Biomedical subjects

Francesco Pallone

Publications and source records attributed to Francesco Pallone.

At least 19 recordsLinked to original sources

PubMed2007

IL-21 counteracts the regulatory T cell-mediated suppression of human CD4+ T lymphocytes.

High expression of IL-21 and/or IL-21R has been described in T cell-mediated inflammatory diseases characterized by defects of counterregulatory mechanisms. CD4(+)CD25(+) regulatory T cells (Treg) are a T cell subset involved in the control of the immune responses. A diminished ability of these cells to inhibit T cell activation has been documented in immune-inflammatory diseases, raising the possibility that inflammatory stimuli can block the regulatory properties of Treg. We therefore examined whether IL-21 controls CD4(+)CD25(+) T cell function. We demonstrate in this study that IL-21 markedly enhances the proliferation of human CD4(+)CD25(-) T cells and counteracts the suppressive activities of CD4(+)CD25(+) T cells on CD4(+)CD25(-) T cells without affecting the percentage of Foxp3(+) cells or survival of Treg. Additionally, CD4(+)CD25(+) T cells induced in the presence of IL-21 maintain the ability to suppress alloresponses. Notably, IL-21 enhances the growth of CD8(+)CD25(-) T cells but does not revert the CD4(+)CD25(+) T cell-mediated suppression of this cell type, indicating that IL-21 makes CD4(+) T cells resistant to suppression rather than inhibiting CD4(+)CD25(+) T cell activity. Finally, we show that IL-2, IL-7, and IL-15, but not IL-21, reverse the anergic phenotype of CD4(+)CD25(+) T cells. Data indicate that IL-21 renders human CD4(+)CD25(-) T cells resistant to Treg-mediated suppression and suggest a novel mechanism by which IL-21 could augment T cell-activated responses in human immune-inflammatory diseases.

CD4-Positive T-Lymphocytes↗

PubMed2007

Interleukin-12 drives the Th1 signaling pathway in Helicobacter pylori-infected human gastric mucosa.

In this study we examined mechanisms that regulate T-helper lymphocyte 1 (Th1) commitment in Helicobacter pylori-infected human gastric mucosa. The levels of gamma interferon (IFN-gamma), interleukin-4 (IL-4), and IL-12 in total extracts of gastric biopsies taken from H. pylori-infected and uninfected patients were determined by an enzyme-linked immunosorbent assay. The levels of signal transducer and activator of transcription 4 (STAT4), STAT6, and T-box expressed in T cells (T-bet) in total proteins extracted from gastric biopsies were determined by Western blotting. Finally, the effect of a neutralizing IL-12 antibody on expression of Th1 transcription factors and the levels of IFN-gamma in organ cultures of H. pylori-infected biopsies was examined. Increased levels of IFN-gamma and IL-12 were found in gastric biopsy samples of H. pylori-infected patients compared to the levels in uninfected patients. In addition, H. pylori-infected biopsies exhibited high levels of expression of phosphorylated STAT4 and T-bet. Higher levels of IFN-gamma and expression of Th1 transcription factors were associated with greater infiltration of mononuclear cells in the gastric mucosa. By contrast, production of IL-4 and expression of phosphorylated STAT6 were not associated with the intensity of mononuclear cell infiltration. In ex vivo organ cultures of H. pylori-infected biopsies, neutralization of endogenous IL-12 down-regulated the expression of phosphorylated STAT4 and T-bet and reduced IFN-gamma production. Our data indicated that IL-12 contributes to the Th1 cell commitment in H. pylori-infected human gastric mucosa.

Adult↗

PubMed2006

Interleukin-12 and Th1 immune response in Crohn's disease: pathogenetic relevance and therapeutic implication.

