Shiho Konno

Wound healing of intestinal epithelial cells.

The intestinal epithelial cells (IECs) form a selective permeability barrier separating luminal content from underlying tissues. Upon injury, the intestinal epithelium undergoes a wound healing process. Intestinal wound healing is dependent on the balance of three cellular events; restitution, proliferation, and differentiation of epithelial cells adjacent to the wounded area. Previous studies have shown that various regulatory peptides, including growth factors and cytokines, modulate intestinal epithelial wound healing. Recent studies have revealed that novel factors, which include toll-like receptors (TLRs), regulatory peptides, particular dietary factors, and some gastroprotective agents, also modulate intestinal epithelial wound repair. Among these factors, the activation of TLRs by commensal bacteria is suggested to play an essential role in the maintenance of gut homeostasis. Recent studies suggest that mutations and dysregulation of TLRs could be major contributing factors in the predisposition and perpetuation of inflammatory bowel disease. Additionally, studies have shown that specific signaling pathways are involved in IEC wound repair. In this review, we summarize the function of IECs, the process of intestinal epithelial wound healing, and the functions and mechanisms of the various factors that contribute to gut homeostasis and intestinal epithelial wound healing.

Ecabet sodium prevents the delay of wound repair in intestinal epithelial cells induced by hydrogen peroxide

BackgroundRecent studies showed that ecabet sodium (ES), a gastro-protective agent, also had a therapeutic effect on inflammation in ulcerative colitis. The aim of this study was to clarify the function of ES in wound repair in intestinal epithelial cells (IECs).MethodsThe activation of signal proteins (ERK1/2 mitogen-activated protein kinase MAPK, and IκB-α) in IEC-6 cells after stimulation with 2.5 mg/ml of ES was assessed by Western blot. The induction of transforming growth factor (TGF)-β1, TGF-α, and cyclooxygenase-2 (COX-2) mRNAs after the stimulation of IEC-6 cells with ES was assessed by reverse transcription ploymerase chain reaction (RT-PCR). IEC-6 cells were wounded and cultured for 24 h with various concentrations of ES in the absence or presence of 20 µM H2O2. Epithelial migration or proliferation was assessed by counting migrated or bromodeoxyuridine (BrdU)-positive cells observed across the wound border. We also assessed apoptotic epithelial cells after the culture.ResultsES clearly activated ERK1/2 MAPK and slightly activated IκB-α. ES also enhanced the expression of TGF-α and COX-2 mRNAs. This enhancement was suppressed by a MAPK/Erk kinase (MEK) inhibitor. ES did not enhance epithelial migration in the absence of H2O2. In contrast, ES significantly decreased the number of apoptotic cells and prevented the reduction of epithelial migration (51.1%; P < 0.01) and proliferation (56%; P < 0.01) induced by H2O2. The function of ES was suppressed by a cyclooxygenase-2 (COX-2) inhibitor and by the MEK inhibitor, and partly suppressed by a nuclear factor (NF)-κB inhibitor.ConclusionsES prevents the delay of wound repair in IEC-6 cells induced by H2O2, probably through the activation of ERK1/2 MAPK and the induction of COX-2.

Rotavirus double-stranded RNA induces apoptosis and diminishes wound repair in rat intestinal epithelial cells

Background:  Recent studies have shown that toll‐like receptor 3 (TLR3) recognizes double‐stranded RNA (dsRNA). Rotaviruses, having a dsRNA genome, infect intestinal epithelial cells (IEC) and cause acute gastroenteritis in young children. The aim of the present study was to clarify the pathophysiological function of rotavirus dsRNA in IEC.

Altered expression of angiogenic factors in the VEGF-Ets-1 cascades in inflammatory bowel disease

