Controles , Adulto , Hepatite B Crônica/genética , Hepatite B Crônica/virologia , Pessoa de Meia-Idade , Mauritânia , Adulto Jovem , Vírus da Hepatite B/genética
4.
Croat Med J ; 65(3): 232-238, 2024 Jun 13.
Artigo em Inglês | MEDLINE | ID: mdl-38868969

RESUMO

AIM: To determine variations in allele and genotype frequencies between keratoacanthoma (KA) and common warts (CW), compared with the control group, in three single nucleotide polymorphisms (SNPs) within the TLR2, TLR3, and TLR9 genes. METHODS: This case-control study involved samples from 161 patients with KA, 152 patients with CW, and 469 controls. DNA was isolated from formalin-fixed paraffin-embedded tissue sections. Three SNPs - rs4696480 in TLR2, rs7657186 in TLR9, and rs35213 in TLR3 - were genotyped with TaqMan Genotyping Assays on the 7500 Real-Time PCR System. RESULTS: TLR2 rs4696480 and TLR3 rs7657186 were significantly overrepresented in KA and CW compared with controls (P<0.001). The association was stronger for CW than for KA, as evidenced by higher frequencies of the A allele and AA genotype for rs4696480. Both KA and CW patients had higher frequencies of the G allele and GG genotype for rs7657186 than controls. rs7657186 was moderately associated with KA and CW, with the G allele and GG genotype being more prevalent in CW cases, where no AA homozygotes were found. CONCLUSION: Genetic variants in TLR2 (rs4696480) and TLR3 (rs7657186) genes may affect KA and CW development, influencing immune responses and susceptibility to these skin lesions. Further research is required to elucidate TLR expression patterns and their role in KA development.


Assuntos
Ceratoacantoma , Polimorfismo de Nucleotídeo Único , Receptor 2 Toll-Like , Receptor 3 Toll-Like , Verrugas , Adolescente , Adulto , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Adulto Jovem , Alelos , Estudos de Casos e Controles , Frequência do Gene , Predisposição Genética para Doença , Genótipo , Ceratoacantoma/genética , Receptor 2 Toll-Like/genética , Receptor 3 Toll-Like/genética , Verrugas/genética
5.
Nat Commun ; 15(1): 3969, 2024 May 10.
Artigo em Inglês | MEDLINE | ID: mdl-38730242

RESUMO

Encephalitis is a rare and potentially fatal manifestation of herpes simplex type 1 infection. Following genome-wide genetic analyses, we identified a previously uncharacterized and very rare heterozygous variant in the E3 ubiquitin ligase WWP2, in a 14-month-old girl with herpes simplex encephalitis. The p.R841H variant (NM_007014.4:c.2522G > A) impaired TLR3 mediated signaling in inducible pluripotent stem cells-derived neural precursor cells and neurons; cells bearing this mutation were also more susceptible to HSV-1 infection compared to control cells. The p.R841H variant increased TRIF ubiquitination in vitro. Antiviral immunity was rescued following the correction of p.R841H by CRISPR-Cas9 technology. Moreover, the introduction of p.R841H in wild type cells reduced such immunity, suggesting that this mutation is linked to the observed phenotypes.


Assuntos
Encefalite por Herpes Simples , Herpesvirus Humano 1 , Mutação , Ubiquitina-Proteína Ligases , Humanos , Ubiquitina-Proteína Ligases/genética , Ubiquitina-Proteína Ligases/metabolismo , Feminino , Encefalite por Herpes Simples/genética , Lactente , Herpesvirus Humano 1/genética , Células-Tronco Pluripotentes Induzidas/metabolismo , Receptor 3 Toll-Like/genética , Receptor 3 Toll-Like/metabolismo , Ubiquitinação , Neurônios/metabolismo , Células-Tronco Neurais/metabolismo , Células-Tronco Neurais/virologia , Sistemas CRISPR-Cas
6.
Immunobiology ; 229(4): 152807, 2024 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-38821752

RESUMO

The study aimed to explore the pontential impact of 10 polymorphisms within IFN-α, IFN-ß1, IFN-γ and TLR3 genes on SLE phenotype and susceptibility and to study the relationship between specific genotypes and clinics. Whole blood samples from SLE patients and healthy controls was obtained. DNA was extracted from the peripheral blood by the QIAamp DNA Blood Mini Kit (Qiagen). The quality and quantity of isolated DNA was estimated by the Quawell Q5000 spectrophotometer. We genotyped SLE patients and healthy subjects using real-time PCR (QuantStudio 5 thermocycler). The study suggests that IFN-γ rs2069705, IFN-γ rs2069718 and IFN-α rs3758236 polymorphisms have a protective role in SLE. We observed relations between TLR3 rs3775292, IFN-ß1 rs7873167, IFN-γ rs2069705, TLR3 rs3775291 and TLR3 rs5743305 polymorphisms and clinical picture of SLE patients. We found associations between the IFN-α rs3758236, IFN-γ rs2069705, IFN-γ rs2069718, IFN-γ rs1861493 and IFN-ß1 rs10964831 polymorphisms and the clinical manifestation of the SLE and/or its comorbidities. We perceived links between IFN-γ rs2069705, IFN-γ rs2069718, IFN-γ rs1861493, TLR3 rs3775291, TLR3 rs3775292 and TLR3 rs5743305 polymorphisms and the occurrence of autoantibodies. Our study presented the relationship between IFN and TLR gene polymorphisms with SLE susceptibility, phenotype and autoantibodies profile. This study propose that polymorphisms within interferons and TLR3 genes can be engaged in the SLE pathogenesis and course.


