Naoki Minami

Analysis of Gene Expression in Mouse 2-Cell Embryos Using Fluorescein Differential Display: Comparison of Culture Environments

Abstract The effect of the oviductal environment on gene expression in 2-cell mouse embryos was examined with mRNA differential display. Embryos used for experiments were cultured in modified Whitten medium with or without oviductal tissue until late 2-cell stage. The results of sequencing indicated that the genes for ATP synthase (ATPase 6), S-adenosylmethionine decarboxylase (S-AMDC) and nuclear autoantigenic sperm protein (NASP) were differentially expressed in embryos cultured in the oviductal environment (nonblocking culture condition). The ATPase 6 gene is encoded by mitochondrial DNA and is essential for the production of ATP. This indicates that the expression of ATP synthesis-related genes at the 2-cell stage may be required to maintain normal development in vitro. S-Adenosylmethionine decarboxylase decarboxylates adenosylmethionine, which is a substrate of DNA methylation. The expression of S-AMDC may be responsible for the low level of methylation of preimplantation development. As NASP is a histone-binding protein that is thought to be testis and sperm specific, its function in embryos remains unclear. On the other hand, the Tcl1 gene and a novel gene, the c-1 gene, were strongly expressed in embryos cultured without oviductal tissue (blocking culture condition). The expression patterns of these genes are quite similar. However, the detailed functions of these genes in embryos remain to be determined.

Diagnosis and Treatment of Ulcerative Colitis with Cytomegalovirus Infection: Importance of Controlling Mucosal Inflammation to Prevent Cytomegalovirus Reactivation

Human cytomegalovirus (HCMV) is a member of the herpesvirus family. HCMV infection persists throughout the host lifespan in a latent state following primary infection. The ability of HCMV to escape control by the host immune system and its resulting reactivation suggests the importance of ongoing immune surveillance in the prevention of HCMV reactivation. HCMV is a common cause of opportunistic infection that causes severe and fatal disease in immune-compromised individuals. In inflammatory bowel disease patients, particularly those with ulcerative colitis (UC), HCMV is often reactivated because these patients are frequently treated with immunosuppressive agents. This reactivation exacerbates colitis. Additionally, HCMV infection can induce severe colitis, even in patients with UC who have never been treated with immunosuppressive agents. However, the role of HCMV in colonic inflammation in patients with UC remains unclear. Here, we present previous and current clinical data on the diagnosis and treatment of HCMV infection in UC. Additionally, our experimental data from a newly established mouse model mimicking UC with concomitant CMV infection clearly demonstrate that inflammation could result in the exacerbation of UC disease activity with induction of HCMV reactivation. In summary, optimal control of colonic inflammation should be achieved in UC patients who are refractory to conventional immunosuppressive therapies and are positive for HCMV.

Efficacy and safety of granulocyte and monocyte adsorption apheresis for ulcerative colitis: A meta-analysis

BACKGROUND Safe and effective treatments are required for patients with ulcerative colitis. It was suggested that granulocyte and monocyte adsorption apheresis might play an important role for ulcerative colitis. Therefore, a meta-analysis was performed. METHODS Medline and the Cochrane controlled trials register were used to identify randomized controlled trials comparing granulocyte and monocyte adsorption apheresis with corticosteroids, and comparing intensive with conventional apheresis in patients with ulcerative colitis. RESULTS Nine randomized trials were eligible for inclusion criteria. According to pooled data, granulocyte and monocyte adsorption apheresis is effective for inducing clinical remission in patients with ulcerative colitis compared with corticosteroids (odds ratio, 2.23; 95% confidence interval: 1.38-3.60). However, the efficacy of granulocyte and monocyte adsorption apheresis was not dependent on the number of apheresis sessions. The intensive apheresis (≥2 sessions per week) is more effective for inducing clinical remission than weekly apheresis (odds ratio, 2.10; 95% confidence interval: 1.12-3.93). The rate of adverse events by apheresis was significantly lower than that by corticosteroids (odds ratio, 0.24; 95% confidence interval: 0.15-0.37). CONCLUSION Our meta-analysis reveals that intensive granulocyte and monocyte adsorption apheresis is a safe and effective treatment with higher rates of clinical remission and response for ulcerative colitis compared with corticosteroids.

