The Experts below are selected from a list of 615 Experts worldwide ranked by ideXlab platform
Daniel K Podolsky - One of the best experts on this subject based on the ideXlab platform.
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Trefoil Peptide expression and goblet cell number in rat intestine effects of kgf and fasting refeeding
American Journal of Physiology-regulatory Integrative and Comparative Physiology, 2003Co-Authors: Concepcion Fernandezestivariz, Daniel K Podolsky, Carolyn R Jonas, Timothy M Wallace, Robert R Pascal, Kathryn L Devaney, Catherine L Farrell, Dean P Jones, Thomas R ZieglerAbstract:The Trefoil factor family Peptides TFF1, TFF2, and TFF3 are important for gut mucosal protection and restitution. Keratinocyte growth factor (KGF) stimulates proliferation and differentiation of ep...
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distinct pathways of cell migration and antiapoptotic response to epithelial injury structure function analysis of human intestinal Trefoil factor
Molecular and Cellular Biology, 2000Co-Authors: Koichi Kinoshita, Douglas Taupin, Hiroshi Itoh, Daniel K PodolskyAbstract:The Trefoil Peptide intestinal Trefoil factor (ITF) plays a critical role in the protection of colonic mucosa and is essential to restitution after epithelial damage. These functional properties are accomplished through coordinated promotion of cell migration and inhibition of apoptosis. ITF contains a unique three-looped Trefoil motif formed by intrachain disulfide bonds among six conserved cysteine residues, which is thought to contribute to its marked protease resistance. ITF also has a seventh cysteine residue, which permits homodimer formation. A series of cysteine-to-serine substitutions and a C-terminally truncated ITF were made by PCR site-directed mutagenesis. Any alteration of the Trefoil motif or truncation resulted in loss of protease resistance. However, neither an intact Trefoil domain nor dimerization was required to promote cell migration. This pro-restitution activity correlated with the ability of the ITF mutants to activate mitogen-activated protein (MAP) kinase independent of phosphorylation of the epidermal growth factor (EGF) receptor. In contrast, only intact ITF retained both phosphatidylinositol 3-kinase and the EGF receptor-dependent antiapoptotic effect in HCT116 and IEC-6 cells. The inability to block apoptosis correlated with a loss of Trefoil Peptide-induced transactivation of the EGF receptor or Akt kinase in HT-29 cells. In addition to defining structural requirements for the functional properties of ITF, these findings demonstrate that distinct intracellular signaling pathways mediate the effects of ITF on cell migration and apoptosis.
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mechanisms of regulatory Peptide action in the gastrointestinal tract Trefoil Peptides
Journal of Gastroenterology, 2000Co-Authors: Daniel K PodolskyAbstract:The Trefoil Peptide family is comprised of three small Peptides (designated pS2, SP, and ITF) exhibiting a unique motif of three intrachain loops formed by disulfide bonds. These highly protease-resistant Peptides are secreted onto the mucosal surface by goblet cells or their equivalents. Most importantly, these factors protect epithelium from injury and promote repair through restitution after injury has occurred. Targeted deletion of the gene encoding ITF results in exquisite sensitivity to colonic injury by standard agents (e.g., dextran sodium sulfate) due to an inability to repair the epithelium. Studies have led to insight into the intracellular responses to Trefoil Peptides, including ras-dependent MAP kinase activation and activation of epidermal growth factor receptor. Among other effects, activation of these pathways is associated with redistribution of E-cadherin from the cell surface to intracellular domains, where it is complexed with catenins, and phosphorylation of akt, inactivating this kinase associated with apoptosis. In addition, Trefoil Peptides appear to block both p53 dependent and p53 independent apoptosis through pathways associated with activation of EGFR and P13 kinase. These observations suggest that Trefoil Peptides elicit a coordinated cellular response enabling cell migration without triggering the programmed cell death response usually precipitated by cell detachment from a stationary anchored state.
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the Trefoil gene family are coordinately expressed immediate early genes egf receptor and map kinase dependent interregulation
Journal of Clinical Investigation, 1999Co-Authors: Douglas Taupin, Timothy C Wang, Woo Kyu Jeon, Kathryn J Devaney, Daniel K PodolskyAbstract:The Trefoil gene family of mucus cell–secreted proteins is a critical mediator of gastrointestinal mucosal restitution. Transcription of Trefoil genes is induced during mucosal repair, but the regulatory mechanisms involved are unknown. Mice deficient in the intestine-specific Peptide intestinal Trefoil factor (ITF), in which colonic restitution is lethally impaired, showed reduced expression of the gastric Trefoil genes SP and pS2, suggesting that Trefoil Peptides may individually regulate transcription of the entire family. In gastric cell lines, the Trefoils were shown to act in a manner suggestive of immediate-early genes capable of auto- and cross-induction through cis-acting regulatory regions. Trefoil-mediated transcriptional regulation required activation of the Ras/MEK/MAP kinase signal transduction pathway. EGF receptor (EGF-R) activation was also necessary for Trefoil auto- and cross-induction, and both spasmolytic polyPeptide (SP) and ITF stimulation of gastric cell lines led to phosphorylation of EGF-R. Nevertheless, ITF and ITF-thioredoxin cell surface binding at 4°C colocalized not with EGF-R, but with CD71, which is found in clathrin-coated pits, suggesting that integration of Trefoil Peptide responses may occur after internalization. As EGF-R expression is itself strongly induced after mucosal damage, the Trefoil/EGF-R relationship may be pivotal in the generation and maintenance of the mucosal repair phenotype.