Assuntos
Predisposição Genética para Doença , Genótipo , Lúpus Eritematoso Sistêmico , Polimorfismo de Nucleotídeo Único , Receptor 3 Toll-Like , Humanos , Lúpus Eritematoso Sistêmico/genética , Receptor 3 Toll-Like/genética , Feminino , Masculino , Adulto , Pessoa de Meia-Idade , Frequência do Gene , Alelos , Estudos de Casos e Controles , Interferons/genética , Estudos de Associação Genética
7.
Int Immunopharmacol ; 134: 112182, 2024 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-38703568

RESUMO

Seipin plays a crucial role in lipid metabolism and is involved in neurological disorders. However, the function and mechanism of action of seipin in acute ischemic stroke have not yet been elucidated. Here, we aimed to investigate the effect of seipin on neuroinflammation induced by oxygen-glucose deprivation/reoxygenation (OGD/R) and further explore the molecular mechanism by functional experiments. Our results revealed a significant decrease in seipin mRNA levels, accompanied by enhanced expression of TNF-α in patients with AIS, and a significant negative correlation between seipin and TNF-α was observed. Additionally, there was a negative correlation between seipin levels and the National Institutes of Health Stroke Scale (NIHSS) score. Furthermore, seipin levels were also decreased in middle cerebral artery occlusion/reperfusion (MCAO/R) mice and OGD/R-treated BV2 cells. RNA sequencing analysis showed that seipin knockdown altered the Toll-like receptor 3 (TLR3) signaling pathway. It was further confirmed in vitro that seipin knockdown caused significantly increased secretion of inflammatory factors including TNF-α, interleukin (IL)-1ß, and interferon (IFN)-ß. Meanwhile, seipin knockdown activated the Tlr3 signal pathway while this effect could be reversed by Tlr3 inhibitor in OGD/R treated BV2 cells. Furthermore, neuroinflammation induced by OGD/R was significantly reduced by seipin overexpression. Overall, our study demonstrate that seipin deficiency aggravates neuroinflammation by activating the TLR3/TRAF3/NF-κB signaling pathway after OGD/R stimuli, and suggest that seipin may be a potential therapeutic target for AIS.


Assuntos
Glucose , NF-kappa B , Doenças Neuroinflamatórias , Oxigênio , Transdução de Sinais , Fator 3 Associado a Receptor de TNF , Receptor 3 Toll-Like , Idoso , Animais , Feminino , Humanos , Masculino , Camundongos , Pessoa de Meia-Idade , Linhagem Celular , Modelos Animais de Doenças , Glucose/metabolismo , Subunidades gama da Proteína de Ligação ao GTP/metabolismo , Subunidades gama da Proteína de Ligação ao GTP/genética , Infarto da Artéria Cerebral Média/metabolismo , Infarto da Artéria Cerebral Média/imunologia , AVC Isquêmico/metabolismo , AVC Isquêmico/imunologia , Camundongos Endogâmicos C57BL , Doenças Neuroinflamatórias/imunologia , Doenças Neuroinflamatórias/metabolismo , NF-kappa B/metabolismo , Oxigênio/metabolismo , Traumatismo por Reperfusão/metabolismo , Traumatismo por Reperfusão/imunologia , Fator 3 Associado a Receptor de TNF/metabolismo , Fator 3 Associado a Receptor de TNF/genética , Receptor 3 Toll-Like/metabolismo , Receptor 3 Toll-Like/genética
8.
Int J Immunogenet ; 51(4): 242-251, 2024 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-38706134

