Masanobu Kayazawa

Fecal eosinophil granule-derived proteins reflect disease activity in inflammatory bowel disease

OBJECTIVES:The aims of this study were: 1) to examine whether the fecal levels of eosinophil granule-derived proteins reflect disease activity in inflammatory bowel disease (IBD); and 2) to examine the extracellular release of these proteins from eosinophils and their stability in feces by an in vitro study.METHODS:We investigated 42 patients with ulcerative colitis (UC), 37 patients with Crohns disease (CD), and 29 control subjects. The stool samples were collected at 4°C over 48 h and were homogenized. The fecal levels of eosinophil cationic protein (ECP) and eosinophil protein X (EPX) were measured by radioimmunoassay. Fecal Hb (Hb), α1-antitrypsin (AT), and lactoferrin (Lf) were also measured by ELISA.RESULTS:Fecal ECP and EPX concentrations were significantly increased in both active UC and active CD compared to inactive UC and inactive CD, respectively. Fecal EPX concentration correlated with the fecal Hb, AT, and Lf concentrations more closely than fecal ECP concentration. Even in the inactive stage, CD patients who relapsed within the following 3 months showed higher fecal ECP and EPX concentrations compared to the patients who did not. EPX was released extracellularly more efficiently than ECP (18.6%vs 6.3%, after incubation for 15 min at 25°C). EPX was more stable in the feces than ECP.CONCLUSIONS:The measurement of eosinophil granule-derived proteins in feces is useful for evaluating disease activity and predicting relapse in patients with IBD. EPX may be more suitable than ECP as a fecal eosinophil marker.

Lactoferrin in whole gut lavage fluid as a marker for disease activity in inflammatory bowel disease: comparison with other neutrophil-derived proteins

OBJECTIVES:We investigated which neutrophil-derived proteins in whole gut lavage fluid (WGLF) most accurately reflect disease activity in inflammatory bowel disease.METHODS:WGLF was obtained from patients undergoing whole gut lavage as a bowel preparation for colonoscopy. Twenty-seven patients with ulcerative colitis (UC), 23 patients with Crohns disease (CD), and 35 control subjects were examined. The concentrations of lactoferrin, polymorphonuclear neutrophil elastase (PMN-E), myeloperoxidase, and lysozyme in WGLF were measured by ELISA. For the assessment of stability, WGLF samples were stored at 37°C for various periods.RESULTS:In UC, the concentrations of lactoferrin, myeloperoxidase, and lysozyme in WGLF had good correlations with colonoscopic grading. Zero, 12, five, and 10 of 28 samples from active UC patients showed normal concentrations of lactoferrin, PMN-E, myeloperoxidase, and lysozyme, respectively. In CD, the concentrations of lactoferrin and myeloperoxidase had good correlations with the Crohns disease activity index. Thirteen and seven of 36 samples from inactive CD patients (Crohns disease activity index ≤ 150) showed high concentrations of lactoferrin and myeloperoxidase, respectively. Most of them (11/13, 6/7) were found to have ulceration by colonoscopy or small bowel x-ray. The ratio of the lactoferrin concentration in the WGLF supernatant to that in total WGLF was highest among these proteins in all disease groups and control subjects. Lactoferrin and myeloperoxidase showed good stability in WGLF, whereas PMN-E and lysozyme did not.CONCLUSION:Lactoferrin is the most suitable of these proteins for use as a neutrophil-derived WGLF marker of intestinal inflammation.

Medium-chain fatty acids stimulate interleukin-8 production in Caco-2 cells with different mechanisms from long-chain fatty acids

Background and Aim: It has been suggested that dietary fat exacerbates intestinal inflammation. We investigated the effect of fatty acids on interleukin (IL)‐8 production in a human intestinal epithelial cell line (Caco‐2).

Left gastric vein hemodynamics and variceal recurrence in patients undergoing prophylactic endoscopic ligation of high-risk esophageal varices.

BACKGROUND Early recurrence of esophageal varices remains problematic after endoscopic variceal ligation. To evaluate the efficacy of prophylactic endoscopic ligation for esophageal varices at high risk for bleeding, the relationship between left gastric vein hemodynamics and variceal recurrence was investigated. METHODS Thirty-five patients with cirrhosis underwent endoscopic variceal ligation. Angiography was performed in all patients before treatment and after eradication of varices to study left gastric vein hemodynamics. RESULTS Before treatment, 12 patients had hepatopetal flow in the left gastric vein (type I), 17 had hepatofugal flow (type II), and 6 had hepatofugal flow with an extra-esophageal shunt (type III). In type I and III patients, the direction of blood flow in the left gastric vein did not change after eradication of varices. Type II patients showed bi-directional flow in the left gastric vein after treatment. Varices recurred in all but one type II patient and in one type I patient during follow-up (mean 36.7 months). The 2-year recurrence-free rate was higher in type I patients (p = 0.0001) and type III patients (p = 0.0002) than in type II patients. CONCLUSIONS Prophylactic ligation seems to be a safe and useful procedure, especially in patients with type I or III hemodynamics in the left gastric vein before treatment.

