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1.
Nutrients ; 16(10)2024 May 19.
Artículo en Inglés | MEDLINE | ID: mdl-38794769

RESUMEN

Several metabolites of the essential amino acid tryptophan have emerged as key players in gut homeostasis through different cellular pathways, particularly through metabolites which can activate the aryl hydrocarbon receptor (AHR). This study aimed to map the metabolism of tryptophan in early life and investigate the effects of specific metabolites on epithelial cells and barrier integrity. Twenty-one tryptophan metabolites were measured in the feces of full-term and preterm neonates as well as in human milk and formula. The ability of specific AHR metabolites to regulate cytokine-induced IL8 expression and maintain barrier integrity was assessed in Caco2 cells and human fetal organoids (HFOs). Overall, higher concentrations of tryptophan metabolites were measured in the feces of full-term neonates compared to those of preterm ones. Within AHR metabolites, indole-3-lactic acid (ILA) was significantly higher in the feces of full-term neonates. Human milk contained different levels of several tryptophan metabolites compared to formula. Particularly, within the AHR metabolites, indole-3-sulfate (I3S) and indole-3-acetic acid (IAA) were significantly higher compared to formula. Fecal-derived ILA and milk-derived IAA were capable of reducing TNFα-induced IL8 expression in Caco2 cells and HFOs in an AHR-dependent manner. Furthermore, fecal-derived ILA and milk-derived IAA significantly reduced TNFα-induced barrier disruption in HFOs.


Asunto(s)
Heces , Leche Humana , Receptores de Hidrocarburo de Aril , Triptófano , Femenino , Humanos , Recién Nacido , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico , Células CACO-2 , Heces/química , Ácidos Indolacéticos/análisis , Ácidos Indolacéticos/metabolismo , Fórmulas Infantiles/química , Recien Nacido Prematuro , Interleucina-8/metabolismo , Leche Humana/metabolismo , Leche Humana/química , Organoides/metabolismo , Receptores de Hidrocarburo de Aril/metabolismo , Triptófano/análisis , Triptófano/metabolismo
2.
J Neuroinflammation ; 19(1): 155, 2022 Jun 17.
Artículo en Inglés | MEDLINE | ID: mdl-35715845

RESUMEN

BACKGROUND: Vagus nerve stimulation has been suggested to affect immune responses, partly through a neuronal circuit requiring sympathetic innervation of the splenic nerve bundle and norepinephrine (NE) release. Molecular and cellular mechanisms of action remain elusive. Here, we investigated the therapeutic value of this neuromodulation in inflammatory bowel disease (IBD) by applying electrical splenic nerve bundle stimulation (SpNS) in mice with dextran sulfate sodium (DSS)-induced colitis. METHODS: Cuff electrodes were implanted around the splenic nerve bundle in mice, whereupon mice received SpNS or sham stimulation. Stimulation was applied 6 times daily for 12 days during DSS-induced colitis. Colonic and splenic tissues were collected for transcriptional analyses by qPCR and RNA-sequencing (RNA-seq). In addition, murine and human splenocytes were stimulated with lipopolysaccharide (LPS) in the absence or presence of NE. Single-cell RNA-seq data from publicly available data sets were analyzed for expression of ß-adrenergic receptors (ß-ARs). RESULTS: Colitic mice undergoing SpNS displayed reduced colon weight/length ratios and showed improved Disease Activity Index scores with reduced Tumor Necrosis Factor α mRNA expression in the colon compared with sham stimulated mice. Analyses of splenocytes from SpNS mice using RNA-seq demonstrated specific immune metabolism transcriptome profile changes in myeloid cells. Splenocytes showed expression of ß-ARs in myeloid and T cells. Cytokine production was reduced by NE in mouse and human LPS-stimulated splenocytes. CONCLUSIONS: Together, our results demonstrate that SpNS reduces clinical features of colonic inflammation in mice with DSS-induced colitis possibly by inhibiting splenic myeloid cell activation. Our data further support exploration of the clinical use of SpNS for patients with IBD.


Asunto(s)
Colitis , Enfermedades Inflamatorias del Intestino , Animales , Colitis/inducido químicamente , Colitis/terapia , Colon/metabolismo , Colon/patología , Sulfato de Dextran/toxicidad , Modelos Animales de Enfermedad , Estimulación Eléctrica , Humanos , Enfermedades Inflamatorias del Intestino/inducido químicamente , Enfermedades Inflamatorias del Intestino/terapia , Lipopolisacáridos/efectos adversos , Ratones , Ratones Endogámicos C57BL
3.
Microorganisms ; 10(4)2022 Mar 29.
Artículo en Inglés | MEDLINE | ID: mdl-35456788