RESUMO

Toll-like receptors (TLRs) play an important role in innate immunity. Previous studies have shown that single nucleotide polymorphisms (SNPs) in the genes coding for these innate immune molecules can affect susceptibility to and the outcome of certain diseases. The aim of the present study was to examine the clinical relevance of well-studied TLR1-4 SNPs in individuals who are prone to infections. Four functional SNPs, TLR1 rs5743618 (1805C > A, Ser602Ile), TLR2 rs5743708 (2258G > A, Arg753Gln), TLR3 rs3775291 (1234C > T, Leu412Phe) and TLR4 rs4986790 (896A > G, Asp299Gly), were analysed in 155 patients with recurrent respiratory infections (n = 84), severe infections (n = 15) or common variable immunodeficiency (n = 56), and in 262 healthy controls, using the High Resolution Melting Analysis method. Polymorphisms of TLR2 rs5743708 (odds ratio [OR] 3.16; 95% confidence interval [CI] 1.45-6.83, p = .004, ap = .016) and TLR4 rs4986790 (OR 1.8; 95% CI 1.05-3.12, p = .028, ap = .112) were more frequent in patients with recurrent or severe infections than in controls. Interestingly, seven patients were found to carry both variant genotypes of TLR2 and TLR4, whereas none of the control group carried such genotypes (p  ≤ .0001). Moreover, TLR2 polymorphism was associated with increased risk for acute otitis media episodes (OR, 3.02; 95% CI 1.41-6.47; p = .012). This study indicates that children and adults who are more prone to recurrent or severe respiratory infections carry one or both variant types of TLR2 and TLR4 more often than control subjects. Genetic variations of TLRs help explain why some children are more susceptible to respiratory infections.


Assuntos
Predisposição Genética para Doença , Polimorfismo de Nucleotídeo Único , Receptor 1 Toll-Like , Receptor 2 Toll-Like , Receptor 3 Toll-Like , Receptor 4 Toll-Like , Humanos , Masculino , Feminino , Receptor 4 Toll-Like/genética , Receptor 2 Toll-Like/genética , Receptor 3 Toll-Like/genética , Receptor 1 Toll-Like/genética , Criança , Adulto , Infecções Respiratórias/genética , Pré-Escolar , Adolescente , Recidiva , Pessoa de Meia-Idade , Genótipo , Frequência do Gene , Estudos de Casos e Controles
9.
Biochem Biophys Res Commun ; 712-713: 149915, 2024 Jun 18.
Artigo em Inglês | MEDLINE | ID: mdl-38663038

RESUMO

Viral infections pose a significant threat to public health, and the production of interferons represents one of the most critical antiviral innate immune responses of the host. Consequently, the screening and identification of compounds or reagents that induce interferon production are of paramount importance. This study commenced with the cultivation of host bacterium 15,597, followed by the infection of Escherichia coli with the MS2 bacteriophage. Utilizing the J2 capture technique, a class of dsRNA mixtures (MS2+15,597) was isolated from the E. coli infected with the MS2 bacteriophage. Subsequent investigations were conducted on the immunostimulatory activity of the MS2+15,597 mixture. The results indicated that the dsRNA mixtures (MS2+15,597) extracted from E. coli infected with the MS2 bacteriophage possess the capability to activate innate immunity, thereby inducing the production of interferon-ß. These dsRNA mixtures can activate the RIG-I and TLR3 pattern recognition receptors, stimulating the expression of interferon stimulatory factors 3/7, which in turn triggers the NF-κB signaling pathway, culminating in the cellular production of interferon-ß to achieve antiviral effects. This study offers novel insights and strategies for the development of broad-spectrum antiviral drugs, potentially providing new modalities for future antiviral therapies.


Assuntos
Escherichia coli , Levivirus , RNA de Cadeia Dupla , Escherichia coli/virologia , Escherichia coli/genética , Escherichia coli/metabolismo , RNA de Cadeia Dupla/metabolismo , Humanos , Levivirus/genética , Receptor 3 Toll-Like/metabolismo , Receptor 3 Toll-Like/genética , Imunidade Inata , Interferon beta/metabolismo , Interferon beta/genética , NF-kappa B/metabolismo , Proteína DEAD-box 58/metabolismo , Proteína DEAD-box 58/genética , Transdução de Sinais , Fator Regulador 7 de Interferon/metabolismo , Fator Regulador 7 de Interferon/genética , Receptores Imunológicos , Fator Regulador 3 de Interferon/metabolismo , Fator Regulador 3 de Interferon/genética
10.
Mol Biol Rep ; 51(1): 550, 2024 Apr 20.
Artigo em Inglês | MEDLINE | ID: mdl-38642183

