RESUMEN
Food allergy is a life-threatening response to specific foods, and microbiota imbalance (dysbiosis) in gut is considered a cause of this disease. Meanwhile, the host immune response also plays an important role in the disease. Notably, interleukin 33 (IL-33) released from damaged or necrotic intestinal epithelial cells facilitates IL-2-producing CD4 helper T (Th2) responses. However, causal relationships between the gut and oral dysbiosis and food allergy remain unknown. In this study, we analyzed effects of gut and oral dysbiosis on development of food allergy. A murine model of food allergy was established via ovalbumin (OVA) injection in BALB/c mice. Viable fecal bacteria were identified using matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS). il33 expression in colon-26 mouse colon cells stimulated by isolated fecal bacteria was quantified by real-time PCR. Intestinal T cells from the mice were analyzed by flow cytometry. Salivary IgA levels were quantified by enzyme-linked immunosorbent assay (ELISA), and IgA-bound oral bacteria were detected by flow cytometry. Among fecal bacteria, the abundance of Citrobacter sp. increased in the feces of allergic mice and induced il33 expression in colon-26 cells. Orally administered Citrobacter koseri JCM1658 exacerbated systemic allergic symptoms and reduced intestinal Th17 cells. Salivary IgA and IgA-bound oral bacteria increased in the allergic mice. Based on the results described above, food allergy induced both gut and oral dysbiosis. Citrobacter sp. aggravated allergy symptoms by inducing IL-33 release from intestinal epithelial cells.
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Disbiosis , Hipersensibilidad a los Alimentos/complicaciones , Tracto Gastrointestinal/microbiología , Inmunoglobulina A/metabolismo , Factores Inmunológicos/metabolismo , Microbiota/efectos de los fármacos , Boca/microbiología , Animales , Citocinas/metabolismo , Modelos Animales de Enfermedad , Células Epiteliales/metabolismo , Ratones Endogámicos BALB CRESUMEN
Phytochromes are red light (R) and far-red light (FR) receptors that play important roles in many aspects of plant growth and development. Phytochromes mainly function in the nucleus and regulate sets of genes by inhibiting negatively acting basic helix-loop-helix transcription factors named PHYTOCHROME INTERACTING FACTORs (PIFs) in Arabidopsis thaliana Although R/FR photoreversible responses and phytochrome genes are well documented in diverse lineages of plants, the extent to which phytochrome signaling is mediated by gene regulation beyond angiosperms remains largely unclear. Here, we show that the liverwort Marchantia polymorpha, an emerging model basal land plant, has only one phytochrome gene, Mp-PHY, and only one PIF gene, Mp-PIF These genes mediate typical low fluence responses, which are reversibly elicited by R and FR, and regulate gene expression. Mp-phy is light-stable and translocates into the nucleus upon irradiation with either R or FR, demonstrating that the single phytochrome Mp-phy exhibits combined biochemical and cell-biological characteristics of type I and type II phytochromes. Mp-phy photoreversibly regulates gemma germination and downstream gene expression by interacting with Mp-PIF and targeting it for degradation in an R-dependent manner. Our findings suggest that the molecular mechanisms for light-dependent transcriptional regulation mediated by PIF transcription factors were established early in land plant evolution.
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Marchantia/metabolismo , Fitocromo/metabolismo , Proteínas de Plantas/metabolismo , Arabidopsis/metabolismo , Arabidopsis/efectos de la radiación , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/metabolismo , Regulación de la Expresión Génica de las Plantas/genética , Regulación de la Expresión Génica de las Plantas/efectos de la radiación , Luz , Marchantia/efectos de la radiación , Proteínas de Plantas/genética , Transducción de Señal/genética , Transducción de Señal/fisiología , Factores de Transcripción/genética , Factores de Transcripción/metabolismoRESUMEN
Small noncoding RNAs (sRNAs) have been identified as important regulators of gene expression in various cellular processes. cia-dependent small RNAs (csRNAs), a group of sRNAs that are controlled by the two-component regulatory system CiaRH, are widely conserved in streptococci, but their targets have been identified only in Streptococcus pneumoniaeStreptococcus sanguinis, a pioneer colonizer of teeth and one of the most predominant bacteria in the early oral biofilm, has been shown to have six csRNAs. Using computational target prediction and the luciferase reporter assay, we identified pilT, a constituent of the type IV pilus operon, as a negative regulatory target for one of the csRNAs, namely, csRNA1-1, in S. sanguinis RNA-RNA electrophoretic mobility shift assay using a nucleotide exchange mutant of csRNA1-1 revealed that csRNA1-1 binds directly to pilT mRNA. In addition, csRNA1-1 and csRNA1-2, a putative gene duplication product of csRNA1-1 that is tandemly located in the S. sanguinis genome, negatively regulated S. sanguinis biofilm formation. These results suggest the involvement of csRNAs in the colonization step of S. sanguinis.