RESUMEN

Fecal microbiota transplantation (FMT) has the potential to restore (bacterial and fungal) microbial imbalance in ulcerative colitis (UC) patients and contribute to disease remission. Here, we aimed to identify fecal fungal species associated with the induction of clinical remission and endoscopic response to FMT for patients with mild-to-moderate ulcerative colitis. We analyzed the internal transcribed spacer 1 (ITS1)-based mycobiota composition in fecal samples from patients (n = 31) and donors (n = 7) that participated previously in a double-blinded randomized control trial evaluating the efficacy of two infusions of donor FMT compared with autologous FMT. The abundance of the yeast genus Filobasidium in fecal material used for transplantation was shown to correlate with clinical remission following FMT, irrespective of its presence in the material of donor or autologous fecal microbiota transfer. The amplified sequence variants within the genus Filobasidium most closely resembled Filobasidium magnum. Monocyte-derived macrophages and HT29 epithelial cells were stimulated with fungal species. Especially Filobasidium floriforme elicited an IL10 response in monocyte-derived macrophages, along with secretion of other cytokines following stimulation with other Filobasidium species. No effect of Filobasidium spp. was seen on epithelial wound healing in scratch assays. In conclusion, the enriched presence of Filobasidium spp. in donor feces is associated with the positive response to FMT for patients with UC and hence it may serve as a predictive fungal biomarker for successful FMT.

4.
Cells ; 11(1)2021 12 25.
Artículo en Inglés | MEDLINE | ID: mdl-35011620

RESUMEN

Antimicrobial responses play an important role in maintaining intestinal heath. Recently we reported that miR-511 may regulate TLR4 responses leading to enhanced intestinal inflammation. However, the exact mechanism remained unclear. In this study we investigated the effect of miR-511 deficiency on anti-microbial responses and DSS-induced intestinal inflammation. miR-511-deficient mice were protected from DSS-induced colitis as shown by significantly lower disease activity index, weight loss and histology scores in the miR-511-deficient group. Furthermore, reduced inflammatory cytokine responses were observed in colons of miR-511 deficient mice. In vitro studies with bone marrow-derived M2 macrophages showed reduced TLR3 and TLR4 responses in miR-511-deficient macrophages compared to WT macrophages. Subsequent RNA sequencing revealed Wdfy1 as the potential miR-511 target. WDFY1 deficiency is related to impaired TLR3/TLR4 immune responses and the expression was downregulated in miR-511-deficient macrophages and colons. Together, this study shows that miR-511 is involved in the regulation of intestinal inflammation through downstream regulation of TLR3 and TLR4 responses via Wdfy1.


Asunto(s)
Proteínas Adaptadoras Transductoras de Señales/metabolismo , Colitis/genética , MicroARNs/metabolismo , Receptor Toll-Like 3/metabolismo , Receptor Toll-Like 4/metabolismo , Animales , Colitis/inducido químicamente , Colitis/microbiología , Colitis/patología , Colon/patología , Sulfato de Dextran , Femenino , Regulación de la Expresión Génica , Inflamación/genética , Inflamación/patología , Lipopolisacáridos , Macrófagos/metabolismo , Macrófagos/microbiología , Masculino , Ratones Endogámicos C57BL , MicroARNs/genética , Monocitos/metabolismo
5.
J Crohns Colitis ; 15(4): 617-630, 2021 Apr 06.
Artículo en Inglés | MEDLINE | ID: mdl-33005945

RESUMEN

BACKGROUND AND AIMS: Tyrosine kinase 2 [TYK2] is required for the signalling of key cytokines in the pathogenesis of inflammatory bowel disease [IBD]. We assessed the efficacy of a novel selective TYK2 inhibitor [TYK2i] in experimental colitis, using pharmacological and genetic tools. METHODS: At onset of T cell transfer colitis, RAG1-/- mice received vehicle or TYK2i daily by oral gavage. T cells lacking TYK2 kinase activity [TYK2KE] were used to confirm selectivity of the inhibitor. To this end, RAG1-/- or RAG1-/-TYK2KE animals were transferred with either wild type [WT] or TYK2KE-CD45RBhigh colitogenic T cells. Loss of body weight, endoscopic disease, the disease activity index [DAI], and histopathology scores were recorded. Tissues were analysed ex vivo for lymphocyte populations by flow cytometry. The impact of TYK2 inhibition on human DC-T cell interactions were studied using autologous Revaxis specific T cell assays. RESULTS: TYK2i [70 mg/kg] prevented weight loss and limited endoscopic activity during T cell transfer colitis. TYK2i [70 mg/kg] decreased DAI. Whereas transfer of WT T cells into RAG-/-TYK2KE hosts induced colitis, TYK2KE T cells transferred into RAG1-/-TYK2KErecipients failed to do so. Ex vivo analysis showed a decrease in colon tissue Th1 cells and an increase in Th17 cells upon transfer of TYK2KE-CD45RBhigh cells. In human antigen-triggered T cells, TYK2i displayed reduced Th1 differentiation, similar to murine Th1 cells. CONCLUSIONS: Oral administration of TYK2i, as well as transfer of T cells lacking TYK2 activity, reduced human Th1 differentiation and ameliorated the course of murine T cell transfer colitis. We conclude that TYK2 is a promising drug target for the treatment of IBD.


