RESUMEN
The intestinal microbiome has emerged as a potential contributor to the severity of sickle cell disease (SCD). We sought to determine whether SCD mice exhibit intestinal barrier dysfunction, inflammation, and dysbiosis. Using the Townes humanized sickle cell mouse model, we found a 3-fold increase in intestinal permeability as assessed via FITC-dextran (4 kDa) assay in SS (SCD) mice compared to AA (wild type) mice (n = 4, p < 0.05). This was associated with 25 to 50% decreases in claudin-1, 3, and 15 and zonula occludens-1 gene expression (n = 8-10, p < 0.05) in the small intestine. Increased Ly6G staining demonstrated more neutrophils in the SS small intestine (3-fold, n = 5, p < 0.05) associated with increased expression of TNFα, IL-17A, CXCL1, and CD68 (2.5 to 5-fold, n = 7-10, p < 0.05). In addition, we observed 30 to 55% decreases in superoxide dismutase-1, glutathione peroxidase-1, and catalase antioxidant enzyme expression (n = 7-8, p < 0.05) concomitant to an increase in superoxide (2-fold, n = 4, p < 0.05). Importantly, all significant observations of a leaky gut phenotype and inflammation were limited to the small intestine and not observed in the colon. Finally, characterization of the composition of the microbiome within the small intestine revealed dysbiosis in SS mice compared to their AA littermates with 47 phyla to species-level significant alterations in amplicon sequence variants. We conclude that the intestinal barrier is compromised in SCD, associated with decreased gene expression of tight junction proteins, enhanced inflammation, oxidative stress, and gut microbiome dysbiosis, all specific to the small intestine.
RESUMEN
Obesity and obesity-related metabolic disorders are linked to the intestinal microbiome. However, the causality of changes in the microbiome-host interaction affecting energy metabolism remains controversial. Here, we show the microbiome-derived metabolite δ-valerobetaine (VB) is a diet-dependent obesogen that is increased with phenotypic obesity and is correlated with visceral adipose tissue mass in humans. VB is absent in germ-free mice and their mitochondria but present in ex-germ-free conventionalized mice and their mitochondria. Mechanistic studies in vivo and in vitro show VB is produced by diverse bacterial species and inhibits mitochondrial fatty acid oxidation through decreasing cellular carnitine and mitochondrial long-chain acyl-coenzyme As. VB administration to germ-free and conventional mice increases visceral fat mass and exacerbates hepatic steatosis with a western diet but not control diet. Thus, VB provides a molecular target to understand and potentially manage microbiome-host symbiosis or dysbiosis in diet-dependent obesity.
Asunto(s)
Metabolismo Energético , Interacciones Microbiota-Huesped , Microbiota , Obesidad/metabolismo , Adiposidad , Animales , Dieta Occidental , Ácidos Grasos/metabolismo , Microbioma Gastrointestinal , Humanos , Metabolismo de los Lípidos , Hígado/metabolismo , Ratones , Mitocondrias/metabolismo , Obesidad/etiología , Oxidación-ReducciónRESUMEN
BACKGROUND & AIMS: In colorectal cancer, approximately 95% of patients are refractory to immunotherapy because of low antitumor immune responses. Therefore, there is an exigent need to develop treatments that increase antitumor immune responses and decrease tumor burden to enhance immunotherapy. METHODS: The gut microbiome has been described as a master modulator of immune responses. We administered the human commensal, Lactobacillus rhamnosus GG (LGG), to mice and characterized the changes in the gut immune landscape. Because the presence of lactobacilli in the gut microbiome has been linked with decreased tumor burden and antitumor immune responses, we also supplemented a genetic and a chemical model of murine intestinal cancer with LGG. For clinical relevance, we therapeutically administered LGG after tumors had formed. We also tested for the requirement of CD8 T cells in LGG-mediated modulation of gut tumor burden. RESULTS: We detected increased colonic CD8 T-cell responses specifically in LGG-supplemented mice. The CD8 T-cell induction was dependent on dendritic cell activation mediated via Toll-like receptor-2, thereby describing a novel mechanism in which a member of the human microbiome induces an intestinal CD8 T-cell response. We also show that LGG decreased tumor burden in the murine gut cancer models by a CD8 T-cell-dependent manner. CONCLUSIONS: These data support the potential use of LGG to augment antitumor immune responses in colorectal cancer patients and ultimately for increasing the breadth and efficacy of immunotherapy.