Crohn's disease (CD) and ulcerative colitis (UC) are chronic inflammatory disorders of the gastrointestinal tract that share clinical and pathological characteristics. The most accredited hypothesis is that both CD and UC result from a deregulated mucosal immune response to normal constituents of the gut microflora. Evidence, however, indicates that the main pathological processes in these two diseases are distinct. In CD, the tissue-damaging inflammatory reaction is driven by activated type 1 helper T-cell (Th1), whereas a humoral response predominates in UC. Consistently, a marked accumulation of macrophages making interleukin (IL)-12, the major Th1-inducing factor, is seen in CD but not in UC mucosa. Preliminary studies also indicate that administration of a monoclonal antibody blocking the IL-12/p40 subunit can be useful to induce and maintain clinical remission in CD patients. Notably, the recently described IL-23 shares the p40 subunit with IL-12, raising the possibility that the clinical benefit of the anti-IL-12/p40 antibody in CD may also be due to the neutralization of IL-23 activity. This review summarizes the current information on the expression and functional role of IL-12 and IL-12-associated signaling pathways both in patients with CD and experimental models of colitis, thus emphasizing major differences between IL-12 and IL-23 activity on the development of intestinal inflammation.

Crohn Disease↗

PubMed2006

Inhibition of Smad7 with a specific antisense oligonucleotide facilitates TGF-beta1-mediated suppression of colitis.

BACKGROUND & AIMS: Defective transforming growth factor (TGF)-beta1 signaling due to high levels of Smad7 is a feature of inflammatory bowel disease (IBD). In this study, we analyzed the effect of reducing Smad7 levels with antisense oligonucleotide on mouse models of colitis. METHODS: Mucosal samples taken from colitic tissue of mice with colitis due to either haptenating reagents (trinitrobenzene sulfonic acid [TNBS] or oxazolone) or to transfer of T cells (SCID transfer colitis) were analyzed for Smad3 and/or Smad7 expression by Western blotting and, in some cases, content of TGF-beta1 by enzyme-linked immunosorbent assay. The effect of oral Smad7 antisense oligonucleotide on mucosal inflammation was assessed. RESULTS: TGF-beta1 levels were increased in the inflamed tissues of mice with colitis induced by either TNBS or oxazolone. Nevertheless, TGF-beta1 did not exert a regulatory effect, probably because TGF-beta1 signaling was blocked, as indicated by the presence of reduced Smad3 phosphorylation and high levels of Smad7. Oral administration of Smad7 antisense oligonucleotide to colitic mice restored TGF-beta1 signaling via Smad3 and ameliorated inflammation in hapten-induced colitis. In addition, Smad7 antisense oligonucleotide had a therapeutic effect on relapsing TNBS-induced colitis but not on cell-transfer colitis. CONCLUSIONS: These data suggest that colitis models associated with high endogenous TGF-beta1 levels and defective TGF-beta1 signaling due to high levels of Smad7 can be ameliorated by down-regulation of Smad7 and by oral administration of Smad7 antisense oligonucleotide. This may represent a new approach to the control of IBD, particularly during active phases when its Smad7 profile resembles that of hapten-induced colitis.

Animals↗

PubMed2006

5-aminosalicylic acid enhances anchorage-independent colorectal cancer cell death.

Resistance to anoikis, the cell death triggered by the loss of anchorage to the substratum, is an essential prerequisite in the proliferation and diffusion of colorectal cancer (CRC) cells. We examined whether 5-aminosalicylic acid (5-ASA), a drug that seems to reduce the risk of colitis-associated CRC, enhances CRC cell anoikis. To this end, Colo205 cells were treated with 5-ASA in the presence or absence of inhibitors of caspases (zVAD-fmk) and reactive oxygen species (ROS). We demonstrate that 5-ASA enhances Colo205 cell death. Although 5-ASA induces dissipation of mitochondrial transmembrane potential and caspase-3 activation, zVAD-fmk does not completely prevent the 5-ASA-induced cell death. 5-ASA also enhances the synthesis of ROS. However, inhibitors of ROS reduce the fraction of 5-ASA-induced Colo205 cell death but do not confer protection. In contrast, the 5-ASA-mediated Colo205 cell death is preventable by Bcl-2 over-expression. These data suggest a mechanism by which 5-ASA interferes with colon carcinogenesis.

Anoikis↗

PubMed2006

Local injection of infliximab in the postoperative recurrence of Crohn's disease.