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Adenosina Trifosfatasas/metabolismo , Proteínas Bacterianas/metabolismo , Biopelículas , Regulación Bacteriana de la Expresión Génica , ARN Bacteriano/genética , ARN Pequeño no Traducido/metabolismo , Infecciones Estreptocócicas/microbiología , Streptococcus sanguis/genética , Adenosina Trifosfatasas/genética , Proteínas Bacterianas/genética , Regulación hacia Abajo , Fimbrias Bacterianas/genética , Fimbrias Bacterianas/metabolismo , ARN Bacteriano/metabolismo , ARN Pequeño no Traducido/genética , Streptococcus sanguis/fisiologíaRESUMEN
We previously reported that inhibition of dipeptidyl peptidase (DPP)-4, the catalytic site of CD26, prevents atherosclerosis in animal models through suppression of inflammation; however, the underlying molecular mechanisms have not been fully elucidated. Caveolin-1 (Cav-1), a major structural protein of caveolae located on the surface of the cellular membrane, has been reported to modulate inflammatory responses by binding to CD26 in T cells. In this study, we investigated the role of Cav-1 in the suppression of inflammation mediated by the DPP-4 inhibitor, teneligliptin, using mouse and human macrophages. Mouse peritoneal macrophages were isolated from Cav-1+/+ and Cav-1-/- mice after stimulation with 3% thioglycolate. Inflammation was induced by the toll-like receptor (TLR)4 agonist, lipopolysaccharide (LPS), isolated from Escherichia coli. The expression of pro-inflammatory cytokines was determined using reverse transcription-polymerase chain reaction. Co-expression of Cav-1 and CD26 was detected using immunohistochemistry in both mouse and human macrophages. Teneligliptin treatment (10 nmol/L) suppressed the LPS-induced expression of interleukin (IL)-6 (70%) and tumor necrosis factor-α (37%) in peritoneal macrophages isolated from Cav-1+/+ mice. However, teneligliptin did not have any effect on the macrophages from Cav-1-/- mice. In human monocyte/macrophage U937 cells, teneligliptin treatment suppressed LPS-induced expression of pro-inflammatory cytokines in a dose-dependent manner (1-10 nmol/L). These anti-inflammatory effects of teneligliptin were mimicked by gene knockdown of Cav-1 or CD26 using small interfering RNA transfection. Furthermore, neutralization of these molecules using an antibody against CD26 or Cav-1 also showed similar suppression. Teneligliptin treatment specifically inhibited TLR4 and TLR5 agonist-mediated inflammatory responses, and suppressed LPS-induced phosphorylation of IL-1 receptor-associated kinase 4, a downstream molecule of TLR4. Next, we determined whether teneligliptin could directly inhibit the physical interaction between Cav-1 and CD26 using the Biacore system. Binding of CD26 to Cav-1 protein was detected. Unexpectedly, teneligliptin also bound to Cav-1, but did not interfere with CD26-Cav-1 binding, suggesting that teneligliptin competes with CD26 for binding to Cav-1. In conclusion, we demonstrated that Cav-1 is a target molecule for DPP-4 inhibitors in the suppression of TLR4-mediated inflammation in mouse and human macrophages.
Asunto(s)
Antiinflamatorios/farmacología , Caveolina 1/inmunología , Dipeptidil Peptidasa 4/inmunología , Inhibidores de la Dipeptidil-Peptidasa IV/farmacología , Macrófagos/efectos de los fármacos , Pirazoles/farmacología , Tiazolidinas/farmacología , Animales , Femenino , Humanos , Mediadores de Inflamación/inmunología , Macrófagos/inmunología , Ratones , Receptor Toll-Like 4/inmunología , Receptor Toll-Like 5/inmunologíaRESUMEN
Oral streptococci including mitis group streptococci are commensal residents and are also the first to colonize the oral cavity. However, various species of these oral streptococci have the potential to invade the host and occasionally lead to severe infectious disease such as cardiovascular diseases. Oral streptococci have close interactions with the host immune system including macrophages at the oral mucosal surface. One notable common trait of oral streptococcus including Streptococcus oralis (S. oralis) is the production of hydrogen peroxide (H2O2). Using a comprehensive microarray approach, we sought to understand the innate immune response profiling affected by H2O2 production from oral streptococci. We compared the gene expression patterns of macrophages infected with S. oralis wild type (WT) and streptococcal pyruvate oxidase knockout (SpxB-KO), a strain that does not produce H2O2. We found that H2O2 from S. oralis suppressed proinflammatory gene expression such as TNF-α, that is induced in response to infection, and activated the cellular stress genes such as Egr-1 in response to oxidative stress. A comparative gene ontology analysis of S. oralis WT and SpxB-KO strains revealed that during infection, down regulated genes were closely related to the processes involved in the host defense reaction and up regulated genes were related with the cellular stress responses. Using qPCR analysis, we also confirmed the same pattern of expression changes such as TNF-α, IL-6 and Egr-1. Furthermore, supernatant from SpxB-KO could not suppress the expression of TNF-α in macrophages stimulated with LPS. These findings suggested that H2O2 production from S. oralis leads to the suppression of inflammatory responses and NF-κB signaling pathways in macrophages as well as the induction of the oxidative stress response. We concluded that streptococcal H2O2 production has the beneficial effects of modulating the innate immune response, thereby stabilizing streptococcal colonization at the mucosal surface and even in the bloodstream leading to cardiovascular disease after invasion, in addition to the commensal role to compete other bacterial species as initial colonizer at oral cavity.