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Trefoil Peptide expression and secretion is regulated by neuroPeptides and acetylcholine
American Journal of Physiology-gastrointestinal and Liver Physiology, 1997Co-Authors: Haruhiko Ogata, Daniel K PodolskyAbstract:Trefoil Peptides are a family of small proteins expressed by goblet cells that are secreted onto the apical gastrointestinal mucosal surface, where they are present in high concentrations. These Peptides appear to both protect the epithelium and promote healing after injury. However, the factors regulating the expression and secretion of these proteins contributing to mucosal defense have not been characterized. To determine the mechanisms controlling production of Trefoil Peptides, the human colon cancer-derived model cell line HT-29 was exposed to a variety of potential secretagogues. Expression and secretion of human intestinal Trefoil factor (hITF) as well as the intestinal apomucin MUC2 were assessed by Northern and Western blot analysis. Carbachol, an analog of acetylcholine, and the neuroendocrine Peptides somatostatin and vasoactive intestinal polyPeptide (VIP) stimulated increased expression of hITF mRNA within 5 min. These same factors stimulated parallel secretion of the hITF Peptide, with maximal stimulation observed at concentrations ranging from 10(-6) M (carbachol and somatostatin) to 10(-7) M (VIP). Expression and secretion of hITF in response to carbachol, VIP, and somatostatin was independent of production of apomucin. hITF was not regulated by other neuroendocrine transmitters including histamine and substance P. Similarly, hITF expression and secretion was not modulated by Peptide growth factors (epidermal growth factor, transforming growth factor-beta, and keratinocyte growth factor), cytokines [interleukin (IL)-1 beta, IL-2, IL-7, and IL-11], or arachidonic acid metabolites (prostaglandin E1/E2 and leukotriene B4). In conclusion, Trefoil Peptides appear to be integrated into mechanisms of mucosal defense and repair through the enteric neuroendocrine system and independent of the classical mucosal immune cytokine network.
N A Wright - One of the best experts on this subject based on the ideXlab platform.
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characterization of monoclonal antibodies raised to c terminal Peptides of ps2 a major Trefoil Peptide and motility factor expressed in adenocarcinomas and regions of mucosal injury
Human Pathology, 1996Co-Authors: R Williams, N A Wright, Gordon Stamp, G Elia, T Oates, E N LalaniAbstract:Two novel monoclonal antibodies, GE1 and GE2 raised against the C-terminal 31 and 28 amino acids of the estrogen-inducible Trefoil Peptide pS2, are described. Both antibodies are able to detect pS2 in formalin-fixed, paraffin-embedded tissues. Conditions are presented under which pS2 can be shown in cell lines by immunohistochemistry that has previously been problematic. The antibodies can specifically show the presence of pS2 in cell lysates by Western blotting and immunoprecipitation. In the form of an affinity column, the GE1 monoclonal antibody can be used to purify pS2 from MCF-7 supernatants. The eluted Peptide from the GE1 affinity column shows a single band at 6,600 Da (predicted size for pS2) on Western blotting. These antibodies are valuable reagents in the analysis of the role of Trefoil Peptides in the maintenance of mucosal integrity, and may have applications in the assessment of pS2 expression in chronic gastrointestinal ulceration and adenocarcinomas that secrete pS2, where it may serve as a prognostic marker.