BACKGROUND: Local injection of infliximab in Crohn's disease (CD) lesions may reduce the risk of rare side effects, reduce the dose, and increase the efficacy of the drug. The objective was to prospectively assess the feasibility and the safety of local injection of infliximab for the postoperative recurrence of patients with CD who were followed for at least 1 year. METHODS: In a pilot, open-label study, 8 patients with CD (3 men; median age 48 years, range 35-82 years) undergoing ileocolonoscopy were prospectively enrolled. Inclusion criteria included the following: (1) localized (<5 cm) recurrence, (2) inflammatory pattern, and (3) clinically inactive CD. At the first endoscopy, lesions were injected with infliximab (median, 30 mg; range, 8-60 mg); a control endoscopy was performed at 2 weeks in 4 patients (3 received a second injection followed by a control endoscopy at 6 weeks) and at 4 weeks in 4 patients (2 received a second injection followed by a control endoscopy at 8 weeks). OBSERVATIONS: No patients showed side effects or clinical relapse in the short term and the long term (median follow-up, 20 months; range, 14-21 months). Endoscopic score improved in 3/8 patients. The histologic scores were reduced in 4 patients, worsened in 3, and were unchanged in one patient with CD. CONCLUSIONS: Local injection of infliximab into patients with CD recurrence is feasible and safe, requiring a low dose. Present findings suggest the need of placebo-controlled trials to assess the efficacy of this new and safe procedure in subgroups of patients with CD.

Adult↗

PubMed2006

A functional role of flip in conferring resistance of Crohn's disease lamina propria lymphocytes to FAS-mediated apoptosis.

BACKGROUND & AIMS: There is evidence that, in Crohn's disease (CD), lamina propria T lymphocytes (LPLs) are resistant to FAS-mediated apoptosis and that this defect contributes to the mucosal T-cell accumulation. In this study we examined the functional role of Flip, a Flice inhibitor protein, in the resistance of CD LPL to FAS-mediated apoptosis. METHODS: Biopsy specimens and LPLs were taken from CD and ulcerative colitis (UC) patients and normal controls and analyzed for Flip by Western blotting. We also examined whether inhibition of Flip by antisense oligonucleotide restored the susceptibility of CD LPLs to FAS-induced apoptosis. LPL apoptosis was assessed by flow cytometry. RESULTS: After FAS stimulation, the rate of apoptosis of CD3+ LPLs was higher in normal controls and patients with UC than in patients with CD. Enhanced expression of both long and short Flip isoforms was seen in biopsy specimens and purified CD3+ and CD45RO+ LPLs of CD patients in comparison with UC patients and normal controls. No increase in Flip was documented in untreated celiac disease mucosa, thus suggesting the possibility that induction of Flip in the gut does not simply rely on the ongoing inflammation. Finally, we showed that inhibition of Flip by antisense oligonucleotide reverted the resistance of CD LPLs to FAS-induced apoptosis. CONCLUSIONS: Data suggest a role for Flip in the resistance of CD LPLs to FAS-mediated apoptosis.

Annexin A5↗

PubMed2006

New mediators of immunity and inflammation in inflammatory bowel disease.

PURPOSE OF REVIEW: In both Crohn's disease and ulcerative colitis, the tissue damage results from an inappropriate or exaggerated immune response to antigens of the gut microflora. This review summarizes current knowledge regarding the role of immune-inflammatory mediators in the pathogenesis of inflammatory bowel disease. RECENT FINDINGS: Despite having a common basis in overresponsiveness to luminal antigens, Crohn's disease and ulcerative colitis are immunologically distinct entities. Crohn's disease is associated with a Th1 T cell-mediated response, characterized by enhanced production of interferon-gamma and tumor necrosis factor-alpha. Interleukin (IL)-12 and, possibly, IL-23 govern the Th1 cell differentiation, but optimal induction and stabilization of polarized Th1 cells would require additional cytokines, such as IL-15, IL-18 and IL-21. In ulcerative colitis, the local immune response is less polarized, but it is characterized by CD1-reactive natural killer T cell production of IL-13. Beyond these differences, Crohn's disease and ulcerative colitis share important end-stage effector pathways of intestinal injury, which are mediated by an active cross-talk between immune and non-immune mucosal cells. SUMMARY: The clarification of the complex network of immune-inflammatory mediators operating in the gut of patients with inflammatory bowel disease has led to the identification of new targets that could, in turn, drive the development of effective biological therapies.