Asunto(s)
Inmunidad , Inmunomodulación , Lacticaseibacillus rhamnosus/inmunología , Neoplasias/inmunología , Animales , Linfocitos T CD8-positivos/inmunología , Linfocitos T CD8-positivos/metabolismo , Comunicación Celular , Colon , Células Dendríticas/inmunología , Células Dendríticas/metabolismo , Modelos Animales de Enfermedad , Microbioma Gastrointestinal , Interacciones Microbiota-Huesped/inmunología , Humanos , Mucosa Intestinal/inmunología , Mucosa Intestinal/metabolismo , Mucosa Intestinal/microbiología , Mucosa Intestinal/patología , Ratones , Neoplasias/metabolismo , Neoplasias/patología , Neoplasias/terapia , Probióticos/administración & dosificación , Transducción de Señal , Receptor Toll-Like 2/metabolismo , Carga TumoralRESUMEN
Genetic factors account for the majority of the variance of human bone mass, but the contribution of non-genetic factors remains largely unknown. By utilizing maternal/offspring transmission, cohabitation, or fecal material transplantation (FMT) studies, we investigated the influence of the gut microbiome on skeletal maturation. We show that the gut microbiome is a communicable regulator of bone structure and turnover in mice. In addition, we found that the acquisition of a specific bacterial strain, segmented filamentous bacteria (SFB), a gut microbe that induces intestinal Th17 cell expansion, was sufficient to negatively impact skeletal maturation. These findings have significant translational implications, as the identification of methods or timing of microbiome transfer may lead to the development of bacteriotherapeutic interventions to optimize skeletal maturation in humans. Moreover, the transfer of SFB-like microbes capable of triggering the expansion of human Th17 cells during therapeutic FMT procedures could lead to significant bone loss in fecal material recipients.
Asunto(s)
Microbioma Gastrointestinal , Esqueleto/crecimiento & desarrollo , Animales , Trasplante de Microbiota Fecal , Heces/microbiología , Femenino , RatonesRESUMEN
The gut microbiota is an important contributor to both health and disease. While previous studies have reported on the beneficial influences of the gut microbiota and probiotic supplementation on bone health, their role in recovery from skeletal injury and resultant systemic sequelae remains unexplored. This study aimed to determine the extent to which probiotics could modulate bone repair by dampening fracture-induced systemic inflammation. Our findings demonstrate that femur fracture induced an increase in gut permeability lasting up to 7 days after trauma before returning to basal levels. Strikingly, dietary supplementation with Bifidobacterium adolescentis augmented the tightening of the intestinal barrier, dampened the systemic inflammatory response to fracture, accelerated fracture callus cartilage remodeling, and elicited enhanced protection of the intact skeleton following fracture. Together, these data outline a mechanism whereby dietary supplementation with beneficial bacteria can be therapeutically targeted to prevent the systemic pathologies induced by femur fracture.
Asunto(s)
Bifidobacterium adolescentis , Fracturas Óseas/terapia , Microbioma Gastrointestinal/fisiología , Inflamación/prevención & control , Probióticos/administración & dosificación , Animales , Fracturas Óseas/complicaciones , Fracturas Óseas/microbiología , Inflamación/etiología , Inflamación/microbiología , Masculino , Ratones , Ratones Endogámicos C57BL , Permeabilidad , Probióticos/farmacologíaRESUMEN
BACKGROUND & AIMS: The intestinal epithelium must be resilient to physiochemical stress to uphold the physiological barrier separating the systemic compartment from the microbial and antigenic components of the gut lumen. Identifying proteins that mediate protection and enhancing their expression is therefore a clear approach to promote intestinal health. We previously reported that oral ingestion of the probiotic Lactobacillus rhamnosus GG not only induced the expression of several recognized cytoprotective factors in the murine colon, but also many genes with no previously described function, including the gene encoding proline-rich acidic protein 1 (PRAP1). PRAP1 is a highly expressed protein in the epithelium of the gastrointestinal tract and we sought to define its function in this tissue. METHODS: Purified preparations of recombinant PRAP1 were analyzed biochemically and PRAP1 antisera were used to visualize localization in tissues. Prap1-/- mice were characterized at baseline and challenged with total body irradiation, then enteroids were generated to recapitulate the irradiation challenge ex vivo. RESULTS: PRAP1 is a 17-kilodalton intrinsically disordered protein with no recognizable sequence homology. PRAP1 expression levels were high in the epithelia of the small intestine. Although Prap1-/- mice presented only mild phenotypes at baseline, they were highly susceptible to intestinal injury upon challenge. After irradiation, the Prap1-/- mice showed accelerated death with a significant increase in apoptosis and p21 expression in the small intestinal epithelium. CONCLUSIONS: PRAP1 is an intrinsically disordered protein highly expressed by the gastrointestinal epithelium and functions at exposed surfaces to protect the barrier from oxidative insult.