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the ulceration associated cell lineage uacl reiterates the brunner s gland differentiation programme but acquires the proliferative organization of the gastric gland
The Journal of Pathology, 1994Co-Authors: Dennis J Ahnen, Richard Poulsom, G Elia, Gordon Stamp, Rosemary Jeffery, Janet M Longcroft, Christine Pike, Mariechristine Rio, Pierre Chambon, N A WrightAbstract:The ulceration-associated cell lineage (UACL) develops in the human gastrointestinal mucosa after ulceration; it grows out from the bases of adjacent crypts and ramifies in the lamina propria to form a new gland, finally giving rise to a duct by which the glandular secretion and indeed cells are carried to the surface. Using immunocytochemistry and in situ hybridization with 35S-labelled riboprobes, we have defined the pattern of Trefoil Peptide gene expression (pS2; human spasmolytic polyPeptide, hSP), epidermal growth factor/urogastrone (EGF/URO), and the distribution of cell proliferation during the development of the UACL, as indicated by immunostaining for proliferating cell nuclear antigen (PCNA). Our studies reveal that the morphogenesis of the UACL shows a marked morphological resemblance to developing Brunner's glands; the pattern of Trefoil Peptide gene expression during UACL development is also very similar. However, Trefoil Peptide gene expression in the mature UACL complex is unique amongst gastrointestinal cells. The mature UACL shows a distinctive proliferative organization: while the early buds and glands are non-proliferative, apparently being fed by cells from the parent crypts, a definitive proliferative zone develops within the duct. This, of course, corresponds to the location of the gastric gland proliferative zone. We propose that while the UACL shows novel features, it shares its differentiation programme with Brunner's glands, but its pattern of cell renewal eventually is that of the gastric gland.
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the production and characterization of a new monoclonal antibody to the Trefoil Peptide human spasmolytic polyPeptide
Histochemical Journal, 1994Co-Authors: G Elia, Rebecca Chinery, Richard Poulsom, Andrew M. Hanby, N A WrightAbstract:Human spasmolytic polyPeptide (hSP) is a member of the growing family of Trefoil Peptides which are expressed in discrete regions of the body, most notably the gastrointestinal tract. Much of the research into the localization of the spasmolytic polyPeptide has relied on hybridization in situ to detect its mRNA, due to the absence of a suitable antibody. The aim of the present study was to develop and characterize a monoclonal antibody against the human spasmolytic polyPeptide, using a combination of immunohistochemistry and hybridization in situ. After immunoblotting, the antibody detected a 14 kDa protein in gastrointestinal tissue extracts from the stomach and small intestine only. Using immunohistochemistry, human spasmolytic polyPeptide showed a distinctive staining pattern in the duodenum which co-localized with its mRNA. The co-localization of the immunoreactive Peptide with its mRNA provides good evidence that the antibody truly recognized human spasmolytic polyPeptide.
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spasmolytic polyPeptide is a major antral Peptide distribution of the Trefoil Peptides human spasmolytic polyPeptide and ps2 in the stomach
Gastroenterology, 1993Co-Authors: Andrew M. Hanby, Richard Poulsom, Rosemary Jeffery, G Elia, Sukhdev Singh, N A WrightAbstract:Abstract Background: The regional differences in the distribution of the Trefoil Peptides pS2 and human spasmolytic polyPeptide (hSP) within the stomach are not known. The aim of this study was to gain insight into these functionally obscure molecules by characterizing their distribution. Methods: Tissue from gastrectomy specimens removed for peptic ulceration was examined to chart the distribution of hSP and pS2, using immuno-histochemistry and in situ hybridization to chart their messenger RNAs (mRNAs). Results: Colocalization of pS2 and hSP was noted in the gastric foveolar and surface epithelium throughout the stomach. In the gastric antrum and pylorus, an extremely strong hSP mRNA signal was present within pyloric-type glands; strong hSP immunostaining was also seen at this site and in mucous-neck cells. Neither pS2 mRNA nor pS2 Peptide were shown within the deep portions of the pyloric glands. Within areas of intestinal metaplasia, a few goblet cells immunostained for pS2 and putative ulcer associated cell lineage was seen with a pattern of Trefoil Peptide localization similar to the ileum. Conclusion: The detailed function of these Trefoil Peptides is unknown, but their distribution suggests involvement in repair-enhancing mechanisms. hSP may be an important antral Peptide and both of these Peptides may play a specific reparative role.
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expression and purification of a Trefoil Peptide motif in a β galactosidase fusion protein and its use to search for Trefoil binding sites
FEBS Journal, 1993Co-Authors: Rebecca Chinery, Richard Poulsom, Andrew M. Hanby, G Elia, N A WrightAbstract:The cysteine-rich Trefoil motif of rat intestinal Trefoil factor (rITF) was cloned and expressed in Escherichia coli. A 270-bp cDNA fragment including the signal sequence and the Trefoil motif was cloned into the expression vector pAX5+ to direct the expression of a β-galactosidase collagenhinged fusion protein in E. coli. Cultures harbouring the recombinant plasmid produced a soluble novel protein with a molecular mass of 134.5 kDa, as predicted for the Trefoil-motif-containing fusion protein. Purification of the rITF moiety was achieved by p-aminophenyl-thio-β-d-galactoside(APTG)-affinity chromatography, collagenase digestion of the hybrid molecule, and removal of the β-galactosidase-hinge molecule by a further APTG-affinity step. It was demonstrated that intrachain disulphide-bond formation in rITF occurred during the procedure, so no refolding steps were required. Analysis by immunoblotting revealed that the fusion protein and the cleaved Trefoil-motif-containing protein were recognised by an antibody raised against the chemically synthesised Peptide. The Trefoil motif present in the fusion protein was used to localise putative Trefoil-binding sites in sections of frozen rat tissue. Binding was demonstrated using the β-galactosidase portion of the fusion protein as a reporter moiety, either directly with 5-bromo-4-chloro-3-indolyl-β-d-galactoside, or indirectly using a monoclonal antibody to β-galactosidase and indirect immunohistochemistry. Binding sites were localised to the foveolar and surface epithelium of rat stomach, the collecting ducts of the kidney and within colonic crypts. The presence of a Trefoil motif was necessary for binding. The use of β-galactosidase fusion proteins for histochemical localisation of Peptide-binding sites should prove more generally useful.
