epidermal-growth-factor and Colitis

The occurrence in the neonatal period and into early infancy of two inflammatory conditions, necrotising enterocolitis (NEC), and allergic colitis, that do not occur in later life highlight the peculiar vulnerability of the gastrointestinal tract in the newborn period to otherwise innocuous insults. The pathogenesis of the relatively benign allergic colitis as a mucosal inflammatory process driven by dietary antigens is relatively well characterised, and its treatment with dietary manipulation is well established. For NEC, hypoxic/ischaemic insult, mucosal immaturity, and its interaction with the intestinal microflora are understood to be the main factors in pathogenesis. Thus far, the most productive interventions have been in preventative approaches, in particular feeding strategies, to reduce the incidence of the condition whilst establishing adequate growth and progression onto enteral feeding. For established NEC, supportive medical therapy or surgical intervention remains the mainstay or treatment, although novel therapies, such as platelet-activating factor (PAF) inhibitors and epidermal growth factor (EGF), have shown some promise in animal models of the condition.

Topics: Colitis; Enterocolitis, Necrotizing; Epidermal Growth Factor; Gastrointestinal Tract; Humans; Infant, Newborn; Platelet Activating Factor

Growth factors and their receptors play important roles in cell proliferation, migration, tissue injury repair and ulcer healing. In gastric mucosa, transforming growth factor alpha (TGF-alpha) and epidermal growth factor (EGF) by activating their common receptor, control cell proliferation. TGF-alpha predominantly plays this role under normal conditions and after acute injury, while EGF exerts its actions mainly during healing of chronic ulcers. During regeneration of injured gastric mucosa, these growth factors serve predominantly to restore the epithelial component. Other growth factors, basic fibroblast growth factor (bFGF) and vascular endothelial growth factor (VEGF) serve to promote restoration of the connective tissue and microvessels (angiogenesis) in injured mucosa. During healing of chronic ulcers, a new epithelial lineage secreting EGF and other growth peptides develops and the majority of cells lining the ulcer margin overexpress the EGF receptor. Activation of the EGF receptor induces dramatic increases in MAP (Erk -1 and -2) kinase activity and phosphorylation levels. Inhibition of this signaling pathway dramatically delays ulcer healing. Granulation connective tissue, which grows under the stimulation of bFGF and VEGF is the major source for regeneration of connective tissue lamina propria and microvessels within the ulcer scar. Other growth factors such as insulin - like growth factor, keratinocyte growth factor, hepatocyte growth factor and trefoil peptides have been implicated in gastrointestinal (gastric ulcers, colitis) regeneration following injury. This paper is intended to provide an overview of the role of growth factors in gastrointestinal mucosal regeneration.

Topics: Animals; Cell Division; Colitis; Epidermal Growth Factor; Gastric Mucosa; Growth Substances; Helicobacter Infections; Helicobacter pylori; Humans; Intestinal Mucosa; Neovascularization, Physiologic; Regeneration; Signal Transduction; Stomach Ulcer; Transforming Growth Factor alpha; Wound Healing

Many adenocarcinomas, including colorectal cancer (CRC), overexpress the MUC13 cell surface mucin, but the functional significance and mechanisms are unknown. Here, we report the roles of MUC13 in colonic tumorigenesis and tumor progression. High-MUC13 expression is associated with poor survival in two independent patient cohorts. In a comprehensive series of in vivo experiments, we identified a critical role for MUC13 in the development of this malignancy, by promoting survival and proliferation of tumor-initiating cells and driving an immunosuppressive environment that protects tumors from checkpoint inhibitor immunotherapy. In Muc13-deficient mice, fewer tumors are generated after exposure to carcinogens and inflammation, they have markedly reduced β-catenin signaling, have more tumor-infiltrating CD103

Topics: Animals; Antigens, Surface; Apoptosis; beta Catenin; Biomarkers, Tumor; Carcinogenesis; Cell Proliferation; Cohort Studies; Colitis; Colorectal Neoplasms; Epidermal Growth Factor; Gene Expression Regulation, Neoplastic; Humans; Mice; Mice, Inbred C57BL; Mice, Knockout; Mucins; Neoplastic Stem Cells; Prognosis; Survival Rate; Tumor Cells, Cultured

Macrophages play a crucial role in maintaining homeostasis in the intestine, but the underlying mechanisms have not yet been elucidated fully. Here, we show for the first time that mature intestinal macrophages in mouse intestine express high levels of αvβ5 integrin, which acts as a receptor for the uptake of apoptotic cells and can activate molecules involved in several aspects of tissue homeostasis such as angiogenesis and remodeling of the ECM. αvβ5 is not expressed by other immune cells in the intestine, is already present on intestinal macrophages soon after birth, and its expression is not dependent on the microbiota. In adults, αvβ5 is induced during the differentiation of monocytes in response to the local environment and it confers intestinal macrophages with the ability to promote engulfment of apoptotic cells via engagement of the bridging molecule milk fat globule EGF-like molecule 8. In the absence of αvβ5, there are fewer monocytes in the mucosa and mature intestinal macrophages have decreased expression of metalloproteases and IL 10. Mice lacking αvβ5 on haematopoietic cells show increased susceptibility to chemical colitis and we conclude that αvβ5 contributes to the tissue repair by regulating the homeostatic properties of intestinal macrophages.