RESUMO

BACKGROUND: The therapeutic efficacy of intra-articular mesenchymal stem cells (MSCs) injection for patients with osteoarthritis (OA) currently exhibits inconsistency, and the underlying mechanism remains elusive. It has been postulated that the immunomodulatory properties and paracrine activity of MSCs might be influenced by the inflammatory micro-environment within osteoarthritic joints, potentially contributing to this observed inconsistency. METHODS: Adipose-derived MSCs (ADSCs) were isolated from SD rats and pre-treated with Toll-like receptor 3 (TLR3) agonist Poly I:C or Toll-like receptor 4 (TLR4) agonist LPS. The pre-treated ADSCs were then co-cultured with IL-1ß-induced osteoarthritic chondrocytes using a Transwell system to analyze the paracrine effect of ADSCs on reversing the osteoarthritic phenotype of chondrocytes. RESULTS: RT-PCR and Western blot analysis revealed that Poly I:C and LPS pre-treatments up-regulated the expression of IL-10 and IL-6 in ADSCs, respectively. Furthermore, only Poly I:C-preconditioned ADSCs significantly promoted proliferation while inhibiting apoptosis in IL-1ß-treated chondrocytes. Additionally, Poly I:C-preconditioned ADSCs downregulated MMP13 expression while upregulating aggrecan and collagen II expression levels in IL-1ß-treated chondrocytes. CONCLUSIONS: TLR3 activation polarizes ADSCs into an immunomodulatory phenotype distinct from TLR4 activation, exerting differential effects on reversing the osteoarthritic phenotype of chondrocytes; thus indicating that MSCs' paracrine effect regulated by TLRs signaling impacts the efficacy of intra-articular MSCs injection.


Assuntos
Condrócitos , Células-Tronco Mesenquimais , Humanos , Ratos , Animais , Condrócitos/metabolismo , Receptor 4 Toll-Like/metabolismo , Receptor 3 Toll-Like/genética , Receptor 3 Toll-Like/metabolismo , Células Cultivadas , Lipopolissacarídeos/farmacologia , Lipopolissacarídeos/metabolismo , Ratos Sprague-Dawley , Células-Tronco Mesenquimais/metabolismo , Receptores Toll-Like/metabolismo , Fenótipo , Poli I/metabolismo , Poli I/farmacologia
11.
Viruses ; 16(4)2024 04 20.
Artigo em Inglês | MEDLINE | ID: mdl-38675983

RESUMO

Human immunodeficiency virus type 1 (HIV-1) infection can result in HIV-associated neurocognitive disorder (HAND), a spectrum of disorders characterized by neurological impairment and chronic inflammation. Combined antiretroviral therapy (cART) has elicited a marked reduction in the number of individuals diagnosed with HAND. However, there is continual, low-level viral transcription due to the lack of a transcription inhibitor in cART regimens, which results in the accumulation of viral products within infected cells. To alleviate stress, infected cells can release accumulated products, such as TAR RNA, in extracellular vesicles (EVs), which can contribute to pathogenesis in neighboring cells. Here, we demonstrate that cART can contribute to autophagy deregulation in infected cells and increased EV release. The impact of EVs released from HIV-1 infected myeloid cells was found to contribute to CNS pathogenesis, potentially through EV-mediated TLR3 (Toll-like receptor 3) activation, suggesting the need for therapeutics to target this mechanism. Three HIV-1 TAR-binding compounds, 103FA, 111FA, and Ral HCl, were identified that recognize TAR RNA and reduce TLR activation. These data indicate that packaging of viral products into EVs, potentially exacerbated by antiretroviral therapeutics, may induce chronic inflammation of the CNS observed in cART-treated patients, and novel therapeutic strategies may be exploited to mitigate morbidity.


Assuntos
Autofagia , Vesículas Extracelulares , Infecções por HIV , HIV-1 , Receptor 3 Toll-Like , Vesículas Extracelulares/metabolismo , Humanos , Receptor 3 Toll-Like/metabolismo , Receptor 3 Toll-Like/genética , HIV-1/fisiologia , Infecções por HIV/virologia , Infecções por HIV/metabolismo , Infecções por HIV/tratamento farmacológico , Autofagia/efeitos dos fármacos , RNA Viral/metabolismo , RNA Viral/genética
12.
Fish Shellfish Immunol ; 149: 109581, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38670412

RESUMO

Deubiquitinating enzyme A (DUBA), a member of the ovarian tumor (OTU) subfamily of deubiquitinases (DUBs), is recognized for its negative regulatory role in type I interferon (IFN) expression downstream of Toll-like receptor 3 (TLR3). However, its involvement in the TLR3 signaling pathway in fish remains largely unexplored. In this study, we investigated the regulatory role of DUBA (OmDUBA) in the TLR3 response in rainbow trout (Oncorhynchus mykiss). OmDUBA features a conserved OTU domain, and its expression increased in RTH-149 cells following stimulation with the TLR3 agonist poly(I:C). Gain- and loss-of-function experiments demonstrated that OmDUBA attenuated the activation of TANK-binding kinase 1 (TBK1), resulting in a subsequent reduction in type I IFN expression and IFN-stimulated response element (ISRE) activation in poly(I:C)-stimulated cells. OmDUBA interacted with TRAF3, a crucial mediator in TLR3-mediated type I IFN production. Under poly(I:C) stimulation, there was an augmentation in the K63-linked polyubiquitination of TRAF3, a process significantly inhibited upon OmDUBA overexpression. These findings suggest that OmDUBA may function similarly to its mammalian counterparts in downregulating the poly(I:C)-induced type I IFN response in rainbow trout by removing the K63-linked ubiquitin chain on TRAF3. Our study provides novel insights into the role of fish DUBA in antiviral immunity.


