Five years of complete remission of gastric diffuse large B cell lymphoma after eradication of Helicobacter pylori infection.

Long term follow up data are not available for cases of diffuse large B cell gastric lymphoma treated by eradicating Helicobacter pylori alone. We present the case of an 82 year old man with diffuse large B cell lymphoma localised to the stomach which responded to H pylori eradication and which has not recurred after more than five years of close follow up. Our patient was not a candidate for other modalities of treatment. This case demonstrates that the option of treating H pylori infection as the initial trial of treatment for localised diffuse large B cell lymphoma is appropriate for consideration. If medical therapy using eradication of H pylori is used, it is essential that the patient undergoes close observation and repeated surveillance endoscopies.

Interleukin-2 receptor beta subunit-dependent and -independent regulation of intestinal epithelial tight junctions.

Interleukin (IL)-15 is able to regulate tight junction formation in intestinal epithelial cells. However, the mechanisms that regulate the intestinal barrier function in response to IL-15 and the involved subunits of the IL-15 ligand-receptor system are unknown. We determined the IL-2Rbeta subunit and IL-15-dependent regulation of tight junction-associated proteins in the human intestinal epithelial cell line T-84. The IL-2Rbeta subunit was expressed and induced signal transduction in caveolin enriched rafts in intestinal epithelial cells. IL-15-mediated tightening of intestinal epithelial monolayers correlated with the enhanced recruitment of tight junction proteins into Triton X-100-insoluble protein fractions. IL-15-mediated up-regulation of ZO-1 and ZO-2 expression was independent of the IL-2Rbeta subunit, whereas the phosphorylation of occludin and enhanced membrane association of claudin-1 and claudin-2 by IL-15 required the presence of the IL-2Rbeta subunit. Recruitment of claudins and hyperphosphorylated occludin into tight junctions resulted in a more marked induction of tight junction formation in intestinal epithelial cells than the up-regulation of ZO-1 and ZO-2 by itself. The regulation of the intestinal epithelial barrier function by IL-15 involves IL-2Rbeta-dependent and -independent signaling pathways leading to the recruitment of claudins, hyperphosphorylated occludin, ZO-1, and ZO-2 into the tight junctional protein complex.

Angioedema of the small bowel due to an angiotensin-converting enzyme inhibitor.

We describe a case of a 72-year-old woman who presented with two episodes of abdominal pain, vomiting, and diarrhea. Abdominal computed tomographic scans done during each episode demonstrated edema of the small bowel. Review of the patient's history revealed that she had been started on a treatment of lisinopril for hypertension 1 month before the first episode and had her prescribed dose increased 24 hours before each presentation. Angiotensin-converting enzyme (ACE) inhibitor-associated angioedema was suspected and the medication was discontinued. The patient has remained symptom-free while not taking the ACE inhibitor for 1 year. Review of the literature reveals only nine similar cases. All cases, including ours, occurred in women. Angioedema of the small bowel associated with ACE inhibitors is rare and often is not recognized before surgical exploration. Angioedema of the gastrointestinal tract should be considered in symptomatic patients taking ACE inhibitors.

Mice with a selective deletion of the CC chemokine receptors 5 or 2 are protected from dextran sodium sulfate-mediated colitis: lack of CC chemokine receptor 5 expression results in a NK1.1+ lymphocyte-associated Th2-type immune response in the intestine.

The chemokine receptors CCR2 and CCR5 and their respective ligands regulate leukocyte chemotaxis and activation. To determine the role of these chemokine receptors in the regulation of the intestinal immune response, we induced colitis in CCR2- and CCR5-deficient mice by continuous oral administration of dextran sodium sulfate (DSS). Both CCR2- and CCR5-deficient mice were susceptible to DSS-induced intestinal inflammation. The lack of CCR2 or CCR5 did not reduce the DSS-induced migration of macrophages into the colonic lamina propria. However, both CCR5-deficient mice and, to a lesser degree, CCR2-deficient mice were protected from DSS-induced intestinal adhesions and mucosal ulcerations. CCR5-deficient mice were characterized by a greater relative infiltration of CD4+ and NK1.1+ lymphocyte in the colonic lamina propria when compared to wild-type and CCR2-deficient mice. In CCR5-deficient mice, mucosal mRNA expression of IL-4, IL-5, and IL-10 was increased, whereas that of IFN-gamma was decreased, corresponding to a Th2 pattern of T cell activation. In CCR2-deficient mice, the infiltration of Th2-type T cells in the lamina propria was absent, but increased levels of IL-10 and decreased levels of IFN-gamma may have down regulated mucosal inflammation. Our data indicate that CCR5 may be critical for the promotion of intestinal Th1-type immune responses in mice.

