RESUMEN
BACKGROUND: Epidemiological studies have demonstrated that periodontitis is an independent risk factor for chronic obstructive pulmonary disease (COPD). However, the mechanism underlying the association between these two diseases remains unclear. The lung microbiota shares similarities with the oral microbiota, and there is growing evidence to suggest that the lung microbiome could play a role in the pathogenesis of COPD. This study aimed to investigate whether periodontal pathogens could contribute to the pathogenesis of COPD in a mouse model. METHODS: We established mouse models with oral infection by typical periodontal pathogens, porphyromonas gingivalis (Pg group) or fusobacterium nucleatum (Fn group), over a three-month period. Mice that did not receive oral infection were set as the control group (C group). We assessed the level of alveolar bone resorption, lung function, and histological changes in the lungs of the mice. Additionally, we measured the levels of inflammatory factors and tissue damage associated factors in the lung tissues. RESULTS: Lung function indices, including airway resistance, peak inspiratory/expiratory flow and expiratory flow-50%, were significantly reduced in the Fn group compared to the C group. Additionally, histological examination revealed an increased number of inflammatory cells and bullae formation in the lung tissue sections of the Fn group. Meanwhile, levels of inflammatory factors such as IL-1ß, IL-6, IFN-γ, and TNF-α, as well as tissue damage associated factors like matrix metalloproteinase-8 and neutrophil elastase, were significantly elevated in the lung tissue of the Fn group in comparison to the C group. The Pg group also showed similar but milder lung changes compared to the Fn group. Pg or Fn could be detected in the lungs of both oral infected groups. CONCLUSION: The results indicated that oral periodontal pathogens infection could induce COPD-like lung changes in mice, and they may play a biological role in the association between periodontitis and COPD.
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Modelos Animales de Enfermedad , Fusobacterium nucleatum , Porphyromonas gingivalis , Enfermedad Pulmonar Obstructiva Crónica , Animales , Enfermedad Pulmonar Obstructiva Crónica/microbiología , Enfermedad Pulmonar Obstructiva Crónica/patología , Enfermedad Pulmonar Obstructiva Crónica/fisiopatología , Enfermedad Pulmonar Obstructiva Crónica/complicaciones , Ratones , Pérdida de Hueso Alveolar/microbiología , Pérdida de Hueso Alveolar/patología , Pulmón/patología , Pulmón/microbiología , Periodontitis/microbiología , Periodontitis/patología , Periodontitis/complicaciones , Masculino , Infecciones por Bacteroidaceae/complicaciones , Infecciones por Bacteroidaceae/microbiología , Infecciones por Bacteroidaceae/patología , Infecciones por Fusobacterium/complicaciones , Infecciones por Fusobacterium/microbiología , Infecciones por Fusobacterium/patología , Ratones Endogámicos C57BLRESUMEN
OBJECTIVE: Implication of human caspase-4 in periodontitis and in sensing periodontal pathogens by gingival epithelial cells (GECs) is unclear. This study aimed to determine caspase-4 and interleukin (IL)-18 expressions in gingival tissues affected with periodontitis and to investigate caspase-4 involvement in mediating innate immune responses in GECs. DESIGN: Ex vivo, caspase-4 and IL-18 expressions in gingival biopsies, obtained from healthy participants with periodontitis or clinically healthy gingiva (N = 20 each), were determined by immunohistochemistry. In vitro, caspase-4 activation in cultured GECs stimulated with Porphyromonas gingivalis or Fusobacterium nucleatum was analyzed by immunoblotting. mRNA expressions of human ß-defensin-2 (hBD-2), IL-8, and IL-18 in stimulated GECs in the presence or absence of a caspase-4 inhibitor were assayed by RT-qPCR. RESULTS: Ex vivo, compared with healthy gingival epithelium, the epithelium affected with periodontitis displayed a significant decrease in caspase-4 expression (P = 0.015), whereas IL-18 expression was significantly increased (P = 0.012). Moreover, the expression of caspase-4, but not IL-18, was found to be a predictor of periodontitis (P = 0.007). In vitro, caspase-4 was activated in cultured GECs challenged with P. gingivalis, but not F. nucleatum. mRNA upregulations of hBD-2, IL-8, and IL-18 upon P. gingivalis stimulation were significantly reduced when caspase-4 was inhibited (P < 0.05), whereas the inhibitor failed to suppress those inductions by F. nucleatum. CONCLUSIONS: Caspase-4 expression is diminished in the epithelium affected with periodontitis while that of IL-18 is enhanced. Caspase-4 activation in P. gingivalis-infected GECs upregulates the three innate immune effector molecules, suggesting a possible sensing mechanism of caspase-4 in GECs in periodontal disease pathogenesis.