Asunto(s)
Administración Oral , Traslado Adoptivo , Enfermedades Inflamatorias del Intestino/enzimología , Enfermedades Inflamatorias del Intestino/inmunología , Linfocitos T/efectos de los fármacos , Linfocitos T/inmunología , TYK2 Quinasa/antagonistas & inhibidores , Animales , Diferenciación Celular/inmunología , Citocinas/inmunología , Modelos Animales de Enfermedad , Femenino , Citometría de Flujo , Proteínas de Homeodominio , Humanos , Ratones , Transducción de Señal , Células TH1/metabolismo
6.
Bioelectron Med ; 6: 20, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-33123616

RESUMEN

BACKGROUND: Recent evidence demonstrated that severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) propagates in intestinal epithelial cells expressing Angiotensin-Converting Enzyme 2 (ACE2), implying that these cells represent an important entry site for the viral infection. Nicotinic receptors (nAChRs) have been put forward as potential regulators of inflammation and of ACE2 expression. As vagus nerve stimulation (VNS) activates nAChRs, we aimed to investigate whether VNS can be instrumental in affecting intestinal epithelial ACE2 expression. METHODS: By using publicly available datasets we qualified epithelial ACE2 expression in human intestine, and assessed gene co-expression of ACE2 and SARS-CoV-2 priming Transmembrane Serine Protease 2 (TMPRSS2) with nAChRs in intestinal epithelial cells. Next, we investigated mouse and human ACE2 expression in intestinal tissues after chronic VNS via implanted devices. RESULTS: We show co-expression of ACE2 and TMPRSS2 with nAChRs and α7 nAChR in particular in intestinal stem cells, goblet cells, and enterocytes. However, VNS did not affect ACE2 expression in murine or human intestinal tissue, albeit in colitis setting. CONCLUSIONS: ACE2 and TMPRSS2 are specifically expressed in epithelial cells of human intestine, and both are co-expressed with nAChRs. However, no evidence for regulation of ACE2 expression through VNS could be found. Hence, a therapeutic value of VNS with respect to SARS-CoV-2 infection risk through ACE2 receptor modulation in intestinal epithelia could not be established.

7.
Front Immunol ; 11: 550769, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-33123128

RESUMEN

Histone deacetylases (HDACs) are a group of enzymes that control histone deacetylation and bear potential to direct expression of large gene sets. We determined the effect of HDAC inhibitors (HDACi) on human monocytes and macrophages, with respect to their polarization, activation, and their capabilities of inducing endotoxin tolerance. To address the role for HDACs in macrophage polarization, we treated monocytes with HDAC3i, HDAC6i or pan-HDACi prior to polarization into M1 or M2 macrophages using IFNγ or IL-4 respectively. To study the HDAC inhibition effect on cytokine expression, macrophages were treated with HDACi prior to LPS-stimulation. TNFα, IL-6, and p40 were measured with ELISA, whereas modifications of Histone 3 and STAT1 were assessed using western blot. To address the role for HDAC3 in repeated LPS challenge induction, HDAC3i or HDAC3 siRNA was added to monocytes prior to incubation with IFNγ, which were then repeatedly challenged with LPS and analyzed by means of protein analyses and transcriptional profiling. Pan-HDACi and HDAC3i reduced cytokine secretion in monocytes and M1 macrophages, whereas HDAC6i yielded no such effect. Notably, neither pan-HDACi nor HDAC3i reduced cytokine secretion in M2 macrophages. In contrast to previous reports in mouse macrophages, HDAC3i did not affect macrophage polarization in human cells. Likewise, HDAC3 was not required for IFNγ signaling or IFNß secretion. Cytokine and gene expression analyses confirmed that IFNγ-treated macrophages consistently develop a cytokine response after LPS repeated challenge, but pretreatment with HDAC3i or HDAC3 siRNA reinstates a state of tolerance reflected by general suppression of tolerizable genes, possibly through decreasing TLRs expression, and particularly TLR4/CD14. The development of endotoxin tolerance in macrophages is important to reduce exacerbated immune response and limit tissue damage. We conclude that HDAC3 is an attractive protein target to mediate macrophage reactivity and tolerance induction in inflammatory macrophages.


Asunto(s)
Histona Desacetilasas/metabolismo , Tolerancia Inmunológica , Lipopolisacáridos/inmunología , Macrófagos/inmunología , Macrófagos/metabolismo , Monocitos/inmunología , Monocitos/metabolismo , Citocinas/genética , Citocinas/metabolismo , Activación Enzimática , Expresión Génica , Inhibidores de Histona Desacetilasas/farmacología , Histona Desacetilasas/genética , Humanos , Tolerancia Inmunológica/efectos de los fármacos , Inmunofenotipificación , Inflamación/tratamiento farmacológico , Inflamación/etiología , Inflamación/metabolismo , Inflamación/patología , Activación de Macrófagos/efectos de los fármacos , Activación de Macrófagos/inmunología , Macrófagos/efectos de los fármacos , Monocitos/efectos de los fármacos , Unión Proteica
8.
Am J Physiol Gastrointest Liver Physiol ; 317(5): G557-G568, 2019 11 01.
Artículo en Inglés | MEDLINE | ID: mdl-31322912

