RESUMEN
BACKGROUND: Elevated systemic antibody responses against gut microbiota flagellins are observed in both Crohn's disease (CD) and myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS), suggesting potential serological biomarkers for diagnosis. However, flagellin-specific antibody repertoires and functional roles in the diseases remain incompletely understood. Bacterial flagellins can be categorized into three types depending on their interaction with toll-like receptor 5 (TLR5): (1) "stimulator" and (2) "silent" flagellins, which bind TLR5 through a conserved N-terminal motif, with only stimulators activating TLR5 (involving a C-terminal domain); (3) "evader" flagellins of pathogens, which entirely circumvent TLR5 activation via mutations in the N-terminal TLR5 binding motif. RESULTS: Here, we show that both CD and ME/CFS patients exhibit elevated antibody responses against distinct regions of flagellins compared to healthy individuals. N-terminal binding to Lachnospiraceae flagellins was comparable in both diseases, while C-terminal binding was more prevalent in CD. N-terminal antibody-bound flagellin sequences were similar across CD and ME/CFS, resembling "stimulator" and "silent" flagellins more than evaders. However, C-terminal antibody-bound flagellins showed a higher resemblance to the stimulator than to silent flagellins in CD, which was not observed in ME/CFS. CONCLUSIONS: These findings suggest that antibody binding to the N-terminal domain of stimulator and silent flagellins may impact TLR5 activation in both CD and ME/CFS patients. Blocking this interaction could lead commensal bacteria to be recognized as pathogenic evaders, potentially contributing to dysregulation in both diseases. Furthermore, elevated antibody binding to the C-terminal domain of stimulator flagellins in CD may explain pathophysiological differences between the diseases. Overall, these results highlight the diagnostic potential of these antibody responses and lay a foundation for deeper mechanistic studies of flagellin/TLR5 interactions and their impact on innate/adaptive immunity balance.
Asunto(s)
Enfermedad de Crohn , Síndrome de Fatiga Crónica , Flagelina , Microbioma Gastrointestinal , Receptor Toll-Like 5 , Flagelina/inmunología , Humanos , Síndrome de Fatiga Crónica/inmunología , Síndrome de Fatiga Crónica/microbiología , Enfermedad de Crohn/inmunología , Enfermedad de Crohn/microbiología , Receptor Toll-Like 5/inmunología , Microbioma Gastrointestinal/inmunología , Femenino , Anticuerpos Antibacterianos/inmunología , Anticuerpos Antibacterianos/sangre , Masculino , Adulto , Formación de Anticuerpos/inmunología , Persona de Mediana Edad , Clostridiales/inmunologíaRESUMEN
Hidradenitis suppurativa (HS) is a chronic inflammatory skin disease characterized by the formation of nodules, abscesses, and fistulae at intertriginous sites. The skin-gut axis is an area of emerging research in inflammatory skin disease and is a potential contributory factor to the pathogenesis of HS. A total of 59 patients with HS provided fecal samples and nasal and skin swabs of affected sites for analysis. A total of 30 healthy controls provided fecal samples, and 20 healthy controls provided nasal and skin swabs. We performed bacterial 16S ribosomal RNA gene amplicon sequencing on total DNA derived from the samples. Microbiome alpha diversity was significantly lower in the fecal, skin, and nasal samples of individuals with HS, which may be secondary to disease biology or related to antibiotic usage. Ruminococcus gnavus was more abundant in the fecal microbiome of individuals with HS, which is also reported in Crohn's disease, suggesting comorbidity due to shared gut microbiota alterations. Finegoldia magna was overabundant in HS skin samples relative to that in the healthy controls. It is possible that local inflammation is driven by F. magna by promoting the formation of neutrophil extracellular traps. These alterations in both the gut and skin microbiome in HS warrant further exploration, and therapeutic strategies, including fecal microbiota transplant or bacteriotherapy, could be of benefit.
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Microbioma Gastrointestinal/inmunología , Hidradenitis Supurativa/microbiología , Piel/microbiología , Adulto , Anciano , Estudios de Casos y Controles , Clostridiales/inmunología , Clostridiales/aislamiento & purificación , Trampas Extracelulares/inmunología , Trasplante de Microbiota Fecal , Heces/microbiología , Femenino , Firmicutes/inmunología , Firmicutes/aislamiento & purificación , Hidradenitis Supurativa/inmunología , Hidradenitis Supurativa/patología , Hidradenitis Supurativa/terapia , Humanos , Masculino , Persona de Mediana Edad , Piel/inmunología , Piel/patología , Adulto JovenRESUMEN
Despite overall success, T cell checkpoint inhibitors for cancer treatment are still only efficient in a minority of patients. Recently, intestinal microbiota was found to critically modulate anti-cancer immunity and therapy response. Here, we identify Clostridiales members of the gut microbiota associated with a lower tumor burden in mouse models of colorectal cancer (CRC). Interestingly, these commensal species are also significantly reduced in CRC patients compared with healthy controls. Oral application of a mix of four Clostridiales strains (CC4) in mice prevented and even successfully treated CRC as stand-alone therapy. This effect depended on intratumoral infiltration and activation of CD8+ T cells. Single application of Roseburia intestinalis or Anaerostipes caccae was even more effective than CC4. In a direct comparison, the CC4 mix supplementation outperformed anti-PD-1 therapy in mouse models of CRC and melanoma. Our findings provide a strong preclinical foundation for exploring gut bacteria as novel stand-alone therapy against solid tumors.
