RESUMO
Herpes simplex virus 1 (HSV-1) or simplexvirus humanalpha 1 is a neurotropic virus that is responsible for orofacial infections in humans. More than 70% of the world's population may have seropositivity for HSV-1, and this virus is a leading cause of sporadic lethal encephalitis in humans. The role of toll-like receptors (TLRs) in defending against HSV-1 infection has been explored, including the consequences of lacking these receptors or other proteins in the TLR pathway. Cell and mouse models have been used to study the importance of these receptors in combating HSV-1, how they relate to the innate immune response, and how they participate in the orchestration of the adaptive immune response. Myeloid differentiation factor 88 (MyD88) is a protein involved in the downstream activation of TLRs and plays a crucial role in this signaling. Mice with functional MyD88 or TLR2 and TLR9 can survive HSV-1 infection. However, they can develop encephalitis and face a 100% mortality rate in a dose-dependent manner when MyD88 or TLR2 plus TLR9 proteins are non-functional. In TLR2/9 knockout mice, an increase in chemokines and decreases in nitric oxide (NO), interferon (IFN) gamma, and interleukin 1 (IL-1) levels in the trigeminal ganglia (TG) have been correlated with mortality.
Assuntos
Encefalite , Herpes Simples , Herpesvirus Humano 1 , Humanos , Animais , Camundongos , Herpesvirus Humano 1/metabolismo , Receptor 2 Toll-Like/metabolismo , Receptor Toll-Like 9/genética , Receptor Toll-Like 9/metabolismo , Fator 88 de Diferenciação Mieloide/genética , Fator 88 de Diferenciação Mieloide/metabolismo , Gânglio Trigeminal/metabolismo , Receptores Toll-Like/metabolismo , Camundongos Knockout , Camundongos Endogâmicos C57BLRESUMO
Although the baculovirus Autographa californica multiple nuclear polyhedrosis virus (AcMNPV) infects lepidopteran invertebrates as natural hosts, represents an efficient vector for vaccine development. Baculovirus surface display induces strong humoral responses against viruses and parasites. A novel strategy based on capsid display carrying foreign antigens in the AcMNPV particle further improved the immune response by eliciting CD8+ T cell activation. In this study, we analyze the intracellular mechanisms and signalling pathways involved in CD8+ T cell activation by capsid display. Our results show that baculovirus can attach to the cell surface, enter dendritic cells (DCs), transit within endocytic vesicles and escape to the cytosol for further degradation by the proteasome. We found that the availability of viral proteins, endosomal acidification, and proteasome activity are needed for efficient Major Histocompatibility Complex class-I presentation by baculovirus carrying Ovalbumin in the viral capsid. Importantly, we demonstrated with this strategy that the induction of cytotoxic T cells and IL-12 production by DCs are TLR9-dependent and STING-independent. Finally, our study shows differential intracellular processing for capsid and surface baculovirus proteins in DCs and highlights the role of different danger receptors during cytotoxic T cell priming through the capsid display delivery system, which could lead to improved baculovirus-based vaccines development.
Assuntos
Antineoplásicos , Baculoviridae , Baculoviridae/genética , Baculoviridae/metabolismo , Capsídeo , Receptor Toll-Like 9/genética , Receptor Toll-Like 9/metabolismo , Complexo de Endopeptidases do Proteassoma/metabolismo , Proteínas do Capsídeo/genéticaRESUMO
Abstract Toll-like receptor 9 (TLR9) is an important component of the innate immune system and have been associated with several autoimmune diseases, such as Systemic Lupus Erythematosus (SLE). The aim of this study was to investigate polymorphisms in TLR9 gene in a Brazilian SLE patients group and their association with clinical manifestation, particularly Jaccoud's arthropathy (JA). We analyzed DNA samples from 204 SLE patients, having a subgroup of them presenting JA (n=24). A control group (n=133) from the same city was also included. TLR9 single nucleotide polymorphisms (SNPs) (−1237 C>T and +2848 G>A) were identified by sequencing analysis. The TLR9 gene genotype frequency was similar both in SLE patients and the control group. In the whole SLE population, an association between the homozygosis of allele C at position −1237 with psychosis and anemia (p < 0.01) was found. Likewise, the homozygosis of allele G at position +2848 was associated with a discoid rash (p < 0.05). There was no association between JA and TLR9 polymorphisms. These data show that TLR9 polymorphisms do not seem to be a predisposing factor for SLE in the Brazilian population, and that SNPs are not associated with JA.