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Perfilación de la Expresión Génica/métodos , Peróxido de Hidrógeno/metabolismo , Macrófagos/metabolismo , Análisis de Secuencia por Matrices de Oligonucleótidos/métodos , Streptococcus oralis/metabolismo , Células 3T3 , Animales , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Western Blotting , Línea Celular , Análisis por Conglomerados , Proteína 1 de la Respuesta de Crecimiento Precoz/genética , Proteína 1 de la Respuesta de Crecimiento Precoz/metabolismo , Ontología de Genes , Interacciones Huésped-Patógeno , Interleucina-6/genética , Interleucina-6/metabolismo , Lipopolisacáridos/farmacología , Macrófagos/efectos de los fármacos , Macrófagos/microbiología , Ratones , Ratones Endogámicos BALB C , Mutación , Piruvato Oxidasa/genética , Piruvato Oxidasa/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Streptococcus oralis/genética , Streptococcus oralis/fisiología , Factor de Necrosis Tumoral alfa/genética , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
Melatonin produced by the pineal gland suppresses inflammatory responses in innate immune cells. However, the mechanism of how melatonin affects inflammatory gene regulation remains unclear. Here we performed comprehensive microarray analysis combined with transcription factor binding site (TFBS) analysis using LPS-induced mouse macrophages to investigate the effect of melatonin treatment. The results showed that melatonin preferentially downregulated interferon regulatory factors (IRFs) and signal transducers and activators of transcription (STATs) related signaling. The results also showed that melatonin strongly suppressed virus infection related gene expression. Furthermore, TFBS analysis implicated that melatonin downregulated the binding activity of hypoxia inducible factors (HIFs), following destabilizing actin cytoskeleton which are indispensable for induction of the TRIF-dependent signaling pathway. Indeed, it was demonstrated that melatonin treatment caused impaired phagocytosis in macrophages. Thus, melatonin regulates inflammatory responses by inhibiting specific subsets of transcription factors (TFs) by disrupting actin dynamics in the macrophage.
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Actinas/metabolismo , Perfilación de la Expresión Génica/métodos , Macrófagos/efectos de los fármacos , Melatonina/farmacología , Análisis de Secuencia por Matrices de Oligonucleótidos/métodos , Animales , Antioxidantes/farmacología , Análisis por Conglomerados , Citocinas/genética , Citocinas/metabolismo , Ensayo de Inmunoadsorción Enzimática , Ontología de Genes , Lipopolisacáridos/farmacología , Macrófagos/inmunología , Macrófagos/metabolismo , Ratones , Microscopía Fluorescente , Polimerizacion/efectos de los fármacos , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Factores de TiempoRESUMEN
OBJECTIVES: Candidalysin (CL), a hydrophobic peptide toxin secreted by Candida albicans, is a key virulence factor that contributes to cytolysis, tissue damage, and immune activation. CL is thought to exert some of its biological activities, including IL-1ß production, through the activation of the NLRP3-inflammasome pathway. To date, the mechanism by which CL affects human NLRP3 is not fully understood. We investigated specific activities of synthetic CL peptides using human-derived NLRP3-deficient cells. METHODS: Two distinct synthetic CL peptide solutions were prepared: CLd, with CL completely solubilized as nanoparticles in dimethyl sulfoxide, and CLw, with CL partly solubilized in water, and including insoluble microparticles. THP-1 human monocytic cells and NLRP3-deficient THP-1 cells were differentiated into macrophages and stimulated with these peptide solutions. Cell membrane damage, lactate dehydrogenase release, IL-1ß production, and caspase-1 activation in stimulated cells were subsequently evaluated. RESULTS: Both CLd and CLw exhibited cytotoxic activities independent of NLRP3. Importantly, CLd induced IL-1ß production and caspase-1 activation in an NLRP3-independent manner, whereas these activities in CLw-stimulated cells were entirely NLRP3-dependent, suggesting that the NLRP3-dependent response might be triggered by insoluble microparticles. CONCLUSIONS: Our results demonstrate that inherent CL activities can cause cell damage and IL-1ß production in an NLRP3-independent manner. Our research advances the elucidation of the role of NLRP3 in CL biological activity, underscoring the necessity for further exploration of the precise mechanisms underlying the NLRP3-independent effects of CL and providing novel insights into the complexity of host-pathogen interactions.