Asunto(s)
Apoptosis/efectos de la radiación , Mucosa Intestinal/metabolismo , Mucosa Intestinal/efectos de la radiación , Proteínas Gestacionales/metabolismo , Animales , Línea Celular , Células Cultivadas , Microbioma Gastrointestinal , Eliminación de Gen , Humanos , Mucosa Intestinal/citología , Mucosa Intestinal/microbiología , Ratones , Ratones Endogámicos C57BL , Proteínas Gestacionales/análisis , Proteínas Gestacionales/genéticaRESUMEN
Many studies have suggested a role for gut-resident microbes (the "gut microbiome") in modulating host health; however, the mechanisms by which they impact systemic physiology remain largely unknown. In this study, metabolomic and transcriptional profiling of germ-free and conventionalized mouse liver revealed an upregulation of the Nrf2 antioxidant and xenobiotic response in microbiome-replete animals. Using a Drosophila-based screening assay, we identified members of the genus Lactobacillus capable of stimulating Nrf2. Indeed, the human commensal Lactobacillus rhamnosus GG (LGG) potently activated Nrf2 in the Drosophila liver analog and the murine liver. This activation was sufficient to protect against two models of oxidative liver injury, acetaminophen overdose and acute ethanol toxicity. Characterization of the portal circulation of LGG-treated mice by tandem mass spectrometry identified a small molecule activator of Nrf2, 5-methoxyindoleacetic acid, produced by LGG. Taken together, these data demonstrate a mechanism by which intestinal microbes modulate hepatic susceptibility to oxidative injury.
Asunto(s)
Lacticaseibacillus rhamnosus/metabolismo , Hígado/metabolismo , Factor 2 Relacionado con NF-E2/metabolismo , Animales , Drosophila , Microbioma Gastrointestinal , Células Hep G2 , Humanos , Hígado/lesiones , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Factor 2 Relacionado con NF-E2/deficiencia , Oxidación-Reducción , Células Tumorales CultivadasRESUMEN
BACKGROUND & AIMS: A Western-style diet, which is high in fat and sugar, can cause significant dyslipidemia and nonalcoholic fatty liver disease; the diet has an especially strong effect in women, regardless of total calorie intake. Dietary supplementation with beneficial microbes might reduce the detrimental effects of a Western-style diet. We assessed the effects of Lactococcus lactis subspecies (subsp) cremoris on weight gain, liver fat, serum cholesterol, and insulin resistance in female mice on a high-fat, high-carbohydrate diet. METHODS: Female C57BL/6 mice were fed either a high-fat, high-carbohydrate (Western-style) diet that contained 40% fat (mostly milk fat) and 43% carbohydrate (mostly sucrose) or a calorie-matched-per-gram control diet. The diets of mice were supplemented with 1 × 109 colony-forming units of L lactis subsp cremoris ATCC 19257 or Lactobacillus rhamnosus GG ATCC 53103 (control bacteria) 3 times per week for 16 weeks. Body weights were measured, and fecal, blood, and liver tissues were collected and analyzed. Livers were analyzed for fat accumulation and inflammation, and blood samples were analyzed for cholesterol and glucose levels. Mice were housed within Comprehensive Lab Animal Monitoring System cages, and respiratory exchange ratio and activity were measured. Hepatic lipid profiles of L lactis subsp cremoris-supplemented mice were characterized by lipidomics mass spectrometry analysis. RESULTS: Mice fed L lactis subsp cremoris while on the Western-style diet gained less weight, developed less hepatic steatosis and inflammation, and had a lower mean serum level of cholesterol and body mass index than mice fed the control bacteria. Mice fed the L lactis subsp cremoris had increased glucose tolerance while on the Western-style diet compared to mice fed control bacteria and had alterations in hepatic lipids, including oxylipins. CONCLUSIONS: Dietary supplementation with L lactis subsp cremoris in female mice on a high-fat, high-carbohydrate (Western-style) diet caused them to gain less weight, develop less liver fat and inflammation, reduce serum cholesterol levels, and increase glucose tolerance compared with mice on the same diet fed control bacteria. L lactis subsp cremoris is safe for oral ingestion and might be developed for persons with metabolic and liver disorders caused by a Western-style diet.