Refractoriness of Intestinal Behçet's Disease with Myelodysplastic Syndrome Involving Trisomy 8 to Medical Therapies - Our Case Experience and Review of the Literature

Background/Aims: Gastrointestinal lesions of Behçets disease (BD) are often refractory to medical therapy and sometimes result in serious comorbidities such as gastrointestinal perforation and massive bleeding. There are several reports of patients with BD comorbid with myelodysplastic syndrome (MDS) involving trisomy 8 that frequently have intestinal lesions refractory to conventional medical therapy. Little is known about the efficacy of infliximab (IFX) for these intestinal lesions. Methods: We present 2 cases of intestinal BD with MDS involving trisomy 8 who did not respond to IFX, and review previous reports of BD with MDS involving trisomy 8 concerning their responsiveness to conventional medical therapy. Results: Among 31 previously reported cases that received medical treatment for BD, 19 (61.3%) showed temporary improvement of the BD symptoms, 9 (29.0%) deteriorated, and 3 (9.7%) showed no change. All of the 9 cases that showed deterioration had intestinal lesions. Our 2 cases failed to respond to IFX, resulting in a poor prognosis. Conclusions: IFX might not be effective for improving intestinal BD comorbid with MDS involving trisomy 8. Trisomy 8 is associated with the BD prognosis and refractoriness to conventional medical therapy.

Tacrolimus or infliximab for severe ulcerative colitis: short-term and long-term data from a retrospective observational study

Objective Treatment of severe ulcerative colitis (UC) is challenging. Although the efficacy of tacrolimus (TAC) and infliximab (IFX) have been evaluated in patients with severe UC, the safety and efficacy levels of sequential therapies (TAC→IFX/IFX→TAC) in these patients remain unclear. The aim of this study was to assess short-term and long-term outcomes in patients with severe UC treated with TAC and IFX. Methods From October 2001 to February 2014, 29 patients with consecutive severe UC treated with TAC or IFX were retrospectively evaluated. Median follow-up duration was 27 months (range 0.5–118 months). The primary end point was short-term outcomes at 8 weeks after induction of TAC (TAC group, n=22) or IFX (IFX group, n=7). The secondary end point included long-term outcomes and colectomy-free survival. The clinical response was evaluated based on a partial Mayo score. Results The clinical remission (CR) rate at 8 weeks in the TAC and IFX groups was 63.6% and 71.4%, respectively. In 13 of the 29 patients (10 in the TAC group, 3 in the IFX group), sequential therapies were used in their clinical courses. In 9 of these 13 patients (6 in the TAC group, 3 in the IFX group), CR was achieved and maintained by sequential therapies. Overall cumulative colectomy-free survival was 79.3% at 118 months. Conclusions TAC and IFX had similar effects on remission induction in patients with severely active UC. Sequential therapies could rescue patients with UC who failed initial treatment with TAC or IFX. In clinical practice, sequential therapies might be deliberately performed.

Osteopontin Deficiency Accelerates Spontaneous Colitis in Mice with Disrupted Gut Microbiota and Macrophage Phagocytic Activity.

Background Osteopontin (OPN) is a multifunctional protein expressed in a variety of tissues and cells. Recent studies revealed increased OPN expression in the inflamed intestinal tissues of patients with inflammatory bowel disease (IBD). The role of OPN in the pathophysiology of IBD, however, remains unclear. Aims To investigate the role of OPN in the development of intestinal inflammation using a murine model of IBD, interleukin-10 knock out (IL-10 KO) mice. Methods We compared the development of colitis between IL-10 KO and OPN/IL-10 double KO (DKO) mice. OPN expression in the colonic tissues of IL-10 KO mice was examined by fluorescence in situ hybridization (FISH) analysis. Enteric microbiota were compared between IL-10 KO and OPN/IL-10 DKO mice by terminal restriction fragment length polymorphism analysis. The effect of OPN on macrophage phagocytic function was evaluated by phagocytosis assay. Results OPN/IL-10 DKO mice had an accelerated onset of colitis compared to IL-10 KO mice. FISH analysis revealed enhanced OPN synthesis in the colonic epithelial cells of IL-10 KO mice. OPN/IL-10 DKO mice had a distinctly different enteric bacterial profile with a significantly lower abundance of Clostridium subcluster XIVa and a greater abundance of Clostridium cluster XVIII compared to IL-10 KO mice. Intracellular OPN deletion in macrophages impaired phagocytosis of fluorescence particle-conjugated Escherichia coli in vitro. Exogenous OPN enhanced phagocytosis by OPN-deleted macrophages when administered at doses of 1 to 100 ng/ml, but not 1000 ng/ml. Conclusions OPN deficiency accelerated the spontaneous development of colitis in mice with disrupted gut microbiota and macrophage phagocytic activity.

