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1.
Cell ; 158(2): 300-313, 2014 Jul 17.
Artículo en Inglés | MEDLINE | ID: mdl-25036630

RESUMEN

Intestinal peristalsis is a dynamic physiologic process influenced by dietary and microbial changes. It is tightly regulated by complex cellular interactions; however, our understanding of these controls is incomplete. A distinct population of macrophages is distributed in the intestinal muscularis externa. We demonstrate that, in the steady state, muscularis macrophages regulate peristaltic activity of the colon. They change the pattern of smooth muscle contractions by secreting bone morphogenetic protein 2 (BMP2), which activates BMP receptor (BMPR) expressed by enteric neurons. Enteric neurons, in turn, secrete colony stimulatory factor 1 (CSF1), a growth factor required for macrophage development. Finally, stimuli from microbial commensals regulate BMP2 expression by macrophages and CSF1 expression by enteric neurons. Our findings identify a plastic, microbiota-driven crosstalk between muscularis macrophages and enteric neurons that controls gastrointestinal motility. PAPERFLICK:


Asunto(s)
Motilidad Gastrointestinal , Tracto Gastrointestinal/citología , Tracto Gastrointestinal/microbiología , Macrófagos/metabolismo , Animales , Proteína Morfogenética Ósea 2/metabolismo , Receptores de Proteínas Morfogenéticas Óseas de Tipo II/metabolismo , Tracto Gastrointestinal/inervación , Tracto Gastrointestinal/fisiología , Técnicas In Vitro , Factor Estimulante de Colonias de Macrófagos , Ratones , Neuronas/metabolismo , Peristaltismo , Receptor de Factor Estimulante de Colonias de Macrófagos/metabolismo , Transducción de Señal
3.
Gastroenterology ; 144(3): 601-612.e1, 2013 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-23232296

RESUMEN

BACKGROUND & AIMS: In healthy individuals, interactions between intestinal epithelial cells and lamina propria lymphocytes give rise to a population of CD8(+) T cells with suppressor functions (Ts cells). Disruption of Ts cell activities can lead to mucosal inflammation. We investigated what factors were required for expansion of the Ts cell population or loss of their activity in patients with Crohn's disease (CD). METHODS: We developed a method to generate Ts cell lines from freshly isolated lamina propria lymphocytes from patients with ulcerative colitis (UC), patients with CD, or healthy individuals (controls). Cells were stimulated with a monoclonal antibody against CD3, interleukin (IL)-7, and IL-15. After 14 days in culture, CD8(+)T cells were purified and cultured with IL-7 and IL-15. The resulting Ts cells were analyzed for suppressor activity, expression of surface markers, and cytokine secretion profiles. RNA was isolated from the 3 groups of Ts cells and used in microarray analyses. RESULTS: Ts cells from patients with UC and controls suppressed proliferation of CD4(+) T cells; the suppression required cell contact. In contrast, Ts cells from patients with CD had a reduced capacity to suppress CD4(+) T-cell proliferation. The difference in suppressive ability was not associated with surface or intracytoplasmic markers or secretion of cytokines. Microarray analysis identified changes in expression of genes regulated by transforming growth factor (TGF)-ß that were associated with the suppressive abilities of Ts cells. We found that TGF-ß or supernatants from Ts cells of patients with CD reduced the suppressor activity of control Ts cells. CONCLUSIONS: Ts cells isolated from patients with CD have a reduced ability to suppress proliferation of CD4(+)T cells compared with control Ts cells. TGF-ß signaling reduces the suppressor activity of Ts cells.


Asunto(s)
Linfocitos T CD8-positivos/inmunología , Tolerancia Inmunológica/inmunología , Mucosa Intestinal/inmunología , Factor de Crecimiento Transformador beta/inmunología , Antígenos CD28/inmunología , Complejo CD3/inmunología , Línea Celular , Separación Celular , Perfilación de la Expresión Génica , Humanos , Tolerancia Inmunológica/genética , Interferón gamma/inmunología , Intestinos/inmunología , Transducción de Señal , Factor de Necrosis Tumoral alfa/inmunología
4.
Gastroenterology ; 140(2): 550-9, 2011 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-21056041