Asunto(s)
Dieta Occidental/efectos adversos , Dislipidemias/prevención & control , Lactococcus , Enfermedad del Hígado Graso no Alcohólico/prevención & control , Probióticos/administración & dosificación , Animales , Colesterol/sangre , Colesterol/metabolismo , Modelos Animales de Enfermedad , Dislipidemias/sangre , Dislipidemias/diagnóstico , Dislipidemias/metabolismo , Femenino , Humanos , Resistencia a la Insulina , Hígado/patología , Ratones , Enfermedad del Hígado Graso no Alcohólico/sangre , Enfermedad del Hígado Graso no Alcohólico/metabolismo , Enfermedad del Hígado Graso no Alcohólico/patología , Aumento de PesoRESUMEN
BACKGROUND & AIMS: Identifying the functional elements that mediate efficient gut epithelial growth and homeostasis is essential for understanding intestinal health and disease. Many of these processes involve the Lactobacillus-induced generation of reactive oxygen species by NADPH oxidase (Nox1). However, the downstream signaling pathways that respond to Nox1-generated reactive oxygen species and mediate these events have not been described. METHODS: Wild-type and knockout mice were fed Lactobacillus rhamnosus GG and the transcriptional and cell signaling pathway responses in the colon measured. Corroboration of data generated in mice was done using in organoid tissue culture and in vivo gut injury models. RESULTS: Ingestion of L rhamnosus GG induces elevated levels of leptin in the gut epithelia, which as well as functioning in the context of metabolism, has pleiotropic activity as a chemokine that triggers cell proliferation. Consistently, using gut epithelial-specific knockout mice, we show that L rhamnosus GG-induced elevated levels of leptin is dependent on a functional Nox1 protein in the colonic epithelium, and that L rhamnosus GG-induced cell proliferation is dependent on Nox1, leptin, and leptin receptor. We also show that L rhamnosus GG induces the JAK-STAT signaling pathway in the gut in a Nox1, leptin, and leptin receptor-dependent manner. CONCLUSIONS: These results demonstrate a novel role for leptin in the response to colonization by lactobacilli, where leptin functions in the transduction of signals from symbiotic bacteria to subepithelial compartments, where it modulates intestinal growth and homeostasis.
Asunto(s)
Células Epiteliales/fisiología , Mucosa Intestinal/metabolismo , Lacticaseibacillus rhamnosus , Leptina/metabolismo , Probióticos/administración & dosificación , Animales , Proliferación Celular , Células Cultivadas , Colon/citología , Colon/metabolismo , Mucosa Intestinal/citología , Quinasas Janus/metabolismo , Ratones , Ratones Noqueados , Modelos Animales , NADPH Oxidasa 1/genética , NADPH Oxidasa 1/metabolismo , Cultivo Primario de Células , Especies Reactivas de Oxígeno/metabolismo , Receptores de Leptina/metabolismo , Factores de Transcripción STAT/metabolismo , Transducción de SeñalRESUMEN
The use of beneficial bacteria to promote health is widely practiced. However, experimental evidence corroborating the efficacy of bacteria promoted with such claims remains limited. We address this gap by identifying a beneficial bacterium that protects against tissue damage and injury-induced inflammation in the gut. We first employed the Drosophila animal model to screen for the capacity of candidate beneficial bacteria to protect the fly gut against injury. From this screen, we identified Lactococcus lactis subsp. cremoris as a bacterium that elicited potent cytoprotective activity. Then, in a murine model, we demonstrated that the same strain confers powerful cytoprotective influences against radiological damage, as well as anti-inflammatory activity in a gut colitis model. In summary, we demonstrate the positive salutary effects of a beneficial bacterium, namely, L. lactis subsp. cremoris on intestinal tissue and propose the use of this strain as a therapeutic to promote intestinal health.