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Terapia Biológica , Clostridiales/inmunología , Neoplasias Colorrectales/inmunología , Neoplasias Colorrectales/terapia , Microbioma Gastrointestinal , Animales , Linfocitos T CD8-positivos/inmunología , Clostridiales/fisiología , Neoplasias Colorrectales/microbiología , Humanos , Inmunidad , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , SimbiosisRESUMEN
Intestinal microbiomes are of vital importance in antagonizing systemic viral infection. However, very little literature has shown whether commensal bacteria play a crucial role in protecting against enteric virus systemic infection from the aspect of modulating host innate immunity. In the present study, we utilized an enteric virus, encephalomyocarditis virus (EMCV), to inoculate mice treated with phosphate-buffered saline (PBS) or given an antibiotic cocktail (Abx) orally or intraperitoneally to examine the impact of microbiota depletion on virulence and viral replication in vivo Microbiota depletion exacerbated the mortality, neuropathogenesis, viremia, and viral burden in brains following EMCV infection. Furthermore, Abx-treated mice exhibited severely diminished mononuclear phagocyte activation and impaired type I interferon (IFN) production and expression of IFN-stimulated genes (ISG) in peripheral blood mononuclear cells (PBMC), spleens, and brains. With the help of fecal bacterial 16S rRNA sequencing of PBS- and Abx-treated mice, we identified a single commensal bacterium, Blautia coccoides, that can restore mononuclear phagocyte- and IFNAR (IFN-α/ß receptor)-dependent type I IFN responses to restrict systemic enteric virus infection. These findings may provide insight into the development of novel therapeutics for preventing enteric virus infection or possibly alleviating clinical diseases by activating host systemic innate immune responses via respective probiotic treatment using B. coccoidesIMPORTANCE While cumulative data indicate that indigenous commensal bacteria can facilitate enteric virus infection, little is known regarding whether intestinal microbes have a protective role in antagonizing enteric systemic infection by modulating host innate immunity. Although accumulating literature has pointed out that the microbiota has a fundamental impact on host systemic antiviral innate immune responses mediated by type I interferon (IFN), only a few specific commensal bacteria species have been revealed to be capable of regulating IFN-I and ISG expression, not to mention the underlying mechanisms. Thus, it is important to understand the cross talk between microbiota and host anti-enteric virus innate immune responses and characterize the specific bacterial species that possess protective functions. Our study demonstrates how fundamental innate immune mediators such as mononuclear phagocytes and type I IFN are regulated by commensal bacteria to antagonize enteric virus systemic infection. In particular, we have identified a novel commensal bacterium, Blautia coccoides, that can restrict enteric virus replication and neuropathogenesis by activating IFN-I and ISG responses in mononuclear phagocytes via an IFNAR- and STAT1-mediated signaling pathway.
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Infecciones por Cardiovirus/prevención & control , Virus de la Encefalomiocarditis/inmunología , Microbioma Gastrointestinal/inmunología , Inmunidad Innata , Interferón Tipo I/inmunología , Viremia/inmunología , Viremia/prevención & control , Animales , Antibacterianos/administración & dosificación , Infecciones por Cardiovirus/inmunología , Clostridiales/inmunología , Virus de la Encefalomiocarditis/patogenicidad , Femenino , Microbioma Gastrointestinal/efectos de los fármacos , Masculino , Ratones , Ratones Endogámicos C57BL , Transducción de Señal , Simbiosis/inmunología , Replicación Viral/inmunologíaRESUMEN
Active inflammatory bowel disease (IBD) often coincides with increases of Ruminococcus gnavus, a gut microbe found in nearly everyone. It was not known how, or if, this correlation contributed to disease. We investigated clinical isolates of R. gnavus to identify molecular mechanisms that would link R. gnavus to inflammation. Here, we show that only some isolates of R. gnavus produce a capsular polysaccharide that promotes a tolerogenic immune response, whereas isolates lacking functional capsule biosynthetic genes elicit robust proinflammatory responses in vitro. Germ-free mice colonized with an isolate of R. gnavus lacking a capsule show increased measures of gut inflammation compared to those colonized with an encapsulated isolate in vivo. These observations in the context of our earlier identification of an inflammatory cell-wall polysaccharide reveal how some strains of R. gnavus could drive the inflammatory responses that characterize IBD.