RESUMEN

BACKGROUND & AIMS: Interactions between lymphocytes and intestinal epithelial cells occur in the subepithelial space of the gastrointestinal tract. Normal human lamina propria lymphocytes (LPLs) induce differentiation of intestinal epithelial cells. The absence of LPLs in mice, such as in RAG1(-/-) mice, results in defects in epithelial cell differentiation. We investigated the role of lymphoepithelial interactions in epithelial differentiation and barrier function. METHODS: We used adoptive transfer to determine if CD4(+) T cells (CD4(+)CD62L(+)CD45Rb(Hi) and/or CD4(+)CD62L(+)CD45Rb(Lo)) could overcome permeability defect (quantified in Ussing chambers). Immunofluorescence staining was performed to determine expression of cleaved Notch-1, villin, and claudin 5 in colon samples from mice and humans. Caco-2 cells were infected with a lentivirus containing a specific Notch-1 or scrambled short hairpin RNA sequence. Tight junction assembly was analyzed by immunoblot and immunofluorescence analyses, and transepithelial resistance was monitored. RESULTS: Expression of cleaved Notch-1, villin, or claudin 5 was not detected in RAG1(-/-) colonocytes; their loss correlated with increased intestinal permeability. Transfer of CD45Rb(Hi) and/or CD45Rb(Lo) cells into RAG1(-/-) mice induced expression of cleaved Notch, villin, and claudin 5 in colonocytes and significantly reduced the permeability of the distal colon. Loss of Notch-1 expression in Caco-2 cells correlated with decreased transepithelial resistance and dysregulated expression and localization of tight junction proteins. Levels of cleaved Notch-1 were increased in colonic epithelium of patients with Crohn's disease. CONCLUSIONS: LPLs promote mucosal barrier function, which is associated with activation of the Notch-1 signaling pathway. LPLs maintain intestinal homeostasis by inducing intestinal epithelial cell differentiation, polarization, and barrier function.


Asunto(s)
Linfocitos T CD4-Positivos/metabolismo , Mucosa Intestinal/fisiología , Receptor Notch1/fisiología , Traslado Adoptivo , Animales , Linfocitos T CD4-Positivos/inmunología , Células CACO-2 , Claudina-5 , Enfermedad de Crohn/patología , Enfermedad de Crohn/fisiopatología , Humanos , Mucosa Intestinal/citología , Mucosa Intestinal/inmunología , Mucosa Intestinal/patología , Selectina L/análisis , Selectina L/inmunología , Antígenos Comunes de Leucocito/análisis , Antígenos Comunes de Leucocito/inmunología , Proteínas de la Membrana/análisis , Ratones , Ratones Endogámicos C57BL , Proteínas de Microfilamentos/metabolismo , Receptor Notch1/análisis , Transducción de Señal/fisiología , Uniones Estrechas/inmunología , Uniones Estrechas/fisiología
5.
J Biol Chem ; 285(2): 1479-89, 2010 Jan 08.
Artículo en Inglés | MEDLINE | ID: mdl-19892711

RESUMEN

The transmembrane glycoprotein CD98 regulates multiple cellular functions, including extracellular signaling, epithelial cell adhesion/polarity, amino acid transport, and cell-cell interactions. MicroRNAs post-transcriptionally regulate gene expression, thereby functioning as modulators of numerous cellular processes, such as cell differentiation, proliferation, and apoptosis. Here, we investigated if microRNAs regulate CD98 expression during intestinal epithelial cell differentiation and inflammation. We found that microRNA-7 repressed CD98 expression in Caco2-BBE cells by directly targeting the 3'-untranslated region of human CD98 mRNA. Expression of CD98 was decreased, whereas that of microRNA-7 was increased in well-differentiated Caco2-BBE cells compared with undifferentiated cells. Undifferentiated crypt cells isolated from mouse jejunum showed higher CD98 levels and lower levels of mmu-microRNA-706, a murine original microRNA candidate for CD98, than well-differentiated villus cells. Importantly, microRNA-7 decreased Caco2-BBE cell attachment on laminin-1, and CD98 overexpression recovered this inhibition, suggesting that microRNA-7 modulates epithelial cell adhesion to extracellular matrix, which in turn could affect proliferation and differentiation during the migration of enterocytes across the crypt-villus axis, by regulating CD98 expression. In a pathological context, the pro-inflammatory cytokine interleukin 1-beta increased CD98 expression in Caco2-BBE cells by decreasing microRNA-7 levels. Consistent with the in vitro findings, microRNA-7 levels were decreased in actively inflamed Crohn disease colonic tissues, where CD98 expression was up-regulated, compared with normal tissues. Together, these results reveal a novel mechanism underlying regulation of CD98 expression during patho-physiological states. This study raises microRNAs as a promising target for therapeutic modulations of CD98 expression in intestinal inflammatory disorders.