Richard Poulsom - One of the best experts on this subject based on the ideXlab platform.
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Trefoil Peptide tff2 treatment reduces vcam 1 expression and leukocyte recruitment in experimental intestinal inflammation
Journal of Leukocyte Biology, 2004Co-Authors: Antonio Sorianoizquierdo, Richard Poulsom, Felicity E. B. May, Lars Thim, Meritxell Gironella, Anna Massaguer, Antonio Salas, Miquel Sans, Josep M Pique, Julian PanesAbstract:There is evidence for a beneficial effect of Trefoil Peptides in animal models of gastric damage and intestinal inflammation, but the optimal treatment strategy and the mechanistic basis have not been explored thoroughly. It has been suggested that these proteins may modulate the inflammatory response. The aims of this study were to compare the protective and curative value of systemic and topical Trefoil factor family (TFF)2 administration in dextran sulfate sodium-induced experimental colitis and to investigate the relationship between the therapeutic effects of TFF2 and modulation of leukocyte recruitment and expression of cell adhesion molecules. Clinical and morphologic severity of colitis was evaluated at the end of the study (Day 10). Leukocyte-endothelial cell interactions were determined in colonic venules by fluorescence intravital microscopy. The expression of cell adhesion molecules vascular cell adhesion molecule 1 (VCAM-1) and mucosal addressin cell adhesion molecule 1 (MAdCAM-1) was measured by the dual radiolabeled monoclonal antibody technique. Pretreatment with TFF2 by subcutaneous or intracolonic (ic) route ameliorated the clinical course of colitis, and the luminal route had a significantly superior effect. This beneficial effect was correlated with significant reductions in endothelial VCAM-1 but not MAdCAM-1 expression and leukocyte adhesion to intestinal venules, which returned to levels similar to those of controls. In established colitis, ic TFF2 treatment did not modify the severity of colonic lesions. In conclusion, TFF2 is useful in the treatment of colitis, and topical administration is superior to the systemic route. Reduction in adhesion molecule expression and leukocyte recruitment into the inflamed intestine contributes to the beneficial effect of this treatment.
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Transgenic mice that overexpress the human Trefoil Peptide pS2 have an increased resistance to intestinal damage
Proceedings of the National Academy of Sciences of the United States of America, 1996Co-Authors: Raymond J. Playford, Tania Marchbank, Robert A. Goodlad, Rebecca Chinery, Richard Poulsom, Andrew M. HanbyAbstract:Abstract pS2 is a member of the Trefoil Peptide family, all of which are overexpressed at sites of gastrointestinal injury. We hypothesized that they are important in stimulating mucosal repair. To test this idea, we have produced a transgenic mice strain that expresses human pS2 (hpS2) specifically within the jejunum and examined the effect of this overexpression on proliferation and susceptibility to indomethacin-induced damage. A transgenic mouse was produced by microinjecting fertilized oocytes with a 1.7-kb construct consisting of rat intestinal fatty acid binding protein promoter (positions -1178 to +28) linked to full-length (490 bp) hpS2 cDNA. Screening for positive animals was by Southern blot analysis. Distribution of hpS2 expression was determined by using Northern and Western blot analyses and immunohistochemical staining. Proliferation of the intestinal mucosa was determined by assessing the crypt cell production rate. Differences in susceptibility to intestinal damage were analyzed in animals that had received indomethacin (85 mg/kg s.c.) 0-30 h previously. Expression of hpS2 was limited to the enterocytes of the villi within the jejunum. In the nondamaged intestine, villus height and crypt cell production rate were similar in transgenic and negative (control) litter mates. However, there was a marked difference in the amount of damage caused by indomethacin in control and transgenic animals in the jejunum (30% reduction in villus height in controls vs. 12% reduction in transgenic animals, P < 0.01) but the damage sustained in the non-hpS2-expressing ileal region was similar in control and transgenic animals. These studies support the hypothesis that Trefoil Peptides are important in stimulating gastrointestinal repair.