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Infecciones por Bacteroidaceae , Caspasas Iniciadoras , Encía , Periodontitis , Porphyromonas gingivalis , Infecciones por Bacteroidaceae/enzimología , Infecciones por Bacteroidaceae/microbiología , Infecciones por Bacteroidaceae/patología , Caspasas Iniciadoras/biosíntesis , Células Cultivadas , Epitelio/enzimología , Epitelio/microbiología , Epitelio/patología , Encía/enzimología , Encía/microbiología , Encía/patología , Humanos , Interleucina-18/biosíntesis , Interleucina-8/biosíntesis , Periodontitis/enzimología , Periodontitis/microbiología , Periodontitis/patología , Porphyromonas gingivalis/metabolismo , ARN Mensajero/metabolismoRESUMEN
Purpose: The present study focused on exploring the associations of Porphyromonas gingivalis (P. gingivalis) infection and low Beclin1 expression with clinicopathological parameters and survival of esophageal squamous cell carcinoma (ESCC) patients, so as to illustrate its clinical significance and prognostic value. Methods: Immunohistochemistry (IHC) was used to detect P. gingivalis infection status and Beclin1 expression in 370 ESCC patients. The chi-square test was adopted to illustrate the relationship between categorical variables, and Cohen's kappa coefficient was used for correlation analysis. Kaplan-Meier survival curves with the log-rank test were used to analyse the correlation of P. gingivalis infection and low Beclin1 expression with survival time. The effects of P. gingivalis infection and Beclin1 downregulation on the proliferation, migration and antiapoptotic abilities of ESCC cells in vitro were detected by Cell Counting Kit-8, wound healing and flow cytometry assays. For P. gingivalis infection of ESCC cells, cell culture medium was replaced with antibiotic-free medium when the density of ESCC cells was 70-80%, cells were inoculated with P. gingivalis at a multiplicity of infection (MOI) of 10. Result: P. gingivalis infection was negatively correlated with Beclin1 expression in ESCC tissues, and P. gingivalis infection and low Beclin1 expression were associated with differentiation status, tumor invasion depth, lymph node metastasis, clinical stage and prognosis in ESCC patients. In vitro experiments confirmed that P. gingivalis infection and Beclin1 downregulation potentiate the proliferation, migration and antiapoptotic abilities of ESCC cells (KYSE150 and KYSE30). Our results provide evidence that P. gingivalis infection and low Beclin1 expression were associated with the development and progression of ESCC. Conclusion: Long-term smoking and alcohol consumption causes poor oral and esophageal microenvironments and ESCC patients with these features were more susceptible to P. gingivalis infection and persistent colonization, and exhibited lower Beclin1 expression, worse prognosis and more advanced clinicopathological features. Our findings indicate that effectively eliminating P. gingivalis colonization and restoring Beclin1 expression in ESCC patients may contribute to preventation and targeted treatment, and yield new insights into the aetiological research on ESCC.
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Infecciones por Bacteroidaceae/microbiología , Beclina-1/metabolismo , Neoplasias Esofágicas/microbiología , Carcinoma de Células Escamosas de Esófago/microbiología , Porphyromonas gingivalis/aislamiento & purificación , Apoptosis , Infecciones por Bacteroidaceae/metabolismo , Infecciones por Bacteroidaceae/mortalidad , Infecciones por Bacteroidaceae/patología , Línea Celular Tumoral , Movimiento Celular , Proliferación Celular , Neoplasias Esofágicas/metabolismo , Neoplasias Esofágicas/mortalidad , Neoplasias Esofágicas/patología , Carcinoma de Células Escamosas de Esófago/metabolismo , Carcinoma de Células Escamosas de Esófago/mortalidad , Carcinoma de Células Escamosas de Esófago/patología , Femenino , Humanos , Masculino , Persona de Mediana Edad , PronósticoRESUMEN
The non-enzymatic addition of glucose (glycation) to circulatory and tissue proteins is a ubiquitous pathophysiological consequence of hyperglycemia in diabetes. Given the high incidence of periodontitis and diabetes and the emerging link between these conditions, it is of crucial importance to define the basic virulence mechanisms employed by periodontopathogens such as Porphyromonas gingivalis in mediating the disease process. The aim of this study was to determine whether glycated proteins are more easily utilized by P. gingivalis to stimulate growth and promote the pathogenic potential of this bacterium. We analyzed the properties of three commonly encountered proteins in the periodontal environment that are known to become glycated and that may serve as either protein substrates or easily accessible heme sources. In vitro glycated proteins were characterized using colorimetric assays, mass spectrometry, far- and near-UV circular dichroism and UV-visible spectroscopic analyses and SDS-PAGE. The interaction of glycated hemoglobin, serum albumin and type one collagen with P. gingivalis cells or HmuY protein was examined using spectroscopic methods, SDS-PAGE and co-culturing P. gingivalis with human keratinocytes. We found that glycation increases the ability of P. gingivalis to acquire heme from hemoglobin, mostly due to heme sequestration by the HmuY hemophore-like protein. We also found an increase in biofilm formation on glycated collagen-coated abiotic surfaces. We conclude that glycation might promote the virulence of P. gingivalis by making heme more available from hemoglobin and facilitating bacterial biofilm formation, thus increasing P. gingivalis pathogenic potential in vivo.
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Infecciones por Bacteroidaceae/metabolismo , Complicaciones de la Diabetes/fisiopatología , Eritrocitos/metabolismo , Hemo/metabolismo , Hemoglobinas/metabolismo , Periodontitis/microbiología , Porphyromonas gingivalis/patogenicidad , Animales , Infecciones por Bacteroidaceae/microbiología , Infecciones por Bacteroidaceae/patología , Glicosilación , Hemoproteínas/química , Hemoglobinas/química , Caballos , Periodontitis/patología , Porphyromonas gingivalis/aislamiento & purificación , Porphyromonas gingivalis/metabolismoRESUMEN
BACKGROUND: Prevotella histicola is a facultative oral pathogen that under certain conditions causes pathologies such as caries and periodontitis in humans. Prevotella spp. also colonize the oral cavity of horses and can cause disease, but P. histicola has not yet been identified. CASE PRESENTATION: A 12-year-old Tinker mare was referred to the clinic for persistent, malodorous purulent nasal discharge and quidding. Conservative antibiotic (penicillin), antiphlogistic (meloxicam), and mucolytic (dembrexine-hydrochloride) treatment prior to referral was unsuccessful and symptoms worsened. Oral examination, radiography, sino-/ rhinoscopy, and standing computed tomography revealed severe apical/ periapical infection of the upper cheek tooth 209 with accompanying unilateral sinonasal inflammation and conchal necrosis. The tooth exhibited extensive subocclusal mesial infundibular cemental hypoplasia and caries, and an occlusal fissure fracture. After mechanical debridement and thermoplastic resin filling of the spacious subocclusal carious infundibular lesion, the tooth was extracted intraorally. The sinusitis and conchal necrosis were treated transendoscopically. Selective bacteriological swab cultures of affected tooth roots and subsequent matrix-assisted laser desorption ionization-time of flight mass spectrometry showed an infection with the obligate anaerobic, Gram-negative bacterium P. histicola. Surgical intervention and adapted antibiotic therapy led to normal healing without complications. CONCLUSIONS: This study provides the first documented case of dental infection in a horse caused by P. histicola at once indicating necessity of more sufficient microbiological diagnostics and targeted antibiotic treatment in equine dental practice. This finding is also conducive to understand species-specific Prevotella diversity and cross-species distribution.
