Search

The role for Helicobacter pylori in the pathogenesis of disease provides the conundrum that only a subset of subjects infected with H. pylori will ever develop peptic ulcer or gastric cancer. Thus, variation in strain as well as environmental or host factors converge in the gastroduodenal milieu and control the final outcome of infection. The host immune and inflammatory response is emerging as an important element in the pathogenesis of these gastric diseases. The ideal host response provides protection to clear an infection without causing excessive amounts of inflammation that could compromise the integrity and function of host cells. This review will cover four main questions: (1) What are the mucosal immune/inflammatory responses that confer protection without damaging the host? (2) How do the gastric immune responses during infection with H. pylori differ from this ideal scenario? (3) Do these responses contribute to autoimmune-mediated damage to gastric tissue? (4) Can immunomodulation through vaccination enhance protective, nondestructive responses that prevent or treat infection or, at least, attenuate inflammation?

Background & Aims: A prominent role for inhibitory molecules PD-L1 and PD-L2 in peripheral tolerance has been proposed. However, the phenotype and function of PD-L–expressing cells in human gut remains unclear. Recent studies suggest that colonic myofibroblasts (CMFs) and fibroblasts are important in the switch from acute inflammation to adaptive immunity. In the normal human colon, CMFs represent a distinct population of major histocompatibility complex class II+ cells involved in the regulation of mucosal CD4+ T-cell responses. Methods: PD-L1 and PD-L2 expression on human CMFs was determined using Western blot, fluorescence-activated cell sorter analysis and confocal microscopy. Lymphoproliferation assays and cytokine enzyme-linked immunosorbent assays were used to evaluate the role of B7 costimulators expressed by CMFs with regard to the regulation of preactivated T-helper cell responses. Results: We demonstrate here the expression of PD-L1/2 molecules by normal human CMF and fibroblasts in situ and in culture. Both molecules support suppressive functions of CMFs in the regulation of activated CD4+ T-helper cell proliferative responses; blocking this interaction reverses the suppressive effect of CMFs on T-cell proliferation and leads to increased production of the major T-cell growth factor, interleukin (IL)-2. PD-L1/2-mediated CMF suppressive functions are mainly due to the inhibition of IL-2 production, because supplementation of the coculture media with exogenous IL-2 led to partial recovery of activated T-cell proliferation. Conclusions: Our data suggest that stromal myofibroblasts and fibroblasts may limit T-helper cell proliferative activity in the gut and, thus, might play a prominent role in mucosal intestinal tolerance.

Background & Aims: Helicobacter pylori infection causes inflammation, accumulation of reactive oxygen species, and oxidative DNA damage in the gastric mucosa. Apurinic/apyrimidinic endonuclease-1 (APE-1)/redox factor-1 (Ref-1) repairs damaged DNA and reductively activates transcription factors, including activator protein-1. Considering that H. pylori generate reactive oxygen species and that reactive oxygen species modulate APE-1/Ref-1 in other cell types, we examined the effect of H. pylori, oxidative stress, and antioxidants on APE-1/Ref-1 expression in human gastric epithelial cells. Methods: Human gastric epithelial cell lines or cells isolated from mucosal biopsy samples were stimulated with H. pylori, Campylobacter jejuni, and/or H2O2 in the presence or absence of antioxidants. APE-1/Ref-1 expression was assayed by Western blot or reverse-transcription polymerase chain reaction, and its cellular distribution was determined by using indirect conventional and confocal immunofluorescence. New protein synthesis was detected by [S35]methionine labeling. APE-1/Ref-1 function was assessed by using a luciferase-linked reporter construct containing 3 activator protein 1 binding sites. Results: APE-1/Ref-1 protein and messenger RNA were detected in resting gastric epithelial cells. APE-1/Ref-1 protein expression was increased after stimulation with H2O2 or live cag pathogenicity island-bearing H. pylori, but not cag pathogenicity island-negative H. pylori or C. jejuni. H. pylori- or reactive oxygen species-mediated increases in APE-1/Ref-1 expression involved de novo protein synthesis that was inhibited by antioxidants. H. pylori or H2O2 also induced nuclear accumulation of APE-1/Ref-1, and overexpression of APE-1/Ref-1 increased activator protein 1 binding activity. Conclusions: The data show that H. pylori or reactive oxygen species enhance APE-1/Ref-1 protein synthesis and nuclear accumulation in human gastric epithelial cells and implicate APE-1/Ref-1 in the modulation of the pathogenesis of H. pylori infection.