BackgroundThe VEGF-Ets-1 cascades play important roles in angiogenesis by converting endothelial cells to an angiogenic phenotype. The aim of this study was to clarify whether the VEGF-Ets-1 cascades are involved in the pathogenesis of inflammatory bowel disease (IBD).MethodsColonic specimens were taken from 42 patients with ulcerative colitis (UC), 37 with Crohn’s disease (CD), 8 with non-IBD colitis, and 21 normal controls. (1) Expression of vascular endothelial growth factor (VEGF), VEGF receptors (Flt-1, KDR), and Ets-1 proteins in colonic mucosa was immunohistochemically examined using specific antibodies. (2) Expression of Ets-1 protein or VEGF, Flt-1, KDR, and Ets-1 mRNA in colonic mucosa was measured by Western blot or RT-PCR.Results(1) The number of VEGF-containing cells was significantly increased in active UC (P < 0.05). The numbers of positive blood vessels (mean ± SE /mm2) to Flt-1, KDR, and Ets-1 antibodies were significantly increased in active UC (Flt-1: 4.0 ± 0.84; KDR: 2.4 ± 0.37; Ets-1: 5.5 ± 0.77) compared to active CD (Flt-1: 0.6 ± 0.30; KDR: 0.77 ± 0.28; Ets-1: 2.0 ± 0.56) (P < 0.01), non-IBD colitis (Flt-1: 1.0 ± 0.45; KDR: 1.83 ± 0.54; Ets-1: 3.0 ± 1.0), and controls (Flt-1: 0.88 ± 0.40; KDR: 0.60 ± 0.22; Ets-1: 1.67 ± 0.47) (P < 0.01). The numbers of positive cells to these antibodies were also increased in active UC. (2) Expression of Ets-1 protein and Flt-1, KDR, and Ets-1 mRNA was increased in active UC.ConclusionsAngiogenic factors in the VEGF-Ets-1 cascades were upregulated in UC, but they were relatively downregulated in CD. These alterations might be involved in the pathogenesis of both diseases.

Infliximab as a treatment for systemic amyloidosis associated with Crohn's disease

Systemic AA amyloidosis is a serious and potentially fatal complication of Crohn’s disease.1 This serious complication is associated with approximately 0.9–2.5% of patients with Crohn’s disease but renal failure due to renal amyloidosis is one of the most common causes of death for patients with Crohn’s disease.2 Although drugs such as azathioprine, colchicines, and dimethylsulphoxide are suggested to delay the progression of renal amyloidosis, the efficacy of these drugs on renal amyloidosis has not been fully elucidated.2 Infliximab is a chimeric anti-tumour necrosis factor α (TNF-α) monoclonal antibody. This drug has been proven …

Novel Evidence Suggesting Clostridium difficile Is Present in Human Gut Microbiota More Frequently than Previously Suspected

Prevalence rate of Clostridium difficile in healthy human adults is believed to be very low. Our RT‐PCR system using glass powder, which can eliminate PCR inhibitors, detected C. difficile toxin B mRNA in 16 of 30 fecal samples (53.3%) from healthy human adults. In contrast, we failed to detect toxin B in the same fecal samples by PCR using DNA templates extracted with phenol‐chloroform. Our results suggest that PCR inhibitors in feces carried through phenol‐chloroform extraction procedure might suppress the sensitivity of PCR and that C. difficile is actually present in human gut microbiota more frequently than previously suspected.

Altered expression of epimorphin in ulcerative colitis.

Background and Aim: Epimorphin is suggested to play a key role in the morphogenesis of epithelial cells. We focused on epimorphin and attempted to clarify the relationship between expression of this molecule and inflammatory bowel disease (IBD).

Efficacy of ecabet sodium enema on steroid resistant or steroid dependent ulcerative colitis

Ecabet sodium (ES), a 12-sulfodehydroabietic acid monosodium salt derived from an ingredient in pine resin, has been shown to topically enhance several gastric mucosal defensive factors.1 ES is widely used for the treatment of gastritis and gastric ulcer in Japan. A recent open label study demonstrated that ES also has therapeutic effects in active ulcerative colitis (UC).2 In that study, the authors demonstrated that six of seven UC patients with proctosigmoiditis achieved complete remission by treatment with ES enema. However, successive clinical trials of the efficacy of ES enema in UC patients have not been reported. Here we demonstrate the efficacy of ES enema in steroid resistant or steroid dependent UC patients. …

Sucralfate prevents the delay of wound repair in intestinal epithelial cells by hydrogen peroxide through NF-κB pathway

BackgroundRecent studies have shown that sucralfate (SF) has therapeutic effects on colonic inflammation in ulcerative colitis. The aim of this study was to clarify the function of SF for wound repair in intestinal epithelial cells (IEC).Methods(1) Activation of signal proteins [ERK1/2 mitogen-activated protein kinase (MAPK), IκB-α] in IEC-6 cells after stimulation with 10−4 M potassium sucrose octasulfate (SOS), which is the functional element of SF, was assessed by Western blot. (2) Induction of transforming growth factor (TGF)-β1, TGF-α, EGF, and cyclooxygenase-2 (COX-2) mRNA after stimulation of IEC-6 cells with SOS was assessed by reverse transcriptase-polymerase chain reaction. (3) IEC-6 cells were wounded and cultured for 24 h with various concentrations of SOS in the absence or presence of 20 μM H2O2. Epithelial migration or proliferation was assessed by counting migrating cells or bromodeoxyuridine (BrdU)-positive cells across the wound border.Results(1) SOS activated IκB-α, but it did not activate ERK1/2 MAPK. (2) SOS enhanced the expression of COX-2 mRNA, but it did not change the mRNA expression of other growth factors. (3) SOS did not enhance wound repair in IEC-6 cells, but it decreased the number of dead cells (maximum, 74%) (P < 0.01) in a dose-dependent manner and prevented the diminishment of epithelial migration (maximum, 61%) (P < 0.01) and proliferation (maximum, 37%) (P < 0.05) induced by H2O2. These functions of SOS were suppressed by the NF-κB and COX-2 inhibitors.ConclusionsSOS prevented the delay of wound repair in IEC-6 cells induced by H2O2, probably through induction of COX-2 and an anti-apoptotic mechanism. These effects of SOS might be given through the activation of the NF-κB pathway.