Resumo O receptor Toll-like 9 (TLR9) é um componente importante do sistema imunológico inato e tem sido associado a várias doenças autoimunes, como o Lúpus Eritematoso Sistêmico (LES). O objetivo deste estudo foi investigar polimorfismos no gene TLR9 em um grupo de pacientes brasileiros com LES e sua associação com a manifestação clínica, particularmente a artropatia de Jaccoud (JA). Foram analisadas amostras de DNA de 204 pacientes com LES, e um subgrupo com JA (n=24). Um grupo de controle (n=133) da mesma cidade também foi incluído. Os polimorfismos de nucleotídeos únicos TLR9 (SNPs) (−1237 C>T e +2848 G>A) foram identificados pela análise de sequenciamento. A frequência do genótipo genético TLR9 foi semelhante tanto em pacientes com LES quanto no grupo controle. Em toda a população de LES, foi encontrada associação entre a homozigose do alelo C na posição −1237 com psicose e anemia (p < 0,01). Da mesma forma, a homozigose do alelo G na posição +2848 foi associada a uma erupção cutânea discoide (p < 0,05). Não houve associação entre polimorfismos JA e TLR9. Esses dados mostram que os polimorfismos TLR9 não parecem ser um fator predisponível para o LES na população brasileira, e que os SNPs não estão associados ao JA.
Assuntos
Humanos , Receptor Toll-Like 9/genética , Lúpus Eritematoso Sistêmico/genética , Brasil , Projetos Piloto , Predisposição Genética para Doença/genética , Frequência do Gene/genéticaRESUMO
Selection of genetically resistant animals is one alternative to reduce the negative impact of gastrointestinal nematodes (GIN) on sheep production. The aim of this study was to identify genomic regions associated with GIN resistance in Corriedale sheep by single-step genome-wide association studies (ssGWAS) using 170, 507 and 50K single nucleotide polymorphisms (SNPs). Analysis included 19,547 lambs with faecal egg counts (FEC) records, a pedigree file of 40,056 animals and 454, 711 and 383 genotypes from 170, 507 and 50K SNPs, respectively. Genomic estimated breeding values (GEBV) were obtained with single-step genomic BLUP methodology (ssGBLUP), using a univariate animal model, which included contemporary group, type of birth and age of dam as class fixed effects and age at FEC recording as covariate. The SNP effects as wells as p-values were estimated with POSTGSF90 program. Significance level was defined by a chromosome-wise False Discovery Rate of 5%. Significant genomic regions were identified in chromosomes 1, 3, 12 and 19 with the 170 SNP set, in chromosomes 7, 12 and 24 using the 507 SNP chip and only in chromosome 7 with the 50K SNP chip. Candidate genes located in these regions, using Oar_v4.0 as reference genome, were TIMP3, TLR5, LEPR and TLR9 (170 SNPs), SYNDIG1L and MGRN1 (507 SNP chip) and INO80, TLN2, TSHR and EIF2AK4 (50K SNP chip). These results validate genomic regions associated with FEC previously identified in Corriedale and other breeds and report new candidate regions for further investigation.
Assuntos
Nematoides , Parasitos , Animais , Estudo de Associação Genômica Ampla , Nematoides/genética , Ovinos/genética , Carneiro Doméstico/genética , Receptor 5 Toll-Like/genética , Receptor Toll-Like 9/genéticaRESUMO
Acute lymphoblastic leukemia (ALL) is the most common hematologic malignancy in children in childhood. Single-nucleotide polymorphism (SNPs) in key molecules of the immune system, such as Toll-like receptors (TLRs) and CD14 molecules, are associated with the development of several diseases. However, their role in ALL is unknown. A case-control study was performed with 152 ALL patients and 187 healthy individuals to investigate the role of SNPs in TLRs and the CD14 gene in ALL. In this study, TLR6 C > T rs5743810 [OR: 3.20, 95% CI: 1.11-9.17, p = 0.003) and TLR9 C > T rs187084 (OR: 2.29, 95% CI: 1.23-4.26, p = 0.000) seems to be a risk for development of ALL. In addition, the TLR1 T > G rs5743618 and TLR6 C > T rs5743810 polymorphisms with protection against death (OR: 0.17, 95% IC: 0.04-0.79, p = 0.008; OR: 0.48, 95% IC: 0.24-0.94, p = 0.031, respectively). Our results show that SNPs in TLRs genes may be involved in the pathogenesis of ALL and may influence clinical prognosis; however, further studies are necessary to elucidate the role of TLR1, TLR4, TLR5, TLR6, TLR9 and CD14 polymorphisms in this disease.