Assuntos
Proteínas de Peixes , Interferon Tipo I , Oncorhynchus mykiss , Poli I-C , Transdução de Sinais , Fator 3 Associado a Receptor de TNF , Animais , Oncorhynchus mykiss/imunologia , Fator 3 Associado a Receptor de TNF/genética , Fator 3 Associado a Receptor de TNF/metabolismo , Fator 3 Associado a Receptor de TNF/imunologia , Interferon Tipo I/imunologia , Interferon Tipo I/genética , Interferon Tipo I/metabolismo , Proteínas de Peixes/genética , Proteínas de Peixes/imunologia , Proteínas de Peixes/metabolismo , Transdução de Sinais/imunologia , Poli I-C/farmacologia , Imunidade Inata , Regulação da Expressão Gênica/imunologia , Ubiquitinação , Receptor 3 Toll-Like/genética , Receptor 3 Toll-Like/metabolismo , Receptor 3 Toll-Like/imunologia
13.
J Biol Chem ; 300(5): 107249, 2024 May.
Artigo em Inglês | MEDLINE | ID: mdl-38556084

RESUMO

Tripartite-motif protein-56 (TRIM56) positively regulates the induction of type I interferon response via the TLR3 pathway by enhancing IRF3 activation and depends on its C-terminal residues 621-750 for interacting with the adaptor TRIF. However, the precise underlying mechanism and detailed TRIM56 determinants remain unclear. Herein, we show ectopic expression of murine TRIM56 also enhances TLR3-dependent interferon-ß promoter activation, suggesting functional conservation. We found that endogenous TRIM56 and TRIF formed a complex early (0.5-2 h) after poly-I:C stimulation and that TRIM56 overexpression also promoted activation of NF-κB by poly-I:C but not that by TNF-α or IL-1ß, consistent with a specific effect on TRIF prior to the bifurcation of NF-κB and IRF3. Using transient transfection and Tet-regulated cell lines expressing various TRIM56 mutants, we demonstrated the Coiled-coil domain and a segment spanning residues ∼434-610, but not the B-box or residues 355-433, were required for TRIM56 augmentation of TLR3 signaling. Moreover, alanine substitution at each putative phosphorylation site, Ser471, Ser475, and Ser710, abrogated TRIM56 function. Concordantly, mutants bearing Ser471Ala, Ser475Ala, or Ser710Ala, or lacking the Coiled-coil domain, all lost the capacity to enhance poly-I:C-induced establishment of an antiviral state. Furthermore, the Ser710Ala mutation disrupted the TRIM56-TRIF association. Using phospho-specific antibodies, we detected biphasic phosphorylation of TRIM56 at Ser471 and Ser475 following TLR3 stimulation, with the early phase occurring at ∼0.5 to 1 h, prior to IRF3 phosphorylation. Together, these data reveal novel molecular details critical for the TRIM56 augmentation of TLR3-dependent antiviral response and highlight important roles for TRIM56 scaffolding and phosphorylation.


Assuntos
Proteínas Adaptadoras de Transporte Vesicular , Imunidade Inata , Receptor 3 Toll-Like , Proteínas com Motivo Tripartido , Animais , Humanos , Camundongos , Proteínas Adaptadoras de Transporte Vesicular/metabolismo , Proteínas Adaptadoras de Transporte Vesicular/genética , Proteínas Adaptadoras de Transporte Vesicular/imunologia , Células HEK293 , Fator Regulador 3 de Interferon/metabolismo , Fator Regulador 3 de Interferon/genética , NF-kappa B/metabolismo , Fosforilação , Poli I-C/farmacologia , Domínios Proteicos , Transdução de Sinais , Receptor 3 Toll-Like/metabolismo , Receptor 3 Toll-Like/genética , Proteínas com Motivo Tripartido/metabolismo , Proteínas com Motivo Tripartido/genética , Ubiquitina-Proteína Ligases/metabolismo , Ubiquitina-Proteína Ligases/genética
14.
J Pathol ; 263(2): 203-216, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38551071