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Infecciones por Bacteroidaceae/diagnóstico por imagen , Infecciones por Bacteroidaceae/veterinaria , Enfermedades de los Caballos/diagnóstico por imagen , Prevotella/aislamiento & purificación , Animales , Infecciones por Bacteroidaceae/patología , Infecciones por Bacteroidaceae/cirugía , Mejilla/diagnóstico por imagen , Mejilla/microbiología , Mejilla/patología , Femenino , Enfermedades de los Caballos/patología , Enfermedades de los Caballos/cirugía , Caballos , Humanos , Tomografía Computarizada por Rayos X/veterinaria , Diente/diagnóstico por imagen , Diente/microbiología , Diente/patología , Extracción Dental/veterinaria , Resultado del TratamientoRESUMEN
Epidemiological data have shown that periodontal bacterial infection, periodontitis, and oral squamous cell carcinoma have close relationship on the disease progress and risk. However, the specific role of periodontal microbes and their mechanism in the development of oral squamous cell carcinoma is not yet clear. In our previous work, metagenomic Illumina Mi-seq analysis was used to identify tstructure and abundance of periodontital microbiome. Accoding to the results, we used Porphyromonas.spp. and Fusobacterium.spp. as the periodontitis positive microbiota; Neisseria.spp and Corynebacterium.spp as periodontitis negative microbiota (their average relative abundance were >5%). These representative strains of the above genus were used to infect OSCC cells to explore their effect on tumor cell biology behavior, and detect the expression level of the gene in related to inflammation, migration, invasion and cell cycle. We find that periodontitis positive correlated microbiota had a promoting effect on the development of oral squamous cell carcinoma in vitro by regulating mRNA and protein expression of IL-6, IL-8, MMP-9 and Cyclin-D1. Periodontitis negative correlated microbiota had suppression effect on the development of oral squamous cell carcinoma in vitro analysis.
Asunto(s)
Neoplasias de Cabeza y Cuello/microbiología , Microbiota , Periodontitis/microbiología , Carcinoma de Células Escamosas de Cabeza y Cuello/microbiología , Infecciones por Bacteroidaceae/complicaciones , Infecciones por Bacteroidaceae/microbiología , Infecciones por Bacteroidaceae/patología , Línea Celular Tumoral , Movimiento Celular/fisiología , Corynebacterium/genética , Corynebacterium/aislamiento & purificación , Infecciones por Fusobacterium/complicaciones , Infecciones por Fusobacterium/microbiología , Infecciones por Fusobacterium/patología , Fusobacterium nucleatum/genética , Fusobacterium nucleatum/aislamiento & purificación , Neoplasias de Cabeza y Cuello/metabolismo , Neoplasias de Cabeza y Cuello/patología , Humanos , Neisseria sicca/genética , Neisseria sicca/aislamiento & purificación , Infecciones por Neisseriaceae/complicaciones , Infecciones por Neisseriaceae/microbiología , Infecciones por Neisseriaceae/patología , Porphyromonas gingivalis/genética , Porphyromonas gingivalis/aislamiento & purificación , Carcinoma de Células Escamosas de Cabeza y Cuello/metabolismo , Carcinoma de Células Escamosas de Cabeza y Cuello/patologíaRESUMEN
Porphyromonas gingivalis (P. gingivalis) is a keystone periodontal pathogen associated with various digestive cancers. However, whether P. gingivalis can promote colorectal cancer and the underlying mechanism associated with such promotion remains unclear. In this study, we found that P. gingivalis was enriched in human feces and tissue samples from patients with colorectal cancer compared with those from patients with colorectal adenoma or healthy subjects. Cohort studies demonstrated that P. gingivalis infection was associated with poor prognosis in colorectal cancer. P. gingivalis increased tumor counts and tumor volume in the ApcMin/+ mouse model and increased tumor growth in orthotopic rectal and subcutaneous carcinoma models. Furthermore, orthotopic tumors from mice exposed to P. gingivalis exhibited tumor-infiltrating myeloid cell recruitment and a proinflammatory signature. P. gingivalis promoted colorectal cancer via NLRP3 inflammasome activation in vitro and in vivo. NLRP3 chimeric mice harboring orthotopic tumors showed that the effect of NLRP3 on P. gingivalis pathogenesis was mediated by hematopoietic sources. Collectively, these data suggest that P. gingivalis contributes to colorectal cancer neoplasia progression by activating the hematopoietic NLRP3 inflammasome. SIGNIFICANCE: This study demonstrates that the periodontal pathogen P. gingivalis can promote colorectal tumorigenesis by recruiting myeloid cells and creating a proinflammatory tumor microenvironment. GRAPHICAL ABSTRACT: http://cancerres.aacrjournals.org/content/canres/81/10/2745/F1.large.jpg.