RESUMEN

Clinical trials suggest that vagus nerve stimulation presents an alternative approach to classical immune suppression in Crohn's disease. T cells capable of producing acetylcholine (ChAT+ T cells) in the spleen are essential mediators of the anti-inflammatory effect of vagus nerve stimulation. Besides the spleen, ChAT+ T cells are found abundantly in Peyer's patches of the small intestine. However, the role of ChAT+ T cells in colitis pathogenesis is unknown. Here, we made use of CD4creChATfl/fl mice (CD4ChAT-/- mice) lacking ChAT expression specifically in CD4+ T cells. Littermates (ChATfl/fl mice) served as controls. In acute dextran sulfate sodium (DSS)-induced colitis (7 days of 2% DSS in drinking water), CD4ChAT-/- mice showed attenuated colitis and lower intestinal inflammatory cytokine levels compared with ChATfl/fl mice. In contrast, in a resolution model of DSS-induced colitis (5 days of 2% DSS followed by 7 days without DSS), CD4ChAT-/- mice demonstrated a worsened colitis recovery and augmented colonic histological inflammation scores and inflammatory cytokine levels as compared with ChATfl/fl mice. In a transfer colitis model using CD4+CD45RBhigh T cells, T cells from CD4ChAT-/- mice induced a similar level of colitis compared with ChATfl/fl T cells. Together, our results indicate that ChAT+ T cells aggravate the acute innate immune response upon mucosal barrier disruption in an acute DSS-induced colitis model, whereas they are supporting the later resolution process of this innate immune-driven colitis. Surprisingly, ChAT expression in T cells seems redundant in the context of T cell-driven colitis.NEW & NOTEWORTHY By using different mouse models of experimental colitis, we provide evidence that in dextran sulfate sodium-induced colitis, ChAT+ T cells capable of producing acetylcholine worsen the acute immune response, whereas they support the later healing phase of this innate immune-driven colitis.


Asunto(s)
Acetilcolina/metabolismo , Linfocitos T CD4-Positivos/inmunología , Colitis Ulcerosa/inmunología , Inmunidad Innata , Animales , Colina O-Acetiltransferasa/genética , Colina O-Acetiltransferasa/metabolismo , Colitis Ulcerosa/etiología , Femenino , Mucosa Intestinal/metabolismo , Ratones , Ratones Endogámicos C57BL , Dodecil Sulfato de Sodio/toxicidad
9.
Mol Med ; 25(1): 1, 2019 01 07.
Artículo en Inglés | MEDLINE | ID: mdl-30616543

RESUMEN

BACKGROUND: Both the parasympathetic and sympathetic nervous system exert control over innate immune responses. In inflammatory bowel disease, sympathetic innervation in intestinal mucosa is reduced. Our aim was to investigate the role of sympathetic innervation to the intestine on regulation of the innate immune responses. METHODS: In lipopolysaccharide (LPS)-stimulated macrophages, we evaluated the effect of adrenergic receptor activation on cytokine production and metabolic profile. In vivo, the effect of sympathetic denervation on mucosal innate immune responses using 6-hydroxydopamine (6-OHDA), or using surgical transection of the superior mesenteric nerve (sympathectomy) was tested in Rag1-/- mice that lack T- and B-lymphocytes. RESULTS: In murine macrophages, adrenergic ß2 receptor activation elicited a dose-dependent reduction of LPS-induced cytokines, reduced LPS-induced glycolysis and increased maximum respiration. Sympathectomy led to a significantly decreased norepinephrine concentration in intestinal tissue. Within 14 days after sympathectomy, mice developed clinical signs of colitis, colon oedema and excess colonic cytokine production. Both 6-OHDA and sympathectomy led to prominent goblet cell depletion and histological damage of colonic mucosa. CONCLUSIONS: We conclude that the sympathetic nervous system plays a regulatory role in constraining innate immune cell reactivity towards microbial challenges, likely via the adrenergic ß2 receptor.


Asunto(s)
Colitis/inmunología , Inmunidad Innata , Inmunidad Mucosa , Mucosa Intestinal/inmunología , Mucosa Intestinal/inervación , Sistema Nervioso Simpático/inmunología , Agonistas de Receptores Adrenérgicos beta 2/farmacología , Albuterol/farmacología , Animales , Células Cultivadas , Colitis/patología , Colon/efectos de los fármacos , Colon/patología , Citocinas/genética , Citocinas/inmunología , Femenino , Humanos , Lipopolisacáridos/farmacología , Macrófagos/efectos de los fármacos , Macrófagos/inmunología , Masculino , Ratones Endogámicos C57BL , Ratones Noqueados , Oxidopamina/farmacología
10.
Am J Physiol Gastrointest Liver Physiol ; 311(5): G920-G933, 2016 11 01.
Artículo en Inglés | MEDLINE | ID: mdl-27514477