Asunto(s)
Diferenciación Celular , Proteína-1 Reguladora de Fusión/biosíntesis , Regulación de la Expresión Génica , Mucosa Intestinal/metabolismo , MicroARNs/metabolismo , Regiones no Traducidas 3'/genética , Animales , Células CACO-2 , Adhesión Celular/genética , Comunicación Celular/genética , Polaridad Celular/genética , Enfermedad de Crohn/genética , Enfermedad de Crohn/metabolismo , Matriz Extracelular/genética , Matriz Extracelular/metabolismo , Proteína-1 Reguladora de Fusión/genética , Humanos , Inflamación/genética , Inflamación/metabolismo , Interleucina-1beta/genética , Interleucina-1beta/metabolismo , Yeyuno/metabolismo , Laminina/genética , Laminina/metabolismo , Masculino , Ratones , MicroARNs/genética
6.
Gastroenterology ; 134(1): 192-203, 2008 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-18045591

RESUMEN

BACKGROUND & AIMS: Intestinal lymphoepithelial interactions occur in the epithelium and the subepithelial space. We asked whether normal, Crohn's disease (CD), or ulcerative colitis (UC) lamina propria lymphocytes (LPL) could promote intestinal epithelial cell (IEC) growth and differentiation. METHODS: T84 cells were cocultured with isolated LPL. IECs were then lysed and subjected to measurement of intestinal alkaline phosphatase (IAP) activity; Western blot analysis for MAPK and Akt activation; and real-time polymerase chain reaction to assess caudal-related homeoprotein 2 (CDX2) messenger RNA (mRNA) levels. Tissue sections were immunostained for evidence of mitogen-activated protein kinase (MAPK) and phosphatidylinositol 3-kinase (PI3K) activation, CDX2, and IAP; and CDX2 mRNA expression was assessed in human colonic biopsy specimens. RESULTS: IAP activity was increased in T84 cells cocultured for 8 days with normal LPL (P < .05) and even greater with CD LPL (P < .001). Crypt IECs in active CD mucosa expressed IAP ex vivo. Phospho-MAPK (extracellular signal-regulated kinase 1/2, p38, and c-Jun-N-terminal kinase) and phospho-Akt were seen as early as 30 minutes after coculture. MAPK activation was greatest in T84 cells cocultured with CD LPL. There was a specific increase in Phospho-p38 MAPK and Phospho-Akt staining in the nuclei of crypt IECs in active vs inactive CD, normal mucosa, and UC mucosa. CDX2 mRNA expression was increased in CD LPL cocultured T84 cells, which did not correlate with CDX2 protein localization ex vivo. CONCLUSIONS: There is cross talk between LPL and IECs, which leads to IEC differentiation. The differentiation is accelerated in CD mucosa.


Asunto(s)
Comunicación Celular/fisiología , Diferenciación Celular/fisiología , Células Epiteliales/citología , Enfermedades Inflamatorias del Intestino/patología , Mucosa Intestinal/patología , Linfocitos/fisiología , Fosfatasa Alcalina , Antígenos de Neoplasias/metabolismo , Factor de Transcripción CDX2 , Técnicas de Cultivo de Célula , Línea Celular Tumoral , Técnicas de Cocultivo , Proteínas Ligadas a GPI , Proteínas de Homeodominio/genética , Proteínas de Homeodominio/metabolismo , Humanos , Enfermedades Inflamatorias del Intestino/metabolismo , Mucosa Intestinal/metabolismo , ARN Mensajero/metabolismo
7.
Oncotarget ; 9(71): 33536-33548, 2018 Sep 11.
Artículo en Inglés | MEDLINE | ID: mdl-30323897

RESUMEN

Increasing evidence links Notch-1 signaling with the maintenance of intestinal architecture and homeostasis. Dysfunction in the common Notch-1 pathway transcription factor recombinant binding protein suppressor of hairless (RBP-J) is associated with loss of epithelial barrier integrity and aberrant conversion of proliferative crypt cells into goblet cells. Furthermore, we have recently discovered that epithelial Notch-1 is indispensable in bridging innate and adaptive immunity in the gut and is required for supporting protective epithelial pro-inflammatory responses. Yet, the epithelial specific function of Notch-1 in intestinal tumorigenesis remains unknown. We generated Villin-Cre/Notch-1 fl/fl (VN -/- ) mice that are selectively deficient in Notch-1 in intestinal epithelial cells. Intestinal epithelial Notch-1 preserved barrier function and integrity, whereas lack of epithelial Notch-1 induced goblet cell hyperplasia, spontaneous serrated lesions, multifocal low- and high-grade dysplasia and colonic mucinous neoplasms in mice. Over time, VN -/- mice displayed high occurrence of colorectal mucinous adenocarcinomas, which correlated with increased levels of mitogenic, angiogenic and pro-tumorigenic gene expression. Finally, we found that the expression of Notch-1 is significantly reduced in human colorectal mucinous adenocarcinoma when compared to colorectal adenocarcinoma. Taken together, our findings reveal a novel and critical protective role for Notch-1 in controlling intestinal tumorigenesis.