Asunto(s)
Carcinogénesis/patología , Neoplasias Colorrectales/patología , Regulación Neoplásica de la Expresión Génica , Inflamasomas/inmunología , Proteína con Dominio Pirina 3 de la Familia NLR/fisiología , Células Madre Neoplásicas/patología , Porphyromonas gingivalis/patogenicidad , Animales , Apoptosis , Infecciones por Bacteroidaceae/complicaciones , Infecciones por Bacteroidaceae/inmunología , Infecciones por Bacteroidaceae/microbiología , Infecciones por Bacteroidaceae/patología , Carcinogénesis/inmunología , Carcinogénesis/metabolismo , Proliferación Celular , Neoplasias Colorrectales/inmunología , Neoplasias Colorrectales/metabolismo , Neoplasias Colorrectales/microbiología , Humanos , Inflamasomas/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Ratones Desnudos , Células Mieloides/inmunología , Células Mieloides/metabolismo , Células Mieloides/microbiología , Células Mieloides/patología , Células Madre Neoplásicas/inmunología , Células Madre Neoplásicas/metabolismo , Células Madre Neoplásicas/microbiología , Pronóstico , Tasa de Supervivencia , Células Tumorales Cultivadas , Microambiente Tumoral/inmunología , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Porphyromonas gingivalis is a major pathogen responsible for severe and chronic manifestations of periodontal disease, which is one of the most common infectious disorders of humans. Although human gingival epithelium prevents intrusions by periodontal bacteria, P. gingivalis is able to invade gingival epithelial cells. To study the dynamics and the fate of intracellular P. gingivalis, confocal laser scanning microscopy (CLSM) is a method of choice. Information gained with CLSM contains not only the number of P. gingivalis associated with gingival epithelial cells but also the bacterial localization on/inside the host cells, morphological change of host cells, and physical interaction between the bacteria and host organelle. In this chapter, we describe the protocols for microscopy techniques to morphologically study gingival epithelial cells infected by P. gingivalis.
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Infecciones por Bacteroidaceae/patología , Células Epiteliales/patología , Encía/patología , Enfermedades Periodontales/patología , Porphyromonas gingivalis/fisiología , Infecciones por Bacteroidaceae/microbiología , Técnicas de Cultivo de Célula/métodos , Línea Celular , Células Epiteliales/microbiología , Encía/citología , Encía/microbiología , Humanos , Microscopía Confocal/métodos , Enfermedades Periodontales/microbiología , Porphyromonas gingivalis/aislamiento & purificación , Coloración y Etiquetado/métodosRESUMEN
Chronic periodontitis is the most common periodontitis observed in adults. Recently, its association with systemic diseases such as ischemic heart-brain disease and diabetes has been pointed out. Porphyromonas gingivalis, a major causative bacterium of chronic periodontitis, has properties of adhering to blood vessels and inducing inflammation, and those properties are involved in the induction of vascular inflammation and promotion of atherosclerosis. Therefore, analysis of the interaction of P. gingivalis with vascular endothelial cells will contribute to an understanding of the link between periodontitis and vascular lesions.
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Infecciones por Bacteroidaceae/patología , Células Endoteliales/patología , Periodontitis/patología , Porphyromonas gingivalis/fisiología , Infecciones por Bacteroidaceae/microbiología , Técnicas de Cultivo de Célula/métodos , Línea Celular , Células Endoteliales/microbiología , Células Endoteliales de la Vena Umbilical Humana , Humanos , Óxido Nítrico/metabolismo , Periodontitis/microbiología , Factor de von Willebrand/metabolismoRESUMEN
Several studies have shown a clear association between periodontal disease and increased risk of cardiovascular disease. Porphyromonas gingivalis (Pg), a key oral pathogen, and its cell surface-expressed gingipains, induce oedema in a zebrafish larvae infection model although the mechanism of these vascular effects is unknown. Here, we aimed to determine whether Pg-induced vascular damage is mediated by gingipains. In vitro, human endothelial cells from different vascular beds were invaded by wild-type (W83) but not gingipain-deficient (ΔK/R-ab) Pg. W83 infection resulted in increased endothelial permeability as well as decreased cell surface abundance of endothelial adhesion molecules PECAM-1 and VE-cadherin compared to infection with ΔK/R-ab. In agreement, when transgenic zebrafish larvae expressing fluorescently labelled PECAM-1 or VE-cadherin were systemically infected with W83 or ΔK/R-ab, a significant reduction in adhesion molecule fluorescence was observed specifically in endothelium proximal to W83 bacteria through a gingipain-dependent mechanism. Furthermore, this was associated with increased vascular permeability in vivo when assessed by dextran leakage microangiography. These data are the first to show that Pg directly mediates vascular damage in vivo by degrading PECAM-1 and VE-cadherin. Our data provide a molecular mechanism by which Pg might contribute to cardiovascular disease.
