Tetsuji Kitahora

Active oxygen species generated by monocytes and polymorphonuclear cells in Crohn's disease

Chemiluminescence (CL) analysis of monocytes and polymorphonuclear cells (PMNs) was performed on 13 patients with Crohns disease (CD) and 10 healthy volunteers. The percentages of monocyte populations in mononuclear cells obtained from the patients with CD were greater than those from the healthy volunteers, but the numbers of PMNs were not different between the two groups. The peak level of phorbol myristate acetate (PMA)-induced CL activity generated by diluted whole blood from the patients with CD was more significantly elevated than that from the healthy volunteers, whereas the peak levels of opsonized zymosan-induced CL activity did not differ between the two groups. In monocytes, the peak levels of both PMA- and opsonized zymosan-induced CL activity were significantly higher in the patients with CD than in the healthy volunteers. CL in PMNs, however, showed no significant difference between CD and controls. It is suggested that monocytes of CD have a large capacity to generate active oxygen species. The present study suggests that excessive active oxygen species released by monocytes and perhaps macrophages may play an important role in formation of the intestinal lesions in CD.

Low-dose azathioprine is effective and safe for maintenance of remission in patients with ulcerative colitis

Background. 6-Mercaptopurine (6-MP) and azathioprine (AZA) have been used in patients with Crohns disease (CD) and ulcerative colitis (UC) for reducing the dose of steroids and maintaining remission. However, some patients treated with 6-MP/AZA develop bone marrow suppression, one of the most serious side effects. The aim of this study was to evaluate the efficacy and safety of low-dose AZA (0.6–1.2 mg/kg per day) for maintaining remission in patients with UC. We also investigated the relationship between bone marrow suppression and thiopurine methyltransferase (TPMT) mutation in the Japanese population. Methods. Study 1. To investigate the frequency of TPMT mutation, findings for 82 patients among 141 patients with UC or CD who were treated with AZA or 6-MP were analyzed retrospectively. Polymerase chain reaction (PCR) methods were used to analyze allele mutations of the TPMT gene. Study 2. A multicenter prospective trial was performed. The subjects were 22 patients with UC with presence of remission for 3 months or more. They were treated with 50 mg/day of AZA, and we evaluated the remission rate at 6 months, adverse side effects, and changes in prednisone doses after the initiation of AZA. Results. Study 1. Seventy-four (91%) of the 82 patients analyzed had no TPMT mutation, 7 (8%) had one mutant allele, and 1 (1%) had two mutant alleles. Of the total of 141 patients, 4 (44%) of the 9 patients who were treated with 50 mg/day of 6-MP or 100 mg/day of AZA developed bone marrow suppression, although no mutation of TPMT was seen in any of these patients. On the other hand, 8 (6%) of the 132 patients who were treated with 30 mg/day of 6-MP or 50 mg/day of AZA developed bone marrow suppression. Seven of 8 patients (88%) who developed bone marrow suppression with a low dose of AZA had a mutant TPMT allele. Study 2. In the 17 patients who could continue taking low-dose AZA for 6 months, 15 (88%) maintained remission. Of 8 patients treated with low-dose prednisone (5–10 mg/day), 3 patients (38%) could discontinue oral prednisone and 4 (50%) could reduce its dose. Six of the 22 patients (27%) had some adverse side effects. These side effects were ameliorated, or disappeared spontaneously, or disappeared with the discontinuation of AZA. Conclusions. A dose of 50 mg/day of AZA is effective and safe for maintenance of remission in the Japanese population. Investigation of the TPMT allele may be useful for predicting the appearance of bone marrow suppression, when low-dose 6-MP or AZA is given.

Dietary fat attenuates the benefits of an elemental diet in active Crohn's disease: a randomized, controlled trial.

Objectives Although an elemental diet has been established as the primary treatment for patients with Crohns disease, the influence of dietary fat on the elemental diet remains unclear. We have designed the first randomized, controlled trial for elemental diets containing different fat percentages in patients with active Crohns disease. Methods Each patient was randomized to receive one of three dose levels of fat in an elemental diet (Elental) for 4 weeks: 10 patients received low fat (3.06 g/day), 10 patients received medium fat (16.56 g/day) and eight patients received high fat (30.06 g/day). The additional fat was composed of long-chain fatty acids. All patients were evaluated using the International Organization of Inflammatory Bowel Disease rating, plus C-reactive protein level and erythrocyte sedimentation rate, which were measured at weekly intervals. Results Although the International Organization of Inflammatory Bowel Disease rating, C-reactive protein level and erythrocyte sedimentation rate in the low-fat group decreased, the values in the medium- and high-fat groups fluctuated during the study. The remission rate after 4 weeks in each group was 80%, 40% and 25% for patients in the low-, medium- and high-fat groups, respectively. Conclusions When the fat consisted of long-chain triglycerides, a high amount of this fat in the elemental diet formula decreased its therapeutic effect against active Crohns disease.

Interleukin-18 overproduction exacerbates the development of colitis with markedly infiltrated macrophages in interleukin-18 transgenic mice

Background and Aim:  The authors have previously shown that production of interleukin (IL)‐18 was increased in the inflamed mucosa of patients with Crohns disease (CD) and blockade of IL‐18 ameliorated the murine model of CD. This demonstrated that IL‐18 plays a significant role during intestinal inflammation. However, the initial role of IL‐18 during intestinal inflammation was unclear; therefore the susceptibility of IL‐18 transgenic (Tg) mice to acute dextran sulfate sodium (DSS)‐induced colitis was examined.