Prophylaxis of rebleeding from isolated gastric fundal varices by balloon-occluded retrograde transvenous obliteration

We describe a case of bleeding gastric fundal varices associated with a gastrorenal shunt that were successfully treated with balloon-occluded retrograde transvenous obliteration. Blood flow in the varices disappeared after treatment. Because of its safety and simplicity, balloon-occluded retrograde transvenous obliteration appears to be a feasible alternative to transjugular intrahepatic portosystemic shunt for the treatment of gastric fundal varices.

Medium-chain fatty acids stimulate interleukin-8 production in Caco-2 cells by different mechanisms from long-chain fatty acids

BACKGROUND AND AIM It has been suggested that dietary fat exacerbates intestinal inflammation. We investigated the effect of fatty acids on interleukin (IL)-8 production in a human intestinal epithelial cell line (Caco-2). METHODS The cells were cultured as monolayers on microporous membranes in culture inserts. Oleic acid (OA), capric acid (CA), docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA) were applied to the apical compartment of Caco-2 cell monolayers. The concentration of IL-8 in the basolateral medium was measured by using enzyme-linked immunosorbent assay, and the expression of IL-8 mRNA was measured by using competitive reverse transcription--polymerase chain reaction. Protein kinase C inhibitors (GF109203X and calphostin C) and H-7 (a protein kinase inhibitor) were used to study the mechanisms by which IL-8 production is stimulated. RESULTS Both OA and CA enhanced IL-8 production (approximately fivefold), whereas DHA and EPA did not. Both OA and CA also enhanced IL-1-induced IL-8 production. The onset of OA-induced IL-8 production was delayed compared with that of CA-induced IL-8 production. Both OA and CA enhanced IL-8 mRNA expression (approximately fivefold) after 6 and 3 h, respectively. The protein kinase inhibitor (H-7) reduced both OA- and CA-induced IL-8 production by 88.0 and 85.9%, respectively. The protein kinase C inhibitors (GF109203X and calphostin C) reduced OA-induced IL-8 production by 29.3 and 54.5%, respectively, but showed no effect on CA-induced IL-8 production. CONCLUSIONS These findings suggest that not only OA but also CA stimulates IL-8 production in intestinal epithelial cells, and the mechanisms of action differ between OA and CA.

The levels and the structure of fecal α 1-antitrypsin in patients with inflammatory bowel disease

Fecal ct 1-antitrypsin (ct 1-AT) clearance is now a standard test for the assessment of intestinal protein loss. In patients with inflammatory bowel disease(IBD), the sources of fecal a 1-AT remains uncleared and molecular heterogeneity of fecal a 1-AT has been reported. AIMS: 1) To examine the levels and the structure of a 1-AT in feces and sera of patients with IBD. 2) To examine the synthesis and secretion of a 1-AT in intestinal epithelial cells and hepatocytes. METHODS: We studied 36 patients with ulcerative colitis(UC), 26 patients with Crohns disease(CD), and 20 control subjects. The stool samples were collected at 4 °C over a period of 48 hours and were homogenized. We also used human intestinal epithelial cell lines, Caco-2 and HT-29, and a hepatocellular cell line, HepG2. These cells were stimulated with interleukin-6 (IL-6), interleukin-1 13 and tumor necrosis factor a. The level of <x 1-AT protein was measured by ELISA, and the expression of <x 1-AT mRNA by RT-PCR. The a 1-AT molecules were examined by Western blotting and the reactivity with Concanavalin A (Con A) was studied by Con Aa 1-AT Ab sandwich EIA. RESULTS: 1) Fecal ct 1-AT concentrations were significantly higher in active UC and CD (medians 1465, and 3482 lag/g respectively) than in inactive UC, CD and control subjects (medians 411,651, and 327 laglg respectively). In all disease groups and control subjects, serum a 1-AT showed an apparent molecular weight (MW) of-52 kDa and fecal a 1-AT showed fragments of various sizes. In active UC and CD, the apparent MW of the main band of fecal ct I-AT was higher than in inactive patients and control subjects. In active UC and CD, ConA-bound ct 1-AT was rich in the feces. 2) The synthesis and secretion of a 1-AT were increased after stimulation with IL-6 (10, 100 ng/ml) in Caco-2 cells, and HT-29 cells as well as HepG2. The percentage of ct I-AT released extracellularly was also increased. The level of ¢t 1-AT mRNA expression reached a peak within 120 mins of stimulation of IL-6. Neither IL-1 13 nor TNF a was a potent stimulant. CONCLUSIONS: An increase of fecal a 1-AT level is at least partly due to an increase of its secretion from intestinal epithelial cells in patients with intestinal inflammation. The structural analysis of fecal a I-AT is useful for assessment of disease activity in IBD.