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Cápsulas Bacterianas/inmunología , Clostridiales/inmunología , Microbioma Gastrointestinal/inmunología , Inmunidad/inmunología , Enfermedades Inflamatorias del Intestino/inmunología , Polisacáridos/inmunología , Adulto , Animales , Antibacterianos/farmacología , Cápsulas Bacterianas/química , Cápsulas Bacterianas/ultraestructura , Células Cultivadas , Niño , Clostridiales/clasificación , Clostridiales/genética , Citocinas/inmunología , Citocinas/metabolismo , Femenino , Microbioma Gastrointestinal/efectos de los fármacos , Microbioma Gastrointestinal/genética , Humanos , Íleon/inmunología , Íleon/metabolismo , Íleon/microbiología , Enfermedades Inflamatorias del Intestino/microbiología , Ratones Endogámicos C57BL , Familia de Multigenes/genética , FilogeniaRESUMEN
BACKGROUND AND AIMS: Crohn's disease and ulcerative colitis are characterized by dysregulated adaptive immune responses to the microbiota in genetically susceptible individuals, but the specificity of these responses remains largely undefined. Therefore, we developed a microbiota antigen microarray to characterize microbial antibody reactivity, particularly to human-derived microbiota flagellins, in inflammatory bowel disease. METHODS: Sera from healthy volunteers (n = 87) at the University of Alabama at Birmingham and from patients recruited from the Kirklin Clinic of University of Alabama at Birmingham Hospital, including patients with Crohn's disease (n = 152) and ulcerative colitis (n = 170), were individually probed against microbiota bacterial flagellins of both mouse and human origin and analyzed for IgG and IgA antibody responses. Circulating flagellin-reactive T effector (CD4+CD154+) and T regulatory (CD4+CD137+) cells were isolated and evaluated in selected patients. Resulting adaptive immune responses were compared with corresponding clinical data to determine relevancy to disease behavior. RESULTS: We show that patients with IBD express selective patterns of antibody reactivity to microbiota flagellins. Patients with Crohn's disease, but not patients with ulcerative colitis, display augmented serum IgG to human ileal-localized Lachnospiraceae flagellins, with a subset of patients having high responses to more than 10 flagellins. Elevated responses to CBir1, a mouse Lachnospiraceae flagellin used clinically to diagnose CD, correlated with multi-Lachnospiraceae flagellin reactivity. In this subset of patients with CD, multi-flagellin reactivity was associated with elevated flagellin-specific CD154+CD45RA- T memory cells, a reduced ratio of flagellin-reactive CD4+ T regulatory to T effector cells, and a high frequency of disease complications. CONCLUSIONS: Patients with Crohn's disease display strong adaptive immune response to human-derived Lachnospiraceae flagellins, which may be targeted for prognosis and future personalized therapies.
Asunto(s)
Inmunidad Adaptativa , Anticuerpos Antibacterianos/sangre , Antígenos Bacterianos/inmunología , Linfocitos T CD4-Positivos/inmunología , Clostridiales/inmunología , Enfermedad de Crohn/inmunología , Flagelina/inmunología , Inmunoglobulina G/sangre , Adulto , Anciano , Anciano de 80 o más Años , Antígenos Bacterianos/metabolismo , Linfocitos B/inmunología , Linfocitos B/metabolismo , Linfocitos B/microbiología , Linfocitos T CD4-Positivos/metabolismo , Linfocitos T CD4-Positivos/microbiología , Estudios de Casos y Controles , Clostridiales/metabolismo , Colitis Ulcerosa/sangre , Colitis Ulcerosa/inmunología , Colitis Ulcerosa/microbiología , Enfermedad de Crohn/sangre , Enfermedad de Crohn/microbiología , Estudios Transversales , Femenino , Flagelina/metabolismo , Humanos , Inmunoglobulina A/sangre , Masculino , Persona de Mediana Edad , Adulto JovenRESUMEN
The immunopathogenesis of inflammatory bowel disease (IBD) has been attributed to a combination of host genetics and intestinal dysbiosis. Previous work in a small cohort of IBD patients suggested that pro-inflammatory bacterial taxa are highly coated with secretory immunoglobulin IgA. Using bacterial fluorescence-activated cell sorting coupled with 16S rRNA gene sequencing (IgA-SEQ), we profiled IgA coating of intestinal microbiota in a large cohort of IBD patients and identified bacteria associated with disease and treatment. Forty-three bacterial taxa displayed significantly higher IgA coating in IBD compared with controls, including 8 taxa exhibiting differential IgA coating but similar relative abundance. Patients treated with anti-TNF-α therapies exhibited dramatically altered microbiota-specific IgA responses compared with controls. Furthermore, increased IgA coating of Oscillospira was associated with a delay in time to surgery. These results demonstrate that investigating IgA responses to microbiota can uncover potential disease-modifying taxa and reveal improved biomarkers of clinical course in IBD.
Asunto(s)
Bacterias/inmunología , Microbioma Gastrointestinal/inmunología , Microbioma Gastrointestinal/fisiología , Inmunoglobulina A Secretora/inmunología , Enfermedades Inflamatorias del Intestino/inmunología , Enfermedades Inflamatorias del Intestino/microbiología , Adulto , Bacterias/crecimiento & desarrollo , Clostridiales/crecimiento & desarrollo , Clostridiales/inmunología , Estudios de Cohortes , Colitis Ulcerosa/tratamiento farmacológico , Colitis Ulcerosa/inmunología , Colitis Ulcerosa/microbiología , Colitis Ulcerosa/fisiopatología , Enfermedad de Crohn/tratamiento farmacológico , Enfermedad de Crohn/inmunología , Enfermedad de Crohn/microbiología , Enfermedad de Crohn/fisiopatología , Progresión de la Enfermedad , Femenino , Humanos , Enfermedades Inflamatorias del Intestino/tratamiento farmacológico , Enfermedades Inflamatorias del Intestino/fisiopatología , Masculino , Factor de Necrosis Tumoral alfa/antagonistas & inhibidoresRESUMEN
There is little known about the effect of the periodontopathogen Filifactor alocis on macrophages as key cells of the innate immune defense in the periodontium. Therefore, the aim of the present study was to investigate the effect of F. alocis and additionally of the pro-inflammatory cytokine tumor necrosis factor-alpha (TNFα) on visfatin and other pro-inflammatory and proteolytic molecules associated with periodontitis in human macrophages. The presence of macrophage markers CD14, CD86, CD68, and CD163 was examined in gingival biopsies from healthy individuals and periodontitis patients. Human macrophages were incubated with F. alocis and TNFα for up to 2 d. The effects of both stimulants on macrophages were determined by real-time PCR, ELISA, immunocytochemistry, and immunofluorescence. F. alocis was able to significantly stimulate the synthesis of visfatin by human macrophages using TLR2 and MAPK pathways. In addition to visfatin, F. alocis was also able to increase the synthesis of cyclooxygenase 2, TNFα, and matrix metalloproteinase 1. Like F. alocis, TNFα was also able to stimulate the production of these proinflammatory and proteolytic molecules. Our results highlight the pathogenetic role of F. alocis in periodontal diseases and also underline the involvement of visfatin in the aetiopathogenesis of periodontitis.