RESUMO

Urothelial damage and barrier dysfunction emerge as the foremost mechanisms in Hunner-type interstitial cystitis/bladder pain syndrome (HIC). Although treatments aimed at urothelial regeneration and repair have been employed, their therapeutic effectiveness remains limited due to the inadequate understanding of specific cell types involved in damage and the lack of specific molecular targets within these mechanisms. Therefore, we harnessed single-cell RNA sequencing to elucidate the heterogeneity and developmental trajectory of urothelial cells within HIC bladders. Through reclustering, we identified eight distinct clusters of urothelial cells. There was a significant reduction in UPK3A+ umbrella cells and a simultaneous increase in progenitor-like pluripotent cells (PPCs) within the HIC bladder. Pseudotime analysis of the urothelial cells in the HIC bladder revealed that cells faced challenges in differentiating into UPK3A+ umbrella cells, while PPCs exhibited substantial proliferation to compensate for the loss of UPK3A+ umbrella cells. The urothelium in HIC remains unrepaired, despite the substantial proliferation of PPCs. Thus, we propose that inhibiting the pivotal signaling pathways responsible for the injury to UPK3A+ umbrella cells is paramount for restoring the urothelial barrier and alleviating lower urinary tract symptoms in HIC patients. Subsequently, we identified key molecular pathways (TLR3 and NR2F6) associated with the injury of UPK3A+ umbrella cells in HIC urothelium. Finally, we conducted in vitro and in vivo experiments to confirm the potential of the TLR3-NR2F6 axis as a promising therapeutic target for HIC. These findings hold the potential to inhibit urothelial injury, providing promising clues for early diagnosis and functional bladder self-repair strategies for HIC patients. © 2024 The Pathological Society of Great Britain and Ireland.


Assuntos
Cistite Intersticial , Receptor 3 Toll-Like , Urotélio , Animais , Feminino , Humanos , Camundongos , Diferenciação Celular , Proliferação de Células , Cistite Intersticial/patologia , Cistite Intersticial/metabolismo , Cistite Intersticial/genética , Camundongos Endogâmicos C57BL , Transdução de Sinais , Análise de Célula Única , Receptor 3 Toll-Like/metabolismo , Receptor 3 Toll-Like/genética , Bexiga Urinária/patologia , Bexiga Urinária/metabolismo , Urotélio/patologia , Urotélio/metabolismo
15.
Arch Med Res ; 55(3): 102985, 2024 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-38520880

RESUMO

BACKGROUND: Toll-like receptors (TLRs) play a critical role in initiating the innate immune response to infection or injury. Recent studies have uncovered their intriguing functions as moonlighting proteins involved in various biological processes, including development, learning, and memory. However, the specific functions of individual TLRs are still largely unknown. AIMS: We investigated the effects of TLR3 and TLR9 receptor deficiency on motor, cognitive, and behavioral functions during development using genetically modified male mice of different ages. METHODS: We evaluated the motor coordination, anxiety-like behavior, spatial learning, and working memory of male mice lacking the TLR3 and TLR9 genes at different ages (two, four, six, and eight months) using the rotarod, open field, water maze, and T-maze tests. RESULTS: We observed that the deletion of either TLR3 or TLR9 resulted in impaired motor performance. Furthermore, young TLR3-deficient mice exhibited reduced anxiety-like behavior and spatial learning deficits; however, their working memory was unaffected. In contrast, young TLR9-knockout mice showed hyperactivity and a tendency toward decreased working memory. CONCLUSIONS: These findings provide valuable insights into the broader roles of the TLR system beyond the innate immune response, revealing its involvement in pathways associated with the central nervous system. Importantly, our results establish a strong association between the endosomal receptors TLR3 and TLR9 and the performance of motor, cognitive, and behavioral tasks that change over time. This study contributes to the growing body of research on the multifaceted functions of TLRs and enhances our understanding of their participation in non-immune-related processes.


Assuntos
Receptor 3 Toll-Like , Receptor Toll-Like 9 , Animais , Masculino , Camundongos , Cognição , Camundongos Knockout , Receptor 3 Toll-Like/genética , Receptor 3 Toll-Like/metabolismo , Receptor Toll-Like 9/genética , Receptores Toll-Like/genética , Receptores Toll-Like/metabolismo
16.
Am J Physiol Lung Cell Mol Physiol ; 326(5): L618-L626, 2024 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-38469627

RESUMO

Thymic stromal lymphopoietin (TSLP) is an epithelial-derived pleiotropic cytokine that regulates T-helper 2 (Th2) immune responses in the lung and plays a major role in severe uncontrolled asthma. Emerging evidence suggests a role for endoplasmic reticulum (ER) stress in the pathogenesis of asthma. In this study, we determined if ER stress and the unfolded protein response (UPR) signaling are involved in TSLP induction in the airway epithelium. For this, we treated human bronchial epithelial basal cells and differentiated primary bronchial epithelial cells with ER stress inducers and the TSLP mRNA and protein expression was determined. A series of siRNA gene knockdown experiments were conducted to determine the ER stress-induced TSLP signaling pathways. cDNA collected from asthmatic bronchial biopsies was used to determine the gene correlation between ER stress and TSLP. Our results show that ER stress signaling induces TSLP mRNA expression via the PERK-C/EBP homologous protein (CHOP) signaling pathway. AP-1 transcription factor is important in regulating this ER stress-induced TSLP mRNA induction, though ER stress alone cannot induce TSLP protein production. However, ER stress significantly enhances TLR3-induced TSLP protein secretion in the airway epithelium. TSLP and ER stress (PERK) mRNA expression positively correlates in bronchial biopsies from participants with asthma, particularly in neutrophilic asthma. In conclusion, these results suggest that ER stress primes TSLP that is then enhanced further upon TLR3 activation, which may induce severe asthma exacerbations. Targeting ER stress using pharmacological interventions may provide novel therapeutics for severe uncontrolled asthma.NEW & NOTEWORTHY TSLP is an epithelial-derived cytokine and a key regulator in the pathogenesis of severe uncontrolled asthma. We demonstrate a novel mechanism by which endoplasmic reticulum stress signaling upregulates airway epithelial TSLP mRNA expression via the PERK-CHOP signaling pathway and enhances TLR3-mediated TSLP protein secretion.