Fractalkine is an epithelial and endothelial cell-derived chemoattractant for intraepithelial lymphocytes in the small intestinal mucosa.

Fractalkine is a unique chemokine that combines properties of both chemoattractants and adhesion molecules. Fractalkine mRNA expression has been observed in the intestine. However, the role of fractalkine in the healthy intestine and during inflammatory mucosal responses is not known. Studies were undertaken to determine the expression and function of fractalkine and the fractalkine receptor CX3CR1 in the human small intestinal mucosa. We identified intestinal epithelial cells as a novel source of fractalkine. The basal expression of fractalkine mRNA and protein in the intestinal epithelial cell line T-84 was under the control of the inflammatory mediator IL-1beta. Fractalkine was shed from intestinal epithelial cell surface upon stimulation with IL-1beta. Fractalkine localized with caveolin-1 in detergent-insoluble glycolipid-enriched membrane microdomains in T-84 cells. Cellular distribution of fractalkine was regulated during polarization of T-84 cells. A subpopulation of isolated human intestinal intraepithelial lymphocytes expressed the fractalkine receptor CX3CR1 and migrated specifically along fractalkine gradients after activation with IL-2. Immunohistochemistry demonstrated fractalkine expression in intestinal epithelial cells and endothelial cells in normal small intestine and in active Crohn's disease mucosa. Furthermore, fractalkine mRNA expression was significantly up-regulated in the intestine during active Crohn's disease. This study demonstrates that fractalkine-CX3CR1-mediated mechanism may direct lymphocyte chemoattraction and adhesion within the healthy and diseased human small intestinal mucosa.

Butyrate switches the pattern of chemokine secretion by intestinal epithelial cells through histone acetylation.

BACKGROUND: Butyrate, a fermentation product of intestinal bacteria, modifies chromatin structure through histone acetylation, thereby altering gene transcription. IL-8 and MCP-1 are chemokines, expressed by intestinal epithelial cells, which attract neutrophils and monocytes, respectively. We hypothesized that butyrate may alter IL-8 and MCP-1 expression by intestinal epithelial cells through histone acetylation. MATERIALS AND METHODS: IL-8 and MCP-1 expression was measured by ELISA and RNA transfer blots. Acetylated histones were separated on acetic acid-urea-triton gels. Butyrate was compared to Trichostatin-A, a specific inhibitor of histone deacetylase and to other short chain fatty acids. RESULTS: Caco-2 cells constitutively secreted MCP-1 but not IL-8. Butyrate reversibly decreased MCP-1 secretion. In contrast, butyrate increased IL-8 production. The effects of butyrate and Trichostatin-A were greater when cells were stimulated with IL-1beta. Butyrate and Trichostatin-A both increased histone acetylation. Trichostatin-A and other short chain fatty acids altered chemokine secretion according to their effect on histone acetylation. CONCLUSIONS: Butyrate reversibly switches chemokine secretion by epithelial cells through histone acetylation. We speculate that butyrate carries information from resident bacteria to epithelial cells. Epithelial cells transduce this signal through histone acetylation, modulating the secretion of chemokines.

Chemokines in the inflammatory bowel diseases.

Ulcerative colitis and Crohn's disease are characterized by chronic intestinal inflammation. Intestinal bacteria initiate the activation of intestinal inflammatory processes, which are mediated by proinflammatory cytokines and chemokines. In inflammatory bowel disease, intestinal inflammation is not downregulated, in part due to defective or absent inhibitory processes. Studies to date have demonstrated that IL-8, MCP-1, and ENA-78 are highly expressed in the intestinal mucosa in areas of active Crohn's disease and ulcerative colitis. Neutrophils and macrophages in the inflamed intestine synthesize and secrete large amounts of chemokines in patients with inflammatory bowel disease. Increased chemokine expression has also been observed in epithelial cells, endothelial cells, and smooth muscle cells. Future trials of specific agents capable of inhibiting chemokine synthesis and secretion or blocking chemokine-chemokine receptor interaction will be important to study in patients with ulcerative colitis and Crohn's disease.