Assuntos
Predisposição Genética para Doença , Leucemia-Linfoma Linfoblástico de Células Precursoras , Brasil/epidemiologia , Estudos de Casos e Controles , Criança , Humanos , Polimorfismo de Nucleotídeo Único , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Receptor 1 Toll-Like/genética , Receptor 6 Toll-Like/genética , Receptor Toll-Like 9/genética , Receptores Toll-Like/genéticaRESUMO
Background: Host genetic factors play a major role with respect to susceptibility to infections. Many polymorphisms of the Toll-like receptors (TLRs), members of the innate immune response, are directly associated with the clinical outcomes following infection. The 2848 G/A variant (rs352140) of the TLR9 gene is associated with increased TLR9 expression. However, the impact of the genotypes of this SNP on HIV+, HCV+, and HCV+/HIV+ individuals is still debated. Materials and Methods: This study investigated the 2848 G/A polymorphism in HCV infection, HIV infection, and HCV/HIV co-infection in a large sample of Brazilians (n = 1,182). Groups were initially compared without considering stratification by ethnicity and subsequently stratifying individuals between whites and non-whites. Results: Considering non-white individuals, a significant difference between the HIV+/HCV+ group and controls was observed with the GG genotype serving as a protective factor (p = 0.023). Additionally, significant allelic differences were observed between the HCV+ group and controls (p = 0.042); between the HIV+/HCV+ group and controls (p = 0.011); and between the HIV+/HCV+ group and HIV+ individuals (p = 0.047). However, all significant results were lost following adjustment for multiple comparisons (p > 0.05). Conclusion: Although our initial results indicated a potential influence of the rs352140 genotype on host altered susceptibility to viral infections, following correction for multiple comparisions the standard (p < 0.05) for statistical association was lost. This may be due to insufficient sample size as we were examining many different associations. Thus, a larger study is warranted to further pursue this topic.
Assuntos
Infecções por HIV , Hepatite C , Predisposição Genética para Doença/genética , Genótipo , Infecções por HIV/genética , Hepatite C/complicações , Hepatite C/genética , Humanos , Polimorfismo de Nucleotídeo Único/genética , Receptor Toll-Like 9/genéticaRESUMO
AIM: To explore the methylation pattern, its role in transcriptional regulation and potential modifiers of methylation of the TLR9 gene in chronic periapical inflammation. METHODOLOGY: In this cross-sectional study, apical lesions of endodontic origin (ALEO, n = 61) and healthy periodontal ligaments (HPL, n = 15) were included. Products from bisulfited and PCR-amplified DNA were analysed for their methylation profiles in the promoter region and at each CpG island. Additionally, TLR9 mRNA levels were quantified by qPCR and bivariate and multiple modelling were performed to better understand the influence of methylation on gene transcription. RESULTS: TLR9 mRNA levels were upregulated in ALEO compared to HPL (p < .001). TLR9 promoter CpG sites and CpG +2086 in the intragenic island 1 were demethylated in ALEO compared to HPL (p < .05). Multivariate analysis, adjusted by smoking and gender, revealed that demethylation of TLR9 promoter sites enhanced transcriptional activity, specifically demethylated CpGs at positions -736 and -683, (p = .02), which are close to CRE binding. Although ALEO reduced the global methylation of the gene promoter and intragenic-island 2 (p < .05) by -42.5 and -9.5 percentage points, respectively, age reduced the global methylation of intragenic-island 3 within the exon 2. CONCLUSIONS: Demethylations of TLR9 promoter CpG sites, along with the intragenic DNA methylation status, were involved in higher transcription in ALEO. Hence, chronic periapical inflammation and ageing modify the methylation status both in the gene promoter and in intragenic CpG islands.
Assuntos
Metilação de DNA , Periodontite Periapical , Receptor Toll-Like 9 , Ilhas de CpG/genética , Estudos Transversais , Humanos , Inflamação , Periodontite Periapical/genética , RNA Mensageiro/metabolismo , Receptor Toll-Like 9/genética , Receptor Toll-Like 9/metabolismoRESUMO
Cancer chemotherapy and radiotherapy may result in mucositis characterized by stem cell damage and inflammation in the gastrointestinal tract. The molecular mechanisms underlying this pathology remain unknown. Based on the assumption that mitochondrial CPG-DNA (mtDNA) released and sensed by TLR9 could underlie mucositis pathology, we analyzed the mtDNA levels in sera as well as inflammatory and disease parameters in the small intestine from wild-type (WT) and TLR9-deficient mice (TLR9-/-) in an experimental model of intestinal mucositis induced by irinotecan. Additionally, we verified the ability of WT and TLR9-/- macrophages to respond to CpG-DNA in vitro. WT mice injected with irinotecan presented a progressive increase in mtDNA in the serum along with increased hematocrit, shortening of small intestine length, reduction of intestinal villus:crypt ratio and increased influx of neutrophils, which were followed by higher expression of Nlrp3 and Casp1 mRNA and increased IL-1ß levels in the ileum when compared to vehicle-injected mice. TLR9-deficient mice were protected in all these parameters when compared to WT mice. Furthermore, TLR9 was required for the production of IL-1ß and NO after macrophage stimulation with CpG-DNA. Overall, our findings show that the amount of circulating free CpG-DNA is increased upon chemotherapy and that TLR9 activation is important for NLRP3 inflammasome transcription and further IL-1ß release, playing a central role in the development of irinotecan-induced intestinal mucositis. We suggest that TLR9 antagonism may be a new therapeutic strategy for limiting irinotecan-induced intestinal inflammation.