RESUMEN
Intestinal homeostasis is regulated in-part by reactive oxygen species (ROS) that are generated in the colonic mucosa following contact with certain lactobacilli. Mechanistically, ROS can modulate protein function through the oxidation of cysteine residues within proteins. Recent advances in cysteine labeling by the Isotope Coded Affinity Tags (ICATs) technique has facilitated the identification of cysteine thiol modifications in response to stimuli. Here, we used ICATs to map the redox protein network oxidized upon initial contact of the colonic mucosa with Lactobacillus rhamnosus GG (LGG). We detected significant LGG-specific redox changes in over 450 proteins, many of which are implicated to function in cellular processes such as endosomal trafficking, epithelial cell junctions, barrier integrity, and cytoskeleton maintenance and formation. We particularly noted the LGG-specific oxidation of Rac1, which is a pleiotropic regulator of many cellular processes. Together, these data reveal new insights into lactobacilli-induced and redox-dependent networks involved in intestinal homeostasis.
Asunto(s)
Mucosa Intestinal/metabolismo , Oxidación-Reducción , Proteoma , Proteómica , Transducción de Señal , Línea Celular , Biología Computacional/métodos , Modelos Biológicos , Estrés Oxidativo , Especies Reactivas de Oxígeno/metabolismo , Espectrometría de Masas en TándemRESUMEN
Nutritional supplementation with probiotics can prevent pathologic bone loss. Here we examined the impact of supplementation with Lactobacillus rhamnosus GG (LGG) on bone homeostasis in eugonadic young mice. Micro-computed tomography revealed that LGG increased trabecular bone volume in mice, which was due to increased bone formation. Butyrate produced in the gut following LGG ingestion, or butyrate fed directly to germ-free mice, induced the expansion of intestinal and bone marrow (BM) regulatory T (Treg) cells. Interaction of BM CD8+ T cells with Treg cells resulted in increased secretion of Wnt10b, a bone anabolic Wnt ligand. Mechanistically, Treg cells promoted the assembly of a NFAT1-SMAD3 transcription complex in CD8+ cells, which drove expression of Wnt10b. Reducing Treg cell numbers, or reconstitution of TCRß-/- mice with CD8+ T cells from Wnt10b-/- mice, prevented butyrate-induced bone formation and bone mass acquisition. Thus, butyrate concentrations regulate bone anabolism via Treg cell-mediated regulation of CD8+ T cell Wnt10b production.
Asunto(s)
Butiratos/farmacología , Osteogénesis/efectos de los fármacos , Linfocitos T Reguladores/metabolismo , Proteínas Wnt/metabolismo , Animales , Butiratos/metabolismo , Linfocitos T CD8-positivos/citología , Linfocitos T CD8-positivos/metabolismo , Comunicación Celular , Proliferación Celular/efectos de los fármacos , Femenino , Lacticaseibacillus rhamnosus/metabolismo , Ratones Endogámicos C57BL , Ratones Noqueados , Osteoblastos/citología , Osteoblastos/efectos de los fármacos , Osteoblastos/metabolismo , Probióticos/administración & dosificación , Probióticos/metabolismo , Linfocitos T Reguladores/citología , Proteínas Wnt/genéticaRESUMEN
Inflammatory bowel disease (IBD) results from aberrant immune stimulation against a dysbiotic mucosal but relatively preserved luminal microbiota and preferentially affects males in early onset disease. However, factors contributing to sex-specific risk and the pattern of dysbiosis are largely unexplored. Core 1 ß3GalT-specific molecular chaperone (Cosmc), which encodes an X-linked chaperone important for glycocalyx formation, was recently identified as an IBD risk factor by genome-wide association study. We deleted Cosmc in mouse intestinal epithelial cells (IECs) and found marked reduction of microbiota diversity in progression from the proximal to the distal gut mucosa, but not in the overlying lumen, as seen in IBD. This loss of diversity coincided with local emergence of a proinflammatory pathobiont and distal gut restricted pathology. Mechanistically, we found that Cosmc regulates host genes, bacterial ligands, and nutrient availability to control microbiota biogeography. Loss of one Cosmc allele in males (IEC-Cosmc-/y) resulted in a compromised mucus layer, spontaneous microbe-dependent inflammation, and enhanced experimental colitis; however, females with loss of one allele and mosaic deletion of Cosmc in 50% of crypts (IEC-Cosmc+/-) were protected from spontaneous inflammation and partially protected from experimental colitis, likely due to lateral migration of normal mucin glycocalyx from WT cells over KO crypts. These studies functionally validate Cosmc as an IBD risk factor and implicate it in regulating the spatial pattern of dysbiosis and sex bias in IBD.