Endoscopic and histopathological study on primary and secondary intestinal lymphangiectasia

Jejunal endoscopy and histopathological study of biopsied specimens were performed to clarify states of jejunal mucosa and the mechanism of enteric protein loss in six patients with protein-losing enteropathy, including four patients with intestinal lymphangiectasia, one patient with constrictive pericarditis associated with dilated lymphatics of the intestine, and one patient with Budd-Chiari syndrome. Three cardinal endoscopic findings, scattered white spots, white villi, and chyle-like substances covering the mucosa, were demonstrated in protein-losing enteropathy. Scattered white spots indicated markedly dilated lymphatics in the stroma of the villi. White villi seemed to be due to fats including chylomicrons or fat droplets in the absorptive cells, interepithelial spaces, and/or stroma, even though the biopsies were obtained in the fasting state. Therefore, white villi suggest impaired transport of fats from intestinal epithelial cells to intestinal lymphatics. These three cardinal findings are thought to be characteristic for protein-losing enteropathy secondary to lymphatic disorders.

Augmented levels of gastric mucosal leucocyte activation by infection with cagA gene‐positive Helicobacter pylori

The possession of the cytotoxin‐associated gene (cagA) of Helicobacter pylori is thought to be highly associated with peptic ulcer disease. However, the pathogenic role of cagA is still unknown. We have emphasized the importance of the interrelationship between H. pylori‐derived ammonia and oxygen radicals from infiltrated leucocytes. The aim of the present study was to explore the relationship between oxygen radical production and H. pylori strain diversity based on cagA possession. An endoscopic examination and gastric mucosal biopsy were performed in 30 H. pylori‐infected patients with gastric ulcer. Myeloperoxidase (MPO) content and the luminol‐dependent chemiluminescence value in the biopsied gastric specimens were measured as an index for leucocyte infiltration and oxygen radical production. From the precipitates of cultured bacterial isolates of biopsied specimens, bacterial DNA was purified and analysed by polymerase chain reaction to characterize the possession of cagA. While all patients had ureC‐positive strains, 22 had cagA‐positive and eight had cagA‐negative strains. In patients with cagA‐positive strains, MPO contents as well as chemiluminescence values in the gastric corpus were significantly higher than those in patients with cagA‐negative strains. Gastric mucosal leucocyte recruitment and activation are suggested to be enhanced by cagA gene‐positive H. pylori infection.

Active oxygen species in formation of acute gastric mucosal lesions induced by thermal injury in rats

Active oxygen species generated by circulating leukocytes and released from the gastric mucosa were measured in the process of acute gastric mucosal lesion formation after thermal injury in rats. Alterations of luminol-dependent chemiluminescence activities generated by leukocytes obtained from the gastric vein and the inferior vena cava were approximately same. A decrease in chemiluminescence activity 15 min after thermal injury and a significant increase in chemiluminescence activity 5 hr after thermal injury were observed in leukocytes from both veins. From 15 min to 12 hr after thermal injury, luciferin-dependent chemiluminescence activities were significantly higher than that of the control group. Oral administration of rebamipide resulted in decreased mucosal lesion formation. Rebamipide, an antiulcer agent that protects the mucosa from damage in various animal models decreased chemiluminescence activities only released from the gastric mucosa but not from circulating leukocytes. These results suggest that two different pathways of active oxygen species formation may exist in the pathogenesis of acute gastric mucosal lesions after thermal injury.

Helicobacter pylori‐eradication therapy decreases the level of neutrophil‐derived oxidants in the ulcerous mucosa of the human stomach: Relationship between ulcer stage and mucosal oxidant level

Background:  The purpose of the present study was to determine the effect of Helicobacter pylori eradication on the intensity of inflammation in the ulcerous mucosa by measuring the level of neutrophil‐associated oxidants and to understand the role of mucosal inflammation in ulcer relapse from the viewpoints of the scarring stage and the H. pylori‐infection status. The level of inflammation in the gastric mucosa after the eradication of H. pylori was examined using biopsy samples obtained from patients with gastric ulcers.

Reactive Oxygen Species and Colonic Epithelial Cell Apoptosis in Ulcerative Colitis

Ulcerative colitis (UC) is one of the inflammatory bowel diseases of unknown etiology characterized by its localization to the large intestine. According to pathological examination, which reveals mucosal inflammatory cell infiltration, goblet cell depletion, and distortion of crypt architecture, it is clear that the target of this disease is epithelial cells. Some reports note that epithelial cell damage is mediated by apoptotic cell death. The aim of this study was to elucidate the role of reactive oxygen species (ROS) in the induction of colonic epithelial cell apoptosis. Colonie biopsies from active and inactive lesions of UC patients and from normal tissues of patients with colon polyps were used. ROS production from the colonic tissues were measured using a chemiluminescence assay. Apoptotic cells were detected by the TUNEL technique. Superoxide dismutase (SOD), Fas, and Bcl-2 were detected by immunohistochemistry. ROS production was increased in active UC compared with inactive UC or normal mucosa. SOD was strongly expressed in the epithelial cells of active UC compared with inactive UC mucosa. The extent of TUNEL-positive epithelial cells was more extensive in active UC compared with inactive UC or nomal mucosa. There was a positive correlation between ROS production and the extent of TUNEL-positive cells. There was a tendency for Fas-positive cells to be fewer in active and inactive UC compared with normal mucosa, but there was no clear tendency for Bcl-2-positive cells. Based on these results, ROS rather than the Fas/Fas ligand system is suggested to have an important role in inducing epithelial cell apoptosis in UC.