Changes in the Expression and Distribution of Claudins, Increased Epithelial Apoptosis, and a Mannan-Binding Lectin-Associated Immune Response Lead to Barrier Dysfunction in Dextran Sodium Sulfate-Induced Rat Colitis.

BACKGROUND/AIMS: This animal study aimed to define the underlying cellular mechanisms of intestinal barrier dysfunction. METHODS: Rats were fed 4% with dextran sodium sulfate (DSS) to induce experimental colitis. We analyzed the sugars in 24-hour urine output by high pressure liquid chromatography. The expression of claudins, mannan-binding lectin (MBL), and MBL-associated serine proteases 2 (MASP-2) were detected in the colonic mucosa by immunohistochemistry; and apoptotic cells in the colonic epithelium were detected by the terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling method assay. RESULTS: The lactulose and sucralose excretion levels in the urine of rats with DSS-induced colitis were significantly higher than those in the control rats. Mannitol excretion was lower and lactulose/mannitol ratios and sucralose/mannitol ratios were significantly increased compared with those in the control group (p<0.05). Compared with the controls, the expression of sealing claudins (claudin 3, claudin 5, and claudin 8) was significantly decreased, but that of claudin 1 was increased. The expression of pore-forming claudin 2 was upregulated and claudin 7 was downregulated in DSS-induced colitis. The epithelial apoptotic ratio was 2.8%±1.2% in controls and was significantly increased to 7.2%±1.2% in DSS-induced colitis. The expression of MBL and MASP-2 in the intestinal mucosa showed intense staining in controls, whereas there was weak staining in the rats with colitis. CONCLUSIONS: There was increased intestinal permeability in DSS-induced colitis. Changes in the expression and distribution of claudins, increased epithelial apoptosis, and the MASP-2-induced immune response impaired the intestinal epithelium and contributed to high intestinal permeability.

Circulating SFRP1 promoter methylation status in gastric adenocarcinoma and esophageal square cell carcinoma.

The secreted frizzled-related protein 1 (SFRP1) gene plays an important role in carcinogenesis of digestive system cancer. Previous studies proved that circulating DNA promoter methylation may be a suitable biomarker for cancer patients. The aim of the present study was to investigate whether the promoter methylation status of serum SFRP1 is a potential biomarker for gastric adenocarcinoma (GAC) and esophageal square cell carcinoma (ESCC). The blood samples obtained from 42 GAC and 36 ESCC patients were detected for the promoter methylation status of SFRP1 by methylation-specific polymerase chain reaction. The control group included 42 benign gastrointestinal disease volunteers (24 benign gastric disease and 18 benign esophageal disease) and 20 healthy volunteers. Serum SFRP1 methylation was evident in 30.95% (13/42) GAC patients and 38.89% (14/36) ESCC patients, which is clearly higher compared to 8.33% (2/24) in benign gastric disease, 11.11% (2/18) in benign esophageal disease and 5% (1/20) in healthy volunteers (P<0.05). The association between the serum SFRP1 promoter methylation status and the clinical pathological features were further analyzed and methylation of the SFRP1 gene was significantly associated with age >60 years in GAC patients (P=0.027). However, no correlations between the SFRP1 methylation status and other clinicopathological parameters were found. In conclusion, the SFRP1 promoter was detected frequently in the serum of GAC and ESCC patients. The detection of circulating methylated SFRP1 in the serum may be a useful biomarker for upper gastrointestinal cancer patients.

CXCR4 antagonist AMD3100 attenuates colonic damage in mice with experimental colitis.

AIM: To investigate the effects of the chemokine stromal cell-derived factor-1 (CXCL12) receptor (CXCR4) antagonist AMD3100 on colonic inflammation and epithelial barrier in dextran sulfate sodium (DSS)-induced colitis in mice. METHODS: Experimental colitis was induced by administration of 5% DSS for 7 d, and assays performed on intestinal segments from the ileocecal valve to the anus. Colonic morphology was examined by hematoxylin and eosin staining. Colonic cytokines were determined by enzyme-linked immunosorbent assay. Myeloperoxidase (MPO) activity (indicator of inflammatory infiltration) was observed spectrophotometrically. Gut permeability was assessed by mucosal-to-serosal clearance of fluorescein isothiocyanate-conjugated dextran 4000 (FD4) in everted gut sacs. The apoptosis of colonic epithelium was assessed by Hoechst-33342 staining. To further elucidate the role of CXCR4 in colonic inflammation, we also investigated the effect of AMD3100 on migration and cytokine production of isolated peripheral blood mononuclear cells (PBMCs). RESULTS: DSS-induced colitis was characterized by morphologic changes, as well as increased colonic cytokines, inflammatory infiltration, epithelial apoptosis, and intestinal permeability in mice. In AMD3100-treated mice, epithelial destruction, inflammatory infiltration, and submucosal edema were markedly reduced; colonic tumor necrosis factor-alpha (TNF-alpha), interleukin-6 (IL-6) and interferon-gamma (IFN-gamma) levels, as well as MPO activity were significantly decreased. Increased intestinal permeability in DSS-treated mice was significantly reduced by AMD3100. The number of apoptotic cells in colitis mice was markedly increased after DSS administration, and decreased when treated with the CXCR4 antagonist AMD3100. In pre-activated PBMCs, CXCL12 stimulation significantly increased the migration of PBMCs, and was inhibited by AMD3100. Moderately increased TNF-alpha, IL-6, and IFN-gamma from CXCL12-treated PBMCs were also reduced by AMD3100. CONCLUSION: The CXCR4 antagonist AMD3100 exerts therapeutic effects on experimental colitis by inhibiting colonic inflammation and enhancing epithelial barrier integrity.