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Infecciones por Bacteroidaceae/etiología , Cardiomegalia/etiología , Edema/etiología , Células Endoteliales/efectos de los fármacos , Cisteína-Endopeptidasas Gingipaínas/toxicidad , Porphyromonas gingivalis/patogenicidad , Animales , Animales Modificados Genéticamente , Antígenos CD/genética , Antígenos CD/metabolismo , Infecciones por Bacteroidaceae/genética , Infecciones por Bacteroidaceae/metabolismo , Infecciones por Bacteroidaceae/patología , Cadherinas/genética , Cadherinas/metabolismo , Permeabilidad Capilar/efectos de los fármacos , Cardiomegalia/genética , Cardiomegalia/metabolismo , Cardiomegalia/patología , Edema/genética , Edema/metabolismo , Edema/patología , Embrión no Mamífero , Células Endoteliales/metabolismo , Células Endoteliales/patología , Angiografía con Fluoresceína , Expresión Génica/efectos de los fármacos , Genes Reporteros , Cisteína-Endopeptidasas Gingipaínas/biosíntesis , Proteínas Fluorescentes Verdes/genética , Proteínas Fluorescentes Verdes/metabolismo , Humanos , Larva/efectos de los fármacos , Larva/microbiología , Molécula-1 de Adhesión Celular Endotelial de Plaqueta/genética , Molécula-1 de Adhesión Celular Endotelial de Plaqueta/metabolismo , Porphyromonas gingivalis/crecimiento & desarrollo , Porphyromonas gingivalis/metabolismo , Cultivo Primario de Células , Transducción de Señal , Pez CebraRESUMEN
Sphingolipids (SLs) are essential structural components of mammalian cell membranes. Our group recently determined that the oral anaerobe Porphyromonas gingivalis delivers its SLs to host cells and that the ability of P. gingivalis to synthesize SLs limits the elicited host inflammatory response during cellular infection. As P. gingivalis robustly produces outer membrane vesicles (OMVs), we hypothesized that OMVs serve as a delivery vehicle for SLs, that the SL status of the OMVs may impact cargo loading to OMVs, and that SL-containing OMVs limit elicited host inflammation similar to that observed by direct bacterial challenge. Transwell cell culture experiments determined that in comparison to the parent strain W83, the SL-null mutant elicited a hyperinflammatory immune response from THP-1 macrophage-like cells with elevated tumor necrosis factor alpha (TNF-α), interleukin 1ß (IL-1ß), and IL-6. Targeted assessment of Toll-like receptors (TLRs) identified elevated expression of TLR2, unchanged TLR4, and elevated expression of the adaptor molecules MyD88 and TRIF (Toll/IL-1 receptor domain-containing adaptor-inducing beta interferon) by SL-null P. gingivalis No significant differences in gingipain activity were observed in our infection models, and both strains produced OMVs of similar sizes. Using comparative two-dimensional gel electrophoresis, we identified differences in the protein cargo of the OMVs between parent and SL-null strain. Importantly, use of purified OMVs recapitulated the cellular inflammatory response observed in the transwell system with whole bacteria. These findings provide new insights into the role of SLs in P. gingivalis OMV cargo assembly and expand our understanding of SL-OMVs as bacterial structures that modulate the host inflammatory response.
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Infecciones por Bacteroidaceae/inmunología , Infecciones por Bacteroidaceae/microbiología , Macrófagos/inmunología , Porphyromonas gingivalis/inmunología , Porphyromonas gingivalis/metabolismo , Esfingolípidos/inmunología , Vesículas Transportadoras/inmunología , Infecciones por Bacteroidaceae/patología , Transporte Biológico , Interacciones Huésped-Patógeno , Inmunomodulación , Mutación , Porphyromonas gingivalis/genética , Proteómica/métodos , Esfingolípidos/metabolismo , Vesículas Transportadoras/metabolismoRESUMEN
Bone destruction in inflammatory osteolytic diseases including periodontitis is related to excessive activity of osteoclasts (OC), which originate from precursor cells of the myeloid lineage, termed osteoclast precursors (OCP). In contrast to ample knowledge that we currently have on mature OC, little is known about OCP and their regulation during bacterial infection. Therefore, this study aimed to identify and characterize OCP following chronic infection with a periodontal bacteria Porphyromonas gingivalis (Pg). We used a micro-osmotic pump to continually release Pg subcutaneously in a murine model. Two weeks after Pg infection, the frequency of CD11b+c-fms+Ly6Chi population is significantly elevated within the bone marrow, spleen and peripheral blood. In vitro and in vivo studies identified these cells as the OCP-containing population and Pg infection significantly enhanced the osteoclastogenic activity of these cells. Furthermore, mRNA sequencing analysis indicated a unique gene and pathway profile in CD11b+c-fms+Ly6Chi population following Pg infection, with changes in genes and pathways related to OC differentiation, cell proliferation and apoptosis, inflammatory response, phagocytosis and immunity, as well as antigen processing and presentation. Moreover, using IL-6 knockout mice, we found that IL-6 is important for Pg-induced accumulation of CD11b+c-fms+Ly6Chi population from the bone marrow and periphery. Our results provide new insights into the characterization and regulation of OCP following a chronic bacterial infection. This knowledge is relevant to the understanding of the pathogenesis of bacteria-induced bone loss, and to the identification of potential therapeutic targets of bone loss diseases.
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Infecciones por Bacteroidaceae/inmunología , Diferenciación Celular/inmunología , Osteoclastos/inmunología , Osteólisis/inmunología , Porphyromonas gingivalis/inmunología , Células Madre/inmunología , Animales , Infecciones por Bacteroidaceae/genética , Infecciones por Bacteroidaceae/patología , Diferenciación Celular/genética , Enfermedad Crónica , Modelos Animales de Enfermedad , Interleucina-6/genética , Interleucina-6/inmunología , Ratones , Ratones Noqueados , Osteoclastos/patología , Osteólisis/genética , Osteólisis/microbiología , Osteólisis/patología , Células Madre/patologíaRESUMEN
Epidemiological studies reveal significant associations between periodontitis and oral cancer. However, knowledge about the contribution of periodontal pathogens to oral cancer and potential regulatory mechanisms involved is limited. Previously, we showed that nisin, a bacteriocin and commonly used food preservative, reduced oral cancer tumorigenesis and extended the life expectancy in tumor-bearing mice. In addition, nisin has antimicrobial effects on key periodontal pathogens. Thus, the purpose of this study was to test the hypothesis that key periodontal pathogens (Porphyromonas gingivalis, Treponema denticola, and Fusobacterium nucleatum) promote oral cancer via specific host-bacterial interactions, and that bacteriocin/nisin therapy may modulate these responses. All three periodontal pathogens enhanced oral squamous cell carcinoma (OSCC) cell migration, invasion, tumorsphere formation, and tumorigenesis in vivo, without significantly affecting cell proliferation or apoptosis. In contrast, oral commensal bacteria did not affect OSCC cell migration. Pathogen-enhanced OSCC cell migration was mediated via integrin alpha V and FAK activation, since stably blocking alpha V or FAK expression abrogated these effects. Nisin inhibited these pathogen-mediated processes. Further, Treponema denticola induced TLR2 and 4 and MyD88 expression. Stable suppression of MyD88 significantly inhibited Treponema denticola-induced FAK activation and abrogated pathogen-induced migration. Together, these data demonstrate that periodontal pathogens contribute to a highly aggressive cancer phenotype via crosstalk between TLR/MyD88 and integrin/FAK signaling. Nisin can modulate these pathogen-mediated effects, and thus has therapeutic potential as an antimicrobial and anti-tumorigenic agent.