Efficacy and Safety of Long-Term Thiopurine Maintenance Treatment in Japanese Patients With Ulcerative Colitis

Background/Aims The long-term clinical outcomes of patients with bio-naive ulcerative colitis (UC) who maintain remission with thiopurine are unclear. The aim of this study was to assess the long-term efficacy and safety of maintenance treatment with thiopurine in UC patients. Methods This was a retrospective observational cohort analysis conducted at a single center. Between December 1998 and August 2013, 59 of 87 patients with bio-naive UC who achieved remission after induction with treatments other than biologics were enrolled. Remission maintenance with thiopurine was defined as no concomitant treatment needed other than 5-aminosalicylate without relapse. We assessed the remission-maintenance rate, mucosal healing rate, colectomy-free rate, and treatment safety in UC patients who received thiopurine as maintenance treatment. Results The 84-month cumulative remission-maintenance and colectomy-free survival rates in the UC patients who were receiving maintenance treatment with thiopurine and 5-aminosalicylate were 43.9% and 88.0%, respectively. Of the 38 patients who underwent colonoscopy during thiopurine maintenance treatment, 23 (60.5%) achieved mucosal healing. Of the 59 patients who achieved clinical remission with thiopurine, 6 patients (10.2%) discontinued the thiopurine therapy because of adverse events. Conclusions Our study demonstrates the long-term efficacy and safety of thiopurine treatment in patients with bio-naive UC.

Lipocalin 2 prevents intestinal inflammation by enhancing phagocytic bacterial clearance in macrophages

Lipocalin 2 (Lcn2), also called neutrophil gelatinase B-associated lipocalin (NGAL), is an anti-microbial peptide originally identified in neutrophil granules. Although Lcn2/NGAL expression is increased in the inflamed intestinal tissues of patients with inflammatory bowel disease, the role of Lcn2/NGAL in the development of intestinal inflammation remains unclear. Here we investigated the role of Lcn2/NGAL in intestinal inflammation using a spontaneous mouse colitis model, interleukin-10 knock out (IL-10 KO) mice. Lcn2 expression in the colonic tissues of IL-10 KO mice increased with the development of colitis. Lcn2/IL-10 double-KO mice showed a more rapid onset and development of colitis compared to IL-10 KO mice. Lcn2 enhanced phagocytic bacterial clearance in macrophages in vitro after infection with Escherichia coli. Transfer of Lcn2-repleted macrophages prevented the development of colitis in Lcn2/IL-10 double-KO mice in vivo. Our findings revealed that Lcn2 prevents the development of intestinal inflammation. One crucial factor seems to be the enhancement of phagocytic bacterial clearance in macrophages by Lcn2.

Unique endoscopic findings of colitis-associated colorectal cancer in a patient with ulcerative colitis and Lynch syndrome

Dear Sir, A 28-year-old woman with coexistent ulcerative colitis (UC) and systemic lupus erythematosus had been treated with prednisolone and salazosulfapyridine. She underwent colonoscopy to evaluate her UC. Endoscopic examination revealed severe colonic inflammation with deep ulcers in the rectum and mild inflammation in the sigmoid colon. Moreover, colonic polyps were observed throughout the entire colon. A semipedunculated polyp (0-Isp) with a central depression, whose endoscopic findings mimicked an inflammatory polyp, was observed at the hepatic flexure (Fig. 1A). Histologic findings of biopsy specimens obtained from this polyp showed signet ring cell carcinoma with …

Ischemic enteritis with intestinal stenosis.

A 75-year-old man was admitted to our hospital with sudden onset of vomiting and abdominal distension. The patient was taking medication for arrhythmia. Computed tomography showed stenosis of the ileum and a small bowel dilatation on the oral side from the region of stenosis. A transnasal ileus tube was placed. Enteroclysis using contrast medium revealed an approximately 6-cm afferent tubular stenosis 10 cm from the terminal ileum and thumbprinting in the proximal bowel. Transanal double-balloon enteroscopy showed a circumferential shallow ulcer with a smooth margin and edema of the surrounding mucosa. The stenosis was so extensive that we could not perform endoscopic balloon dilation therapy. During hospitalization, the patients nutritional status deteriorated. In response, we surgically resected the region of stenosis. Histologic examination revealed disappearance of the mucosal layer and transmural ulceration with marked fibrosis, especially in the submucosal layer. Hemosiderin staining revealed sideroferous cells in the submucosal layers. Based on the pathologic findings, the patient was diagnosed with ischemic enteritis. The patients postoperative course was uneventful.