Asunto(s)
Microbioma Gastrointestinal , Genes Ligados a X , Enfermedades Inflamatorias del Intestino/genética , Chaperonas Moleculares/genética , Factores Sexuales , Alelos , Animales , Colitis/microbiología , Femenino , Eliminación de Gen , Ligamiento Genético , Estudio de Asociación del Genoma Completo , Glicocálix , Inflamación , Enfermedades Inflamatorias del Intestino/microbiología , Mucosa Intestinal/microbiología , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Mosaicismo , Factores de Riesgo , Cromosoma XRESUMEN
A eubiotic microbiota influences many physiological processes in the metazoan host, including development and intestinal homeostasis. Here, we have shown that the intestinal microbiota modulates inflammatory responses caused by sex steroid deficiency, leading to trabecular bone loss. In murine models, sex steroid deficiency increased gut permeability, expanded Th17 cells, and upregulated the osteoclastogenic cytokines TNFα (TNF), RANKL, and IL-17 in the small intestine and the BM. In germ-free (GF) mice, sex steroid deficiency failed to increase osteoclastogenic cytokine production, stimulate bone resorption, and cause trabecular bone loss, demonstrating that the gut microbiota is central in sex steroid deficiency-induced trabecular bone loss. Furthermore, we demonstrated that twice-weekly treatment of sex steroid-deficient mice with the probiotics Lactobacillus rhamnosus GG (LGG) or the commercially available probiotic supplement VSL#3 reduces gut permeability, dampens intestinal and BM inflammation, and completely protects against bone loss. In contrast, supplementation with a nonprobiotic strain of E. coli or a mutant LGG was not protective. Together, these data highlight the role that the gut luminal microbiota and increased gut permeability play in triggering inflammatory pathways that are critical for inducing bone loss in sex steroid-deficient mice. Our data further suggest that probiotics that decrease gut permeability have potential as a therapeutic strategy for postmenopausal osteoporosis.
Asunto(s)
Microbioma Gastrointestinal/fisiología , Hormonas Esteroides Gonadales/deficiencia , Osteoporosis Posmenopáusica/etiología , Osteoporosis Posmenopáusica/prevención & control , Probióticos/farmacología , Animales , Remodelación Ósea , Fenómenos Fisiológicos del Sistema Digestivo , Modelos Animales de Enfermedad , Femenino , Vida Libre de Gérmenes , Humanos , Interleucina-17/metabolismo , Lacticaseibacillus rhamnosus , Ratones , Ratones Endogámicos C57BL , Osteoporosis Posmenopáusica/patología , Permeabilidad , Ligando RANK/metabolismo , Células Th17/inmunología , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
An optimal gut microbiota influences many beneficial processes in the metazoan host. However, the molecular mechanisms that mediate and function in symbiont-induced host responses have not yet been fully characterized. Here, we report that cellular ROS enzymatically generated in response to contact with lactobacilli in both mice and Drosophila has salutary effects against exogenous insults to the intestinal epithelium via the activation of Nrf2 responsive cytoprotective genes. These data show that the xenobiotic-inducible Nrf2 pathway participates as a signaling conduit between the prokaryotic symbiont and the eukaryotic host. Indeed, our data imply that the capacity of lactobacilli to induce redox signaling in epithelial cells is a highly conserved hormetic adaptation to impel cellular conditioning to exogenous biotic stimuli. These data also highlight the role the microbiota plays in eukaryotic cytoprotective pathways and may have significant implications in the characterization of a eubiotic microbiota.