8.
Nat Commun ; 8(1): 932, 2017 10 13.
Artículo en Inglés | MEDLINE | ID: mdl-29030607

RESUMEN

Inflammatory bowel disease (IBD) involves interaction between host genetic factors and environmental triggers. CCDC88B maps within one IBD risk locus on human chromosome 11q13. Here we show that CCDC88B protein increases in the colon during intestinal injury, concomitant with an influx of CCDC88B+lymphoid and myeloid cells. Loss of Ccdc88b protects against DSS-induced colitis, with fewer pathological lesions and reduced intestinal inflammation in Ccdc88b-deficient mice. In a T cell transfer model of colitis, Ccdc88b mutant CD4+ T cells do not induce colitis in immunocompromised hosts. Expression of human CCDC88B RNA and protein is higher in IBD patient colons than in control colon tissue. In human CD14+ myeloid cells, CCDC88B is regulated by cis-acting variants. In a cohort of patients with Crohn's disease, CCDC88B expression correlates positively with disease risk. These findings suggest that CCDC88B has a critical function in colon inflammation and the pathogenesis of IBD.Hook-related protein family member CCDC88b is encoded by a locus that has been associated with inflammatory bowel disease. Here the authors show that Ccdc88b inactivation in T cells prevents colitis in a transfer model, and detect high colonic levels of CCDC88b in patients with Crohn disease or ulcerative colitis, identifying that expression correlates with disease risk.


Asunto(s)
Proteínas Portadoras/genética , Proteínas Portadoras/metabolismo , Colitis/patología , Enfermedades Inflamatorias del Intestino/patología , Animales , Colitis/inducido químicamente , Colitis/metabolismo , Colon/metabolismo , Colon/patología , Enfermedad de Crohn/metabolismo , Enfermedad de Crohn/patología , Sulfato de Dextran/toxicidad , Regulación de la Expresión Génica , Proteínas de Homeodominio/genética , Humanos , Enfermedades Inflamatorias del Intestino/metabolismo , Receptores de Lipopolisacáridos/metabolismo , Masculino , Ratones Endogámicos C57BL , Ratones Mutantes , Células Mieloides/metabolismo , Células Mieloides/patología , Polimorfismo de Nucleótido Simple , Linfocitos T/metabolismo , Linfocitos T/patología
9.
Res Microbiol ; 157(5): 456-65, 2006 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-16487684

RESUMEN

Induction of apoptosis and necrosis by enterohemorrhagic Escherichia coli (EHEC) has been reported in vivo and in vitro, but features of cell death were not noted in those reports. Since tumor necrosis factor-alpha (TNF-alpha) has been implicated in the apoptosis of invasive bacteria, we investigated the role of this cytokine in EHEC-induced apoptosis. We hypothesize that the probiotic yeast strain Saccharomyces boulardii that interferes with EHEC-induced pro-inflammatory pathways delays EHEC-induced apoptosis. By 6 h of infection, flow cytometry analysis of T84 cells demonstrated that 40% of cells were FITC-annexin-V-positive and 40% of cells incorporated both annexin and propidium iodide (PI). Simultaneously, western blot analysis demonstrated that procaspases-8 and -3 were cleaved. Fragmentation of internucleosomal DNA revealed evidence of apoptotic leader formation after 8 and 9 h of infection. Procaspase-9 activation and 3',3-dihexyloxacarbocyanine iodide (DiOC(6)) incorporation were observed at 3 h of infection. In cells preincubated with S. boulardii and infected with EHEC in the presence of yeast, the quantities of procaspases-8, -9 and -3 did not vary, and no DNA fragmentation was observed. The TNF-alpha transcript level and the level of secreted TNF-alpha increased considerably (P<0.001vs control cells) at 6 h of infection in EHEC-alone-infected cells, but were significantly reduced in cells infected in the presence of S. boulardii (P<0.001vs EHEC-alone-infected cells). The presence of anti-TNF-alpha antibody during infection reduced by 30% the level of FITC-annexin V-positive cells. Altogether, these findings demonstrated that: (i) EHEC infection stimulated TNF-alpha synthesis that is implicated in apoptosis of T84 cells; and (ii) S. boulardii induced a decrease in TNF-alpha and related apoptosis in EHEC-infected T84 cells.