Assuntos
Asma , Citocinas , Estresse do Retículo Endoplasmático , Células Epiteliais , Linfopoietina do Estroma do Timo , Receptor 3 Toll-Like , Resposta a Proteínas não Dobradas , Humanos , Citocinas/metabolismo , Receptor 3 Toll-Like/metabolismo , Receptor 3 Toll-Like/genética , Asma/metabolismo , Asma/patologia , Asma/genética , Células Epiteliais/metabolismo , Células Epiteliais/patologia , Fator de Transcrição CHOP/metabolismo , Fator de Transcrição CHOP/genética , Transdução de Sinais , Mucosa Respiratória/metabolismo , Mucosa Respiratória/patologia , Brônquios/metabolismo , Brônquios/patologia , eIF-2 Quinase/metabolismo , eIF-2 Quinase/genética , Células Cultivadas , Feminino , RNA Mensageiro/genética , RNA Mensageiro/metabolismo
17.
Biomed Pharmacother ; 173: 116450, 2024 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-38503239

RESUMO

Cisplatin not only targets DNA but also RNA. However, it is largely unknown whether platinated RNA (Pt-RNA) causes apoptosis and thus contributes to the cytotoxic effects of cisplatin. Consequently, cellular RNA was isolated from HepG2 and LS180 cells, exposed to cisplatin, and the resulting Pt-RNA (20 ng Pt/µg RNA) was transfected into these cancer cell lines or used to treat an apoptosis reporter Caenorhabditis elegans (C. elegans) strain (MD701, expressing CED-1::GFP). Cellular and molecular effects of Pt-RNA were evaluated by luminogenic caspase 3/7 assays, PCR array analysis, and fluorescence microscopy-based quantification of apoptosis in C. elegans gonads. Assuming RNA cross-linking (pseudo double-stranded RNA), the contribution of the Toll-like receptor 3 (TLR3, a sensor of double-stranded RNA) to apoptosis induction in cancer cell lines was investigated by pharmacological TLR3 inhibition and overexpression. In contrast to controls, Pt-RNA significantly enhanced apoptosis in C. elegans (2-fold) and in the cancer cell lines (2-fold to 4-fold). TLR3 overexpression significantly enhanced the pro-apoptotic effects of Pt-RNA in HepG2 cells. TLR3 inhibition reduced the pro-apoptotic effects of Pt-RNA and cisplatin, but not of paclitaxel (off-target control). Gene expression analysis showed that Pt-RNA (but not RNA) significantly enhanced the mRNA levels of nuclear factor kappa B subunit 2 and interleukin-8 in HepG2 cells, suggesting that Pt-RNA is a damage-associated molecular pattern that additionally causes pro-inflammatory responses. Together, this data suggests that not only DNA but also cellular RNA is a functionally relevant target of cisplatin, leading to pro-apoptotic and immunogenic effects.


Assuntos
Cisplatino , Neoplasias , Animais , Cisplatino/farmacologia , Caenorhabditis elegans/genética , Caenorhabditis elegans/metabolismo , Receptor 3 Toll-Like/genética , Receptor 3 Toll-Like/metabolismo , RNA de Cadeia Dupla/genética , RNA de Cadeia Dupla/farmacologia , Apoptose , Linhagem Celular Tumoral , DNA , Neoplasias/tratamento farmacológico , Neoplasias/genética
18.
Mol Biol Rep ; 51(1): 417, 2024 Mar 14.
Artigo em Inglês | MEDLINE | ID: mdl-38483660