Asunto(s)
Drosophila/metabolismo , Mucosa Intestinal/metabolismo , Lacticaseibacillus rhamnosus/metabolismo , Lactobacillus plantarum/metabolismo , Factor 2 Relacionado con NF-E2/metabolismo , Animales , Drosophila/microbiología , Mucosa Intestinal/microbiología , Lactobacillus plantarum/patogenicidad , Lacticaseibacillus rhamnosus/patogenicidad , Ratones , Factor 2 Relacionado con NF-E2/genética , Estrés Oxidativo , Transducción de SeñalRESUMEN
BACKGROUND: The aim of this study was to examine the effect of colitis and anti-inflammatory therapies on the healing of colonic anastomoses in mice. METHODS: Female C57BL/6 mice were randomized into eight groups; four groups receiving plain tap-water and four groups receiving dextran sulfate sodium. Intra-peritoneal treatment was given therapeutically for 14 days with placebo, prednisolone, azathioprine, or infliximab (IFX). Colonic anastomoses were performed and bursting pressure (BP) measurements were recorded and the inflammation evaluated with histology and zymography. RESULTS: The mice with colitis had a more active inflammation based on histology and bowel weight compared with the tap water group, 8.3 (7.6-9.5) mg/mm and 5.5 (4.8-6.2) mg/mm respectively (p < 0.0001). Similarly mice with colitis receiving placebo had a more active inflammation, 12.8 (10.6-15.0) mg/mm, which differed significantly from all the other therapy arms among the colitic mice; prednisolone 8.1 (7.5-9.1) mg/mm (p = 0.014), azathioprine 8.2 (7.0-8.5) mg/mm (p = 0.0046), IFX 6.7 (6.4-7.9) mg/mm (p = 0.0055). BP for the placebo group was 90.0 (71.5-102.8) mmHg and did not differ from azathioprine or IFX groups, 84.4 (70.5-112.5) and 92.3 (75.8-122.3) mmHg respectively. In contrast BP for the prednisolone group was significantly decreased compared to placebo, 55.5 (42.8-73.0) mmHg (p = 0.0004). CONCLUSIONS: All therapies had a beneficial effect on the colitis. An impaired BP of colonic anastomoses was noted after preoperative steroids but not after azathioprine or IFX in this model.
Asunto(s)
Antiinflamatorios/uso terapéutico , Azatioprina/uso terapéutico , Colitis/tratamiento farmacológico , Colitis/patología , Infliximab/uso terapéutico , Prednisolona/uso terapéutico , Anastomosis Quirúrgica/efectos adversos , Animales , Colitis/inducido químicamente , Sulfato de Dextran , Modelos Animales de Enfermedad , Femenino , Ratones , Ratones Endogámicos C57BL , Presión , Cicatrización de Heridas/efectos de los fármacosRESUMEN
Inflammasomes are a large family of multiprotein complexes recognizing pathogen-associated molecular pattern molecules (PAMPs) and damage-associated molecular patterns (DAMPs). This leads to caspase-1 activation, promoting the secretion of mature IL-1ß, IL-18 and under certain conditions even induce pyroptosis. Inflammatory Bowel Diseases (IBD) is associated with alterations in microbiota composition, inappropriate immune responses and genetic predisposition associated to bacterial sensing and autophagy. Besides their acknowledged role in mounting microbial induced host responses, a crucial role in maintenance of intestinal homeostasis was revealed in inflammasome deficient mice. Further, abnormal activation of these functions appears to contribute to the pathology of intestinal inflammation including IBD and colitis-associated cancer. Herein, the current literature implicating the inflammasomes, microbiota and IBD is comprehensively reviewed.
Asunto(s)
Inflamasomas/inmunología , Enfermedades Inflamatorias del Intestino/inmunología , Animales , Bacterias/inmunología , Modelos Animales de Enfermedad , Humanos , Inflamasomas/genética , Enfermedades Inflamatorias del Intestino/genética , Enfermedades Inflamatorias del Intestino/microbiología , Interleucina-18/genética , Interleucina-18/inmunología , Interleucina-1beta/genética , Interleucina-1beta/inmunología , Ratones , Microbiota/inmunologíaRESUMEN
There is now convincing evidence that liver X receptor (LXR) is an important modulator of the inflammatory response; however, its mechanism of action remains unclear. This study aimed to examine the effect of LXR on the IL-12 family of cytokines and examined the mechanism by which LXR exerted this effect. We first demonstrated that activation of murine-derived dendritic cells (DC) with a specific agonist to LXR enhanced expression of LXR following activation with LPS, suggesting a role in inflammation. Furthermore, we showed LXR expression to be increased in vivo in dextrane sulphate sodium-induced colitis. LXR activation also suppressed production of IL-12p40, IL-12p70, IL-27 and IL-23 in murine-derived DC following stimulation with LPS, and specifically targeted the p35, p40 and EBI3 subunits of the IL-12 cytokine family, which are under the control of the NF-κB subunit p50 (NF-κBp50). Finally, we demonstrated that LXR can associate with NF-κBp50 in DC and that LXR activation prevents translocation of the p50 subunit into the nucleus. In summary, our study indicates that LXR can specifically suppress the IL-12 family of cytokines though its association with NF-κBp50 and highlights its potential as a therapeutic target for chronic inflammatory diseases.