Asunto(s)
Apoptosis , Infecciones por Escherichia coli/microbiología , Escherichia coli/crecimiento & desarrollo , Saccharomyces/fisiología , Factor de Necrosis Tumoral alfa/metabolismo , Apoptosis/efectos de los fármacos , Línea Celular Tumoral , Regulación hacia Abajo , Infecciones por Escherichia coli/patología , Humanos , Probióticos , Factor de Necrosis Tumoral alfa/farmacología
10.
Oncotarget ; 7(29): 46384-46400, 2016 Jul 19.
Artículo en Inglés | MEDLINE | ID: mdl-27344176

RESUMEN

Necroptosis is a programmed form of non-apoptotic cell death that requires the kinase activity of the receptor interacting protein kinase 3 (RIPK3). Although in vitro data suggests that cancer cells lacking expression of RIPK3 are invasive, the physiological role of RIPK3 in a disease-relevant setting remains unknown. Here we provide evidence that RIPK3 has a critical role in suppressing colorectal cancer (CRC). RIPK3-deficient mice were highly susceptible to colitis-associated CRC and exhibited greater production of pro-inflammatory mediators and tumor promoting factors. Tumorigenesis in RIPK3-deficiency resulted from uncontrolled activation of NF-κB, STAT3, AKT and Wnt-ß-catenin signaling pathways that enhanced the ability of intestinal epithelial cells (IECs) to aberrantly proliferate in the face of the sustained inflammatory microenvironment and promote CRC. We found that RIPK3 expression is reduced in tumors from patients with inflammatory bowel diseases, and further confirmed that expression of RIPK3 is downregulated in human CRC and correlated with cancer progression. Thus, our results reveal that the necroptosis adaptor RIPK3 has key anti-inflammatory and anti-tumoral functions in the intestine, and define RIPK3 as a novel colon tumor suppressor.


Asunto(s)
Transformación Celular Neoplásica/metabolismo , Neoplasias del Colon/patología , Proteína Serina-Treonina Quinasas de Interacción con Receptores/metabolismo , Animales , Muerte Celular , Colitis/complicaciones , Humanos , Ratones , Ratones Noqueados
11.
Inflamm Bowel Dis ; 20(1): 144-53, 2014 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-24252977

RESUMEN

BACKGROUND: Crohn's disease (CD) is a chronic inflammatory disease with increasing incidence in children. Current medications have potentially serious side effects, hence increasing interest in alternative therapies. We previously developed an herbal formula, FAHF-2, based on a classical traditional Chinese herbal formula Wu Mei Wan that has long been used in China to treat colitis. We investigated FAHF-2's potential anti-inflammatory effects. METHODS: FAHF-2 efficacy was tested in vivo in the CD45RbRAG1 transfer colitis model. Weight loss, colonic histology, and cytokine production from mesenteric lymph nodes were assessed. Human peripheral blood mononuclear cells (PBMCs) and colonic biopsies were obtained from children newly diagnosed with CD and controls and cultured with or without FAHF-2. Cytokine levels were measured by multiplex immunoassay. The effect of FAHF-2 on TNF-α-producing cells was determined by flow cytometry. NF-κB signaling was investigated in human lamina propria mononuclear cells upon FAHF-2 treatment by In-Cell Western. RESULTS: FAHF-2-treated mice had decreased weight loss, improved histology, and reduced TNF-α, IL-17, IL-6, and IFN-γ production. In vitro treated PBMCs produced less TNF-α, IFN-γ, and IL-12. FAHF-2 reduced the TNF-α-producing monocytes and T cells. Inflamed CD biopsies produced less TNF-α, IL-17, IL-6, and IL-1ß. These effects are because of decreased NF-κB activation. CONCLUSIONS: FAHF-2 inhibited both adaptive and innate immune proinflammatory cytokine responses in PBMCs and inflamed CD mucosa due in part to blockage of NF-κB activation. FAHF-2 was effective in halting progression of colitis in a murine model. This study shows that FAHF-2 has potential as a novel treatment of CD.