RESUMEN

The cholinergic anti-inflammatory pathway reduces systemic tumor necrosis factor (TNF) via acetylcholine-producing memory T cells in the spleen. These choline acetyltransferase (ChAT)-expressing T cells are also found in the intestine, where their function is unclear. We aimed to characterize these cells in mouse and human intestine and delineate their function. We made use of the ChAT-enhanced green fluorescent protein (eGFP) reporter mice. CD4Cre mice were crossed to ChATfl/fl mice to achieve specific deletion of ChAT in CD4+ T cells. We observed that the majority of ChAT-expressing T cells in the human and mouse intestine have characteristics of Th17 cells and coexpress IL17A, IL22, and RORC The generation of ChAT-expressing T cells was skewed by dendritic cells after activation of their adrenergic receptor ß2 To evaluate ChAT T cell function, we generated CD4-specific ChAT-deficient mice. CD4ChAT-/- mice showed a reduced level of epithelial antimicrobial peptides lysozyme, defensin A, and ang4, which was associated with an enhanced bacterial diversity and richness in the small intestinal lumen in CD4ChAT-/- mice. We conclude that ChAT-expressing T cells in the gut are stimulated by adrenergic receptor activation on dendritic cells. ChAT-expressing T cells may function to mediate the host AMP secretion, microbial growth and expansion.


Asunto(s)
Acetilcolina/metabolismo , Defensinas/metabolismo , Microbioma Gastrointestinal/fisiología , Mucosa Intestinal/metabolismo , Muramidasa/metabolismo , Ribonucleasa Pancreática/metabolismo , Linfocitos T/metabolismo , Animales , Colina O-Acetiltransferasa/genética , Colina O-Acetiltransferasa/metabolismo , Humanos , Ratones , Ratones Noqueados , Ratones Transgénicos
11.
Am J Physiol Gastrointest Liver Physiol ; 310(11): G1138-46, 2016 06 01.
Artículo en Inglés | MEDLINE | ID: mdl-27151945

RESUMEN

In the intestinal mucosa, retinoic acid (RA) is a critical signaling molecule. RA is derived from dietary vitamin A (retinol) through conversion by aldehyde dehydrogenases (aldh). Reduced levels of short-chain fatty acids (SCFAs) are associated with pathological microbial dysbiosis, inflammatory disease, and allergy. We hypothesized that SCFAs contribute to mucosal homeostasis by enhancing RA production in intestinal epithelia. With the use of human and mouse epithelial cell lines and primary enteroids, we studied the effect of SCFAs on the production of RA. Functional RA conversion was analyzed by Adlefluor activity assays. Butyrate (0-20 mM), in contrast to other SCFAs, dose dependently induced aldh1a1 or aldh1a3 transcript expression and increased RA conversion in human and mouse epithelial cells. Epithelial cell line data were replicated in intestinal organoids. In these organoids, butyrate (2-5 mM) upregulated aldh1a3 expression (36-fold over control), whereas aldh1a1 was not significantly affected. Butyrate enhanced maturation markers (Mucin-2 and villin) but did not consistently affect stemness markers or other Wnt target genes (lgr5, olfm4, ascl2, cdkn1). In enteroids, the stimulation of RA production by SCFA was mimicked by inhibitors of histone deacetylase 3 (HDAC3) but not by HDAC1/2 inhibitors nor by agonists of butyrate receptors G-protein-coupled receptor (GPR)43 or GPR109A, indicating that butyrate stimulates RA production via HDAC3 inhibition. We conclude that the SCFA butyrate inhibits HDAC3 and thereby supports epithelial RA production.


Asunto(s)
Butiratos/farmacología , Inhibidores de Histona Desacetilasas/farmacología , Histona Desacetilasas/metabolismo , Mucosa Intestinal/efectos de los fármacos , Tretinoina/metabolismo , Aldehído Oxidorreductasas/genética , Aldehído Oxidorreductasas/metabolismo , Animales , Células CACO-2 , Células Cultivadas , Humanos , Mucosa Intestinal/metabolismo , Ratones , Ratones Endogámicos C57BL , Proteínas de Microfilamentos/genética , Proteínas de Microfilamentos/metabolismo , Mucina 2/genética , Mucina 2/metabolismo
12.
Am J Gastroenterol ; 110(9): 1289-97, 2015 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-26303134

RESUMEN

OBJECTIVES: The esophageal mucosal integrity is impaired in patients with eosinophilic esophagitis (EoE). We aimed to evaluate the effect of fluticasone propionate on inflammation and functional and structural markers of esophageal mucosal barrier integrity in adult patients with EoE. METHODS: In this prospective study, we included 15 EoE patients (median age (IQR), 43 (30-45) years). Patients underwent upper endoscopy before and after an 8-week course of swallowed fluticasone propionate 500 µg BID. Several parameters of esophageal mucosal barrier integrity were evaluated: esophageal electrical tissue impedance in vivo during endoscopy, transepithelial electrical resistance (TER) and transepithelial molecule flux in Ussing chambers using esophageal biopsy specimens, and intercellular spaces as a structural marker of permeability using electron microscopy. Esophageal eosinophils and mast cells were counted, and expression of inflammatory cytokines and barrier integrity proteins was investigated using qPCR. Esophageal symptoms and signs were also assessed. RESULTS: Peak eosinophil and mast cell counts decreased significantly after fluticasone propionate treatment. The esophageal mucosal integrity increased substantially during treatment, as shown by increased extracellular impedance and TER (both P<0.01) and decreased transepithelial molecule flux in Ussing chambers (P<0.05). Whereas expression of genes encoding for inflammatory cytokines (IL5, IL13, eotaxin-3, periostin, TSLP) decreased after treatment, expression of genes encoding for barrier integrity proteins (filaggrin and desmoglein-1) increased. CONCLUSIONS: Fluticasone propionate treatment decreases eosinophilic inflammation and improves the esophageal mucosal barrier integrity in adult EoE patients. Improvement of the mucosal barrier integrity correlates with normalization of expression of desmoglein-1 and filaggrin marker genes.