Asunto(s)
Interleucina-12/biosíntesis , Subunidad p50 de NF-kappa B/antagonistas & inhibidores , Receptores Nucleares Huérfanos/efectos de los fármacos , Animales , Células de la Médula Ósea/efectos de los fármacos , Células de la Médula Ósea/inmunología , Núcleo Celular/metabolismo , Núcleo Celular/patología , Colitis/inducido químicamente , Colitis/metabolismo , Citocinas/biosíntesis , Citoplasma/metabolismo , Citoplasma/patología , Inflamación/metabolismo , Inflamación/patología , Receptores X del Hígado , Ratones , Ratones Endogámicos BALB C , Transporte de Proteínas , ARN Mensajero/biosíntesis , ARN Mensajero/genéticaRESUMEN
Mutations that result in loss of function of Nod2, an intracellular receptor for bacterial peptidoglycan, are associated with Crohn's disease. Here we found that the E3 ubiquitin ligase Pellino3 was an important mediator in the Nod2 signaling pathway. Pellino3-deficient mice had less induction of cytokines after engagement of Nod2 and had exacerbated disease in various experimental models of colitis. Furthermore, expression of Pellino3 was lower in the colons of patients with Crohn's disease. Pellino3 directly bound to the kinase RIP2 and catalyzed its ubiquitination. Loss of Pellino3 led to attenuation of Nod2-induced ubiquitination of RIP2 and less activation of the transcription factor NF-κB and mitogen-activated protein kinases (MAPKs). Our findings identify RIP2 as a substrate for Pellino3 and Pellino3 as an important mediator in the Nod2 pathway and regulator of intestinal inflammation.
Asunto(s)
Colitis/metabolismo , Proteína Adaptadora de Señalización NOD2/metabolismo , Proteína Serina-Treonina Quinasas de Interacción con Receptores/metabolismo , Transducción de Señal , Ubiquitina-Proteína Ligasas/genética , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Animales , Citrobacter rodentium/inmunología , Colitis/genética , Colitis/inmunología , Colitis/microbiología , Enfermedad de Crohn/genética , Enfermedad de Crohn/inmunología , Enfermedad de Crohn/metabolismo , Modelos Animales de Enfermedad , Femenino , Expresión Génica , Humanos , Masculino , Ratones , Ratones Noqueados , Persona de Mediana Edad , Unión Proteica , Dominios y Motivos de Interacción de Proteínas , Proteína Serina-Treonina Quinasa 2 de Interacción con Receptor , Ubiquitina-Proteína Ligasas/química , Ubiquitina-Proteína Ligasas/metabolismo , Ubiquitinación , Adulto JovenRESUMEN
Lactobacillus ruminis is one of at least twelve motile but poorly characterized species found in the genus Lactobacillus. Of these, only L. ruminis has been isolated from mammals, and this species may be considered as an autochthonous member of the gastrointestinal microbiota of humans, pigs and cows. Nine L. ruminis strains were investigated here to elucidate the biochemistry and genetics of Lactobacillus motility. Six strains isolated from humans were non-motile while three bovine isolates were motile. A complete set of flagellum biogenesis genes was annotated in the sequenced genomes of two strains, ATCC25644 (human isolate) and ATCC27782 (bovine isolate), but only the latter strain produced flagella. Comparison of the L. ruminis and L. mali DSM20444(T) motility loci showed that their genetic content and gene-order were broadly similar, although the L. mali motility locus was interrupted by an 11.8 Kb region encoding rhamnose utilization genes that is absent from the L. ruminis motility locus. Phylogenetic analysis of 39 motile bacteria indicated that Lactobacillus motility genes were most closely related to those of motile carnobacteria and enterococci. Transcriptome analysis revealed that motility genes were transcribed at a significantly higher level in motile L. ruminis ATCC27782 than in non-motile ATCC25644. Flagellin proteins were isolated from L. ruminis ATCC27782 and from three other Lactobacillus species, while recombinant flagellin of aflagellate L. ruminis ATCC25644 was expressed and purified from E. coli. These native and recombinant Lactobacillus flagellins, and also flagellate L. ruminis cells, triggered interleukin-8 production in cultured human intestinal epithelial cells in a manner suppressed by short interfering RNA directed against Toll-Like Receptor 5. This study provides genetic, transcriptomic, phylogenetic and immunological insights into the trait of flagellum-mediated motility in the lactobacilli.