Asunto(s)
Antiinflamatorios/uso terapéutico , Colitis/prevención & control , Colon/efectos de los fármacos , Enfermedades Inflamatorias del Intestino/prevención & control , Leucocitos Mononucleares/efectos de los fármacos , Extractos Vegetales/uso terapéutico , Adolescente , Adulto , Animales , Western Blotting , Estudios de Casos y Controles , Células Cultivadas , Niño , Preescolar , China , Colitis/etiología , Citocinas/metabolismo , Citometría de Flujo , Estudios de Seguimiento , Humanos , Enfermedades Inflamatorias del Intestino/inmunología , Enfermedades Inflamatorias del Intestino/metabolismo , Macrófagos/efectos de los fármacos , Macrófagos/inmunología , Macrófagos/metabolismo , Ratones Endogámicos C57BL , Pronóstico , Adulto Joven
12.
Immunol Res ; 54(1-3): 69-74, 2012 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-22427015

RESUMEN

The epithelium of the gastrointestinal tract, which represents the largest surface area of the body, is constantly exposed to the contents of its surrounding environment. The intestinal epithelium forms barriers that are essential in maintaining equilibrium within the human body. This barrier supports nutrient and water transport while preventing microbial invasion. Intestinal epithelial cells (IECs) sit at the interface between an antigen-rich lumen and a lymphocyte-rich lamina propria (LP). IECs have the capability to discriminate between "peaceful" and "harmful" antigens. The epithelium is constantly sampling luminal contents and making molecular adjustments accordingly. These molecular changes influence the actions of innate and adaptive immune cells. The crosstalk that occurs between the epithelium and the immune compartments serves to maintain intestinal homeostasis. A better understanding of the nature of the interactions between normal LP lymphocytes (LPLs) and IECs will ultimately provide insights into the defects occurring in inflammatory bowel disease patients.


Asunto(s)
Enfermedades Inflamatorias del Intestino/inmunología , Mucosa Intestinal/inmunología , Receptores Notch/inmunología , Animales , Humanos , Mucosa Intestinal/citología , Transducción de Señal
13.
Ann N Y Acad Sci ; 1165: 323-6, 2009 May.
Artículo en Inglés | MEDLINE | ID: mdl-19538323

RESUMEN

Intestinal lymphoepithelial interactions occur in the epithelium and subepithelial space. We asked whether or not lamina propria lymphocytes (LPL) could promote intestinal epithelial cell (IEC) differentiation. In contrast to epithelial cells in UC mucosa, which do not differentiate because of rapid turnover, differentiation of epithelial cells in CD mucosa occurs in the crypts. This dysregulation is driven by alterations in the crosstalk between CD LPL and CD IECs, leading to an acceleration of their differentiation. This alteration seems to involve the transcription factor CDX2 via the activation of the PI3K and MAPK pathways and provides new insights into the dysfunction of the epithelial barrier in CD versus UC. The absence of lymphocytes in Rag1-deficient mice was associated with a defect in colonic IEC differentiation, restored by co-transfer of naïve and regulatory T cells. Interestingly, the transfer of naïve T cells alone induced an acceleration of IEC differentiation similar to what was seen in the colonic mucosa of CD patients. Thus, there is a crosstalk between LPL and IECs that is altered in CD, which leads to an absorptive phenotype of IEC differentiation.


Asunto(s)
Células Epiteliales/metabolismo , Linfocitos/metabolismo , Animales , Diferenciación Celular , Células Cultivadas , Células Epiteliales/citología , Humanos , Mucosa Intestinal/citología , Mucosa Intestinal/metabolismo , Linfocitos/citología , Ratones , Ratones Endogámicos , Transducción de Señal
14.
Inflamm Bowel Dis ; 15(12): 1775-83, 2009 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-19637360

RESUMEN

BACKGROUND: CEACAM1, CEACAM5, and CEACAM6 represent 3 of the CEACAM (carcinoembryonic antigen-related cell adhesion molecule) subfamily members expressed on intestinal epithelial cells (IECs). Deficiency in their expression, as seen in inflammatory bowel disease (IBD), results in the lack of activation of CD8+ regulatory T cells in the mucosa. Since CEACAM expression was shown to be regulated by the transcription factor SOX9, we sought to determine whether the defect in CEACAM expression in IBD was related to aberrant SOX9 expression. METHODS: IECs and lamina propria lymphocytes (LPLs) were freshly isolated from colonic tissues. T84 and HT29 16E cells were cocultured with LPLs. SOX9 and CEACAM subfamily member expression was assessed by real-time polymerase chain reaction (PCR), Western blot, immunohistochemistry, and immunofluorescence. RESULTS: In Crohn's disease (CD) but not in ulcerative colitis (UC), a significant reduction in mRNA and protein expression for CEACAM1 and 5 was noted; in contrast, no difference in SOX9 mRNA expression was seen. However, nuclear SOX9 immunostaining was increased in CD IECs. Furthermore, SOX9 protein was reduced in the cytoplasm of LPL-stimulated T84 and HT29 16E cells, while CEACAM5 expression was increased. CONCLUSIONS: The defect in CEACAM family members in CD IECs appears to be related to the aberrant nuclear localization of SOX9. Changes in SOX9 expression in the CD mucosa relate to the local microenvironment and altered IEC:LPL crosstalk.