RESUMO

BACKGROUND: Bronchial epithelial cells are at the front line of viral infections. Toll-like receptor 3 (TLR3) cascade causes the expression of interferon (IFN)-ß and IFN-stimulated genes (ISGs), which in turn induce an antiviral response. Members of the transmembrane protein (TMEM) family are expressed in various cell types. Although the prognostic value of TMEM2 in various cancers has been reported, its association with infectious diseases remains unknown. In this study, we investigated the effects of TMEM2 on antiviral immunity in BEAS-2B bronchial epithelial cells. METHODS AND RESULTS: TMEM2 protein was found in the cytoplasm of normal human bronchial epithelial cells and differed between organs using immunohistochemistry. Cultured BEAS-2B cells were transfected with TMEM2 siRNA, followed by administration of TLR3 ligand polyinosinic-polycytidylic acid (poly IC) or recombinant human (r(h)) IFN-ß. The expression of TMEM2, IFN-ß, ISG56, C-X-C motif chemokine ligand 10 (CXCL10) and hyaluronan were evaluated appropriately by western blotting, quantitative reverse transcription-polymerase chain reaction and enzyme-linked immunosorbent assay. TMEM2 expression was not altered by poly IC stimulation. Knockdown of TMEM2 increased poly IC-induced expression of IFN-ß, CXCL10, and ISG56, while IFN-ß-induced expression of ISG56 and CXCL10 were not changed by TMEM2 knockdown. The hyaluronan concentration in the medium was decreased by either TMEM2 knockdown or poly IC, but additive or synergistic effects were not observed. CONCLUSIONS: TMEM2 knockdown enhanced TLR3-mediated IFN-ß, CXCL10, and ISG56 expression in BEAS-2B cells. This implies that TMEM2 suppresses antiviral immune responses and prevents tissue injury in bronchial epithelial cells.


Assuntos
Ácido Hialurônico , Receptor 3 Toll-Like , Humanos , Receptor 3 Toll-Like/genética , Receptor 3 Toll-Like/metabolismo , Ligantes , Poli I-C/farmacologia , Células Epiteliais/metabolismo , Células Cultivadas , Quimiocina CXCL10/genética
19.
EMBO J ; 43(5): 780-805, 2024 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-38316991

RESUMO

Inflammation is a common condition of prostate tissue, whose impact on carcinogenesis is highly debated. Microbial colonization is a well-documented cause of a small percentage of prostatitis cases, but it remains unclear what underlies the majority of sterile inflammation reported. Here, androgen- independent fluctuations of PSA expression in prostate cells have lead us to identify a prominent function of the Transient Receptor Potential Cation Channel Subfamily M Member 8 (TRPM8) gene in sterile inflammation. Prostate cells secret TRPM8 RNA into extracellular vesicles (EVs), which primes TLR3/NF-kB-mediated inflammatory signaling after EV endocytosis by epithelial cancer cells. Furthermore, prostate cancer xenografts expressing a translation-defective form of TRPM8 RNA contain less collagen type I in the extracellular matrix, significantly more infiltrating NK cells, and larger necrotic areas as compared to control xenografts. These findings imply sustained, androgen-independent expression of TRPM8 constitutes as a promoter of anticancer innate immunity, which may constitute a clinically relevant condition affecting prostate cancer prognosis.


Assuntos
Neoplasias da Próstata , Canais de Cátion TRPM , Humanos , Masculino , Androgênios , Inflamação/genética , Fator Regulador 3 de Interferon , Proteínas de Membrana , NF-kappa B/genética , Neoplasias da Próstata/genética , Receptor 3 Toll-Like/genética , Canais de Cátion TRPM/genética , Animais
20.
Bioorg Med Chem Lett ; 101: 129672, 2024 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-38387691

RESUMO

Influenza and COVID-19 continue to pose global threats to public health. Classic antiviral drugs have certain limitations, coupled with frequent viral mutations leading to many drugs being ineffective, the development of new antiviral drugs is urgent. Meanwhile, the invasion of influenza virus can cause an immune response, and an excessive immune response can generate a large number of inflammatory storms, leading to tissue damage. Toll-like receptor 3 (TLR3) recognizes virus dsRNA to ignite the innate immune response, and inhibit TLR3 can block the excess immune response and protect the host tissues. Taking TLR3 as the target, SMU-CX1 was obtained as the specific TLR3 inhibitor by high-throughput screening of 15,700 compounds with IC50 value of 0.11 µM. Its anti-influenza A virus activity with IC50 ranged from 0.14 to 0.33 µM against multiple subtypes of influenza A virus and also showed promising anti-SARS-CoV-2 activity with IC50 at 0.43 µM. Primary antiviral mechanism study indicated that SMU-CX1 significantly inhibited PB2 and NP protein of viruses, it can also inhibit inflammatory factors in host cells including IFN-ß, IP-10 and CCL-5. In conclusion, this study demonstrates the potential of SMU-CX1 in inhibiting IAV and SARS-CoV-2 activity, thereby offering a novel approach for designing antiviral drugs against highly pathogenic viruses.


Assuntos
COVID-19 , Elipticinas , Vírus da Influenza A , Humanos , Vírus da Influenza A/metabolismo , SARS-CoV-2/metabolismo , Receptor 3 Toll-Like/genética , Receptor 3 Toll-Like/metabolismo , Antivirais/farmacologia , Antivirais/uso terapêutico
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