Topics: Animals; Cells, Cultured; Colitis; Epidermal Growth Factor; Gene Expression Regulation; Homeostasis; Humans; Integrin alpha5; Intestinal Mucosa; Intestines; Macrophages; Metalloproteases; Mice; Mice, Inbred C57BL; Mice, Knockout; Phagocytosis; Receptors, Vitronectin; Transplantation Chimera

Current treatments for inflammatory bowel disease (IBD) target the overactive immune response of the intestinal mucosa. However, epidermal growth factor (EGF), an activating ligand of the EGF receptor (EGFR), has been shown to induce disease remission through direct targeting of intestinal mucosal healing. Despite promising preclinical and clinical results, this EGFR-activating therapy has not progressed, in part due to the potential for carcinogenesis associated with long-term use and the increased risk of colitis-associated cancer (CAC) in IBD. Here we tested whether pharmacological modulation of EGFR altered outcomes of CAC in the murine azoxymethane/dextran sulfate sodium model. We found that administering EGF during the period of maximum colitis severity ("early"), coincident with the initiation and early promotion of tumors, improved outcomes of colitis and reduced tumor size. In contrast, daily EGF administration beginning ~2 months after tumor initiation ("late") increased tumor size. Administration of the EGFR kinase inhibitor gefitinib increased the tumor size when the drug was given early and decreased the tumor size when the drug was administered late. EGF administration not only reduced colonic cytokine and chemokine expression during injury, but also baseline chemokine expression in homeostasis. These results suggest that EGFR activation during acute bouts of colitis may reduce the long-term burden of CAC.

Topics: Animals; Azoxymethane; Colitis; Colonic Neoplasms; Dextran Sulfate; Epidermal Growth Factor; ErbB Receptors; Mice; Neoplasm Proteins; Neoplasms, Experimental; Signal Transduction

We studied changes in the expression of mRNA for mucins and claudins in the medial part of the colon in male C57Bl/6 mice on the model of acute and chronic colitis induced by substitution of drinking water with 1% solution of dextran sodium sulphate for 5 days. In acute colitis, the expression of the main structural component of glycocalyx, mucin Muc3, decreased and expression of pore-forming claudin Cldn2 increased, which reflected enhanced permeability of tight junctions. In the chronic colitis group, in comparison with the normal group, we observed an increase in expression of mRNA of main structural mucus component Muc2, enhanced of expression of Muc1 associated with carcinogenesis, and reduced expression of Muc13, which led to a more severe course of colitis; the expression of pore-forming claudin Cldn2 was elevated. These findings indicate that the imbalance in the expression of mucins and claudins plays an important role in the mechanisms of development of acute and chronic colitis.

Topics: Animals; Antigens, Surface; Claudins; Colitis; Colon; Epidermal Growth Factor; Male; Mice; Mice, Inbred C57BL; Mucin-1; Mucin-2; Mucins

Intestinal homeostasis is maintained by a hierarchy of immune defenses acting in concert to minimize contact between luminal microorganisms and the intestinal epithelial cell surface. The intestinal mucus layer, covering the gastrointestinal tract epithelial cells, contributes to mucosal homeostasis by limiting bacterial invasion. In this study, we used γδ T-cell-deficient (TCRδ(-/-)) mice to examine whether and how γδ T-cells modulate the properties of the intestinal mucus layer. Increased susceptibility of TCRδ(-/-) mice to dextran sodium sulfate (DSS)-induced colitis is associated with a reduced number of goblet cells. Alterations in the number of goblet cells and crypt lengths were observed in the small intestine and colon of TCRδ(-/-) mice compared with C57BL/6 wild-type (WT) mice. Addition of keratinocyte growth factor to small intestinal organoid cultures from TCRδ(-/-) mice showed a marked increase in crypt growth and in both goblet cell number and redistribution along the crypts. There was no apparent difference in the thickness or organization of the mucus layer between TCRδ(-/-) and WT mice, as measured in vivo. However, γδ T-cell deficiency led to reduced sialylated mucins in association with increased gene expression of gel-secreting Muc2 and membrane-bound mucins, including Muc13 and Muc17. Collectively, these data provide evidence that γδ T cells play an important role in the maintenance of mucosal homeostasis by regulating mucin expression and promoting goblet cell function in the small intestine.

Topics: Animals; Antigens, Surface; Colitis; Dextran Sulfate; Disease Models, Animal; Epidermal Growth Factor; Gene Expression Regulation; Glycosylation; Goblet Cells; Homeostasis; Immunity, Mucosal; Intestine, Small; Mice; Mice, Inbred C57BL; Mice, Knockout; Mucin-2; Mucins; Mucus; Organoids; Receptors, Antigen, T-Cell, gamma-delta; Time Factors; Tissue Culture Techniques

Molecular mechanisms specific to colitis-associated cancers have been poorly characterized. Using comparative whole-genome expression profiling, we observed differential expression of epiregulin (EREG) in mouse models of colitis-associated, but not sporadic, colorectal cancer. Similarly, EREG expression was significantly upregulated in cohorts of patients with colitis-associated cancer. Furthermore, tumor-associated fibroblasts were identified as a major source of EREG in colitis-associated neoplasms. Functional studies showed that Ereg-deficient mice, although more prone to colitis, were strongly protected from colitis-associated tumors. Serial endoscopic studies revealed that EREG promoted tumor growth rather than initiation. Additionally, we demonstrated that fibroblast-derived EREG requires ERK activation to induce proliferation of intestinal epithelial cells (IEC) and tumor development in vivo. To demonstrate the functional relevance of EREG-producing tumor-associated fibroblasts, we developed a novel system for adoptive transfer of these cells via mini-endoscopic local injection. It was found that transfer of EREG-producing, but not Ereg-deficient, fibroblasts from tumors significantly augmented growth of colitis-associated neoplasms in vivo. In conclusion, our data indicate that EREG and tumor-associated fibroblasts play a crucial role in controlling tumor growth in colitis-associated neoplasms.