Animals↗

PubMed2006

Protease-activated receptor-2 activation in gastric cancer cells promotes epidermal growth factor receptor trans-activation and proliferation.

Dysregulated epidermal growth factor receptor (EGFR) signaling is involved in gastric cancer (GC) cell growth. However, the mechanism that sustains EGFR signaling in GC remains unknown. Since protease-activated receptor-2 (PAR-2), a G protein-coupled receptor, has been shown to trans-activate EGFR in several cell types, we examined the role of PAR-2 in GC. We show here that in vitro activation of PAR-2 enhances the growth of two GC cell lines, AGS and MKN28. In both these cell lines, PAR-2 trans-activated EGFR and inhibition of EGFR tyrosine kinase activity by AG1478 or specific EGFR siRNA completely prevented PAR-2-driven proliferation. Antibody blockade of EGF-like ligands to EGFR did not modify EGFR signaling or cell growth induced by PAR-2 activation. In contrast, PAR-2 promoted Src activation and interaction of this kinase with EGFR. In support of this, inhibition of Src kinase activity by PP1 or siRNA blocked PAR-2-induced EGFR signaling cascade and cell growth. Finally, PAR-2 was detectable in both normal and GC specimens, but its expression was more pronounced in GC than controls and correlated with activated EGFR. These data show that PAR-2 is overexpressed in GC and suggest a role of PAR-2 in EGFR trans-activation and cell growth.

Adenocarcinoma↗

PubMed2005

Interleukin-21 enhances T-helper cell type I signaling and interferon-gamma production in Crohn's disease.

BACKGROUND & AIMS: T-helper (Th)1 cells play a central role in the pathogenesis of tissue damage in Crohn's disease (CD). Interleukin (IL)-12/STAT4 signaling promotes Th1 cell commitment in CD, but other cytokines are needed to maintain activated Th1 cells in the mucosa. In this study, we examined the expression and role of IL-21, a T-cell-derived cytokine of the IL-2 family; in tissues and cells isolated from patients with inflammatory bowel disease. METHODS: IL-21 was examined by Western blotting in whole mucosa and lamina propria mononuclear cells (LPMCs) from patients with CD, ulcerative colitis (UC), and controls. We also examined the effects of exogenous IL-12 on IL-21 production, as well as the effects of blocking IL-21 with an IL-21-receptor Ig fusion protein. Interferon (IFN)-gamma was measured in the culture supernatants by enzyme-linked immunosorbent assay, and phosphorylated STAT4 and T-bet were examined by Western blotting. RESULTS: IL-21 was detected in all samples, but its expression was higher at the site of disease in CD in comparison with UC and controls. Enhanced IL-21 was seen in both ileal and colonic CD and in fibrostenosing and nonfibrostenosing disease. IL-12 enhanced IL-21 in normal lamina propria lymphocytes through an IFN-gamma-independent mechanism, and blocking IL-12 in CD LPMCs decreased anti-CD3-stimulated IL-21 expression. Neutralization of IL-21 in CD LPMC cultures decreased phosphorylated STAT4 and T-bet expression, thereby inhibiting IFN-gamma production. CONCLUSIONS: Our data suggest that IL-21 contributes to the ongoing Th1 mucosal response in CD.

Blotting, Western↗

PubMed2005

Post-transcriptional regulation of Smad7 in the gut of patients with inflammatory bowel disease.