Asunto(s)
Antineoplásicos , Proteína con Dominio Pirina 3 de la Familia NLR , Humanos , Proteína con Dominio Pirina 3 de la Familia NLR/metabolismo , Inflamasomas/metabolismo , Inflamasomas/farmacología , Macrófagos/metabolismo , Antineoplásicos/metabolismo , Antineoplásicos/farmacología , Caspasas/metabolismo , Caspasas/farmacologíaRESUMEN
OBJECTIVES: The possibility of leakage of CD34+ bone marrow stem cells from the peripheral blood into the coelomic cavity and the capability of coelomic fluid factors to induce their non-hematogenous differentiation were examined by immunocytochemistry (ICC). STUDY DESIGN: Body cavity fluid smears from 12 and 18 patients with and without cancer, respectively, were processed for double immunoperoxidase or double fluorescent ICC methods using antibodies against CD34, CD14, CD16, CD68, AE1/AE3, epidermal growth factor receptor (EGFR), D2-40, and CA125. RESULTS: Heavily irritated exudative fluid from 6 patients with or without cancer contained a few small round cells positive for CD34. Some of them co-expressed myeloid or monocytic markers such as CD14, CD68 or CD16. Some of the CD34+ cells also co-expressed AE1/AE3 or EGFR. In addition, D2-40 and CA125 were also demonstrated though the expression of the latter was quite sporadic. CONCLUSION: These findings support the concept that CD34+ stem cells can be released into irritated body cavity fluid and the possibility of subsequent differentiation to a non-hematogenous lineage under the influence of local humoral factors, in agreement with our previous in vitro experiments. The possibility of such a phenomenon should be kept in mind when body cavity fluid specimens are analyzed by ICC for diagnostic purposes.
Asunto(s)
Líquidos Corporales/citología , Células Madre Hematopoyéticas/citología , Neoplasias/patología , Adulto , Anciano , Anciano de 80 o más Años , Antígenos CD34/metabolismo , Femenino , Células Madre Hematopoyéticas/metabolismo , Humanos , Inmunohistoquímica , Inmunofenotipificación , Masculino , Persona de Mediana EdadRESUMEN
Candidalysin, a peptide toxin produced specifically from hyphae of Candida albicans, plays a crucial role in C. albicans pathogenesis in the oral cavity and vagina. Synthetic peptides have been widely used in previous studies to investigate the bioactivity of candidalysin. Although the solubility of the peptide, which is expected to have a hydrophobic property, has not been well characterized, candidalysin solutions are usually prepared in water. In this study, we prepared the synthetic peptide candidalysin in water (CLw) or in dimethyl sulfoxide (CLd) and compared their cytotoxicity and interleukin (IL)-1ß-producing activity to determine whether the activity of the peptide would be affected. In addition, we evaluated whether the NOD-like receptor family pyrin domain-containing 3 (NLRP3) inflammasome pathway or other pathways were involved in their activities. Unexpectedly, we found that CLw was not completely solubilized and contained abundant insoluble microparticles. CLw was active at comparably high concentrations (≥ 10 µM). In contrast, CLd is completely solubilized and sufficiently active at low concentrations, that is, 1 µM or less. CLw showed weak cytotoxicity and NLRP3-dependent and cathepsin B-dependent IL-1ß-producing activity, whereas CLd showed strong cytotoxicity and cathepsin B-dependent IL-1ß-producing activity. Fractionation of CLw revealed that NLRP3-dependent activity was caused by insoluble microparticles. Furthermore, nanoparticle tracking of CLd revealed that the peptide was present as nanoparticles with a size of 96 nm. CLw contained a small amount of such nanoparticles. Thus, the bioactivities of the synthetic peptide candidalysin, especially the IL-1ß-producing activity, are affected by the solubility of the peptide depending on the solvent employed. The NLRP3-dependent activity of the synthetic peptide is caused by insoluble microparticles and may not be the intrinsic activity of candidalysin.
Asunto(s)
Catepsina B , Proteína con Dominio Pirina 3 de la Familia NLR , Candida albicans/metabolismo , Catepsina B/metabolismo , Proteínas Fúngicas , Inflamasomas/metabolismo , Interleucina-1beta/metabolismo , Proteína con Dominio Pirina 3 de la Familia NLR/metabolismo , Solubilidad , Agua/metabolismoRESUMEN
Almost cases of Posterolateral Corner (PLC) injuries are combined injuries involving the anterior or posterior cruciate ligament. Although numerous techniques of PLC reconstruction have been reported, it is unknown whether these techniques reconstruct PLC sufficiently. The anatomy of PLC is complex of tendon-muscle and ligament. The major structures are the fibular collateral ligament (FCL) and the Popliteus Tendon Complex. The latter comprises the popliteus tendon-muscle and the popliteo-fibular ligament (PFL). The FCL and PFL are static stabilizers, whereas the popliteus tendon-muscle is a dynamic stabilizer. The most of current PLC reconstructions statically restore all component parts, therefore not true "anatomically". We describe an operative technique to reconstruct PLC anatomically. Our technique preserves dynamic stability of the popliteus tendon-muscle and reconstructs the PFL and FCL selectively.Semitendinosus tendon is harvested, and almost used for the anterior or posterior cruciate ligament reconstruction. Gracilis tendon or contralateral semitendinosus tendon is used for PLC. Femoral bone tunnel for FCL is prepared at anatomical insertion. Fibular bone tunnel is prepared to connect PFL insertion with FCL insertion. One end of the graft is sutured to the popliteus tendon. The other end is passed though the fibular tunnel, and fixed at the femoral tunnel. The interference screws are used at each tunnel. One half of the graft composes PFL part, the other half composes FCL part. Advantages of this technique are preservation of dynamic popliteus tendon-muscle function, and simplifying preparation.