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Clostridiales/inmunología , Encía/metabolismo , Macrófagos/metabolismo , Nicotinamida Fosforribosiltransferasa/biosíntesis , Periodontitis/inmunología , Receptor Toll-Like 2/metabolismo , Factor de Necrosis Tumoral alfa/farmacología , Antígenos CD/genética , Antígenos CD/metabolismo , Antígenos de Diferenciación Mielomonocítica/genética , Antígenos de Diferenciación Mielomonocítica/metabolismo , Antígeno B7-2/genética , Antígeno B7-2/metabolismo , Ciclooxigenasa 2/biosíntesis , Ciclooxigenasa 2/genética , Ciclooxigenasa 2/metabolismo , Encía/citología , Encía/patología , Humanos , Inmunohistoquímica , Receptores de Lipopolisacáridos/genética , Receptores de Lipopolisacáridos/metabolismo , Sistema de Señalización de MAP Quinasas/inmunología , Macrófagos/efectos de los fármacos , Metaloproteinasa 1 de la Matriz/biosíntesis , Metaloproteinasa 1 de la Matriz/genética , Metaloproteinasa 1 de la Matriz/metabolismo , Nicotinamida Fosforribosiltransferasa/genética , Nicotinamida Fosforribosiltransferasa/metabolismo , Periodontitis/metabolismo , Periodontitis/microbiología , Receptores de Superficie Celular/genética , Receptores de Superficie Celular/metabolismo , Factor de Necrosis Tumoral alfa/biosíntesis , Factor de Necrosis Tumoral alfa/genética , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
Paneth cells are the primary source of C-type lysozyme, a ß-1,4-N-acetylmuramoylhydrolase that enzymatically processes bacterial cell walls. Paneth cells are normally present in human cecum and ascending colon, but are rarely found in descending colon and rectum; Paneth cell metaplasia in this region and aberrant lysozyme production are hallmarks of inflammatory bowel disease (IBD) pathology. Here, we examined the impact of aberrant lysozyme production in colonic inflammation. Targeted disruption of Paneth cell lysozyme (Lyz1) protected mice from experimental colitis. Lyz1-deficiency diminished intestinal immune responses to bacterial molecular patterns and resulted in the expansion of lysozyme-sensitive mucolytic bacteria, including Ruminococcus gnavus, a Crohn's disease-associated pathobiont. Ectopic lysozyme production in colonic epithelium suppressed lysozyme-sensitive bacteria and exacerbated colitis. Transfer of R. gnavus into Lyz1-/- hosts elicited a type 2 immune response, causing epithelial reprograming and enhanced anti-colitogenic capacity. In contrast, in lysozyme-intact hosts, processed R. gnavus drove pro-inflammatory responses. Thus, Paneth cell lysozyme balances intestinal anti- and pro-inflammatory responses, with implications for IBD.
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Clostridiales/inmunología , Colitis Ulcerosa/patología , Muramidasa/genética , Muramidasa/metabolismo , Células de Paneth/metabolismo , Animales , Clostridiales/genética , Colitis Ulcerosa/microbiología , Enfermedad de Crohn/patología , Femenino , Microbioma Gastrointestinal/genética , Células Caliciformes/citología , Humanos , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Ratones Noqueados , Factor de Transcripción STAT6/genéticaRESUMEN
The gut pathogen Clostridium bolteae has been associated with the onset of autism spectrum disorder (ASD). To create vaccines against C. bolteae, it is important to identify exact protective epitopes of the immunologically active capsular polysaccharide (CPS). Here, a series of C. bolteae CPS glycans, up to an octadecasaccharide, was prepared. Key to achieving the total syntheses is a [2+2] coupling strategy based on a ß-d-Rhap-(1â3)-α-d-Manp repeating unit that in turn was accessed by a stereoselective ß-d-rhamnosylation. The 4,6-O-benzylidene-induced conformational locking is a powerful strategy for forming a ß-d-mannose-type glycoside. An indirect strategy based on C2 epimerization of ß-d-quinovoside was efficiently achieved by Swern oxidation and borohydride reduction. Sequential glycosylation, and regioselective and global deprotection produced the disaccharide and tetrasaccharide, up to the octadecasaccharide. Glycan microarray analysis of sera from rabbits immunized with inactivated C. bolteae bacteria revealed a humoral immune response to the di- and tetrasaccharide, but none of the longer sequences. The tetrasaccharide may be a key motif for designing glycoconjugate vaccines against C. bolteae.