Topics: Animals; Cell Line, Tumor; Cell Movement; Cell Proliferation; Colitis; Colon; Colorectal Neoplasms; Epidermal Growth Factor; Epiregulin; Extracellular Signal-Regulated MAP Kinases; Fibroblasts; Humans; Intestinal Mucosa; Ki-67 Antigen; MAP Kinase Signaling System; Mice; Mice, Inbred C57BL; Mice, Knockout; Oligonucleotide Array Sequence Analysis; Transcriptome; Tumor Burden

Selenium (Se) is an essential micronutrient that exerts its functions via selenoproteins. Little is known about the role of Se in inflammatory bowel disease (IBD). Epidemiological studies have inversely correlated nutritional Se status with IBD severity and colon cancer risk. Moreover, molecular studies have revealed that Se deficiency activates WNT signaling, a pathway essential to intestinal stem cell programs and pivotal to injury recovery processes in IBD that is also activated in inflammatory neoplastic transformation. In order to better understand the role of Se in epithelial injury and tumorigenesis resulting from inflammatory stimuli, we examined colonic phenotypes in Se-deficient or -sufficient mice in response to dextran sodium sulfate (DSS)-induced colitis, and azoxymethane (AOM) followed by cyclical administration of DSS, respectively. In response to DSS alone, Se-deficient mice demonstrated increased morbidity, weight loss, stool scores, and colonic injury with a concomitant increase in DNA damage and increases in inflammation-related cytokines. As there was an increase in DNA damage as well as expression of several EGF and TGF-β pathway genes in response to inflammatory injury, we sought to determine if tumorigenesis was altered in the setting of inflammatory carcinogenesis. Se-deficient mice subjected to AOM/DSS treatment to model colitis-associated cancer (CAC) had increased tumor number, though not size, as well as increased incidence of high grade dysplasia. This increase in tumor initiation was likely due to a general increase in colonic DNA damage, as increased 8-OHdG staining was seen in Se-deficient tumors and adjacent, non-tumor mucosa. Taken together, our results indicate that Se deficiency worsens experimental colitis and promotes tumor development and progression in inflammatory carcinogenesis.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Animals; Azoxymethane; Carcinogenesis; Colitis; Colonic Neoplasms; Deoxyguanosine; Dextran Sulfate; Diet; DNA Damage; Epidermal Growth Factor; Gene Expression Regulation; Inflammation; Mice; Mice, Inbred C57BL; Selenium; Signal Transduction; Transforming Growth Factor beta; Weight Loss

Ulcerative colitis is a chronic inflammatory disease and involves multiple etiological factors. Acetic acid (AA)-induced colitis is a reproducible and simple model, sharing many characteristics with human colitis. N-acetylcysteine (NAC) has been widely used as an antioxidant in vivo and in vitro. NAC can affect several signaling pathways involving in apoptosis, angiogenesis, cell growth and arrest, redox-regulated gene expression, and inflammatory response. Therefore, NAC may not only protect against the direct injurious effects of oxidants, but also beneficially alter inflammatory events in colitis. This study was conducted to investigate whether NAC could alleviate the AA-induced colitis in a porcine model.. Weaned piglets were used to investigate the effects of NAC on AA-induced colitis. Severity of colitis was evaluated by colon histomorphology measurements, histopathology scores, tissue myeloperoxidase activity, as well as concentrations of malondialdehyde and pro-inflammatory mediators in the plasma and colon. The protective role of NAC was assessed by measurements of antioxidant status, growth modulator, cell apoptosis, and tight junction proteins. Abundances of caspase-3 and claudin-1 proteins in colonic mucosae were determined by the Western blot method. Epidermal growth factor receptor, amphiregulin, tumor necrosis factor-alpha (TNF-α), and toll-like receptor 4 (TLR4) mRNA levels in colonic mucosae were quantified using the real-time fluorescent quantitative PCR.. Compared with the control group, AA treatment increased (P < 0.05) the histopathology scores, intraepithelial lymphocyte (IEL) numbers and density in the colon, myeloperoxidase activity, the concentrations of malondialdehyde and pro-inflammatory mediators in the plasma and colon, while reducing (P < 0.05) goblet cell numbers and the protein/DNA ratio in the colonic mucosa. These adverse effects of AA were partially ameliorated (P < 0.05) by dietary supplementation with NAC. In addition, NAC prevented the AA-induced increase in caspase-3 protein, while stimulating claudin-1 protein expression in the colonic mucosa. Moreover, NAC enhanced mRNA levels for epidermal growth factor and amphiregulin in the colonic mucosa.. Dietary supplementation with NAC can alleviate AA-induced colitis in a porcine model through regulating anti-oxidative responses, cell apoptosis, and EGF gene expression.