Asunto(s)
Deglución/fisiología , Esofagitis Eosinofílica/patología , Eosinófilos/patología , Fluticasona/administración & dosificación , Mucosa Intestinal/ultraestructura , Recuperación de la Función , Adulto , Antiinflamatorios/administración & dosificación , Biopsia , Citocinas/biosíntesis , Citocinas/genética , Relación Dosis-Respuesta a Droga , Impedancia Eléctrica , Esofagitis Eosinofílica/tratamiento farmacológico , Esofagitis Eosinofílica/fisiopatología , Eosinófilos/efectos de los fármacos , Esofagoscopía , Femenino , Proteínas Filagrina , Estudios de Seguimiento , Regulación de la Expresión Génica , Humanos , Mucosa Intestinal/metabolismo , Mucosa Intestinal/fisiopatología , Recuento de Leucocitos , Masculino , Microscopía Electrónica , Persona de Mediana Edad , Estudios Prospectivos , ARN/genética , Reacción en Cadena en Tiempo Real de la Polimerasa , Resultado del Tratamiento
13.
Am J Physiol Gastrointest Liver Physiol ; 307(3): G323-9, 2014 Aug 01.
Artículo en Inglés | MEDLINE | ID: mdl-24924748

RESUMEN

Increased esophageal sensitivity and impaired mucosal integrity have both been described in patients with gastroesophageal reflux disease, but the relationship between hypersensitivity and mucosal integrity is unclear. The aim of the present study was to investigate acid sensitivity in patients with erosive and nonerosive reflux disease and control subjects to determine the relation with functional esophageal mucosal integrity changes as well as to investigate cellular mechanisms of impaired mucosal integrity in these patients. In this prospective experimental study, 12 patients with nonerosive reflux disease, 12 patients with esophagitis grade A or B, and 11 healthy control subjects underwent an acid perfusion test and upper endoscopy. Mucosal integrity was measured during endoscopy by electrical tissue impedance spectroscopy and biopsy specimens were analyzed in Ussing chambers for transepithelial electrical resistance, transepithelial permeability and gene expression of tight junction proteins and filaggrin. Patients with nonerosive reflux disease and esophagitis were more sensitive to acid perfusion compared with control subjects, having a shorter time to perception of heartburn and higher perceived intensity of heartburn. In reflux patients, enhanced acid sensitivity was associated with impairment of in vivo and vitro esophageal mucosal integrity. Mucosal integrity was significantly impaired in patients with esophagitis, displaying higher transepithelial permeability and lower extracellular impedance. Although no significant differences in the expression of tight junction proteins were found in biopsies among patient groups, mucosal integrity parameters in reflux patients correlated negatively with the expression of filaggrin. In conclusion, sensitivity to acid is enhanced in patients with gastroesophageal reflux disease, irrespective of the presence of erosions, and is associated with impaired esophageal mucosal integrity. Mucosal integrity of the esophagus is associated with the expression of filaggrin.


Asunto(s)
Esofagitis/metabolismo , Esófago/metabolismo , Reflujo Gastroesofágico/metabolismo , Pirosis/metabolismo , Percepción del Dolor , Adulto , Anciano , Biopsia , Estudios de Casos y Controles , Impedancia Eléctrica , Monitorización del pH Esofágico , Esofagitis/genética , Esofagitis/patología , Esofagitis/fisiopatología , Esofagoscopía , Esófago/inervación , Esófago/patología , Femenino , Proteínas Filagrina , Reflujo Gastroesofágico/genética , Reflujo Gastroesofágico/patología , Reflujo Gastroesofágico/fisiopatología , Regulación de la Expresión Génica , Pirosis/genética , Pirosis/patología , Pirosis/fisiopatología , Humanos , Concentración de Iones de Hidrógeno , Proteínas de Filamentos Intermediarios/genética , Proteínas de Filamentos Intermediarios/metabolismo , Masculino , Persona de Mediana Edad , Membrana Mucosa/inervación , Membrana Mucosa/metabolismo , Membrana Mucosa/patología , Dimensión del Dolor , Estudios Prospectivos , Índice de Severidad de la Enfermedad , Análisis Espectral , Uniones Estrechas/metabolismo , Uniones Estrechas/patología , Adulto Joven
14.
Clin Gastroenterol Hepatol ; 12(11): 1815-23.e2, 2014 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-24657840