RESUMEN
OBJECTIVES: Sjögren's syndrome (SS) is a chronic autoimmune disease characterized by inflammatory lesions in the salivary and lacrimal glands, which are caused by distinct lymphocytic infiltrates. Female non-obese diabetic (NOD) mice spontaneously develop inflammatory lesions of the salivary glands with SS-like pathological features. Previous studies have shown that MyD88, a crucial adaptor protein that activates innate immune signaling, affects lymphocytic infiltration, but its detailed role remains unclear. In this study, we investigated the role of MyD88 through gene expression profiling in the early phase of pathogenesis in the salivary glands of female NOD mice. METHODS: Submandibular glands collected from 10-week-old female wild-type and Myd88-deficient NOD mice were used for RNA preparation, followed by microarray analysis. The microarray dataset was analyzed to identify Myd88-dependent differentially expressed genes (DEGs). Data generated were used for GO enrichment, KEGG pathway, STRING database, and INTERFEROME database analyses. RESULTS: Myd88 deficiency was found to affect 230 DEGs, including SS-associated genes, such as Cxcl9 and Bpifa2. Most of the DEGs were identified as being involved in immunological processes. KEGG pathway analysis indicated that the DEGs were putatively involved in autoimmune diseases, such as systemic lupus erythematosus and rheumatoid arthritis. Furthermore, the DEGs included 149 interferon (IFN)-regulated genes. CONCLUSIONS: MyD88 is involved in the expression of specific genes associated with IFN-associated immunopathological processes in the salivary glands of NOD mice. Our findings are important for understanding the role of MyD88-dependent innate immune signaling in SS manifestation.
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Diabetes Mellitus , Glándulas Salivales , Animales , Modelos Animales de Enfermedad , Femenino , Perfilación de la Expresión Génica , Ratones , Ratones Endogámicos NOD , Análisis por Micromatrices , Factor 88 de Diferenciación Mieloide/genética , Enfermedades de Inmunodeficiencia Primaria , Glándulas Salivales/metabolismoRESUMEN
Introduction. Food allergies (FAs) occur due to intestinal immune dysfunction elicited by dysbiotic conditions. It was previously determined by us that Citrobacter species propagate in the faeces of mice with FAs and worsen allergic symptoms by inducing the allergenic cytokine IL-33. Dendritic cells can play important roles in regulation of FA responses.Hypothesis. Citrobacter species propagating in intestines of mice worsen allergic symptoms by stimulating dendritic cells to induce IL-33 expression.Aim. The aim of the present study was to analyse whether C. koseri stimulates dendritic cells to induce IL-33 expression.Methodology. IL-33 expression was evaluated in a DC2.4 mouse dendritic cell line stimulated by live or heat-inactivated C. koseri JCM1658, ATP, LPS extracted from C. koseri JCM1658 or other enterobacteria by real-time PCR. The ATP concentration and number of live bacteria in the culture supernatant were measured simultaneously.Results. Live C. koseri JCM1658 induced higher levels of IL-33 expression than other enterobacteria tested, but such a response was not elicited by heat-inactivated C. koseri JCM1658. LPS extracted from C. koseri JCM1658 did not induce IL-33 expression and suppressed live C. koseri JCM1658-induced IL-33 expression via the activation of Toll-like receptor 4 signalling. Furthermore, ATP produced by C. koseri JCM1658 stimulated dendritic cells to induce IL-33 expression by stimulating the P2X7 receptor, and LPS attenuated extracellular ATP-induced IL-33 expression. C. koseri JCM1658 was observed to proliferate more vigorously and produce more ATP than other enterobacteria.Conclusion. C. koseri acts as an allergenic bacterium through ATP production, stimulating dendritic cells to induce IL-33 expression, while LPS released from inactivated C. koseri JCM1658 attenuates this allergenicity.