Asunto(s)
Antígenos CD/biosíntesis , Antígeno Carcinoembrionario/biosíntesis , Moléculas de Adhesión Celular/biosíntesis , Enfermedad de Crohn/metabolismo , Mucosa Intestinal/metabolismo , Factor de Transcripción SOX9/metabolismo , Línea Celular Tumoral , Células Cultivadas , Técnicas de Cocultivo , Colon/metabolismo , Enfermedad de Crohn/patología , Proteínas Ligadas a GPI/biosíntesis , Humanos , Mucosa Intestinal/patología , Linfocitos/metabolismo , Membrana Mucosa/metabolismo , Membrana Mucosa/patología
15.
Immunol Rev ; 215: 243-53, 2007 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-17291293

RESUMEN

The mucosal immune system is governed by a unique set of rules and regulations. The local microenvironment dictates the necessity for these differences. The intestinal epithelial cell (IEC) sits at the interface between an antigen-rich lumen and a lymphocyte-rich lamina propria (LP). The cross talk that occurs between these compartments serves to maintain intestinal homeostasis. IECs have the capacity to talk to LP lymphocytes, activating populations of unique regulatory T cells. These cells have the capacity to talk back to the epithelium, influencing epithelial cell growth and differentiation. This review looks at this cross talk and places it in the context of mucosal immunoregulation.


Asunto(s)
Comunicación Celular/inmunología , Células Epiteliales/inmunología , Inmunidad Mucosa , Mucosa Intestinal/inmunología , Receptor Cross-Talk/inmunología , Linfocitos T/inmunología , Animales , Células Presentadoras de Antígenos/inmunología , Células Presentadoras de Antígenos/metabolismo , Células Epiteliales/metabolismo , Humanos , Mucosa Intestinal/citología , Activación de Linfocitos/inmunología , Linfocitos T/metabolismo
16.
Gastroenterology ; 131(1): 47-58, 2006 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-16831589

RESUMEN

BACKGROUND & AIMS: Secreted immunoglobulins play an integral role in host defense at mucosal surfaces, and recent evidence shows that IgG can participate in antigen sampling from the intestinal lumen. We examined whether IgE also could facilitate transepithelial antigen sampling. METHODS: Stool samples from food-allergic patients undergoing oral food challenge were analyzed for CD23 and food-specific IgE. CD23 isoform expression on primary human intestinal epithelial cells (IEC) was analyzed by polymerase chain reaction. The role of CD23 isoforms in transcytosis of antigen and IgE-antigen complexes was assessed using polarized human T84 cells retrovirally transfected with CD23a or CD23b. RESULTS: CD23 was expressed constitutively on IECs, and food-allergic patients had increased levels of soluble CD23 and food-specific IgE in the stool after challenge. CD23a, but not CD23b, was expressed by primary human IECs. We show in transcytosis assays that CD23a, but not CD23b, acts as a bidirectional transporter of IgE. In addition, specific IgE facilitated the uptake of antigen from the apical surface of an epithelial monolayer by diverting antigen from delivery to lysosomes. Finally, delivery of antigen-IgE complexes across the epithelial barrier could induce the degranulation of rat basophil leukemia cells transfected with the human high-affinity IgE receptor. CONCLUSIONS: These studies show that CD23a is expressed normally on human IECs, and in the presence of IgE can function as an antigen-sampling mechanism capable of activating subepithelial mast cells. IgE may serve as a secretory immunoglobulin that in concert with CD23 participates in food-induced pathophysiology of the gastrointestinal tract.


Asunto(s)
Células Epiteliales/inmunología , Hipersensibilidad a los Alimentos/metabolismo , Inmunoglobulina E/metabolismo , Absorción Intestinal/inmunología , Mucosa Intestinal/inmunología , Receptores de IgE/inmunología , Adolescente , Antígenos CD/genética , Antígenos CD/inmunología , Antígenos CD/metabolismo , Línea Celular , Niño , Preescolar , Células Epiteliales/metabolismo , Células Epiteliales/patología , Heces/química , Hipersensibilidad a los Alimentos/inmunología , Hipersensibilidad a los Alimentos/patología , Expresión Génica , Humanos , Inmunoglobulina E/inmunología , Mucosa Intestinal/metabolismo , Mucosa Intestinal/patología , ARN Mensajero/genética , Receptores de IgE/genética , Receptores de IgE/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa
17.
Infect Immun ; 70(5): 2304-10, 2002 May.
Artículo en Inglés | MEDLINE | ID: mdl-11953364