RESUMEN

BACKGROUND & AIMS: Histologic analysis is used to distinguish patients with proton pump inhibitor-responsive eosinophilia (PPI-REE) from those with eosinophilic esophagitis (EoE). It is not clear whether these entities have different etiologies. Exposure to acid reflux can impair the integrity of the esophageal mucosal. We proposed that patients with EoE and PPI-REE might have reflux-induced esophageal mucosal damage that promotes transepithelial flux of allergens. We therefore assessed the integrity of the esophageal mucosal in these patients at baseline and after PPI. METHODS: We performed a prospective study of 16 patients with suspected EoE and 11 controls. Patients had dysphagia, endoscopic signs of EoE, and esophageal eosinophilia (>15 eosinophils/high-power field [eos/hpf]). All subjects underwent endoscopy at baseline; endoscopy was performed again on patients after 8 weeks of treatment with high-dose esomeprazole. After PPI treatment, patients were diagnosed with EoE (>10 eos/hpf; n = 8) or PPI-REE (≤10 eos/hpf; n = 8). We evaluated the structure (intercellular spaces) and function (electrical tissue impedance, transepithelial electrical resistance, transepithelial molecule flux) of the esophageal mucosal barrier. RESULTS: Compared with controls, electrical tissue impedance and transepithelial electrical resistance were reduced in patients with EoE (P < .001 and P < .001, respectively) and PPI-REE (P = .01 and P = .06, respectively), enabling transepithelial small-molecule flux. PPI therapy partially restored these changes in integrity and inflammation in patients with PPI-REE, but not in those with EoE. CONCLUSIONS: The integrity of the esophageal mucosa is impaired in patients with EoE and PPI-REE, allowing transepithelial transport of small molecules. PPI therapy partially restores mucosal integrity in patients with PPI-REE, but not in those with EoE. Acid reflux might contribute to transepithelial allergen flux in patients with PPI-REE. Trialregister.nl number: NTR3480.


Asunto(s)
Eosinofilia/tratamiento farmacológico , Esofagitis Eosinofílica/tratamiento farmacológico , Esófago/patología , Membrana Mucosa/patología , Membrana Mucosa/fisiopatología , Inhibidores de la Bomba de Protones/uso terapéutico , Adulto , Femenino , Humanos , Masculino , Persona de Mediana Edad , Estudios Prospectivos , Resultado del Tratamiento
15.
Eur J Gastroenterol Hepatol ; 24(8): 965-70, 2012 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-22664939

RESUMEN

BACKGROUND AND AIM: The balance between microbes and host defence mechanisms at the mucosal frontier plays an important, yet unclarified role in the pathogenesis of inflammatory bowel disease (IBD). The importance of microorganisms in IBD is supported by the association of IBD with mutations in pattern recognition receptors (PRRs) such as NOD2 and TLR4. We aimed to examine whether polymorphisms in another type of PRRs, the so-called C-type lectin receptors (CLRs), are associated with IBD. Growing insights into the pathogenetic role of NOD2 mutations in Crohn's disease (CD) and the fact that the majority of CLR-encoding genes are located in IBD susceptibility loci provide strong arguments for further exploration of the role of CLRs in IBD. METHODS: In this study, we selected four single nucleotide polymorphisms (SNPs) in different CLRs to determine whether there could be a role for these CLRs in IBD. Functional SNPs in the genes coding for the candidate CLRs DC-SIGN, LLT1, DCIR and MGL were examined. Genotyping of all SNPs was performed at the Academic Medical Center. In this study, around 1572 samples were included from a maximum of 621 CD patients, 457 ulcerative colitis (UC) patients and 586 healthy controls (HCs). RESULTS AND CONCLUSION: No association was found between our IBD cohort and the candidate SNPs for DC-SIGN (CD/HC: P=0.25 and UC/HC: P=0.36), DCIR (CD/HC: P=0.22 and UC/HC: P=0.41) and MGL (CD/HC: P=0.37 and UC/HC: P=0.25). However, one polymorphism in LLT1 was found to be associated with our CD population (P<0.034). Our UC cohort was not associated with the variation in LLT1 (P=0.33). LLT1 is a ligand for the recently discovered CD161. CD161 is a new surface marker for human interleukin (IL)-17-producing Th17 cells. The Th17 phenotype has been linked to CD by the fact that IL-22, IL-17 and IL-23 receptor levels are increased in CD. The signal transduction pathways involving LLT1 and CD161 are not completely clarified and are currently under investigation in our laboratory.