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Adenosina Trifosfato/metabolismo , Citrobacter koseri/patogenicidad , Infecciones por Enterobacteriaceae , Hipersensibilidad a los Alimentos , Interleucina-33/inmunología , Animales , Línea Celular , Células Dendríticas/microbiología , Infecciones por Enterobacteriaceae/inmunología , Infecciones por Enterobacteriaceae/microbiología , Hipersensibilidad a los Alimentos/inmunología , Hipersensibilidad a los Alimentos/microbiología , Ratones , Transducción de SeñalRESUMEN
Toll-like receptors (TLRs) are important innate immune receptors that sometimes cause excessive inflammatory responses and a perpetuated inflammatory loop that can be involved in inflammatory and autoimmune diseases. TLR2 recognizes bacterial lipoproteins in association with TLR1 or TLR6, and triggers inflammatory responses through activation of the transcription factor NF-κB. Naringenin, a type of citrus flavonoid, has been shown to possess anti-inflammatory properties, but its detailed action against TLR2 remains to be fully elucidated. The present study was designed to determine whether naringenin affects the inflammatory responses triggered by TLR2. Naringenin inhibited proinflammatory cytokine production and attenuated NF-κB activation in cells stimulated with a synthetic triacylated-type lipopeptide known as a TLR2/TLR1 ligand, as well as a synthetic diacylated-type lipopeptide known as a TLR2/TLR6 ligand. Moreover, a similar inhibitory effect was observed in cells stimulated with a crude lipophilic fraction extracted from Staphylococcus aureus cell walls and in cells stimulated with S. aureus cells. Furthermore, we showed that such an effect is caused by inhibition of TLR2 clustering in lipid rafts on the cell membrane. These results suggest that naringenin suppresses the inflammatory responses induced by TLR2 signal transduction. Our findings indicate a novel anti-inflammatory property of naringenin, mediated through the regulation of cell surface TLR2 functioning.
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Mast cells and basophils are central players in allergic reactions triggered by immunoglobulin E (IgE). They have intracellular granules containing allergic mediators (e.g., histamine, serotonin, inflammatory cytokines, proteases and ß-hexosaminidase), and stimulation by IgE-allergen complex leads to the release of such allergic mediators from the granules, that is, degranulation. Mast cells are residents of mucosal surfaces, including those of nasal and oral cavities, and play an important role in the innate defense system. Members of the mitis group streptococci such as Streptococcus oralis, are primary colonizers of the human oral cavity. They produce hydrogen peroxide (H2O2) as a by-product of sugar metabolism. In this study, we investigated the effects of streptococcal infection on RBL-2H3 mast cell/basophil cell line. Infection by oral streptococci did not induce degranulation of the cells. Stimulation of the RBL-2H3 cells with anti-dinitrophenol (DNP) IgE and DNP-conjugated human serum albumin triggers degranulation with the release of ß-hexosaminidase. We found that S. oralis and other mitis group streptococci inhibited the IgE-triggered degranulation of RBL-2H3 cells. Since mitis group streptococci produce H2O2, we examined the effect of S. oralis mutant strain deficient in producing H2O2, and found that they lost the ability to suppress the degranulation. Moreover, H2O2 alone inhibited the IgE-induced degranulation. Subsequent analysis suggested that the inhibition of degranulation was related to the cytotoxicity of streptococcal H2O2. Activated RBL-2H3 cells produce interleukin-4 (IL-4); however, IL-4 production was not induced by streptococcal H2O2. Furthermore, an in vivo study using the murine pollen-induced allergic rhinitis model suggested that the streptococcal H2O2 reduces nasal allergic reaction. These findings reveal that H2O2 produced by oral mitis group streptococci inhibits IgE-stimulated degranulation by inducing cell death. Consequently, streptococcal H2O2 can be considered to modulate the allergic reaction in mucosal surfaces.