BACKGROUND & AIMS: Transforming growth factor (TGF)-beta1 is one of the most powerful endogenous negative regulators of inflammation. In patients with inflammatory bowel disease, despite abundant local TGF-beta1, there is a failure of TGF-beta-mediated negative regulation of nuclear factor kappaB activation and proinflammatory cytokine production because of increased intracellular expression of the endogenous inhibitor of TGF-beta1 signaling, Smad7. In this study, we examined the molecular mechanism underlying the induction of Smad7 in the human gut. METHODS: Whole intestinal mucosal and lamina propria mononuclear cell samples were analyzed for Smad7 by real-time polymerase chain reaction and Western blotting. Smad7 ubiquitination and acetylation, and interaction of Smad7 with the intrinsic histone acetyltransferase, p300, were examined by immunoprecipitation and Western blotting. The effect of p300 silencing on Smad7 expression was determined in Crohn's disease lamina propria mononuclear cells. RESULTS: We showed that Smad7 is not transcriptionally regulated in human gut but that its increase in patients with inflammatory bowel disease is due to posttranscriptional acetylation and stabilization by p300, which prevents Smad7 ubiquitination and degradation in the proteasome. Hence, Smad7 protein in cells from normal gut is ubiquitinated and rapidly degraded. In contrast, in inflamed gut, Smad7 is acetylated and not ubiquitinated, is not degraded, and can be decreased by short interfering RNA to p300. CONCLUSIONS: These results identify posttranslational protein modification as of importance in chronic gut inflammation in humans.

Acetylation↗

PubMed2004

Cutting edge: TGF-beta induces a regulatory phenotype in CD4+CD25- T cells through Foxp3 induction and down-regulation of Smad7.

CD4(+)CD25(+) regulatory cells are a subpopulation of T lymphocytes of thymic origin. However, recent data suggest an alternative commitment of regulatory T cells in the periphery, although the precise mechanism is unknown. In the present work, we demonstrate that TGF-beta is able to induce Foxp3 expression and subsequently a regulatory phenotype in CD4(+)CD25(-) peripheral murine T cells. Similarly, TGF-beta induced Foxp3 in human CD4(+)CD25(-) T cells. Moreover, we show that the inhibitory Smad7 protein that is normally induced by TGF-beta and limits TGF-beta signaling, is strongly down-regulated by Foxp3 at the transcriptional level. Foxp3-mediated down-regulation of Smad7 subsequently rendered CD4(+)CD25(-) T cells highly susceptible to the morphogenic and regulatory effects of TGF-beta signaling via Smad3/4. In summary, we demonstrate that TGF-beta induces a regulatory phenotype in CD4(+)CD25(-) T cells through the induction of Foxp3 and a positive autoregulatory loop of TGF-beta signaling due to the absence of Smad7.

Animals↗

PubMed2004

Smad7 in TGF-beta-mediated negative regulation of gut inflammation.

Mice with targeted disruptions of the transforming growth factor-beta1 (TGF-beta1) gene or TGF-beta1 intracellular signalling pathways develop intestinal inflammation. Conversely, TGF-beta1-producing regulatory T cells protect against experimental colitis. Paradoxically, however, TGF-beta1 production is high in the gut of patients with chronic inflammatory intestinal disease, and yet inflammation proceeds unchecked. Here we discuss the functional role of Smad7, an intracellular inhibitor of TGF-beta1 signalling, in the control of gut inflammation by TGF-beta1. In particular, we delineate a scenario in which the high expression of Smad7 in inflammatory cells renders them unresponsive to TGF-beta1 and propose that control of Smad7, not TGF-beta1 production, is a key determinant in understanding how TGF-beta1 negatively regulates gut inflammation.

Animals↗

PubMed2004

Induction and regulation of Smad7 in the gastric mucosa of patients with Helicobacter pylori infection.