A case of pseudoobstruction of the intestine associated with scleroderma dramatically responding to antibiotics.

To the Editor: Scleroderma is a systemic disease of unknown etiology characterized by excessive deposition of collagen and other connective tissue components in the skin and multiple organs.1,2 Scleroderma is commonly complicated by gastrointestinal involvement, which is characterized by dilatation of the gastrointestinal tract and reduction of peristalsis, sometimes resulting in pseudoobstruction of the intestine.3 Because pseudoobstruction is intractable and relapses are frequent despite careful treatment, it is difficult to manage patients with pseudoobstruction. Malnutrition due to pseudoobstruction has been reported to be the primary causes of death in scleroderma.3 Thus, it is very important to manage the pseudoobstruction and rescue such patients. Here, we describe a patient with intractable pseudoobstruction of the intestine associated with scleroderma, who significantly responded to treatment with antibiotics. In April 2001, a 65-year-old man visited our hospital with swelling of the skin of the fingers and breast. He was diagnosed as having scleroderma on the basis of findings of macroscopic and microscopic sclerosis cutanea and Raynaud’s phenomenon. In September 2004, he became conscious of abdominal fullness, vomiting, and inability to defecate. He visited another hospital, which suggested that he suffered from pseudoobstruction of the intestine associated with scleroderma. He underwent decompression with a long intestinal tube and total parenteral nutrition with administration of metoclopramide. His clinical symptoms of the patient soon improved, but he experienced frequent relapses of pseudoobstruction. Thus, he was referred to our hospital on 4 December 2004. The physical examination on admission revealed abdominal distension, a decrease in bowel sounds, and stiffness and swelling of the skin of the hand and foot. Hematologic and serum chemistry showed hypoalbuminemia (3.0g/dl) and anemia (RBC, 245 × 104/μl, Hb 7.0 g/dl). Antinuclear antibody was strongly positive (1 : 320), with a speckled pattern. Antiscleroderma 70 antibody and anticentromere antibody were also positive (1 : 22.7 and 1 : 35.1, respectively). Fecal culture showed normal gut flora and no pathogenic bacteria. The plain film of the abdomen on admission demonstrated significantly dilated loops of small intestine with air–fluid levels (Fig. 1A). Abdominal ultrasonography also showed dilatation of loops of the small bowel. Endoscopic examination of the upper gastrointestinal tract and the colon and terminal ileum showed no obstructive or inflammatory lesions. The small intestine series demonstrated significant dilatation and hypomotility of the small intestine, but no mechanical obstruction was found (Fig. 1B,C). Based on these findings, we confirmed the diagnosis of pseudoobstruction of the intestine associated with scleroderma. Therefore, the patient was treated with dimeticon and a Chinese herbal drug (dai-kenchu-to; Tsumura, Tokyo, Japan), but the abdominal symptoms and X-ray findings did not improve. Taking into account the association of bacterial overgrowth with his symptoms, we added a treatment with kanamycin and lactulose. In response, the abdominal symptoms of the patient soon improved. However, abdominal symptoms recurred after 2 weeks. We changed antibiotics from kanamycin to metronidazole on the 28th hospital day because Lock et al.4 recommended treatment with metronidazole for pseudoobstruction associated with scleroderma. After the treatment with metronidazole (750 mg/day), the abdominal symptoms of the patient dramatically improved in 3 days, and the X-ray findings also significantly improved (Fig. 1D). He started oral ingestion of food on the 47th hospital day, but abdominal symptoms did not recur. He was discharged from our hospital on the 65th hospital day. Pseudoobstruction in the patient recurred 6 months later, when levofloxacin (300 mg/day) significantly improved his abdominal symptoms. This case suggests that bacterial overgrowth in the intestine might be closely involved with abdominal symptoms on the part of patients with intractable pseudoobstruction associated with scleroderma. For such patients, treatment with specific antibiotics such as metronidazole or new quinolones might improve their abdominal symptoms even when pathogenic bacteria cannot be identified in the patient’s feces.