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Trastorno Autístico/microbiología , Clostridiales/inmunología , Epítopos/inmunología , Polisacáridos/metabolismo , Secuencia de Carbohidratos , Clostridiales/metabolismo , Humanos , Polisacáridos/químicaRESUMEN
Colonization by the microbiota causes a marked stimulation of B cells and induction of immunoglobulin, but mammals colonized with many taxa have highly complex and individualized immunoglobulin repertoires1,2. Here we use a simplified model of defined transient exposures to different microbial taxa in germ-free mice3 to deconstruct how the microbiota shapes the B cell pool and its functional responsiveness. We followed the development of the immunoglobulin repertoire in B cell populations, as well as single cells by deep sequencing. Microbial exposures at the intestinal mucosa generated oligoclonal responses that differed from those of germ-free mice, and from the diverse repertoire that was generated after intravenous systemic exposure to microbiota. The IgA repertoire-predominantly to cell-surface antigens-did not expand after dose escalation, whereas increased systemic exposure broadened the IgG repertoire to both microbial cytoplasmic and cell-surface antigens. These microbial exposures induced characteristic immunoglobulin heavy-chain repertoires in B cells, mainly at memory and plasma cell stages. Whereas sequential systemic exposure to different microbial taxa diversified the IgG repertoire and facilitated alternative specific responses, sequential mucosal exposure produced limited overlapping repertoires and the attrition of initial IgA binding specificities. This shows a contrast between a flexible response to systemic exposure with the need to avoid fatal sepsis, and a restricted response to mucosal exposure that reflects the generic nature of host-microbial mutualism in the mucosa.
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Linfocitos B/citología , Linfocitos B/inmunología , Inmunidad Mucosa/inmunología , Mucosa Intestinal/inmunología , Mucosa Intestinal/microbiología , Simbiosis/inmunología , Administración Intravenosa , Administración Oral , Animales , Clostridiales/inmunología , Clostridiales/aislamiento & purificación , Escherichia coli/inmunología , Escherichia coli/aislamiento & purificación , Femenino , Vida Libre de Gérmenes , Inmunoglobulina A/química , Inmunoglobulina A/inmunología , Inmunoglobulina G/química , Inmunoglobulina G/inmunología , Cadenas Pesadas de Inmunoglobulina/inmunología , Memoria Inmunológica/inmunología , Masculino , Ratones , Ratones Endogámicos C57BL , Células Plasmáticas/citología , Células Plasmáticas/inmunología , Memoria ImplícitaRESUMEN
The microbiome provides resistance to infection. However, the underlying mechanisms are poorly understood. We demonstrate that colonization with the intestinal bacterium Clostridium scindens protects from Entamoeba histolytica colitis via innate immunity. Introduction of C. scindens into the gut microbiota epigenetically altered and expanded bone marrow granulocyte-monocyte progenitors (GMPs) and resulted in increased intestinal neutrophils with subsequent challenge with E. histolytica. Introduction of C. scindens alone was sufficient to expand GMPs in gnotobiotic mice. Adoptive transfer of bone marrow from C. scindens-colonized mice into naive mice protected against amebic colitis and increased intestinal neutrophils. Children without E. histolytica diarrhea also had a higher abundance of Lachnoclostridia. Lachnoclostridia C. scindens can metabolize the bile salt cholate, so we measured deoxycholate and discovered that it was increased in the sera of C. scindens-colonized specific pathogen-free and gnotobiotic mice, as well as in children protected from amebiasis. Administration of deoxycholate alone increased GMPs and provided protection from amebiasis. We elucidated a mechanism by which C. scindens and the microbially metabolized bile salt deoxycholic acid alter hematopoietic precursors and provide innate protection from later infection with E. histolytica.
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Médula Ósea/inmunología , Clostridiales/inmunología , Disentería Amebiana/inmunología , Entamoeba histolytica/inmunología , Microbioma Gastrointestinal/inmunología , Animales , Médula Ósea/patología , Modelos Animales de Enfermedad , Susceptibilidad a Enfermedades/inmunología , Susceptibilidad a Enfermedades/microbiología , Disentería Amebiana/microbiología , Disentería Amebiana/patología , Humanos , Intestinos/inmunología , Intestinos/microbiología , Intestinos/patología , RatonesRESUMEN
Periodontitis is an irreversible, bacteria-induced, chronic inflammatory disease that compromises the integrity of tooth-supporting tissues and adversely affects systemic health. As the immune system's first line of defense against bacteria, neutrophils use their microbicidal functions in the oral cavity to protect the host against periodontal disease. However, periodontal pathogens have adapted to resist neutrophil microbicidal mechanisms while still propagating inflammation, which provides essential nutrients for the bacteria to proliferate and cause disease. Advances in sequencing technologies have recognized several newly appreciated bacteria associated with periodontal lesions such as the Gram-positive anaerobic rod, Filifactor alocis. With the discovery of these oral bacterial species, there is also a growing need to assess their pathogenic potential and determine their contribution to disease progression. Currently, few studies have addressed the pathogenic mechanisms used by oral bacteria to manipulate the neutrophil functional responses at the level of the transcriptome. Thus, this study aims to characterize the global changes at the gene expression level in human neutrophils during infection with F. alocis. Our results indicate that the challenge of human neutrophils with F. alocis results in the differential expression of genes involved in multiple neutrophil effector functions such as chemotaxis, cytokine and chemokine signaling pathways, and apoptosis. Moreover, F. alocis challenges affected the expression of components from the TNF and MAPK kinase signaling pathways. This resulted in transient, dampened p38 MAPK activation by secondary stimuli TNFα but not by fMLF. Functionally, the F. alocis-mediated inhibition of p38 activation by TNFα resulted in decreased cytokine production but had no effect on the priming of the respiratory burst response or the delay of apoptosis by TNFα. Since the modulatory effect was characteristic of viable F. alocis only, we propose this as one of F. alocis' mechanisms to control neutrophils and their functional responses.