BACKGROUND & AIMS: Helicobacter pylori (Hp) infection causes a chronic gastric inflammation, which can lead to peptic ulceration and cancer. The inflammatory response is multifactorial and is characterized by exaggerated Th1 cytokine production. How the Th1 response is induced and maintained in the stomach of Hp-infected patients remains unclear. Transforming growth factor (TGF)-beta 1 negatively regulates Th1 cell development, and TGF-beta 1-deficient mice spontaneously develop gastritis. Here, we examined TGF-beta 1 signaling in Hp-associated gastritis. METHODS: Gastric biopsy specimens taken from patients with or without Hp infection were analyzed for the content of activated TGF-beta1 by ELISA and Smad3 and 7 expression by Western blotting. Induction of Smad7 by interferon (IFN)-gamma was examined in normal gastric mucosal biopsy specimens, whereas the effect of Smad7 inhibition on the ongoing Th1 response was analyzed in Hp-colonized biopsy specimens. RESULTS: Activated TGF-beta 1 was abundant in the mucosa of controls and Hp-infected patients, with no significant difference between the 2 groups. Despite this, in whole biopsy specimens and isolated mucosal cells from Hp-infected patients, there was defective TGF-beta 1-associated Smad3 phosphorylation, which was associated with high expression of the inhibitor Smad7. Blocking Smad7 with antisense oligonucleotides restored TGF-beta 1 signaling in biopsy specimens from Hp-infected patients and concomitantly reduced interferon-gamma and T-bet. Smad7 was inducible in normal gastric biopsy specimens by interferon-gamma through a STAT1-dependent mechanism, and neutralization of interferon-gamma in biopsy specimens from Hp-infected patients reduced Smad7 expression. CONCLUSIONS: These data suggest that, in Hp-infected gastric mucosa, interferon-gamma induces the expression of Smad7, which then prevents endogenous TGF-beta 1 from down-regulating the ongoing tissue-damaging Th1 response.

Adult↗

PubMed2004

CARD15 mutation analysis in an Italian population: Leu1007fsinsC but neither Arg702Trp nor Gly908Arg mutations are associated with Crohn's disease.

BACKGROUND: CARD15 gene mutations have been demonstrated to confer a high risk of Crohn's disease (CD). Despite this, recent studies reported variable associations between CD and CARD15 mutations in distinct ethnic groups, thus raising the hypothesis that genetic and/or allelic heterogeneity may influence the relationship between CARD15 and CD. The purpose of this study was to evaluate the frequency of the main mutations of the CARD15 gene (Leu 1007fsinsC, Arg702Trp, and Gly908Arg) in Italian CD patients and to establish possible genotype-phenotype correlations. METHODS: One hundred sixty-five CD patients and 125 healthy subjects were consecutively enrolled from January to November 2001. The Leu1007fsinsC mutation was assessed by denaturing high-performance liquid chromatography and Arg702Trp and Gly908Arg mutations by Pyrosequencing technology. RESULTS: Among the CARD15 gene mutations tested, only the Leu1007fsinsC was associated with CD (30/165 CD patients, 18%, versus 3/125 healthy subjects, 2.4%; p < 0.001). In particular, 23 CD patients were heterozygotes and 7 were homozygotes. No healthy subject exhibited the mutant homozygous genotype. Odds ratios for CD were 6.9 for heterozygotes and 41.0 for homozygotes. The genotype-phenotype analysis revealed that a fibrostenosing CD of the distal ileum was more frequent in patients carrying the Leu1007fsinsC mutation. CONCLUSIONS: This study confirms the association between CARD15 gene mutations and CD and shows that only the Leu1007fsinsC mutation is a risk factor of CD in an Italian population.

Adolescent↗

PubMed2004

Psoriatic arthritis and CARD15 gene polymorphisms: no evidence for association in the Italian population.

Psoriatic arthritis (PsA) has been defined as an inflammatory arthritis associated with psoriasis that may affect as many as 30% of psoriasis patients. Epidemiological study reported strong familial clustering of PsA although the precise etiology of PsA is poorly understood. Recently, a genomewide linkage scan in PsA revealed a LOD score of 2.17 on chromosome 16q and provided strong evidence for a paternal imprinting effect. That region surrounds a psoriasis susceptibility locus including the CARD15 gene which has convincingly been shown to confer susceptibility to Crohn's disease. The existence of a common susceptibility gene for psoriasis/PsA and Crohn disease was recently demonstrated by evidence of association of CARD15 polymorphisms with PsA. To confirm these results in an independent population, we analyzed a data set of 193 Italian PsA patients and 150 controls for CARD15 polymorphisms (R702W, G908R and leu1007finsC) previously demonstrated associated with PsA. Here we report no evidence for association in the examined population for CARD15 polymorphisms, suggesting that the positive association previously reported in a genetically isolated population was the result of a linkage disequilibrium due to a founder effect.