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the gene encoding human intestinal Trefoil factor tff3 is located on chromosome 21q22 3 clustered with other members of the Trefoil Peptide family
Genomics, 1996Co-Authors: Rebecca Chinery, Jill Williamson, Richard PoulsomAbstract:The gene coding for human intestinal Trefoil factor (hITF), a recently described cellular motogen produced by gastrointestinal goblet cells and epithelia elsewhere, is a member of the rapidly growing Trefoil Peptide family. In a rodent-human somatic cell hybrid panel, the hITF (HGMW-approved symbol TFF3) genomic locus segregated with human chromosome 21q. Fluorescence in situ hybridization with a 2.1-kb genomic probe of the hITF gene mapped this locus more precisely to the q22.3 region. Triple fluorescence in situ hybridization, together with physical mapping of human genomic DNA using pulsed-field gel electrophoresis, revealed that the hITF gene is tightly linked to those encoding the other known human Trefoil Peptides, namely the breast cancer estrogen-inducable gene pS2 (BCEI) and human spasmolytic polyPeptide (hSP/SML1). This gene family could become a useful marker for the genetic and physical mapping of chromosome 21 and for a better definition of the region involved in the clinical phenotype of several genetic diseases.
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the ulceration associated cell lineage uacl reiterates the brunner s gland differentiation programme but acquires the proliferative organization of the gastric gland
The Journal of Pathology, 1994Co-Authors: Dennis J Ahnen, Richard Poulsom, G Elia, Gordon Stamp, Rosemary Jeffery, Janet M Longcroft, Christine Pike, Mariechristine Rio, Pierre Chambon, N A WrightAbstract:The ulceration-associated cell lineage (UACL) develops in the human gastrointestinal mucosa after ulceration; it grows out from the bases of adjacent crypts and ramifies in the lamina propria to form a new gland, finally giving rise to a duct by which the glandular secretion and indeed cells are carried to the surface. Using immunocytochemistry and in situ hybridization with 35S-labelled riboprobes, we have defined the pattern of Trefoil Peptide gene expression (pS2; human spasmolytic polyPeptide, hSP), epidermal growth factor/urogastrone (EGF/URO), and the distribution of cell proliferation during the development of the UACL, as indicated by immunostaining for proliferating cell nuclear antigen (PCNA). Our studies reveal that the morphogenesis of the UACL shows a marked morphological resemblance to developing Brunner's glands; the pattern of Trefoil Peptide gene expression during UACL development is also very similar. However, Trefoil Peptide gene expression in the mature UACL complex is unique amongst gastrointestinal cells. The mature UACL shows a distinctive proliferative organization: while the early buds and glands are non-proliferative, apparently being fed by cells from the parent crypts, a definitive proliferative zone develops within the duct. This, of course, corresponds to the location of the gastric gland proliferative zone. We propose that while the UACL shows novel features, it shares its differentiation programme with Brunner's glands, but its pattern of cell renewal eventually is that of the gastric gland.
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the production and characterization of a new monoclonal antibody to the Trefoil Peptide human spasmolytic polyPeptide
Histochemical Journal, 1994Co-Authors: G Elia, Rebecca Chinery, Richard Poulsom, Andrew M. Hanby, N A WrightAbstract:Human spasmolytic polyPeptide (hSP) is a member of the growing family of Trefoil Peptides which are expressed in discrete regions of the body, most notably the gastrointestinal tract. Much of the research into the localization of the spasmolytic polyPeptide has relied on hybridization in situ to detect its mRNA, due to the absence of a suitable antibody. The aim of the present study was to develop and characterize a monoclonal antibody against the human spasmolytic polyPeptide, using a combination of immunohistochemistry and hybridization in situ. After immunoblotting, the antibody detected a 14 kDa protein in gastrointestinal tissue extracts from the stomach and small intestine only. Using immunohistochemistry, human spasmolytic polyPeptide showed a distinctive staining pattern in the duodenum which co-localized with its mRNA. The co-localization of the immunoreactive Peptide with its mRNA provides good evidence that the antibody truly recognized human spasmolytic polyPeptide.
Andrew S Giraud - One of the best experts on this subject based on the ideXlab platform.