Assuntos
Mucosite , Animais , DNA Mitocondrial/genética , Inflamação/metabolismo , Irinotecano/toxicidade , Ligantes , Camundongos , Camundongos Knockout , Mucosite/induzido quimicamente , Mucosite/tratamento farmacológico , Mucosite/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR , Receptor Toll-Like 9/genética , Receptor Toll-Like 9/metabolismoRESUMO
The innate and acquired immune response induced by a commercial inactivated vaccine against Bovine Herpesvirus-1 (BoHV-1) and protection conferred against the virus were analyzed in cattle. Vaccination induced high levels of BoHV-1 antibodies at 30, 60, and 90 days post-vaccination (dpv). IgG1 and IgG2 isotypes were detected at 90 dpv, as well as virus-neutralizing antibodies. An increase of anti-BoHV-1 IgG1 in nasal swabs was detected 6 days post-challenge in vaccinated animals. After viral challenge, lower virus excretion and lower clinical score were observed in vaccinated as compared to unvaccinated animals, as well as BoHV-1-specific proliferation of lymphocytes and production of IFNγ, TNFα, and IL-4. Downregulation of the expression of endosome Toll-like receptors 8-9 was detected after booster vaccination. This is the first thorough study of the immunity generated by a commercial vaccine against BoHV-1 in cattle.
Assuntos
Anticorpos Neutralizantes/biossíntese , Herpesvirus Bovino 1/imunologia , Vacinas contra Herpesvirus/administração & dosagem , Imunoglobulina G/biossíntese , Rinotraqueíte Infecciosa Bovina/prevenção & controle , Receptor 8 Toll-Like/imunologia , Receptor Toll-Like 9/imunologia , Imunidade Adaptativa/efeitos dos fármacos , Animais , Anticorpos Antivirais , Bovinos , Proliferação de Células , Endossomos/imunologia , Endossomos/metabolismo , Expressão Gênica , Herpesvirus Bovino 1/patogenicidade , Imunidade Inata/efeitos dos fármacos , Imunização Secundária/métodos , Rinotraqueíte Infecciosa Bovina/genética , Rinotraqueíte Infecciosa Bovina/imunologia , Rinotraqueíte Infecciosa Bovina/virologia , Interferon gama/genética , Interferon gama/imunologia , Interleucina-4/genética , Interleucina-4/imunologia , Linfócitos/imunologia , Linfócitos/virologia , Masculino , Cavidade Nasal/imunologia , Cavidade Nasal/virologia , Receptor 8 Toll-Like/agonistas , Receptor 8 Toll-Like/genética , Receptor Toll-Like 9/agonistas , Receptor Toll-Like 9/genética , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/imunologia , Vacinação/métodos , Vacinas de Produtos InativadosRESUMO
BACKGROUND AND PURPOSE: Severe diarrhoea, a common gastrointestinal manifestation of anticancer treatment with irinotecan, might involve single nucleotide polymorphisms (SNPs) of toll-like receptors (TLRs), described as critical bacterial sensors in the gut. Here, colorectal cancer patients carrying missense TLR4 A896G (rs4986790) or C1,196T (rs4986791) SNPs and Tlr4 knockout (Tlr4-/-) mice were given irinotecan to investigate the severity of the induced diarrhoea. EXPERIMENTAL APPROACH: Forty-six patients treated with irinotecan-based regimens had diarrhoea severity analysed according to TLR4 genotypes. In the experimental setting, wild-type (WT) or Tlr4-/- mice were given irinotecan (45 or 75 mg·kg-1 , i.p.) or saline (3 ml·kg-1 ). Diarrhoea severity was evaluated by measuring intestinal injury and inflammatory markers expression after animals were killed. KEY RESULTS: All patients with TLR4 SNPs chemotherapy-treated presented diarrhoea, whereas gastrointestinal toxicity was observed in 50% of the wild homozygous individuals. Mice injected with irinotecan presented systemic bacterial translocation and increased TLR4 immunostaining in the intestine. In line with the clinical findings, Tlr4 gene deficiency enhanced irinotecan-related diarrhoea and TLR9 expression in mice. An increased myeloperoxidase activity and Il-18 expression along with IL-10 decreased production in Tlr4-/- mice also indicated an intensified intestinal damage and inflammatory response. CONCLUSION AND IMPLICATIONS: TLR4 deficiency upregulates TLR9 expression and enhances intestinal damage and the severity of late-onset diarrhoea during irinotecan-based treatment. Identifying patients genetically predisposed to chemotherapy-associated diarrhoea is a strategy toward precision medicine.