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Clostridiales/inmunología , Neutrófilos/fisiología , Periodontitis/inmunología , Activación Enzimática , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Perfilación de la Expresión Génica , Humanos , Estallido Respiratorio , Transducción de Señal , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
BACKGROUND & AIMS: Bacterial flagellin is an important antigen in inflammatory bowel disease, but the role of flagellin-specific CD4+ T cells in disease pathogenesis remains unclear. Also unknown is how changes in intestinal microbiome intersect with those in microbiota-specific CD4+ T cells. We aimed to quantify and characterize flagellin-specific CD4+ T cells in Crohn's disease (CD) and ulcerative colitis (UC) patients and study their relationship with intestinal microbiome diversity. METHODS: Blood was collected from 3 cohorts that included CD patients, UC patients, and healthy controls. Flow cytometry analyzed CD4+ T cells specific for Lachnospiraceae-derived A4-Fla2 and Escherichia coli H18 FliC flagellins, or control vaccine antigens. Serum antiflagellin IgG and IgA antibodies were detected by enzyme-linked immunosorbent assay and stool samples were collected and subjected to 16S ribosomal DNA sequencing. RESULTS: Compared with healthy controls, CD and UC patients had lower frequencies of vaccine-antigen-specific CD4+ T cells and, as a proportion of vaccine-specific cells, higher frequencies of flagellin-specific CD4+ T cells. The proportion of flagellin-specific CD4+ T cells that were CXCR3negCCR4+CCR6+ Th17 cells was reduced in CD and UC patients, with increased proportions of CD39+, PD-1+, and integrin ß7+ cells. Microbiome analysis showed differentially abundant bacterial species in patient groups that correlated with immune responses to flagellin. CONCLUSIONS: Both CD and UC patients have relative increases in the proportion of circulating Fla2-specific CD4+ T cells, which may be associated with changes in the intestinal microbiome. Evidence that the phenotype of these cells strongly correlate with disease severity provides insight into the potential roles of flagellin-specific CD4+ T cells in inflammatory bowel disease.
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Anticuerpos Antibacterianos/sangre , Colitis Ulcerosa/inmunología , Enfermedad de Crohn/inmunología , Disbiosis/complicaciones , Proteínas de Escherichia coli/inmunología , Flagelina/inmunología , Inmunidad Adaptativa , Adolescente , Adulto , Anciano , Anticuerpos Antibacterianos/inmunología , Antígenos Bacterianos/inmunología , Linfocitos T CD4-Positivos/inmunología , Clostridiales/genética , Clostridiales/inmunología , Colitis Ulcerosa/sangre , Colitis Ulcerosa/microbiología , Enfermedad de Crohn/sangre , Enfermedad de Crohn/microbiología , Estudios Transversales , ADN Bacteriano/aislamiento & purificación , Disbiosis/diagnóstico , Disbiosis/inmunología , Disbiosis/microbiología , Ensayo de Inmunoadsorción Enzimática , Heces/microbiología , Femenino , Microbioma Gastrointestinal/inmunología , Voluntarios Sanos , Humanos , Masculino , Persona de Mediana Edad , ARN Ribosómico 16S/genética , Subgrupos de Linfocitos T/inmunología , Adulto JovenRESUMEN
INTRODUCTION: Histopathological alterations in the ileum and colon in irritable bowel syndrome (IBS) are controversial, and normal values are poorly established. We hypothesized that changes in mucosal immune cells characterize IBS and key changes in immune composition are associated with the mucosa-associated microbiota (MaM). METHODS: A nested case-control study (48 IBS and 106 controls included) from 745 colonoscopy participants in a random population sample. Intraepithelial lymphocytes (IELs)/100 enterocytes and eosinophils/5 nonoverlapping high-power fields counted; mast cells identified by immunocytochemistry (CD117)/5 high-power fields. Paneth cells quantified per 5 crypts. 16S rRNA gene amplicon sequencing performed on available sigmoid MaM, n = 55 and fecal microbiota, n = 20. Microbiota profiles compared between samples with high and low IEL counts. RESULTS: IBS had increased IELs in the terminal ileum (relative risk ratio = 1.70, 95% confidence interval 1.08-2.76, P = 0.022 adjusted for age, sex, and smoking). Cecal IELs were increased in IBS-diarrhea (relative risk ratio = 2.03, 95% confidence interval 1.13-3.63, P = 0.017). No difference was observed in alpha diversity of MaM or fecal microbiota based on IEL count. There was no difference in beta diversity of the MaM according to IEL count in the terminal ileal (TI) (P = 0.079). High TI IEL counts associated with a significant expansion of the genus Blautia (P = 0.024) and unclassified Clostridiales (P = 0.036) in colon MaM. DISCUSSION: A modest but significant increase in IELs was observed in IBS vs. controls in a population-based setting. Subtle TI and cecal inflammation may play a pathogenic role in IBS but needs confirmation. Modest but discernible differences in the colonic MaM were seen according to TI IEL count but not IBS status.