RESUMEN

Enterohemorrhagic Escherichia coli (EHEC) infections are associated with hemorrhagic colitis and the hemolytic-uremic syndrome (HUS). In vivo, elevated plasma levels of the proinflammatory cytokine interleukin-8 (IL-8) in EHEC-infected children are correlated with a high risk of developing HUS. As IL-8 gene transcription is regulated by the transcription factors NF-kappaB and AP-1, we analyzed the role of these factors in the regulation of IL-8 production after infection of the epithelial intestinal T84 cell line by EHEC. By 6 h of infection, EHEC had induced significant secretion of IL-8 (35.84 +/- 6.76 ng/ml versus 0.44 +/- 0.04 ng/ml in control cells). EHEC induced AP-1 and NF-kappaB activation by 3 h of infection. Moreover, the three mitogen-activated protein kinases (MAPK) (ERK1/2, p38, and JNK) were phosphorylated in EHEC-infected T84 cells concomitant with induction of AP-1 DNA binding activity, and IkappaB-alpha was phosphorylated and then degraded concomitant with induction of NF-kappaB DNA binding activity. Pretreatment of cells with the highly specific MEK1/2 inhibitor U0126, the p38 inhibitor SB203580, and/or the proteasome inhibitor ALLN led to inhibition of the IL-8 secretion induced in EHEC-infected T84 cells. These findings demonstrate that (i) EHEC can induce in vitro a potent proinflammatory response by secretion of IL-8 and (ii) the secretion of IL-8 is due to the involvement of MAPK, AP-1, and NF-kappaB signaling pathways.


Asunto(s)
Infecciones por Escherichia coli/inmunología , Escherichia coli O157 , Interleucina-8/biosíntesis , Proteínas Quinasas Activadas por Mitógenos/metabolismo , FN-kappa B/metabolismo , Factor de Transcripción AP-1/metabolismo , Línea Celular , Activación Enzimática , Humanos , Lipopolisacáridos/toxicidad , Fosforilación
18.
Infect Immun ; 71(2): 766-73, 2003 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-12540556

RESUMEN

Enterohemorrhagic Escherichia coli (EHEC) infections are associated with the modification of tight-junction permeability and synthesis of proinflammatory cytokine interleukin-8 (IL-8). In a previous study, it was demonstrated that EHEC-induced IL-8 secretion is due to the involvement of the mitogen-activated protein kinase (MAPK), AP-1, and NF-kappaB pathways. In this study, we investigated the effect of the yeast Saccharomyces boulardii on EHEC infection in T84 cells. For this purpose, cells were (i) incubated with bacteria and yeast at the same time or (ii) incubated overnight with yeast cells that were maintained during infection or eliminated by several washes before infection. Coincubation is sufficient to maintain the transmonolayer electrical resistance (TER) of EHEC-infected cells, whereas the preincubation of cells with the yeast without elimination of the yeast during infection is necessary to significantly decrease IL-8 secretion. We thus analyzed the mechanisms of S. boulardii action. We showed that S. boulardii has no effect on EHEC growth or on EHEC adhesion. Kinetics studies revealed that EHEC-induced myosin light chain (MLC) phosphorylation precedes the decrease of TER. ML-7, an MLC kinase inhibitor, abolishes the EHEC-induced MLC phosphorylation and decrease of TER. Studies show that S. boulardii also abolishes EHEC-induced MLC phosphorylation. We demonstrated that the preincubation of cells with S. boulardii without washes before EHEC infection inhibits NF-kappaB DNA binding activity, phosphorylation and degradation of IkappaB-alpha, and activation of the three members of a MAPK group (extracellular signal-regulated protein kinases 1 and 2, p38, and c-jun N-terminal kinase). These findings demonstrate that S. boulardii exerts a preventive effect on EHEC infection by (i) interfering with one of the transduction pathways implicated in the control of tight-junction structure and (ii) decreasing IL-8 proinflammatory secretion via inhibition of the NF-kappaB and MAPK signaling pathways in infected T84 cells.


Asunto(s)
Escherichia coli O157/patogenicidad , Saccharomyces/fisiología , Transducción de Señal , Adhesión Bacteriana , Línea Celular , Permeabilidad de la Membrana Celular , Colon/microbiología , Impedancia Eléctrica , Escherichia coli O157/crecimiento & desarrollo , Humanos , Interleucina-8/biosíntesis , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Cadenas Ligeras de Miosina/metabolismo , FN-kappa B/metabolismo , Fosforilación
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