Arthritis, Psoriatic↗

PubMed2004

Extracellular signal-regulated protein kinase mediates interleukin 17 (IL-17)-induced IL-8 secretion in Helicobacter pylori-infected human gastric epithelial cells.

Helicobacter pylori-induced mucosal inflammation results in high production of interleukin 17 (IL-17), a potent inducer of IL-8 in gastric epithelial cells. The aim of this study was to investigate signaling pathways by which IL-17 regulates IL-8 production in human gastric epithelial cells. Activation of mitogen-activated protein (MAP) kinases in both IL-17-stimulated MKN28 cells and epithelial cells isolated from H. pylori-colonized gastric mucosa was assessed by Western blotting. In IL-17-stimulated MKN28 cells the activation of activatior protein 1 (AP-1), nuclear factor (NF)-IL-6, and NF-kappaB was also assessed by electrophoretic mobility shift assay. IL-8 production was evaluated by reverse transcription-PCR and enzyme-linked immunosorbent assay (ELISA) both for IL-17-stimulated MKN28 cells treated with specific MAP kinase inhibitors and gastric biopsy cultures treated with a neutralizing IL-17 antibody. Serum from H. pylori-infected patients was tested for immunoglobulin G response to CagA by ELISA. Treatment of MKN28 cells with IL-17 caused activation of extracellular signal-regulated protein kinase 1/2 (ERK 1/2) but not other MAP kinases and had the downstream effects of AP-1 and NF-kappaB activation and IL-8 synthesis. Blocking ERK 1/2 activity inhibited AP-1-mediated, but not NF-kappaB-mediated, IL-8 induction. Enhanced activation of ERK 1/2 was seen in gastric epithelial cells isolated from H. pylori-infected patients in comparison to uninfected controls, and this was associated with high IL-8. These effects were even more pronounced in patients seropositive for CagA than in seronegative ones. In gastric biopsy cultures, the addition of a neutralizing IL-17 antibody decreased ERK 1/2 activation, thus resulting in a significant inhibition of IL-8. In H. pylori-colonized gastric epithelial cells, IL-17-induced IL-8 synthesis is associated with and depends at least in part on the activation of ERK 1/2 MAP kinase.

Adult↗

PubMed2003

A failure of transforming growth factor-beta1 negative regulation maintains sustained NF-kappaB activation in gut inflammation.

Immunologically mediated tissue damage in the gut is associated with increased production of proinflammatory cytokines, which activate the transcription factor NF-kappaB in a variety of different cell types. The mechanisms/factors that negatively regulate NF-kappaB in the human gut and the pathways leading to the sustained NF-kappaB activation in gut inflammation remain to be identified. Pretreatment of normal human intestinal lamina propria mononuclear cells (LPMC) with transforming growth factor-beta1 (TGF-beta1) resulted in a marked suppression of TNF-alpha-induced NF-kappaB p65 accumulation in the nucleus, NF-kappaB binding DNA activity, and NF-kappaB-dependent gene activation. TGF-beta1 also increased IkappaBalpha transcripts and protein in normal LPMC. In marked contrast, treatment of LPMC from patients with inflammatory bowel disease with TGF-beta1 did not reduce TNF-induced NF-kappaB activation due to the overexpression of Smad7. Indeed inhibiting Smad7 by specific antisense oligonucleotides increased IkappaBalpha expression and reduced NF-kappaB p65 accumulation in the nucleus. This effect was due to endogenous TGF-beta1. TGF-beta1 directly stimulated IkappaBalpha promoter transcriptional activity in gut fibroblasts in vitro, and overexpression of Smad7 blocked this effect. These data show that TGF-beta1 is a negative regulator of NF-kappaB activation in the gut and that Smad7 maintains high NF-kappaB activity in gut inflammation by blocking TGF-beta1 signaling.

Active Transport, Cell Nucleus↗