Increased interleukin-8 (IL-8) in rectal dialysate from patients with ulcerative colitis: evidence for a biological role for IL-8 in inflammation of the colon.

OBJECTIVE: Infiltration of neutrophils and their release of toxic reactive oxygen species (ROS) in the colonic mucosa are associated with tissue damage in ulcerative colitis (UC). This neutrophil migration may be induced by chemoattractants, such as cytokines, in the colonic milieu. One such chemoattractant is interleukin-8 (IL-8), a neutrophil chemokine that is present at high concentrations in inflamed mucosa. However, the functional significance of IL-8 in neutrophil attraction and activation in UC has not been established. We hypothesized that IL-8 in the colonic lumen of patients with UC primes neutrophils, leading to their attraction and activation. METHODS: The colonic milieu was sampled by rectal dialysis. Using a semi-permeable membrane with a molecular weight cut-off of 12 kDa, dialysis solution was placed in the rectum and allowed to equilibrate over a 4-h period with the colonic milieu of controls or of patients with UC. IL-8 concentrations were measured by ELISA. Two functions of healthy neutrophils (PMN) were measured: expression of CD11-b surface adhesion molecules (by flow cytometry), and production of ROS (by both chemiluminescence and cytochrome C reduction assays). Neutrophil functions after exposure to rectal dialysates or n-formyl-methionyl-leucyl-phenylalanine (fMLP) were assessed before and after adding anti-IL-8 antibody or the fMLP blocker BMLP. RESULTS: IL-8 concentrations in dialysates from patients with active UC were significantly higher than in controls and correlated with disease activity. UC dialysates significantly increased ROS production and CD11-b expression by neutrophils and anti-IL-8 antibody partially (50%) inhibited these stimulatory effects of UC dialysates. Preincubation of neutrophils with UC dialysates significantly potentiated the fMLP-induced rise in ROS and anti-IL-8 antibody completely abolished this priming effect. CONCLUSIONS: The colonic milieu, sampled by rectal dialysis, from patients with active UC can both activate and prime neutrophils in vitro. High concentrations of IL-8 in the colonic lumen of UC patients are partially responsible for the activating effects of rectal dialysates, and account for all of its priming effects. These findings provide direct evidence for a role for IL-8 in inflammatory bowel disease.

Short-chain fatty acids regulate IGF-binding protein secretion by intestinal epithelial cells.

Gastrointestinal epithelial cells secrete insulin-like growth factor (IGF)-binding proteins (IGFBPs), which modulate the actions of IGFs on cell proliferation and differentiation. Short-chain fatty acids are bacterial metabolites from unabsorbed carbohydrate (including fiber). We hypothesized that they may alter the pattern of IGFBPs secreted by epithelial cells as part of a wider phenomenon by which luminal molecules regulate gastrointestinal epithelial cell signaling. The intestinal epithelial cell line, Caco-2, predominantly secretes IGFBP-3; however, butyrate increased the secretion of IGFBP-2 in a dose-dependent and reversible manner. Butyrate decreased the secretion of IGFBP-3. Butyrate altered only the synthesis and not the cell sorting of IGFBPs because 1) the secretion of IGFBPs remained polarized despite changes in their rates of production, and 2) IGFBP secretion corresponded to mRNA accumulation. The ability of short-chain fatty acids or the fungicide trichostatin A to stimulate IGFBP-2 correlated with their actions on histone acetylation. In conclusion, intestinal epithelial cells respond to short-chain fatty acids by altering secretion of IGFBPs.

The central role of chemokines (chemotactic cytokines) in the immunopathogenesis of ulcerative colitis and Crohn's disease.