Background & Aims: Studies have shown that gastric T cells are increased during Helicobacter pylori infection. The purpose of this study was to characterize the human gastric T-cell responses in the presence or absence of H. pylori . Methods: T-cell surface antigens were examined by immunohistochemistry or after isolation for evaluation of surface antigens and cytoplasmic cytokines using flow cytometry. Results: CD4 + and CD8 + T cells were increased in situ during infection with H. pylori . Freshly isolated gastric T cells expressed cytoplasmic interferon gamma (IFN-γ) and interleukin (IL)-2 after a brief stimulation. Simultaneous four-color flow cytometry demonstrated that both CD8 + and CD4 + T cells expressed IFN-γ. Because stimulation through CD30 favors the induction of IL-5 and Th2 cells, gastric and colonic T cells were examined for CD30 expression. Consistent with the notion that Th2 cells are found in the intestine, CD30 was evident throughout the lamina propria of the colon but was virtually absent in the stomach. Furthermore, freshly isolated gastric T cells produced little IL-4 and virtually no IL-5 or tumor necrosis factor β. Conclusions: These observations show that gastric T cells resemble the Th1 type, which may explain their failure to induce immunity to H. pylori and their ability to contribute to the pathogenesis of gastric disease. GASTROENTEROLOGY 1998;114:482-492

STATUS: Sessioned Background: Esophageal adenocarcinoma (EAC) continues to be one of the fastest rising incident cancers. However, the overall risk of progression to HGD and EAC in pts with BE is low. Identifying pts at risk of progression may help tailor management. Aim: In a large multi-center cohort of BE pts undergoing screening/surveillance to: -Identify predictors for HGD/EAC development. -Formulate an equation/score capable of predicting pts likely to harbor or progress to HGD/ EAC Methods: This is a prospective, multicenter outcomes project (5 centers) involving BE pts. BE diagnosis required intestinal metaplasia on histology. Demographics, medication use, family history, and endoscopy results were recorded. Time of LGD, HGD, and EAC detection was recorded. Assuming that the clinical and demographic factors of pts who develop HGD/ EAC while in surveillance should be no different than pts presenting with HGD/EAC, the endpoint included both prevalent and incident cases. Univariate analysis using Chi-square and Mann-Whitney U test identified predictors for prevalent and incident HGD/EAC. LRM was developed with factors significant on univariate analysis. Missing Variable Analysis was performed (Little's MCAR test ,0.01) and missing values were computed by multiple imputation. Receiver Operating Characteristic (ROC) curve of the prediction score was plotted to determine the best cut-off value. Results: 3677 pts were included [87.6% males, 94.9% Caucasians, mean BMI 28.3 (SD 5.7), mean age 60.8 (SD 12.3), and mean BE length 3.5 cm (SD 3.1)]. 348 prevalent and 75 incident cases of HGD/ EAC were recorded and compared to pts without HGD/EAC. On univariate analysis age, BMI, BE length, current/past smoking history, lack of aspirin or NSAID use, hiatal hernia, number of consecutive EGDs showing nondysplastic Barrett's Esophagus (NDBE), and presence of visible lesions within the BE segment were significantly associated with HGD/EAC. A statistically significant LRM (Nagelkerke R square = 0.31) resulted in the following equation for prediction score (Table): Y = (0 .019*Age)+(0 .864*Caucasian race)+(0 .538*Male gender)+(0.050*BE length)+(1.139*visible lesion)+(0.322*Presence of hiatal hernia)+(-0.674*Number of consecutive EGDs showing NDBE)-4.416. A cut-off for optimal performance of the prediction model was determined by plotting an ROC curve using the prediction score [AUROC = 0.877 (95% CI 0.86\u20130.90),p=0.01]. Prediction score cut off at 0.0937 yielded the best sensitivity (90%) and specificity (69%) (Figure). Conclusions: Using demographic and endoscopic risk factors from a large cohort of more than 3500 patients, we have developed a prediction model with an AUC of 0.877. If this prediction model is successfully externally validated, using a progression score, high risk BE patients could be identified leading to tailored management of these patients. A G A A b st ra ct s

G A A b st ra ct s round-shaped ones in tumor glands (stromal Twist1+cells, A/R = 2.69, glandular Twist1+ cells, A/R= 1.67, A/R p