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Colon/patología , Microbioma Gastrointestinal/inmunología , Íleon/patología , Mucosa Intestinal/patología , Síndrome del Colon Irritable/inmunología , Adulto , Biopsia , Estudios de Casos y Controles , Clostridiales/genética , Clostridiales/inmunología , Clostridiales/aislamiento & purificación , Colon/diagnóstico por imagen , Colon/inmunología , Colon/microbiología , ADN Bacteriano/aislamiento & purificación , Heces/microbiología , Femenino , Microbioma Gastrointestinal/genética , Humanos , Íleon/diagnóstico por imagen , Íleon/inmunología , Íleon/microbiología , Mucosa Intestinal/diagnóstico por imagen , Mucosa Intestinal/inmunología , Mucosa Intestinal/microbiología , Síndrome del Colon Irritable/diagnóstico , Síndrome del Colon Irritable/microbiología , Síndrome del Colon Irritable/patología , Masculino , Mastocitos/inmunología , Persona de Mediana Edad , Células de Paneth/inmunología , ARN Ribosómico 16S/genéticaRESUMEN
Periodontitis is a multifactorial chronic inflammatory infectious disease that compromises the integrity of tooth-supporting tissues. The disease progression depends on the disruption of host-microbe homeostasis in the periodontal tissue. This disruption is marked by a shift in the composition of the polymicrobial oral community from a symbiotic to a dysbiotic, more complex community that is capable of evading killing while promoting inflammation. Neutrophils are the main phagocytic cell in the periodontal pocket, and the outcome of the interaction with the oral microbiota is an important determinant of oral health. Novel culture-independent techniques have facilitated the identification of new bacterial species at periodontal lesions and induced a reappraisal of the microbial etiology of periodontitis. In this chapter, we discuss how neutrophils interact with two emerging oral pathogens, Filifactor alocis and Peptoanaerobacter stomatis, and the different strategies deploy by these organisms to modulate neutrophil effector functions, with the goal to outline a new paradigm in our knowledge about neutrophil responses to putative periodontal pathogens and their contribution to disease progression.
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Neutrófilos , Periodontitis , Clostridiales/inmunología , Disbiosis , Humanos , Microbiota/inmunología , Neutrófilos/inmunología , Neutrófilos/microbiología , Periodontitis/inmunología , Periodontitis/microbiología , Periodoncio/microbiologíaRESUMEN
Given the immense antigenic load present in the microbiome, we hypothesized that microbiota mimotopes can be a persistent trigger in human autoimmunity via cross-reactivity. Using antiphospholipid syndrome (APS) as a model, we demonstrate cross-reactivity between non-orthologous mimotopes expressed by a common human gut commensal, Roseburia intestinalis (R. int), and T and B cell autoepitopes in the APS autoantigen ß2-glycoprotein I (ß2GPI). Autoantigen-reactive CD4+ memory T cell clones and an APS-derived, pathogenic monoclonal antibody cross-reacted with R. int mimotopes. Core-sequence-dependent anti-R. int mimotope IgG titers were significantly elevated in APS patients and correlated with anti-ß2GPI IgG autoantibodies. R. int immunization of mice induced ß2GPI-specific lymphocytes and autoantibodies. Oral gavage of susceptible mice with R. int induced anti-human ß2GPI autoantibodies and autoimmune pathologies. Together, these data support a role for non-orthologous commensal-host cross-reactivity in the development and persistence of autoimmunity in APS, which may apply more broadly to human autoimmune disease.
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Anticuerpos Antibacterianos/inmunología , Antígenos Bacterianos/inmunología , Autoinmunidad , Linfocitos B/inmunología , Clostridiales/inmunología , Reacciones Cruzadas , Linfocitos T/inmunología , Adulto , Anciano , Animales , Síndrome Antifosfolípido/patología , Autoanticuerpos/sangre , Autoanticuerpos/inmunología , Femenino , Tracto Gastrointestinal/microbiología , Humanos , Inmunoglobulina G/sangre , Masculino , Ratones , Persona de Mediana Edad , Modelos Animales , Adulto Joven , beta 2 Glicoproteína I/inmunologíaRESUMEN
BACKGROUND: Anaerobic Clostridial spores (CG) cause significant oncolysis in hypoxic tumour microenvironment and result in tumour regression in both animal models and clinical trials. The immune mediated response plays a critical role in the antitumour effect by the anaerobic spore treatment. METHOD: Human papillomavirus 16 E6/E7 transformed TC-1 tumour bearing mice were intravenously administered with low (1 × 108 CFU/kg) or high dosage (3 × 108 CFU/kg) of Derivative Clostridial spore (DCG). RESULTS: Intravenous administration of the derivative of Clostridial ghonii (DCG) spores leads to both tumour and systemic inflammatory responses characterized by increased IFNγ/IL-9 secreting T cells in the spleen and the tumour. Low numbers of antigen specific T cells (<20/106 spleen cells) in the spleen of the tumour bearing mice are also detected after intravenous DCG delivery. Interestingly, our results showed that a mixed IL-9/IFNγ secreting T cell response was induced when the tumour bearing mice received a low dose of DCG spore (1 × 108 CFU/kg), while a strong IFNγ response was elicited with a high dosage of DCG spore (3 × 108 CFU/kg). CONCLUSION: The dosage of DCG spore will determine the types of the DCG induced immune responses.