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biliary epithelial Trefoil Peptide expression is increased in biliary diseases
Histopathology, 2002Co-Authors: G Srivatsa, Meera Ulaganathan, Neville D Yeomans, Andrew S Giraud, C Dow, Amanda NicollAbstract:Biliary epithelial Trefoil Peptide expression is increased in biliary diseases Aims: Maintenance of the cellular integrity of the biliary epithelium may involve the production of mucins and mucin-associated Peptides. In the luminal gastrointestinal tract, mucins and the mucin-associated Trefoil Peptides (TFF) are integral to cytoprotection and cellular repair of the mucosa. Methods and results: Samples of normal and diseased human liver tissue were examined using histological and immunohistochemical techniques, for the expression of TFF and mucins. Bile ducts were classified as small, medium or large depending upon the number of biliary epithelial cells. TFF expression was demonstrated in biliary epithelial cells of both normal and diseased liver tissue. TFF expression was greatest in the large bile ducts. In normal liver tissue, expression of at least one TFF was demonstrated in 2–7% of small bile ducts, 5–31% of medium bile ducts and 31–85% of large bile ducts. Seventy-seven percent of large bile ducts secreted mucins and all three TFF concurrently, compared with 3% of medium bile ducts and no small bile ducts. Biliary disease resulted in an increased expression of TFF1 and TFF3 in the medium bile ducts. Conclusions: The biliary epithelial cells in normal and diseased human liver tissue express TFF, particularly in the larger bile ducts. TFF expression may be up-regulated or induced in biliary diseases as a response to injury, as is seen in epithelial damage elsewhere in the gastrointestinal tract.
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effect of omeprazole induced achlorhydria on Trefoil Peptide expression in the rat stomach
Journal of Gastroenterology and Hepatology, 2001Co-Authors: Baolin Kang, Amanda Nicoll, Barbara M Alderman, G A Cook, Andrew S GiraudAbstract:Background: Omeprazole is an inhibitor of the H+K+ ATPase of the gastric parietal cell, which is used clinically to suppress gastric acid secretion. It has also been found to inhibit gastric mucin production; however, its effects on the synthesis and secretion of the Trefoil Peptides, which are also expressed by mucus cells, and which play a key role in cytoprotection and epithelial repair, are unknown. Methods: Rats (n = 8) were given either omeprazole (30 mg/kg per day; p.o.) or inert carrier for 1 week, and the effects on synthesis and Peptide expression of the gastric Trefoil Peptides, TFF1/pS2 and TFF2/SP, were compared. Results: As expected, omeprazole treatment abolished H+ ion production with a mean gastric juice pH of 7.2 compared with 2.4 for controls. The omeprazole group had elevated total protein levels of 35-fold and TFF1/pS2 Peptide levels elevated fourfold, respectively, but not TFF2/SP Peptide in gastric juice, suggesting that the increased pH reduced the viscosity of adherent mucus, thereby increasing gastric juice concentrations by dissolution of adherent TFF1/pS2 and increased secretion. Concomitant with increased TFF1/pS2 secretion was a fall in predominantly antral mucosal Trefoil Peptide concentrations. In contrast to Trefoil secretory rates, the steady-state synthesis of both TFF1/pS2 and TFF2/SP was unchanged after omeprazole treatment, implying both a large cellular pool of processed Peptide and rapid secretion. Conclusion: The increase in the concentration of TFF1/pS2 in gastric secretions during chronic omeprazole-induced achlorhydria may be important in preventing tissue injury and promoting repair in response to an increased luminal bacterial population.
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spatio temporal expression of Trefoil Peptide following severe gastric ulceration in the rat implicates it in late stage repair processes
Journal of Gastroenterology and Hepatology, 2001Co-Authors: Meera Ulaganathan, Mary Familari, Neville D Yeomans, Andrew S Giraud, Gregory A CookAbstract:Background: The Trefoil Peptide (TFF1) is a member of a family of mucin-associated regulatory Peptides that are widely distributed in gastrointestinal tissues and have been implicated in the maintenance of the gastric mucosa. The role of TFF1 in gastric mucosal repair was examined by analysis of the spatio-temporal expression of TFF1 following gastric ulceration in the rat. Methods: Gastric ulcers were induced in rats by application of glacial acetic acid to the serosa of the fundus. At various time points post injury (0–28 days), macroscopic and microscopic examination of the gastric mucosa was performed. In addition, the spatio-temporal expression of TFF1 protein and proliferating cell nuclear antigen were identified by immunohistochemistry, TFF1 message by in situ hybridization, and acidic/neutral secreting mucins by Alcian blue-periodic acid–Schiff staining. Results: In normal rat gastric tissue, TFF1 Peptide and mRNA were expressed in mucosal cells of the superficial epithelium. Trefoil Peptide and mRNA were significantly induced between 4 and 28 days post ulceration, with expression extending beyond the superficial epithelium and being localized to acidic mucin-producing cells deep within the repairing mucosa. Conclusions: Spatio-temporal expression of TFF1 mRNA and Peptide following macroscopic repair implicates TFF1 as a potential mediator of late stage-repair processes. Whether this is through direct stimulation of cellular differentiation or the enhancement of mucosal protective properties through an interaction with gastric mucins remains to be elucidated.