Asunto(s)
Colitis Ulcerosa/genética , Enfermedad de Crohn/genética , Predisposición Genética a la Enfermedad/genética , Lectinas Tipo C/genética , Polimorfismo de Nucleótido Simple/genética , Adulto , Moléculas de Adhesión Celular/genética , Estudios de Cohortes , Femenino , Genotipo , Humanos , Enfermedades Inflamatorias del Intestino/genética , Masculino , Glicoproteínas de Membrana/genética , Persona de Mediana Edad , Receptores de Superficie Celular/genética , Receptores Inmunológicos/genética
16.
Neuroimmunomodulation ; 19(4): 255-65, 2012.
Artículo en Inglés | MEDLINE | ID: mdl-22441542

RESUMEN

OBJECTIVE: Smoking is generally accepted as a factor that affects the disease course in inflammatory bowel disease patients. Whether these effects can be contributed to the immunomodulatory effects of nicotine via nicotinic acetylcholine receptor (nAChR) activation is unclear. As previous data suggest that the α7 nicotinic acetylcholine receptor (CHRNA7) and its duplicated variant CHRFAM7A may specifically participate in the inflammatory response of monocytes, we evaluated whether repeated nicotine exposure or smoking affects monocyte CHRNA7 and CHRFAM7A expression and cholinergic immunomodulation. METHODS: The human monocyte cell line THP-I was incubated with nicotine for different time points before endotoxin exposure. In a pilot volunteer study using smoking (n = 4) and nonsmoking (n = 7) individuals, vagal output was stimulated by olive oil administration after which monocytes were analyzed for nicotinic receptor expression. Serum tumor necrosis factor (TNF) levels were determined using ELISA and expression levels of the nAChR subunits CHRNA7, CHRNB2 or CHRFAM7A were analyzed using QPCR. RESULTS: Repeated nicotine exposure upregulated CHRNA7 expression on THP-I monocytes and led to an enhanced potential of α7 nAChR agonist GSK1345038A to reduce TNF levels. Furthermore, CHRNA7 was only detectable in isolated blood monocytes of smokers. On the other hand, the expression of CHRFAM7A and CHRNB2 was not affected by nicotine exposure. Lipopolysaccharides-induced TNF secretion was inhibited by nicotinic receptor activation in THP-I monocytes, but this response was not consistently seen in blood monocytes from smoking individuals. CONCLUSIONS: We conclude that CHRNA7 expression on blood monocytes is upregulated in smoking individuals, which may contribute to cholinergic immunomodulation.


Asunto(s)
Inmunomodulación/efectos de los fármacos , Monocitos/efectos de los fármacos , Receptores Nicotínicos/metabolismo , Fumar/inmunología , Factor de Necrosis Tumoral alfa/efectos de los fármacos , Adulto , Línea Celular , Femenino , Estimulantes Ganglionares/inmunología , Estimulantes Ganglionares/farmacología , Humanos , Masculino , Monocitos/inmunología , Nicotina/inmunología , Nicotina/farmacología , Receptores Nicotínicos/efectos de los fármacos , Receptores Nicotínicos/inmunología , Factor de Necrosis Tumoral alfa/inmunología , Regulación hacia Arriba , Receptor Nicotínico de Acetilcolina alfa 7
17.
Gastroenterology ; 137(3): 1029-39, 1039.e1-4, 2009 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-19427310

RESUMEN

BACKGROUND & AIMS: The vagus nerve negatively regulates macrophage cytokine production via the release of acetylcholine (ACh) and activation of nicotinic acetylcholine receptors (nAChR). In various models of intestinal inflammation, vagus nerve efferent stimulation ameliorates disease. Given the actively constrained cytokine responses of intestinal macrophages, we explored the effect of nAChR activation on endocytosis and phagocytosis by macrophages residing in the peritoneal and mucosal compartment. METHODS: The phagocytic uptake by intestinal and peritoneal macrophages was measured by fluorescence-activated cell sorter analysis, and the nAChR involved was determined by pharmacologic blockade, short hairpin RNA-assisted gene knockdown, and the use of specific nAChR knockout mice. The effect of electrical vagus nerve stimulation on epithelial translocation and macrophage uptake of luminal particles was studied in mice. RESULTS: In isolated intestinal and peritoneal macrophages, nAChR activation enhanced endocytosis and phagocytosis. This effect was mediated via stimulated recruitment of GTPase Dynamin-2 to the forming phagocytic cup. These effects involve nAChR alpha4/beta2, rather than nAChR alpha7. Despite enhanced bacterial uptake, acetylcholine reduced NF-kappaB activation and pro-inflammatory cytokine production, while stimulating anti-inflammatory interleukin-10 production. Vagus nerve stimulation in mice altered mucosal immune responses by augmenting epithelial transport and uptake of luminal bacteria by lamina propria macrophages. CONCLUSIONS: ACh enhances phagocytic potential while inhibiting immune reactivity via nAChR alpha4/beta2 in mouse macrophages. Hence, vagus nerve efferent activity may stimulate surveillance in the intestinal mucosa and peritoneal compartment.


Asunto(s)
Mucosa Intestinal/citología , Macrófagos/fisiología , Fagocitosis , Receptores Nicotínicos/metabolismo , Nervio Vago/fisiología , Acetilcolina/farmacología , Animales , Células Cultivadas , Agonistas Colinérgicos/farmacología , Dinamina II/metabolismo , Endocitosis/fisiología , Técnicas In Vitro , Macrófagos/inmunología , Macrófagos Peritoneales/fisiología , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Ratones Noqueados , Nicotina/farmacología
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