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Alérgenos/metabolismo , Hipersensibilidad/inmunología , Inmunoglobulina E/inmunología , Infecciones Estreptocócicas/tratamiento farmacológico , Alérgenos/inmunología , Animales , Basófilos/inmunología , Basófilos/microbiología , Basófilos/patología , Degranulación de la Célula/inmunología , Supervivencia Celular/inmunología , Dinitrofenoles/farmacología , Humanos , Peróxido de Hidrógeno/metabolismo , Hipersensibilidad/tratamiento farmacológico , Hipersensibilidad/patología , Inmunoglobulina E/metabolismo , Interleucina-4/genética , Interleucina-4/metabolismo , Mastocitos/inmunología , Mastocitos/microbiología , Mastocitos/patología , Ratones , Extractos Vegetales/química , Extractos Vegetales/farmacología , Albúmina Sérica Humana/inmunología , Albúmina Sérica Humana/metabolismo , Infecciones Estreptocócicas/inmunología , Streptococcus oralis/inmunología , Streptococcus oralis/patogenicidad , Azúcares/metabolismoRESUMEN
STUDY DESIGN: A preliminary report of a new operative method termed selective laminoplasty after the preoperative diagnosis of the responsible level using spinal cord evoked potentials (SCEPs) in elderly patients with cervical spondylotic myelopathy. OBJECTIVE: To introduce the method and clinical results for selective laminoplasty. SUMMARY OF BACKGROUND DATA: Clinical results for conventional laminoplasty and anterior decompression and fusion guided by SCEPs have been reported. However, there have been no reports that consider SCEP results for selecting the optimal level in lamioplasty for cervical spondylotic myelopathy. METHODS: Seven elderly patients who underwent selective laminoplasty were followed for a minimum of 12 months. The T2-high-intensity area on magnetic resonance imaging, the responsible level detected by SCEPs, and the laminoplasty level were recorded. The operative time, intraoperative bleeding, clinical results including the Japanese Orthopaedic Association score, recovery rate, Nurick grading scale, and visual analog scale of axial pain were investigated preoperatively and postoperatively. RESULTS: The responsible intervertebral levels were at C3-C4 in 3 patients and at C4-C5 in 4 patients. These were identical for SCEP recorded after median nerve stimulation and transcranial electric stimulation. High-intensity area on T2-weighted magnetic resonance imaging was seen in 6 patients (3 at C3-C4 and 3 at C4-C5). The average operative time was 106 minutes and the average amount of bleeding was 20 mL. Neurologic recovery was achieved in all patients except 1 who had severe myelopathy. Visual analog scales of axial pain were 41.3 + or - 33.9 before surgery and 18.0 + or - 19.4 at final follow-up. The Japanese Orthopaedic Association score and the Nurick grade improved in 6 patients but did not change in 1 patient. CONCLUSIONS: Preliminary clinical results for selective laminoplasty were satisfactory in all but 1 case. Although long-term results are not yet available, we consider this method to be less invasive and capable of giving satisfactory clinical results and benefits for elderly patients.
Asunto(s)
Electrodiagnóstico/métodos , Potenciales Evocados/fisiología , Cuidados Preoperatorios/métodos , Compresión de la Médula Espinal/diagnóstico , Compresión de la Médula Espinal/cirugía , Espondilosis/cirugía , Factores de Edad , Anciano , Anciano de 80 o más Años , Vértebras Cervicales/patología , Vértebras Cervicales/fisiopatología , Vértebras Cervicales/cirugía , Descompresión Quirúrgica , Estimulación Eléctrica/métodos , Electrofisiología/métodos , Femenino , Humanos , Laminectomía/métodos , Imagen por Resonancia Magnética , Masculino , Nervio Mediano/fisiología , Procedimientos Quirúrgicos Mínimamente Invasivos/métodos , Procedimientos Neuroquirúrgicos/métodos , Proyectos Piloto , Valor Predictivo de las Pruebas , Compresión de la Médula Espinal/fisiopatología , Espondilosis/complicaciones , Vertebroplastia/métodosRESUMEN
A recent study of a rat model treated with grafted collagen filament (CF) after spinal cord transection showed dramatic recovery of motor function but did not report on the acute-stage phenomenon. In the present study, we describe molecular and histological aspects of the axonal regeneration process during the acute stage following spinal cord transection. The spinal cord of 8-week-old rats was completely transected, and a scaffold of almost the same size as the resected portion was implanted in the gap. Changes in the mRNA expression of four neurotrophic factors [nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), NT-3, and glial cell-derived neurotrophic factor (GDNF)] were analyzed after 72 h. The expression of BDNF and NT-3 mRNA increased significantly in the CF-grafted group compared to the nongrafted group. Immunostaining for BDNF and NT-3 revealed that cells positive for these neurotrophic factors extended along the collagen filaments in the CF-grafted group. Similarly, astrocytes extended into the collagen filament scaffold together with the neurotrophic factors and partly across a border line. These findings indicate that collagen filament helps to reduce scar tissue, supports the expression of neurotrophic factors, and serves as a scaffold for the outgrowth of regenerating axons.
Asunto(s)
Regeneración Tisular Dirigida/métodos , Regeneración Nerviosa/fisiología , Traumatismos de la Médula Espinal/fisiopatología , Animales , Astrocitos/metabolismo , Colágeno/metabolismo , Femenino , Implantes Experimentales , Factores de Crecimiento Nervioso/genética , Factores de Crecimiento Nervioso/metabolismo , Ratas , Ratas Endogámicas F344RESUMEN
Cervical flexion myelopathy is thought to arise following compression of the spinal cord by vertebrae or intervertebral discs and dura mater, or from overstretching of the spinal cord induced by cervical spinal flexion. However, the influence of spinal flexion on the spinal cord and the detailed origins of this disease are unknown. In this article the authors report a case of cervical flexion myelopathy in which dynamic electrophysiological examination was performed using an epidural electrode. This investigation showed the real-time influence of flexion of the cervical spine on spinal cord function. This technique was considered to be useful for diagnosis and in decision making for treatment. Anterior fusion was the optimal surgical method for treating this disease.