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x Trefoil Peptide and egf receptor ligand transgenic mice
American Journal of Physiology-gastrointestinal and Liver Physiology, 2000Co-Authors: Andrew S GiraudAbstract:The use of genetically engineered mice with both gain-of-function and loss-of-function mutations has been particularly informative about the normal and pathophysiological actions of a number of regulatory Peptides of the gastrointestinal tract. This review highlights some of the major findings pertinent to the epidermal growth factor (EGF) receptor and its ligands, particularly the major gut ligand transforming growth factor-α, as well as the Trefoil Peptides. Both of these Peptide families have important local actions in maintaining tissue homeostasis and repair after injury, and when mechanisms governing their regulation are disrupted they may contribute to disease progression. Future applications of transgenic technology to these areas are likely to be productive in furthering our understanding of the biology of these Peptides in health and disease.
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lessons from genetically engineered animal models x Trefoil Peptide and egf receptor ligand transgenic mice
American Journal of Physiology, 2000Co-Authors: Andrew S GiraudAbstract:The use of genetically engineered mice with both gain-of-function and loss-of-function mutations has been particularly informative about the normal and patho-physiological actions of a number of regulatory Peptides of the gastrointestinal tract. This review highlights some of the major findings pertinent to the epidermal growth factor (EGF) receptor and its ligands, particularly the major gut ligand transforming growth factor-a, as well as the Trefoil Peptides. Both of these Peptide families have important local actions in maintaining tissue homeostasis and repair after injury, and when mechanisms governing their regulation are disrupted they may contribute to disease progression. Future applications of transgenic technology to these areas are likely to be productive in furthering our understanding of the biology of these Peptides in health and disease.
Rebecca Chinery - One of the best experts on this subject based on the ideXlab platform.
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Transgenic mice that overexpress the human Trefoil Peptide pS2 have an increased resistance to intestinal damage
Proceedings of the National Academy of Sciences of the United States of America, 1996Co-Authors: Raymond J. Playford, Tania Marchbank, Robert A. Goodlad, Rebecca Chinery, Richard Poulsom, Andrew M. HanbyAbstract:Abstract pS2 is a member of the Trefoil Peptide family, all of which are overexpressed at sites of gastrointestinal injury. We hypothesized that they are important in stimulating mucosal repair. To test this idea, we have produced a transgenic mice strain that expresses human pS2 (hpS2) specifically within the jejunum and examined the effect of this overexpression on proliferation and susceptibility to indomethacin-induced damage. A transgenic mouse was produced by microinjecting fertilized oocytes with a 1.7-kb construct consisting of rat intestinal fatty acid binding protein promoter (positions -1178 to +28) linked to full-length (490 bp) hpS2 cDNA. Screening for positive animals was by Southern blot analysis. Distribution of hpS2 expression was determined by using Northern and Western blot analyses and immunohistochemical staining. Proliferation of the intestinal mucosa was determined by assessing the crypt cell production rate. Differences in susceptibility to intestinal damage were analyzed in animals that had received indomethacin (85 mg/kg s.c.) 0-30 h previously. Expression of hpS2 was limited to the enterocytes of the villi within the jejunum. In the nondamaged intestine, villus height and crypt cell production rate were similar in transgenic and negative (control) litter mates. However, there was a marked difference in the amount of damage caused by indomethacin in control and transgenic animals in the jejunum (30% reduction in villus height in controls vs. 12% reduction in transgenic animals, P < 0.01) but the damage sustained in the non-hpS2-expressing ileal region was similar in control and transgenic animals. These studies support the hypothesis that Trefoil Peptides are important in stimulating gastrointestinal repair.
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the gene encoding human intestinal Trefoil factor tff3 is located on chromosome 21q22 3 clustered with other members of the Trefoil Peptide family
Genomics, 1996Co-Authors: Rebecca Chinery, Jill Williamson, Richard PoulsomAbstract:The gene coding for human intestinal Trefoil factor (hITF), a recently described cellular motogen produced by gastrointestinal goblet cells and epithelia elsewhere, is a member of the rapidly growing Trefoil Peptide family. In a rodent-human somatic cell hybrid panel, the hITF (HGMW-approved symbol TFF3) genomic locus segregated with human chromosome 21q. Fluorescence in situ hybridization with a 2.1-kb genomic probe of the hITF gene mapped this locus more precisely to the q22.3 region. Triple fluorescence in situ hybridization, together with physical mapping of human genomic DNA using pulsed-field gel electrophoresis, revealed that the hITF gene is tightly linked to those encoding the other known human Trefoil Peptides, namely the breast cancer estrogen-inducable gene pS2 (BCEI) and human spasmolytic polyPeptide (hSP/SML1). This gene family could become a useful marker for the genetic and physical mapping of chromosome 21 and for a better definition of the region involved in the clinical phenotype of several genetic diseases.