Topics: Acetic Acid; Acetylcysteine; Amphiregulin; Animals; Apoptosis; Caspase 3; Claudin-1; Colitis; Colitis, Ulcerative; Colon; Dietary Supplements; Dinoprostone; Disease Models, Animal; EGF Family of Proteins; Epidermal Growth Factor; ErbB Receptors; Free Radical Scavengers; Glycoproteins; Intercellular Signaling Peptides and Proteins; Interleukin-6; Intestinal Mucosa; Swine; Toll-Like Receptor 4; Transforming Growth Factor alpha; Tumor Necrosis Factor-alpha

Intestinal trefoil factor (ITF) has been proved to be effective in treatment of ulcerative colitis. However, the mechanisms of it remain unclear. In this study, we observed the effects of combined treatment with 5-aminosalicylic acid (5-ASA) and recombinant human ITF (rhITF) on the expression of Myeloperoxidase (MPO), nuclear factor-κB (NF-κB) and epidermal growth factor (EGF) in trinitrobenzene sulphonic acid (TNBS) induced colitis in rats. Forty Sprague-Dawley (SD) male rats which were induced to distal colitis by the colonic administration of TNBS, were randomly divided into four groups and colonically treated with normal saline (A), 5-ASA (B), rhITF (C), respectively. The macroscopic and histological changes of the colon, activities of MPO, expressions of serum EGF and tissue NF-κB were detected. The results showed that manifestation, colonic damage score and MPO activities of the rats treated with 5-ASA or/and rhITFs were improved, serum EGF production was augmented and expression of tissue NF-κB was down-regulated. Single usage of 5-ASA or rhITF had no significant difference, but combined using of them had more significant and noticeable effects compared to any single treatment. It could be concluded that topical treatment with 5-ASA and rhITF had beneficial effects in treating TNBS-induced colitis of rats and combined treatment was better than single treatment. It was possibly related to suppression of neutrophil infiltration, down-regulation expression of NF-κB and up-regulation expression of EGF.

Topics: Animals; Colitis; Epidermal Growth Factor; Male; NF-kappa B; Peptides; Peroxidase; Rats; Rats, Sprague-Dawley; Recombinant Proteins; Time Factors; Trefoil Factor-2; Trinitrobenzenesulfonic Acid

The MUC13 transmembrane mucin is highly and constitutively expressed in the small and large intestine. Although MUC13 polymorphisms have been associated with human inflammatory bowel diseases and susceptibility to Escherichia coli infection in pigs, the biological functions of MUC13 are unknown. This study aimed to explore whether MUC13 modulates intestinal inflammation.. Muc13(-/-) mice were generated, phenotyped and challenged with the colitis-inducing agent, dextran sodium sulphate (DSS). Colitis was assessed by clinical symptoms and intestinal histopathology. Intestinal epithelial cell apoptosis and proliferation, macrophage infiltration and cytokine production were also quantified. Apoptosis of human LS513 intestinal epithelial cells in response to apoptotic agents, including DSS, was also measured, following knockdown of MUC13 with siRNA.. Muc13(-/-) mice were viable, fertile and developed normally, with no spontaneous intestinal pathology except mild focal neutrophilic inflammation in the small and large intestines of old mice. In response to DSS challenge, Muc13(-/-) mice developed more severe acute colitis, as reflected by increased weight loss, rectal bleeding, diarrhoea and histological colitis scores compared with wild-type mice. Increased numbers of F4/80(+) macrophages in inflamed mucosa of Muc13(-/-) mice were accompanied by increased expression of intestinal IL-1β and TNFα mRNA. Muc13(-/-) mice had significantly increased intestinal epithelial cell apoptosis within 3 days of DSS exposure. LS513 cells were more susceptible to DSS, actinomycin-D, ultraviolet irradiation and TRAIL-induced apoptosis when MUC13 was knocked down by siRNA.. These novel findings indicate a protective role for Muc13 in the colonic epithelium by inhibiting toxin-induced apoptosis and have important implications for intestinal infections, inflammatory diseases and the development of intestinal cancer.

Topics: Animals; Antigens, Surface; Apoptosis; Cell Line; Cell Proliferation; Colitis; Cytokines; Dextran Sulfate; Epidermal Growth Factor; Humans; Inflammatory Bowel Diseases; Intestinal Mucosa; Macrophages, Peritoneal; Mice; Mice, Knockout; Mucins; Real-Time Polymerase Chain Reaction

Epiregulin (EPI) and amphiregulin (AR) are epidermal growth factor receptor (EGFR) ligands implicated in mucosal repair and tumorigenesis. We have shown that Toll-like receptor 4 (TLR4) induces intestinal epithelial cell (IEC) proliferation by activating EGFR through AR expression. We examined whether TLR4 differentially regulates expression of EGFR ligands in response to mucosal injury. The human IEC line SW480 was examined expression of EGFR ligands, EGFR phosphorylation, and proliferation in response to lipopolysaccharide (LPS). Small-interfering RNA (siRNA) was used to block TLR4. Neutralizing antibodies to EGFR ligands were used to examine inhibition of LPS-dependent EGFR activation. Acute colitis and recovery were examined in the mice given 2.5% dextran sodium sulfate (DSS). Colonic secretion of EPI and AR was analyzed by enzyme-linked immunosorbent assay. LPS selectively induces EPI and AR but not other EGFR ligands. LPS induced early EPI mRNA expression between 30 min and 24 h. The neutralizing antibodies to EPI and AR prevented activation of EGFR by LPS. LPS induces IEC proliferation (200%, P=0.01) in 24 h but blocking EPI and AR significantly decreased proliferation. In vivo, mucosal EPI and AR expression are significantly decreased in TLR4(-/-) mice (P=0.02) compared to wild-type mice during acute colitis. EPI and AR exhibit different kinetics in response to mucosal damage: EPI expression is upregulated acutely at day 7 of DSS, but falls during recovery at day 14. By contrast, a sustained upregulation of AR expression is seen during mucosal injury and repair. We show that TLR4 regulates EPI and AR expression and that both these EGFR ligands are necessary for optimal proliferation of IEC. The diverse kinetics of EPI and AR expression suggest that they function in distinct roles with respect to acute injury vs repair. Our results highlight the role of bacterial sensing for IEC homeostasis and may lead to targeted therapy for mucosal healing and prevention of tumorigenesis.