Asunto(s)
Compresión de la Médula Espinal/diagnóstico , Adulto , Electrofisiología , Humanos , Masculino , Compresión de la Médula Espinal/diagnóstico por imagen , Compresión de la Médula Espinal/cirugía , Tomografía Computarizada por Rayos XRESUMEN
OBJECT: The goal of this study was to perform a biomechanical study of cervical flexion myelopathy (CFM) using a finite element method. METHODS: A 3D finite element model of the spinal cord was established consisting of gray matter, white matter, and pia mater. After the application of semi-static compression, the model underwent anterior flexion to simulate CFM. The flexion angles used were 5 degrees and 10 degrees , and stress distributions inside the spinal cord were then evaluated. RESULTS: Stresses on the spinal cord were very low under semi-static compression but increased after 5 degrees of flexion was applied. Stresses were concentrated in the gray matter, especially the anterior and posterior horns. The stresses became much higher after application of 10 degrees of flexion and were observed in the gray matter, posterior funiculus, and a portion of the lateral funiculus. CONCLUSIONS: The 5 degrees model was considered to represent the mild type of CFM. This type corresponds to the cases described in the original report by Hirayama and colleagues. The main symptom of this type of CFM is muscle atrophy and weakness caused by the lesion of the anterior horn. The 10 degrees model was considered to represent a severe type of CFM and was associated with lesions in the posterior fand lateral funiculi. This type of CFM corresponds to the more recently reported clinical cases with combined long tract signs and sensory disturbance.
Asunto(s)
Vértebras Cervicales/fisiopatología , Análisis de Elementos Finitos , Compresión de la Médula Espinal/etiología , Enfermedades de la Columna Vertebral/complicaciones , Algoritmos , Células del Asta Anterior/fisiopatología , Fenómenos Biomecánicos , Simulación por Computador , Humanos , Imagenología Tridimensional/métodos , Modelos Neurológicos , Piamadre/fisiopatología , Células del Asta Posterior/fisiopatología , Médula Espinal/fisiopatología , Estrés MecánicoRESUMEN
Oral streptococci, including cariogenic bacterium Streptococcus mutans, comprise a large percentage of human supragingival plaque, which contacts both tooth surfaces and gingiva. Eukaryotic cells are able to take up macromolecules and particles, including bacteria, by endocytosis. Increasing evidence indicates endocytosis may be used as an entry process by bacteria. We hypothesized that some endocytosed bacteria might survive and obtain nutrients, such as amino acids, until they are killed. To verify this hypothesis, we focused on bacterial utilization of branched-chain amino acids (BCAAs; isoleucine, leucine and valine) in host cells. A branched-chain aminotransferase, IlvE (EC 2.6.1.42), has been suggested to play an important role in internal synthesis of BCAAs in S. mutans UA159. Therefore, we constructed an ilvE-deficient S. mutans 109c strain and confirmed that it had similar growth behavior as reported previously. 14C radioactive leucine uptake assays showed that ilvE-deficient S. mutans took up more leucine both inside and outside of host cells. We further clarified that a relative decrease of BCAAs in host cells caused enhanced endocytic and autophagic activity. In conclusion, S. mutans is endocytosed by host cells and may survive and obtain nutrients, such as BCAAs, inside the cells, which might affect cellular functions of host cells.
Asunto(s)
Aminoácidos de Cadena Ramificada/metabolismo , Infecciones Estreptocócicas/microbiología , Infecciones Estreptocócicas/patología , Streptococcus mutans/metabolismo , Autofagosomas/metabolismo , Proteínas Bacterianas/genética , Línea Celular Tumoral , Endocitosis , Células Epiteliales/microbiología , Células Epiteliales/patología , Humanos , Leucina/metabolismo , Mutación , Streptococcus mutans/genética , Streptococcus mutans/crecimiento & desarrollo , Transaminasas/genéticaRESUMEN
A significant amount of evidence has been accumulated to show that Toll-like receptors (TLRs) function as sensors for microbial invasion. However, little is known about how signalling triggered by TLRs leads to the phagocytosis of pathogens. This study was designed to determine whether stimulation of TLR2 mainly with the lipopeptide FSL-1 plays a role in the phagocytosis of pathogens by macrophages. FSL-1 enhanced the phagocytosis of Escherichia coli to a markedly greater extent than it did that of Staphylococcus aureus, but did not enhance the phagocytosis of latex beads. FSL-1 stimulation resulted in enhanced phagocytosis of bacteria by macrophages from TLR2(+/+) mice but not by those from TLR2(-/-) mice. Chinese hamster ovary cells stably expressing TLR2 failed to phagocytose these bacteria, but the cells expressing CD14 did. FSL-1 induced upregulation of the expression of phagocytic receptors, including MSR1, CD36, DC-SIGN and Dectin-1 in THP-1 cells. Human embryonic kidney 293 cells transfected with DC-SIGN and MSR1 phagocytosed these bacteria. These results suggest that the FSL-1-induced enhancement of phagocytosis of bacteria by macrophages may be explained partly by the upregulation of scavenger receptors and the C-type lectins through TLR2-mediated signalling pathways, and that TLR2 by itself does not function as a phagocytic receptor.