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the production and characterization of a new monoclonal antibody to the Trefoil Peptide human spasmolytic polyPeptide
Histochemical Journal, 1994Co-Authors: G Elia, Rebecca Chinery, Richard Poulsom, Andrew M. Hanby, N A WrightAbstract:Human spasmolytic polyPeptide (hSP) is a member of the growing family of Trefoil Peptides which are expressed in discrete regions of the body, most notably the gastrointestinal tract. Much of the research into the localization of the spasmolytic polyPeptide has relied on hybridization in situ to detect its mRNA, due to the absence of a suitable antibody. The aim of the present study was to develop and characterize a monoclonal antibody against the human spasmolytic polyPeptide, using a combination of immunohistochemistry and hybridization in situ. After immunoblotting, the antibody detected a 14 kDa protein in gastrointestinal tissue extracts from the stomach and small intestine only. Using immunohistochemistry, human spasmolytic polyPeptide showed a distinctive staining pattern in the duodenum which co-localized with its mRNA. The co-localization of the immunoreactive Peptide with its mRNA provides good evidence that the antibody truly recognized human spasmolytic polyPeptide.
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expression and purification of a Trefoil Peptide motif in a β galactosidase fusion protein and its use to search for Trefoil binding sites
FEBS Journal, 1993Co-Authors: Rebecca Chinery, Richard Poulsom, Andrew M. Hanby, G Elia, N A WrightAbstract:The cysteine-rich Trefoil motif of rat intestinal Trefoil factor (rITF) was cloned and expressed in Escherichia coli. A 270-bp cDNA fragment including the signal sequence and the Trefoil motif was cloned into the expression vector pAX5+ to direct the expression of a β-galactosidase collagenhinged fusion protein in E. coli. Cultures harbouring the recombinant plasmid produced a soluble novel protein with a molecular mass of 134.5 kDa, as predicted for the Trefoil-motif-containing fusion protein. Purification of the rITF moiety was achieved by p-aminophenyl-thio-β-d-galactoside(APTG)-affinity chromatography, collagenase digestion of the hybrid molecule, and removal of the β-galactosidase-hinge molecule by a further APTG-affinity step. It was demonstrated that intrachain disulphide-bond formation in rITF occurred during the procedure, so no refolding steps were required. Analysis by immunoblotting revealed that the fusion protein and the cleaved Trefoil-motif-containing protein were recognised by an antibody raised against the chemically synthesised Peptide. The Trefoil motif present in the fusion protein was used to localise putative Trefoil-binding sites in sections of frozen rat tissue. Binding was demonstrated using the β-galactosidase portion of the fusion protein as a reporter moiety, either directly with 5-bromo-4-chloro-3-indolyl-β-d-galactoside, or indirectly using a monoclonal antibody to β-galactosidase and indirect immunohistochemistry. Binding sites were localised to the foveolar and surface epithelium of rat stomach, the collecting ducts of the kidney and within colonic crypts. The presence of a Trefoil motif was necessary for binding. The use of β-galactosidase fusion proteins for histochemical localisation of Peptide-binding sites should prove more generally useful.
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Trefoil Peptide gene expression in small intestinal crohn s disease and dietary adaptation
Journal of Clinical Gastroenterology, 1993Co-Authors: Richard Poulsom, Rebecca Chinery, C Sarraf, Elnasir Lalani, G Elia, Susan Van Noorden, Gordon Stamp, N A WrightAbstract:We examined the patterns of Trefoil Peptide gene expression in the ulcer-associated cell lineage (UACL) and mucosa adjacent to Crohn's disease in humans and during gastrointestinal adaptation to enteral feeding in rats. In the UACL, human spasmolytic polyPeptide (hSP) mRNA and Peptide are present in the acinar and proximal duct cells, whereas pS2 mRNA and Peptide are found in the distal duct cells and in the surface cells. In mucosa adjacent to UACL, pS2 mRNA and Peptide are expressed ectopically by goblet cells and neuroendocrine cells. Intestinal crypts associated with the UACL showed marked neuroendocrine cell hyperplasia. Ultrastructural immunolocalization showed pS2 to be copackaged in the mucous cell and neuroendocrine granules. The copackaging of a secretory protein in both mucous and neuroendocrine granules, which have different functions, is unusual and indicates an important role for pS2 in the secretory process itself or as a ligand delivered to its receptor via multiple routes. We also cloned the newest Trefoil Peptide, intestinal Trefoil factor (ITF), from human and rat intestinal mucosa. Using in situ hybridization we demonstrated its synthesis by normal rat intestinal goblet cells. RNAse protection analysis revealed that the level of mRNA for rat ITF in small and large intestine was affected by the process of enteral feeding. We conclude that Trefoil Peptides are widely distributed in the intestine in human inflammatory bowel disease and are of considerable potential functional importance.