The final composition of leukocytes present in a site of inflammation in response to chemokine stimulation and activation may depend on both the nature of the secreted chemokines as well as the relative expression of the multitude of specific chemokine cell surface receptors on many different cell types. Because related receptors with different affinities and cross-reactive binding capabilities are present on each type of leukocyte, relative differences in receptor distribution and receptor affinity for specific chemokines may significantly influence which cells are ultimately attracted to and activated by each individual chemokine. Production of IL-8, MCP-1, and ENA-78 by endothelial cells, LPMNC, and epithelial cells in IBD could establish a chemotactic gradient capable of influencing the increased migration of monocytes/macrophages, granulocytes, and lymphocytes from the blood stream through the endothelium into both the mucosa and submucosa during chronic IBD. The ability of chemokines to induce chemotaxis, leukocyte activation, granule exocytosis, increased production of metalloenzymes, and up-regulation of respiratory burst activity indicates that there may be a variety of different mechanisms by which chemokines could markedly increase chronic inflammation and chronic intestinal tissue destruction in IBD.

Pouchitis associated with primary cytomegalovirus infection.

We report a patient with a history of ulcerative colitis status after total proctocolectomy with an ileoanal J pouch who presented with marked, refractory pouchitis associated with a primary cytomegalovirus (CMV) infection. The patient had atypical lymphocytosis in the blood and serology consistent with primary CMV infection. Biopsies of the pouch revealed CMV inclusion bodies and yielded positive CMV cultures. The patient improved clinically with resolution of pouchitis after a 10-day course of therapy with gancyclovir and has remained in remission for over 5 yr. This is the first report of pouchitis associated with a primary CMV infection. This case demonstrates that CMV infection is in the differential diagnosis for causes of pouchitis, and it suggests that the pouch, like the colon, is a potential site for a primary CMV infection in an immunocompetent host.

Butyrate enhances interleukin (IL)-8 secretion by intestinal epithelial cells in response to IL-1beta and lipopolysaccharide.

Intestinal epithelial (Caco-2) cells secrete the chemokine, IL-8, after stimulation with IL-1beta, but not after lipopolysaccharide. Butyrate is a short chain fatty acid derived from the metabolism of intestinal contents by gut bacteria. Butyrate concentrations reflect, therefore, the bacterial microenvironment established within the intestine. We hypothesized that butyrate may alter the secretion of IL-8 by intestinal epithelial cells in response to stimulation by IL-1beta or lipopolysaccharide. Caco-2 cells were incubated in varying concentrations of sodium butyrate (0-20 mM) for 24 h before stimulation with lipopolysaccharide or IL-1beta. IL-8 secretion was measured over 24 h by ELISA. IL-8 mRNA accumulation was detected by Northern blots. Lipopolysaccharide induced the secretion of IL-8 only after Caco-2 cells cells had been cultured with sodium butyrate. Furthermore, butyrate significantly enhanced IL-8 secretion by cells stimulated with IL-1beta. Butyrate also increased IL-8 mRNA accumulation in stimulated Caco-2 cells. Intestinal epithelial cells can, therefore, be primed by butyrate to become activated by lipopolysaccharide and proinflammatory cytokines. This may represent a mechanism by which intestinal epithelial cells can regulate intestinal inflammation in response to changes in the intestinal milieu.

Macrophage inflammatory protein-2: chromosomal regulation in rat small intestinal epithelial cells.

Nonpathogenic, resident bacteria participate in the pathogenesis of inflammation in the small intestine, but the molecular messages produced by such bacteria are unknown. Inflammatory responses involve the recruitment of specific leukocyte subsets. We, therefore, hypothesized that butyrate, a normal bacterial metabolite, may modulate chemokine secretion by epithelial cells, by amplifying their response to proinflammatory signals. We studied the expression of the chemokine, macrophage inflammatory protein-2 (MIP-2) by the rat small intestinal epithelial cell line, IEC-6. Cells were stimulated with lipopolysaccharide or with interleukin 1beta (IL-1beta) and incubated with sodium butyrate. Acetylation of histones was examined in Triton X acetic acid-urea gels by PAGE. Unstimulated IEC-6 cells did not secrete MIP-2. However, lipopolysaccharide and IL-1beta induced MIP-2 expression. Butyrate enhanced MIP-2 secretion both in lipopolysaccharide-stimulated and IL-1beta-stimulated enterocytes; but butyrate alone did not induce MIP-2 expression. Butyrate increased the acetylation of histones extracted from the nuclei of IEC-6 cells. Furthermore, acetylation of histones (induced by trichostatin A, a specific inhibitor of histone deacetylase) enhanced MIP-2 expression by cells stimulated with IL-1beta. In conclusion, trichostatin A reproduced the effects of butyrate on MIP-2 secretion. Butyrate, therefore, increases MIP-2 secretion in stimulated cells by increasing histone acetylation. We speculate that butyrate carries information from bacteria to epithelial cells. Epithelial cells transduce this signal through histone deacetylase, modulating the secretion of chemokines.