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Infecciones por Bacteroidaceae/tratamiento farmacológico , Carcinoma de Células Escamosas/tratamiento farmacológico , Neoplasias de la Boca/tratamiento farmacológico , Periodontitis/tratamiento farmacológico , Porphyromonas gingivalis/efectos de los fármacos , Probióticos/farmacología , Animales , Apoptosis , Infecciones por Bacteroidaceae/metabolismo , Infecciones por Bacteroidaceae/microbiología , Infecciones por Bacteroidaceae/patología , Carcinoma de Células Escamosas/metabolismo , Carcinoma de Células Escamosas/microbiología , Carcinoma de Células Escamosas/patología , Movimiento Celular , Proliferación Celular , Quinasa 1 de Adhesión Focal/genética , Quinasa 1 de Adhesión Focal/metabolismo , Humanos , Integrinas/genética , Integrinas/metabolismo , Ratones , Ratones Desnudos , Neoplasias de la Boca/metabolismo , Neoplasias de la Boca/microbiología , Neoplasias de la Boca/patología , Factor 88 de Diferenciación Mieloide/genética , Factor 88 de Diferenciación Mieloide/metabolismo , Periodontitis/metabolismo , Periodontitis/microbiología , Periodontitis/patología , Porphyromonas gingivalis/patogenicidad , Receptor Toll-Like 2/genética , Receptor Toll-Like 2/metabolismo , Células Tumorales Cultivadas , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Microbial dysbiosis in the upper digestive tract is linked to an increased risk of esophageal squamous cell carcinoma (ESCC). Overabundance of Porphyromonas gingivalis is associated with shorter survival of ESCC patients. We investigated the molecular mechanisms driving aggressive progression of ESCC by P. gingivalis. Intracellular invasion of P. gingivalis potentiated proliferation, migration, invasion, and metastasis abilities of ESCC cells via transforming growth factor-ß (TGFß)-dependent Drosophila mothers against decapentaplegic homologs (Smads)/Yes-associated protein (YAP)/Transcriptional coactivator with PDZ-binding motif (TAZ) activation. Smads/YAP/TAZ/TEA domain transcription factor1 (TEAD1) complex formation was essential to initiate downstream target gene expression, inducing an epithelial-mesenchymal transition (EMT) and stemness features. Furthermore, P. gingivalis augmented secretion and bioactivity of TGFß through glycoprotein A repetitions predominant (GARP) up-regulation. Accordingly, disruption of either the GARP/TGFß axis or its activated Smads/YAP/TAZ complex abrogated the tumor-promoting role of P. gingivalis. P. gingivalis signature genes based on its activated effector molecules can efficiently distinguish ESCC patients into low- and high-risk groups. Targeting P. gingivalis or its activated effectors may provide novel insights into clinical management of ESCC.
Asunto(s)
Infecciones por Bacteroidaceae/complicaciones , Neoplasias Esofágicas/patología , Carcinoma de Células Escamosas de Esófago/patología , Porphyromonas gingivalis/fisiología , Factor de Crecimiento Transformador beta/fisiología , Aciltransferasas , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Adulto , Anciano , Animales , Infecciones por Bacteroidaceae/metabolismo , Infecciones por Bacteroidaceae/mortalidad , Infecciones por Bacteroidaceae/patología , Células Cultivadas , Progresión de la Enfermedad , Drosophila , Neoplasias Esofágicas/metabolismo , Neoplasias Esofágicas/microbiología , Neoplasias Esofágicas/mortalidad , Carcinoma de Células Escamosas de Esófago/metabolismo , Carcinoma de Células Escamosas de Esófago/microbiología , Carcinoma de Células Escamosas de Esófago/mortalidad , Femenino , Estudios de Seguimiento , Células HCT116 , Humanos , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Desnudos , Persona de Mediana Edad , Transducción de Señal/fisiología , Proteínas Smad/metabolismo , Análisis de Supervivencia , Factores de Transcripción/metabolismo , Factor de Crecimiento Transformador beta/metabolismo , Proteínas Señalizadoras YAPRESUMEN
Recent epidemiological studies link Periodontal disease(PD) to age-related macular degeneration (AMD). We documented earlier that Porphyromonas gingivalis(Pg), keystone oral-pathobiont, causative of PD, efficiently invades human gingival epithelial and blood-dendritic cells. Here, we investigated the ability of dysbiotic Pg-strains to invade human-retinal pigment epithelial cells(ARPE-19), their survival, intracellular localization, and the pathological effects, as dysfunction of RPEs leads to AMD. We show that live, but not heat-killed Pg-strains adhere to and invade ARPEs. This involves early adhesion to ARPE cell membrane, internalization and localization of Pg within single-membrane vacuoles or cytosol, with some nuclear localization apparent. No degradation of Pg or localization inside double-membrane autophagosomes was evident, with dividing Pg suggesting a metabolically active state during invasion. We found significant downregulation of autophagy-related genes particularly, autophagosome complex. Antibiotic protection-based recovery assay further confirmed distinct processes of adhesion, invasion and amplification of Pg within ARPE cells. This is the first study to demonstrate invasion of human-RPEs, begin to characterize intracellular localization and survival of Pg within these cells. Collectively, invasion of RPE by Pg and its prolonged survival by autophagy evasion within these cells suggest a strong rationale for studying the link between oral infection and AMD pathogenesis in individuals with periodontitis.