Topics: Amphiregulin; Animals; Antibodies, Neutralizing; Cell Line; Cell Proliferation; Colitis; Dextran Sulfate; EGF Family of Proteins; Enzyme-Linked Immunosorbent Assay; Epidermal Growth Factor; Epiregulin; Epithelial Cells; ErbB Receptors; Glycoproteins; Humans; Intercellular Signaling Peptides and Proteins; Lipopolysaccharides; Mice; Mice, Knockout; Mucous Membrane; Toll-Like Receptor 4; Up-Regulation

Intestinal smooth muscle cells are normally quiescent, but in the widely studied model of trinitrobenzene sulfonic acid (TNBS)-induced colitis in the rat, the onset of inflammation causes proliferation that leads to increased cell number and an altered phenotype. The factors that drive this are unclear and were studied in primary cultures of circular smooth muscle cells (CSMC) from the rat colon. While platelet-derived growth factor (PDGF)-AA, fibroblast growth factor (FGF), and epidermal growth factor (EGF) were ineffective, PDGF-BB and insulin-like growth factor-1 (IGF-1) caused significant increase in [(3)H]thymidine incorporation, bromodeoxyuridine uptake, and increased CSMC number, with PDGF-BB (≥0.2 nM) substantially more effective than IGF-1. Surprisingly, CSMC lacked expression of PDGF receptor-β (PDGF-Rβ) upon isolation but by 4 days in vitro, CSMC gained expression of PDGF-Rβ as shown by quantitative PCR, Western blot analysis, and immunocytochemistry; these CSMC responded to PDGF-BB but not IGF-1. PDGF-BB caused PDGF-Rβ phosphorylation and mobilization from the surface membrane, leading to activation of both Akt and ERK signaling pathways, which were essential for subsequent proliferation. In contrast, PDGF-AA, FGF, EGF, and IGF-1 were ineffective. In vivo, control CSMC lacked expression of PDGF-Rβ. However, this changed rapidly with TNBS-colitis, and by day 2 when CSMC proliferation in vivo is maximal, freshly isolated CSMC showed on-going PDGF-Rβ phosphorylation that was further increased by exogenous PDGF-BB. This suggests that the onset of PDGF-Rβ expression is a key factor in CSMC growth in vitro and in vivo, where inflammation may damage intrinsic inhibitory mechanisms and thus lead to hyperplasia.

Topics: Animals; Becaplermin; Cell Proliferation; Cells, Cultured; Colitis; Epidermal Growth Factor; Humans; Insulin-Like Growth Factor I; Intercellular Signaling Peptides and Proteins; Intestines; Male; Myocytes, Smooth Muscle; Platelet-Derived Growth Factor; Proto-Oncogene Proteins c-sis; Rats; Rats, Sprague-Dawley; Receptor, Platelet-Derived Growth Factor beta; Signal Transduction; Trinitrobenzenesulfonic Acid

Toll-like receptors (TLRs) trigger intestinal inflammation when the epithelial barrier is breached by physical trauma or pathogenic microbes. Although it has been shown that TLR-mediated signals are ultimately protective in models of acute intestinal inflammation [such as dextran sulfate sodium (DSS)-induced colitis], it is less clear which cells mediate protection. Here we demonstrate that TLR signaling in the nonhematopoietic compartment confers protection in acute DSS-induced colitis. Epithelial cells of MyD88/Trif-deficient mice express diminished levels of the epidermal growth factor receptor (EGFR) ligands amphiregulin (AREG) and epiregulin (EREG), and systemic lipopolysaccharide administration induces their expression in the colon. N-ethyl-N-nitrosourea (ENU)-induced mutations in Adam17 (which is required for AREG and EREG processing) and in Egfr both produce a strong DSS colitis phenotype, and the Adam17 mutation exerts its deleterious effect in the nonhematopoietic compartment. The effect of abrogation of TLR signaling is mitigated by systemic administration of AREG. A TLR→MyD88→AREG/EREG→EGFR signaling pathway is represented in nonhematopoietic cells of the intestinal tract, responds to microbial stimuli once barriers are breached, and mediates protection against DSS-induced colitis.

Topics: Adaptor Proteins, Vesicular Transport; Amphiregulin; Animals; Colitis; Dextran Sulfate; EGF Family of Proteins; Epidermal Growth Factor; Epiregulin; ErbB Receptors; Glycoproteins; Hematopoietic System; Intercellular Signaling Peptides and Proteins; Ligands; Lipopolysaccharides; Metagenome; Mice; Mice, Inbred C57BL; Mutation; Myeloid Differentiation Factor 88; Phenotype; Signal Transduction; Toll-Like Receptors