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Clostridiales/genética , Interferón gamma/genética , Interleucina-9/genética , Neoplasias del Cuello Uterino/genética , Animales , Clostridiales/inmunología , Femenino , Papillomavirus Humano 16/genética , Humanos , Ratones , Proteínas Oncogénicas Virales/genética , Proteínas Represoras/genética , Esporas Bacterianas/genética , Esporas Bacterianas/inmunología , Linfocitos T/inmunología , Linfocitos T/microbiología , Transformación Bacteriana/genética , Microambiente Tumoral/inmunología , Neoplasias del Cuello Uterino/inmunología , Neoplasias del Cuello Uterino/microbiología , Neoplasias del Cuello Uterino/virologíaRESUMEN
AIMS: NOD2 and CD1d play a key role in innate immunity by recognizing conserved molecular patterns of pathogens. While NOD2-/- and CD1d-/- mice display structural and functional alterations in Paneth cells, animal studies have reported no impact of NOD2 or CD1d deficiency on experimental colitis. NOD2 mutations increase the susceptibility to inflammatory bowel diseases and the CD1d bound to α-galactosylceramide [α-GalCer] alleviates intestinal inflammation. We evaluated the effect of CD1d modulation on experimental colitis in NOD2-/- mice. METHODS: The effect of CD1d augmentation and depletion in NOD2-/- mice was assessed in a dextran sodium sulphate [DSS]-induced colitis model via administration of α-GalCer and construction of NOD2-/-CD1d-/- mice. The structural and functional changes in Paneth cells were evaluated using transmission electron microscopy and pilocarpine administration. Colitogenic taxa were analysed in the faeces of NOD2-/-CD1d-/- mice using 16S rRNA gene sequencing. RESULTS: In NOD2-/- mice, α-GalCer alleviated and CD1d depletion [NOD2-/-CD1d-/- mice] aggravated colitis activity and histology compared with co-housed littermates NOD2-/-, CD1d-/- and wild-type mice after administration of 3% DSS. In NOD2-/-CD1d-/- mice, the ultrastructure and degranulation ability of secretary granules in Paneth cells were altered and the intestinal microbial composition differed from that of their littermates. Faecal microbiota transplantation [FMT] with NOD2-/-CD1d-/- mice faeces into wild-type mice aggravated DSS-induced colitis, while FMT with wild-type mice faeces into NOD2-/-CD1d-/- mice alleviated DSS-induced colitis. Acetatifactor muris was identified only in NOD2-/-CD1d-/- mice faeces and the oral gavage of A. muris in wild-type mice aggravated DSS-induced colitis. CONCLUSION: CD1d modulates colonic inflammation in NOD2-/- mice by altering the intestinal microbial composition comprising A. muris.
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Antígenos CD1d/inmunología , Clostridiales , Microbioma Gastrointestinal/inmunología , Enfermedades Inflamatorias del Intestino/inmunología , Proteína Adaptadora de Señalización NOD2/inmunología , Células de Paneth/inmunología , Animales , Clostridiales/inmunología , Clostridiales/aislamiento & purificación , Modelos Animales de Enfermedad , Inflamación/inmunología , Enfermedades Inflamatorias del Intestino/patología , RatonesRESUMEN
BACKGROUND/PURPOSE: To search for a transmissible agent involved in lupus pathogenesis, we investigated the faecal microbiota of patients with systemic lupus erythematosus (SLE) for candidate pathobiont(s) and evaluated them for special relationships with host immunity. METHODS: In a cross-sectional discovery cohort, matched blood and faecal samples from 61 female patients with SLE were obtained. Faecal 16 S rRNA analyses were performed, and sera profiled for antibacterial and autoantibody responses, with findings validated in two independent lupus cohorts. RESULTS: Compared with controls, the microbiome in patients with SLE showed decreased species richness diversity, with reductions in taxonomic complexity most pronounced in those with high SLE disease activity index (SLEDAI). Notably, patients with SLE had an overall 5-fold greater representation of Ruminococcus gnavus (RG) of the Lachnospiraceae family, and individual communities also displayed reciprocal contractions of a species with putative protective properties. Gut RG abundance correlated with serum antibodies to only 1/8 RG strains tested. Anti-RG antibodies correlated directly with SLEDAI score and antinative DNA levels, but inversely with C3 and C4. These antibodies were primarily against antigen(s) in an RG strain-restricted pool of cell wall lipoglycans. Novel structural features of these purified lipoglycans were characterised by mass spectrometry and NMR. Highest levels of serum anti-RG strain-restricted antibodies were detected in those with active nephritis (including Class III and IV) in the discovery cohort, with findings validated in two independent cohorts. CONCLUSION: These findings suggest a novel paradigm in which specific strains of a gut commensal may contribute to the immune pathogenesis of lupus nephritis.