Asunto(s)
Autofagosomas , Autofagia , Infecciones por Bacteroidaceae , Citosol , Porphyromonas gingivalis , Epitelio Pigmentado de la Retina , Vacuolas , Autofagosomas/metabolismo , Autofagosomas/microbiología , Autofagosomas/ultraestructura , Infecciones por Bacteroidaceae/metabolismo , Infecciones por Bacteroidaceae/microbiología , Infecciones por Bacteroidaceae/patología , Línea Celular , Citosol/metabolismo , Citosol/microbiología , Citosol/ultraestructura , Humanos , Porphyromonas gingivalis/metabolismo , Porphyromonas gingivalis/ultraestructura , Epitelio Pigmentado de la Retina/metabolismo , Epitelio Pigmentado de la Retina/microbiología , Epitelio Pigmentado de la Retina/ultraestructura , Vacuolas/microbiología , Vacuolas/patología , Vacuolas/ultraestructuraRESUMEN
The relationship of Porphyromonas gingivalis and oral squamous cell carcinoma (OSCC) has been studied for several years. Previous studies have focused on the direct effect of P. gingivalis on the activities of primary epithelial cells and OSCC cells. However, the immune system is responsible for mediating cancer development, whether P. gingivalis can affect oral cancer immunity has seldom been explored to date. In this study, we investigated the role of P. gingivalis in the immunoevasion of OSCC. We evaluated the effect of P. gingivalis on the phagocytosis of Cal-27 cells (OSCC cell line) by bone marrow-derived macrophages in vitro and studied the effect of P. gingivalis on the growth of OSCC and the polarization of tumor-associated macrophages in vivo. We found that P. gingivalis was able to inhibit the phagocytosis of Cal-27 cells by macrophages, and membrane-component molecules of P. gingivalis, such as proteins, were speculated to be the effector components. In addition, sustained infection with antibiotics-inactivated P. gingivalis promoted OSCC growth in mice and induced the polarization of macrophages into M2 tumor-associated macrophages, which mainly display protumor properties. Transcriptome analysis and quantitative RT-PCR revealed that P. gingivalis infection upregulated the expression of genes encoding protumor molecules in Cal-27 cells (suprabasin, IL-1R2, and CD47) and in macrophages (IL-1α, CCL-3, and CCL-5). Our in vitro and in vivo data suggest that P. gingivalis can promote immunoevasion of oral cancer by protecting cancer from macrophage attack. To our knowledge, the present study reveals a novel mechanism by which P. gingivalis promotes OSCC development.
Asunto(s)
Infecciones por Bacteroidaceae/inmunología , Macrófagos/inmunología , Neoplasias de la Boca/inmunología , Porphyromonas gingivalis/inmunología , Carcinoma de Células Escamosas de Cabeza y Cuello/inmunología , Escape del Tumor , Animales , Infecciones por Bacteroidaceae/microbiología , Infecciones por Bacteroidaceae/patología , Carcinogénesis/inmunología , Línea Celular Tumoral , Femenino , Regulación Neoplásica de la Expresión Génica/inmunología , Interacciones Huésped-Patógeno/inmunología , Humanos , Macrófagos/metabolismo , Ratones , Mucosa Bucal/inmunología , Mucosa Bucal/microbiología , Mucosa Bucal/patología , Neoplasias de la Boca/microbiología , Neoplasias de la Boca/patología , Fagocitosis/inmunología , RNA-Seq , Organismos Libres de Patógenos Específicos , Carcinoma de Células Escamosas de Cabeza y Cuello/microbiología , Carcinoma de Células Escamosas de Cabeza y Cuello/patología , Regulación hacia Arriba , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
OBJECTIVE: To analyse the citrulline level in the periodontium in association with the presence of or antibody levels against Aggregatibacter actinomycetemcomitans and Porphyromonas gingivalis. DESIGN: Gingival crevicular fluid (GCF), subgingival biofilm and blood serum were sampled from 98 subjects (26 with RA, 72 without RA (NoRA)). GCF was analyzed for the level of citrulline, for interleukin (IL)-1ß, IL-17, IL-10 and monocyte-chemoattractant protein (MCP)-1. Microorganisms were identified in subgingival biofilms. Antibodies againstP. gingivalis, and Aggregatibacter actinomycetemcomitans were quantified in serum. RESULTS: GCF citrulline level was the lowest (by trend) in NoRA group without periodontitis. In NoRA, but not in RA an association between GCF citrulline level and P. gingivalis antibody levels was found and the GCF citrulline levels were higher in P. gingivalis positive samples. Any association of A. actinomycetemcomitans with GCF citrulline level did not exist. A model of univariate variance analysis (p = 0.001) showed a dependence of GCF citrulline level from the number of sites with PD (probing depth) ≥5 mm (p = 0.003) and the GCF MCP-1/CCL2 level (p = 0.019). Compared with NoRA in RA the number of teeth was lower, the number of sites with PD ≥ 5 mm was less, GCF levels of interleukin-17 and MCP-1/CCL2 were higher and those of IL-10 lower. Yeasts were only cultured in 15 RA patients (p < 0.001). CONCLUSION: Citrullination in periodontium might be associated with P. gingivalis supporting the potential role as a trigger in the development of RA. Pathogenesis of periodontal disease in RA patients seems to differ from that in NoRA and should be investigated further.