Epidermal growth factor (EGF) and transforming growth factor alpha (TGFalpha) protect gastrointestinal mucosa against injury. Having shown earlier, that TGFalpha but not EGF is locally increasingly expressed after mucosal injury in the colon, we now wanted to explore the pattern of expression of EGF and TGFalpha in the remaining gastrointestinal tract and to infer from the pattern of expression, to possible signals for the induction of the growth factor expression and further mechanisms for mucosal protection.. The trinitrobenzene sulfonic acid/ethanol-induced model of colitis in rats was used. TGFalpha-mRNA and EGF-mRNA expression was evaluated in inflamed and noninflamed colon, in the ileum, jejunum, duodenum, stomach, and in the submandibulary glands.. A significant increase of TGFalpha-mRNA and EGF-mRNA expressions was detected in the duodenal mucosa and a significant increase in TGFalpha-mRNA expression was observed in the inflamed colonic mucosa after mucosal injury in the colon within the first hours of colitis.. The increased expression of EGF and TGFalpha in the duodenum may lead to neutralization of gastric acid and proteolytic enzymes in the upper gastrointestinal tract during the course of colitis. Possible signals for the increased expression of EGF and TGFalpha presumably are fasting, parasympathetic, or adrenergic parts of the enteric nervous system or yet unknown mechanisms.

Topics: Animals; Colitis; Colon; Duodenum; Epidermal Growth Factor; Ethanol; Gene Expression; Intestinal Mucosa; Male; Models, Animal; Rats; Rats, Sprague-Dawley; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Transforming Growth Factor alpha; Trinitrobenzenesulfonic Acid

Adiponectin (APN) is an adipokine that regulates insulin sensitivity and is anti-inflammatory in atherosclerosis. The goal of this study was to investigate the role of APN in intestinal inflammation.. APN knockout (KO) mice and their wild-type (WT) littermates received dextran sulfate sodium (DSS) or trinitrobenzene sulfonic acid (TNBS) to induce intestinal inflammation. Clinical and histologic scores and proliferation of epithelial cells were assessed. Cytokines and APN levels were measured. Expression of APN and heparin binding epidermal growth factor (HB-EGF) was analyzed by immunohistochemistry. Expression of APN and its receptors, HB-EGF, and basic fibroblast growth factor (bFGF) messenger RNA was assessed by reverse-transcription polymerase chain reaction. Association of serum APN with HB-EGF and bFGF was studied by coimmunoprecipitation.. APN KO mice are protected from chemically induced colitis; administration of APN restores inflammation. APN is expressed in the colon, luminal APN associates with colonic epithelial cells. In vitro, APN increases production of proinflammatory cytokines from colonic tissue. Expression of colonic APN overlaps with that of bFGF and HB-EGF, which play a protective role in colitis. Circulating APN binds to bFGF and HB-EGF, likely inhibiting their protective activity. Inhibition of EGF receptor signaling, which is required for biologic activity of HB-EGF, restores inflammation in APN KO mice.. APN deficiency is associated with protection from chemically induced colitis. APN exerts proinflammatory activities in the colon by inducing production of proinflammatory cytokines and inhibiting bioactivity of protective growth factors. Thus, in colitis, APN exerts an opposite role compared with atherosclerosis.

Topics: Adiponectin; Animals; Chemokine CXCL2; Chemokines; Colitis; Colon; Cytokines; Dextran Sulfate; Disease Models, Animal; Epidermal Growth Factor; Female; Fibroblast Growth Factor 2; Heparin-binding EGF-like Growth Factor; Intercellular Signaling Peptides and Proteins; Interleukin-6; Intestinal Mucosa; Mice; Mice, Inbred C57BL; Mice, Knockout; Organ Culture Techniques; Receptors, Adiponectin; Receptors, Cell Surface; Severity of Illness Index; Time Factors; Trinitrobenzenesulfonic Acid

The flavonoid luteolin has anti-inflammatory properties both in vivo and in vitro. However, the impact of luteolin on experimental models of colitis is unknown.. To address the therapeutic impact of luteolin, NF-kappaB(EGFP) transgenic mice were fed a chow diet containing 2% luteolin- or isoflavone-free control chow (AIN-76), and acute colitis was induced using 3% dextran sodium sulfate (DSS). Additionally, development of spontaneous colitis was evaluated in IL-10(-/-);NF-kappaB(EGFP) transgenic mice fed 2% luteolin chow diet or control chow diet. Interestingly, NF-kappaB(EGFP) transgenic mice exposed to luteolin showed worse DSS-induced colitis (weight loss, histological scores) compared to control-fed mice, whereas spontaneous colitis in IL-10(-/-);NF-kappaB(EGFP) mice was significantly attenuated. Macroscopic imaging of live resected colon showed enhanced EGFP expression (NF-kappaB activity) in luteolin-fed mice as compared to control-fed animals after DSS exposure, while cecal EGFP expression was attenuated in luteolin-fed IL-10(-/-) mice. Interestingly, confocal microscopy showed that EGFP positive cells were mostly located in the lamina propria and not in the epithelium. Caspase 3 activation was significantly enhanced whereas COX-2 gene expression was reduced in luteolin-fed, DSS-exposed NF-kappaB(EGFP) transgenic mice as assessed by Western blot and immunohistochemical analysis. In vitro, luteolin sensitized colonic epithelial HT29 cells to TNFalpha-induced apoptosis, caspase 3 activation, DNA fragmentation and reduced TNFalpha-induced C-IAP1, C-IAP2 and COX-2 gene expression.. We conclude that while luteolin shows beneficial effects on spontaneous colitis, it aggravates DSS-induced experimental colitis by blocking NF-kappaB-dependent protective molecules in enterocytes.