Intestinal epithelial cells both express and respond to interleukin 15.

BACKGROUND & AIMS: Interleukin (IL)-15 exerts functional effects on lymphocytes similar to those of IL-2. IL-15 is expressed by nonlymphoid cells and may integrate these cells into classical immune responses. The aim of this study was to characterize the expression of IL-15 by intestinal epithelial cells and determine the functional roles of IL-15 within the mucosal immune system. METHODS: Rat IL-15 was cloned from a rat jejunal library. Expression of IL-15 in rat and human intestinal epithelial cells was assessed by Northern and Western blotting. Tyrosine kinase activation in response to IL-15 in intestinal epithelial cells was determined by immunoprecipitation. RESULTS: Rat and human intestinal epithelial cells express IL-15 messenger RNA. IL-15 activates Stat3 and stimulates the proliferation of intestinal epithelial cells. The relevance of the observations for intestinal epithelial cell function in vivo was supported by the demonstration of transcripts for IL-15 in primary human intestinal epithelial cells. CONCLUSIONS: IL-15 is expressed by intestinal epithelial cells function. These experiments suggest that IL-15 is an important mediator that could integrate intestinal epithelial cell function with the intestinal immune system.

Alterations of the mucosal immune system in inflammatory bowel disease.

The normal intestinal immune system is under a balance in which proinflammatory and anti-inflammatory cells and molecules are carefully regulated to promote a normal host mucosal defense capability without destruction of intestinal tissue. Once this careful regulatory balance is disturbed, nonspecific stimulation and activation can lead to increased amounts of potent destructive immunologica and inflammatory molecules being produced and released. The concept of balance and regulation of normal mucosal immune and inflammatory events is indicative of how close the intestine is to developing severe inflammation. The normal intestinal mucosal immune system is constantly stimulated by lumenal contents and bacteria. The stimulatory molecules present in the intestinal lumen that activate and induce subsequent mucosal immunologic and inflammatory events include bacterial cell wall products, such as peptidoglycans and lipopolysaccharides, as well as other chemotactic and toxic bacterial products that are produced by the many different types of bacteria within the gastrointestinal tract. These highly stimulatory bacterial cell wall products are capable of activating macrophages and T lymphocytes to release potent proinflammatory cytokines, including interleukin-1 (IL-1), interleukin-6 (IL-6), and tumor necrosis factor alpha (TNF-alpha). IL-1, IL-6, and TNF-alpha increase the presence of human leukocyte antigen (HLA) class II antigen-presenting molecules on the surfaces of epithelial cells, endothelial cells, macrophages, and B cells, thus increasing their ability to present lumenal antigens and bacterial products. The proinflammatory cytokines IL-1 and TNF-alpha also increase the ability of epithelial cells, endothelial cells, macrophages, and fibroblasts to secrete potent chemotactic cytokines, such as interleukin-8 (IL-8) and monocyte chemoattractant protein-1 (MCP-1), which serve to increase the movement of macrophages and granulocytes from the circulation into the inflamed mucosa. Thus, through lumenal exposure to potent, nonspecific stimulatory bacterial products, the state of activation of the intestinal immune system and mucosal inflammatory pathways are markedly up-regulated. This raises the question of whether there is a deficiency in effective down-regulation through the absence of normally suppressive cytokines such as interleukin-10 (IL-10), transforming growth factor-beta (TGF-beta), interleukin-4 (IL-4), and IL-1 receptor antagonist. Normally, the turning off of the active and destructive immunologic and inflammatory events should occur following the resolution of a bacterial or viral infection that has been appropriately defended against and controlled by the mucosal immune system. In inflammatory bowel disease (IBD), however, the down-regulatory events and processes that should turn off the immunologic and inflammatory protective processes, once the pathogenic agent has been cleared, appear to be deficient or only partially effective. We may find that we ultimately are dealing with disease processes that have more than one genetic or cellular basis. The improved understanding of the immunopathophysiology of IBD will allow exploration of novel immunologic and genetic approaches, such as gene replacement therapy, administration of a suppressor cytokine or an altered cell surface antigen, the administration of humanized monoclonal antibodies directed against proinflammatory cytokines, or the development of newer strategies against fundamental cell biologic mechanisms such as adhesion molecules.