Asunto(s)
Artritis Reumatoide/complicaciones , Citrulinación , Citrulina/análisis , Periodontitis/microbiología , Periodoncio/química , Aggregatibacter actinomycetemcomitans , Infecciones por Bacteroidaceae/patología , Líquido del Surco Gingival , Humanos , Periodoncio/microbiología , Porphyromonas gingivalis/patogenicidadRESUMEN
BACKGROUND AND OBJECTIVE: Excessive osteoclast activity is a major characteristic of pathogenic bone loss in inflammatory bone diseases including periodontitis. However, beyond the knowledge that osteoclasts are differentiated from the monocyte/macrophage lineage and share common ancestry with macrophages and DC, the nature and function of osteoclast precursors are not completely understood. Furthermore, little is known about how osteoclast precursors respond to bacterial infection in vivo. We have previously demonstrated in vitro that the periodontal pathogen Porphyromonas gingivalis (Pg) plays a biphasic role on the receptor activator of nuclear factor kappa B ligand (RANKL)-induced osteoclast differentiation. In this study, we investigated the in vivo effect of Pg infection on the regulation of osteoclast precursors, using a mouse calvarial infection model. METHODS AND RESULTS: C57BL/6 wild-type and the myeloid differentiation factor 88 knockout (MyD88-/- ) mice were infected with Pg by calvarial injection. Local and systemic bone loss, and the number and function of CD11b+ c-fms+ cells from bone marrow and spleen were analyzed. Our results show that Pg infection induces localized inflammatory infiltration and osteoclastogenesis, as well as increased number and osteoclastogenic potential of CD11b+ c-fms+ osteoclast precursors in the bone marrow and periphery. We also show that CD11b+ c-fms+ RANK+ and CD11b+ c-fms+ RANK- are precursors with similar osteoclastogenic and pro-inflammatory potentials. In addition, CD11b+ c-fms+ cells exhibit an antigen-specific T-cell immune-suppressive activity, which are increased with Pg infection. Moreover, we demonstrate that MyD88 is involved in the regulation of osteoclast precursors upon Pg infection. CONCLUSIONS: In this study, we demonstrate an enhanced dual function of osteoclast precursors following calvarial Pg infection. Based on our findings, we propose the following model: Pg infection increases a pool of precursor cells that can be shunted toward osteoclast formation at the infection/inflammation sites, while at the same time dampening host immune responses, which is beneficial for the persistence of infection and maintenance of the characteristic chronic nature of periodontitis. Understanding the nature, function, and regulation of osteoclast precursors will be helpful for identifying therapeutic interventions to aid in the control and prevention of inflammatory bone loss diseases including periodontitis.
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Infecciones por Bacteroidaceae/patología , Osteoclastos/citología , Cráneo/microbiología , Animales , Diferenciación Celular , Ratones , Ratones Endogámicos C57BL , Osteoclastos/microbiología , Porphyromonas gingivalis , Ligando RANKRESUMEN
PURPOSE: Clinical and in vitro studies showed selected oral microorganisms to be related to delayed wound healing and ulcerative oral mucositis. However, it is not known whether this effect is due to reduced metabolism and/or the reduced reproductive capacity of epithelial cells. Therefore, we studied the influence of the oral microorganisms Porphyromonas gingivalis, Candida glabrata, and Candida kefyr on cell metabolism and reproductive capacity of oral epithelial cells, aimed to further unravel the pathogenesis of oral mucositis. METHODS: Oral epithelial cells were exposed to different concentrations of P. gingivalis, C. glabrata, and C. kefyr as mono-infections or mixed together. An MTT assay was performed to determine the effect on cell metabolism. A clonogenic assay was used to study the effect on the reproductive capacity of oral epithelial cells. RESULTS: The metabolism of oral epithelial cells was reduced when the microorganisms were present in high concentrations: P. gingivalis at a multiplicity of infection (MOI) of 1000 and the Candida spp. at MOI 100. No statistical difference was observed in the ability of a single epithelial cell to grow into a colony of cells between control and P. gingivalis, C. glabrata, and C. kefyr, independent of the concentrations and combinations used. CONCLUSION: P. gingivalis, C. glabrata, and C. kefyr lowered the metabolic activity of oral epithelial cells in high concentrations, yet they did not influence the reproductive capacity of epithelial cells. Their impact on ulcerative oral mucositis is likely due to an effect on the migration, proliferation, and metabolism of epithelial cells.
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Candida/fisiología , Porphyromonas gingivalis/fisiología , Estomatitis/microbiología , Infecciones por Bacteroidaceae/metabolismo , Infecciones por Bacteroidaceae/microbiología , Infecciones por Bacteroidaceae/patología , Candida glabrata/fisiología , Candidiasis/metabolismo , Candidiasis/microbiología , Candidiasis/patología , Línea Celular , Células Epiteliales/metabolismo , Células Epiteliales/microbiología , Células Epiteliales/patología , Humanos , Técnicas In Vitro , Estomatitis/metabolismo , Estomatitis/patologíaRESUMEN
Bone destruction in inflammatory osteolytic diseases including periodontitis is related to excessive activity of osteoclasts (OC), which originate from precursor cells of the myeloid lineage, termed osteoclast precursors (OCP). In contrast to ample knowledge that we currently have on mature OC, little is known about OCP and their regulation during bacterial infection. Therefore, this study aimed to identify and characterize OCP following chronic infection with a periodontal bacteria Porphyromonas gingivalis (Pg). We used a microosmotic pump to continually release Pg subcutaneously in a murine model. Two weeks after Pg infection, the frequency of CD11b+c-fms+Ly6Chi population is significantly elevated within the bone marrow, spleen, and peripheral blood. In vitro and in vivo studies identified these cells as the OCP-containing population and Pg infection significantly enhanced the osteoclastogenic activity of these cells. Furthermore, mRNA sequencing analysis indicated a unique gene and pathway profile in CD11b+c-fms+Ly6Chi population following Pg infection, with changes in genes and pathways related to OC differentiation, cell proliferation and apoptosis, inflammatory response, phagocytosis, and immunity, as well as antigen processing and presentation. Moreover, using IL-6 knockout mice, we found that IL-6 is important for Pg-induced accumulation of CD11b+c-fms+Ly6Chi population from the bone marrow and periphery. Our results provide new insight into the characterization and regulation of OCP following a chronic bacterial infection. This knowledge is relevant to the understanding of the pathogenesis of bacteria-induced bone loss, and to the identification of potential therapeutic targets of bone loss diseases.