Topics: Adenoviridae; Animals; Caspase Inhibitors; Caspases; Colitis; Dinoprostone; Epidermal Growth Factor; Gene Expression Regulation; Genetic Vectors; HT29 Cells; Humans; Interleukin-10; Luciferases; Luteolin; Mice; Mice, Transgenic; NF-kappa B; Reference Values; Reverse Transcriptase Polymerase Chain Reaction; RNA; RNA, Neoplasm; Tumor Necrosis Factor-alpha

Novel therapies for the treatment of colitis are required. We therefore examined the potential value of the trefoil factor family 1 (TFF1) peptide and epidermal growth factor (EGF) alone and in combination. Effects of TFF1- Cys58 +/- EGF on an in vitro HT29 cell wounding model of restitution showed synergistic activity when used in combination. In addition, animals had colitis induced by adding 4% dextran sulphate sodium (DSS) to the drinking water for 7 days and they also received twice daily subcutaneous injections of test peptides. Treatment with TFF1-Cys58 alone (100 microg/kg) reduced histological colitis score by 22%, but the TFF1-Ser58 variant was ineffective. In a second study, TFF1-Cys58 reduced histological colitis score by 15%, EGF (600 microg/kg) by 26%, and an additive response (42% reduction) was demonstrated when used together (P < 0.01 versus either peptide given alone). Similar results were found using tissue myeloperoxidase (MPO) activity as a marker of inflammation. Where clinical risk/benefit seems justified, these initial studies suggest that combination therapy of systemic EGF and TFF peptides may prove useful for treatment of colitis in patients with disease extending beyond the reach of topical (enema) therapy.

Topics: Animals; Colitis; Colon; Dextran Sulfate; Drug Synergism; Epidermal Growth Factor; HT29 Cells; Humans; Lipopolysaccharides; Male; Mucins; Muscle Proteins; Peptides; Peroxidase; Proteins; Rats; Rats, Sprague-Dawley; Recombinant Proteins; Trefoil Factor-1; Trefoil Factor-2; Tumor Suppressor Proteins; Wound Healing

There have been several reports implying a benefit for heparin therapy in patients with refractory ulcerative colitis. Although this effect has been attributed to the anti-inflammatory properties of heparin, other mechanisms have not been excluded. Heparin is a potent modulator of receptor binding of growth factors such as fibroblast growth factor (FGF), vascular endothelial growth factor, and heparin-binding epidermal growth factor (HB-EGF), that play a role in wound repair. We examined the effect of heparin on the functional levels of FGF and HB-EGF in a model of experimental colitis. Fifty-six Wistar rats were divided into four groups: group 1 was the control group, group 2 received s.c. heparin 50 units/kg/d, group 3 underwent induction of 3% iodoacetamide colitis, and group 4 underwent induction of colitis and heparin treatment. Rats were killed and evaluated for severity of colitis by macroscopic and microscopic colitis scores, area of inflammation, and myeloperoxidase levels. FGF and HB-EGF levels were functionally assessed in colonic tissue in each group. Heparin therapy resulted in significant improvement in macroscopic and microscopic features of colitis (p < 0.05), accompanied by a partial reduction in myeloperoxidase levels. FGF receptor binding activity was identical in groups 1 and 2 but increased more than 3-fold after colitis induction in group 3 (p < 0.05). Treatment with heparin caused a significant decrease in FGF concentration. Levels of HB-EGF binding activity were similar in groups 1 and 2 and decreased in group 3 (p < 0.01). Heparin caused a significant increase in HB-EGF content in group 4 (p < 0.05). Levels of growth factors are altered differently in experimental colitis. Colonic FGF binding activity increases with colitis, whereas HB-EGF binding decreases with colitis. These trends were reversed by heparin, concomitant with a clinical and pathologic improvement in colitis. We suggest that one mechanism of heparin-mediated improvement in colitis may involve tissue healing associated with changes in functional levels of colonic growth factors.

Topics: Animals; Colitis; Drug Evaluation, Preclinical; Epidermal Growth Factor; Fibroblast Growth Factors; Heparin; Heparin-binding EGF-like Growth Factor; Intercellular Signaling Peptides and Proteins; Intestinal Mucosa; Iodoacetamide; Male; Peroxidase; Protein Binding; Rats; Rats, Wistar

It has been reported that epithelial growth factor (EGF) and transforming growth factor-alpha (TGF-alpha) play an important role in colonic mucosal defense and repair. Waved-2 (wa-2) mice harboring a defect EGF-R and phenotypically similar to TGF-alpha knockout mice provide a novel approach to study the role of EGF-R ligands in the maintenance and repair of colonic mucosa.. Acute colonic mucosal injury was induced by oral administration of dextran sodium sulfate (DSS: 5 g%) given for 6 days ad libitum to wa-2 homozygotes and their genetic controls (n = 10, each group), as well as to wa-2 mice with and without exogenous EGF administration. Severity of colonic injury was assessed histologically of the entire colon and graded. A crypt damage score (CDS) reflecting all three grades of mucosal pathology was calculated. Decrease in total body weight, colon length and colonic blood content was determined for all groups.. Thirty-eight percent of the entire colonic mucosa was destroyed in wa-2 animals compared to 15% in control mice. The CDS was 16.0 +/- 1.4 and 9.6 +/- 0.8 in wa-2 and control mice, respectively. EGF application to wa-2 mice did not reduce the severity of mucosal injury (CDS: 18.9 +/- 1.7 and 19.4 +/- 2.1 in EGF and vehicle injected mice, respectively).. The increased susceptibility of wa-2 mice to DSS demonstrates the pivotal role of EGF-R ligands such as EGF and TGF-alpha in preserving the integrity of the colonic mucosa against mucosal injury. The missing beneficial effect of exogenous EGF administration in these mice further underlines the importance of an intact ligand/EGF-R pathway.