Perinuclear anti-neutrophil cytoplasmic antibodies are spontaneously produced by mucosal B cells of ulcerative colitis patients.

Approximately 60% of sera from ulcerative colitis (UC) patients contains Igs reactive with neutrophil components, raising the question of the origin of these anti-neutrophil cytoplasmic Abs (ANCA). Our assertion that ANCA is a marker for a mucosal disease-related immune response predicts the existence of ANCA producing B cell clones in the lamina propria lymphocyte (LPL) fraction of UC patients. This hypothesis was tested by examining 12-day culture supernatants of LPL ANCA expression. LPL were isolated from surgically removed mucosa from patients with UC, Crohn's disease (CD), and diverticulitis. Normal mucosa was obtained from accident victims or normal margins of colon cancer resections. Supernatants were assayed by a fixed neutrophil ELISA. The ANCA staining pattern of supernatants expressing ANCA, as determined by ELISA, was assessed by indirect immunofluorescent staining of alcohol-fixed neutrophils. ANCA was found in 70% of culture supernatants from UC LPL fractions. In contrast, only approximately 11% of supernatants from CD and diverticulitis/normal (noninflammatory bowel disease (IBD)) LPL displayed ANCA binding. A perinuclear (pANCA) staining pattern was obtained with 70% of ANCA-expressing UC LPL supernatants, whereas ANCA-expressing CD and non-IBD LPL supernatants displayed a cytoplasmic reaction. PBL and mesenteric lymph node lymphocytes lacked spontaneous pANCA production, and pANCA production from PBL was not inducible. These findings indicate the existence of pANCA-producing B cell clones in mucosal lesions of UC patients and support our hypothesis that pANCA production is a consequence of a mucosal immune response specific to UC.

Monocyte-chemoattractant protein 1 gene expression in intestinal epithelial cells and inflammatory bowel disease mucosa.

BACKGROUND: Monocyte-chemoattractant protein 1 (MCP-1) activates macrophages and increases the migration of monocytes into tissue during inflammation. It was hypothesized that MCP-1 expression is involved in intestinal inflammation. METHODS: MCP-1 protein was detected by immunohistochemistry and immunoprecipitation. Biological activity of MCP-1 was assessed using a chemotactic assay. MCP-1 messenger RNA (mRNA) levels were measured by quantitative reverse-transcription polymerase chain reaction. RESULTS: In normal mucosa, MCP-1 was predominantly present in surface epithelium. In contrast, inflamed mucosa from patients with ulcerative colitis or Crohn's disease contained multiple cells immunoreactive for MCP-1, including spindle cells, mononuclear cells, and endothelial cells. Furthermore, MCP-1 mRNA expression was markedly increased in inflamed intestinal biopsy specimens from patients with inflammatory bowel disease. MCP-1 was detected in isolated intestinal epithelial cells and in conditioned media from Caco-2 cells. Caco-2 cell-conditioned media stimulated monocyte chemotaxis activity that was inhibited by anti-MCP-1 antibodies. Constituitive MCP-1 mRNA levels in Caco-2 cells were up-regulated by interleukin 1 beta and down-regulated by dexamethasone. CONCLUSIONS: In addition to lamina propria macrophages, endothelial cells, and spindle cells, intestinal epithelial cells are able to produce MCP-1. MCP-1 is expressed constitutively in the intestinal colonic mucosa and is up-regulated during inflammation.