BACKGROUND: Activation of TLR3 triggers specific signaling pathways leading to the activation of NF-kB and/or IRF3 transcription factors, enabling the host to mount innate immune responses against different dsRNA viruses and serve as a “danger” receptor in nonviral diseases. However, TLR3-mediated organ specific activation of the B7-coinhibitory pathway PD-L1/PD-1 can be exploited locally leading to the impairment in antiviral T effector cell responses promoting successful viral invasion of the host. Despite significant advances in understanding of the process of TLR3 signaling, the mechanisms by which TLR3 influence homeostasis and contribute to the B7 negative molecules-mediated viral invasion in the gut remain obscure. Human colonic stromal cells, A.k.a. myofibroblasts/fibroblasts (CMFs), are one of the important cell types taking part in innate immunity. CMFs express TLR3, and represent a major cell population expressing B7 negative costimulators PD-L1 and PD-L2 in normal colon. CMFs suppress activated CD4+ effector T cell responses in a PD-L1/2 dependent manner. We hypothesized that in human colonic mucosa, CMFs might be among the first cells to respond to TLR3 mediated stimuli via modulation of PD-L1/2 expression and, thus, may contribute to the impaired T effector cell responses associated with viral invasion. METHODS: TLR3, PD-L1/2 expression on human CMFs isolated from normal mucosa in response to the synthetic analog of TLR3 ligand, poly(I:C), was quantified using real-time RT-PCR, Western blot and FACS analysis. RESULTS: TLR3 stimulation on CMFs by poly(I:C) resulted in robust upregulation of PD-L1 and PD-L2 mRNA and cell surface protein. The expression of IFN-β precedes the upregulation of PD-L1 and PD-L2 mRNA. Stimulation of CMFs with IFN-β leads to the upregulation of the PD-L1 and PD-L2 protein expression (Westen blot analysis) suggesting involvement of IFN-β in the TLR3-mediated PD-L1/2 upregulation. Blocking of the MyD88 adaptor expression in CMFs by using specific siRNA did not affect PD-L2 upregulation in response to the TLR3 stimulation, but reduced the induction of the PD-L1. It is possible that TLR3mediated upregulation of PD-L1 on CMFs may be synergized by activation of heat shock proteins that are known to signal through the TLR4MyD88 adaptor pathway. CONCLUSION: The presented results indicate that CMFs are among the key players in innate immune responses in colonic mucosa. Our data suggest that TLR3 mediated PD-L1/2 upregulation on CMFs may favor suppression of T effector cells responses and this may allow the progression of host invasion during intestinal viral infection. Supported by AGA, CCFA foundations and NIDDK.

BACKGROUND: The B7 co-stimulator PD-L1 is critical in suppression of activated T cell responses in peripheral tolerance. However, overexpression of PD-L1 may contribute to progression of chronic inflammatory diseases. The role of PD-L1 and the phenotype of PDL1 expressing cells in inflammatory bowel diseases is unclear. Human colonic myofibroblasts and fibroblasts (CMFs) are major PD-L1 expressing cells in normal colonic mucosa and may suppress activated CD4+T cell proliferation and production of the Th1 acute inflammatory cytokine IFN-γ in a PD-L1 dependent manner. We noted a significant increase in PD-L1 surface expression on CMFs isolated from ulcerative colitis (UC). Thus, we hypothesize that overexpression of PD-L1 by CMFs may have a critical role in the chronic inflammatory responses during UC pathogenesis. METHODS: PD-L1 expression in colonic tissue from normal (N), UC and Crohn's Disease (CD) patients was determined by immunostaining followed by confocal microscopy. PD-L1, Tbet and GATA3 expression was analyzed by realtime RT-PCR, Western blot and flow cytometry. Lymphoproliferation assays and ELISA were used to evaluate the role of PD-L1 in the regulation of Th cell activity. RESULTS: Confocal microscopy analysis revealed strong upregulation of PD-L1 in inflamed colonic mucosa from UC when compared to CD or normal controls. The PD-L1 upregulation in UC colon was mostly associated with cells bearing fibroblast/myofibroblast phenotype (CD90+). PD-L1 on CMFs isolated from UC (UC-CMFs) was functional: UC-CMFs were capable of suppressing CD3/CD28-activated Th cell proliferation in a PD-L1 dependent manner. Both Nand UCCMFs inhibited expression of a Th1 transcriptional factor, Tbet, in Th cells. PD-L1 blocking led to increased expression of Tbet in Th cells, but did not affect expression of Th2 transcriptional factor GATA3. Treatment of N-CMFs with IFN-γ (10-1000 U/mL) for 7 days resulted in a dose dependent upregulation of PD-L1. Treatment of N-CMFs with IL-13 (25 ng/mL), the major cytokine associated with the UC atypical Th2 response, also upregulate PD-L1. Our data suggest that increased secretion of IFN-γ by activated T cells during acute intestinal inflammation may contribute to an increase in PD-L1 expression on CMFs resulting in the negative feedback on the Th1 responses, in turn, promoting IL-13 production by Th2 and NK(T) cells, which will further upregulate PD-L1 expression on CMFs. CONCLUSIONS: Overexpression of PD-L1 on CMFs in the UC colonic mucosa may play an important immunoregulatory role promoting the chronicity of the Th2 inflammatory responses during progression of UC. Supported by NIDDK, CCFA and Sealy foundation.