Topics: Animals; Colitis; Colon; Dextran Sulfate; Epidermal Growth Factor; ErbB Receptors; Female; Genetic Predisposition to Disease; Homozygote; Intestinal Mucosa; Mice; Mice, Knockout; Mice, Mutant Strains; Transforming Growth Factor alpha

Epidermal growth factor (EGF) and transforming growth factor alpha (TGF-alpha), members of the EGF family of growth factors, protect rat gastric and colonic mucosa against injury. Having shown previously that exogenously applied EGF protects rat colonic mucosa against injury, the aim of the present study was to evaluate the endogenously expressed ligand mediating the protective effect of EGF/TGF-alpha in vivo.. In an experimental model of trinitrobenzene sulphonic acid (TNBS)/ethanol induced colitis in rats EGF and TGF-alpha expression was evaluated using a ribonuclease protection assay, northern blot analysis, western blot analysis, and immunohistochemistry.. TGF-alpha mRNA increased 3-4 times at 4-8 hours after induction of colitis and returned to control levels within 24 hours. TGF-alpha immunoreactive protein with a molecular size of about 28 kDa representing TGF-alpha precursors increased markedly after induction of colitis with a peak at 8-12 hours. No fully processed 5.6 kDa TGF-alpha protein was detected in normal or inflamed colon tissue. Only a weak signal for EGF mRNA expression was detected in the rat colon and no EGF protein was observed by immunohistochemistry or western blot analysis.. TGF-alpha precursors are the main ligands for the EGF receptor in acute colitis. It is hypothesised that TGF-alpha precursors convey the biological activity of endogenous TGF-alpha peptides during mucosal defence and repair.

Topics: Acute Disease; Animals; Blotting, Northern; Blotting, Western; Colitis; DNA Primers; Epidermal Growth Factor; Ethanol; Gene Expression Regulation; Immunohistochemistry; Male; Protein Precursors; Rats; Rats, Sprague-Dawley; RNA, Messenger; Transforming Growth Factor alpha; Trinitrobenzenesulfonic Acid

The role of epidermal growth factor (EGF) in the maintenance of mucosal integrity in the lower gastrointestinal tract is unknown. The aim of this study was to determine the effect of EGF in experimental colitis.. Colitis was induced with 2,4,6-trinitrobenzenesulfonic acid/ethanol enemas. Rats were pretreated with intraperitoneal administration of recombinant human EGF (600 micrograms/kg) or vehicle 1 hour before induction of colitis and daily thereafter until killed at 8 hours, 48 hours, and 1 week. A separate group received an identical dosage and administration of EGF or vehicle for 1 week with treatment initiated 24 hours after the induction of colitis. Colonic tissue was evaluated macroscopically, histologically, and for myeloperoxidase activity.. Pretreatment with EGF reduced microscopic erosions at 8 and 48 hours by 74% and 54%, respectively (P < 0.05). At 1 week, microscopic ulcerations and myeloperoxidase activity were reduced by 65% in the EGF-pretreated group (P < 0.05). No significant difference in macroscopic injury, histological damage, or myeloperoxidase activity was noted when EGF treatment was initiated after the induction of colitis.. Systemic EGF administration reduces mucosal damage and inflammation in a trinitrobenzenesulfonic acid/ethanol model of colitis in rats through a mechanism involving mucosal protection.

Topics: Animals; Colitis; Disease Models, Animal; Epidermal Growth Factor; Ethanol; Gastric Mucosa; Male; Peroxidase; Rats; Rats, Sprague-Dawley; Trinitrobenzenesulfonic Acid

The effect of epidermal growth factor on the rate of healing was examined in a rat model of colitis. Ulceration and inflammation of the distal colon were induced by a single intracolonic administration of 0.25 ml of 30% ethanol containing 30 mg of trinitrobenzenesulfonic acid. Epidermal growth factor was delivered intracolonically via the rectum or s.c. for 7 days after induction of colitis. Repeated s.c. injections of epidermal growth factor (25 and 100 micrograms/kg/12 hr) or continuous s.c. delivery with Alzet osmotic pumps (50 and 200 micrograms/kg/24 hr) significantly reduced colonic ulceration and inflammation. Epidermal growth factor significantly reduced myeloperoxidase activity in colonic tissue and there was restitution of the glandular mucosa after epidermal growth factor treatment. In contrast, daily intracolonic treatment with epidermal growth factor (25, 100 and 200 micrograms/kg/24 hr) did not significantly reduce the colonic damage. However, intracolonic delivery of 5-aminosalicylic acid (100 and 200 mg/kg/24 hr) dose-dependently reduced the colonic damage as assessed macroscopically and histologically. We conclude that systemic and not intracolonic administration of epidermal growth factor can accelerate healing of colonic ulceration and is effective in reducing inflammation in this model of colitis.

Topics: Aminosalicylic Acids; Animals; Colitis; Colon; Disease Models, Animal; Eicosanoids; Epidermal Growth Factor; Ethanol; Intestinal Mucosa; Male; Mesalamine; Peroxidase; Prednisolone; Rats; Rats, Sprague-Dawley; Trinitrobenzenesulfonic Acid