Yoshikazu Kinoshita

Architectural roles of multiple chromatin insulators at the human apolipoprotein gene cluster

Long‐range regulatory elements and higher‐order chromatin structure coordinate the expression of multiple genes in cluster, and CTCF/cohesin‐mediated chromatin insulator may be a key in this regulation. The human apolipoprotein (APO) A1/C3/A4/A5 gene region, whose alterations increase the risk of dyslipidemia and atherosclerosis, is partitioned at least by three CTCF‐enriched sites and three cohesin protein RAD21‐enriched sites (two overlap with the CTCF sites), resulting in the formation of two transcribed chromatin loops by interactions between insulators. The C3 enhancer and APOC3/A4/A5 promoters reside in the same loop, where the APOC3/A4 promoters are pointed towards the C3 enhancer, whereas the APOA1 promoter is present in the different loop. The depletion of either CTCF or RAD21 disrupts the chromatin loop structure, together with significant changes in the APO expression and the localization of transcription factor hepatocyte nuclear factor (HNF)‐4α and transcriptionally active form of RNA polymerase II at the APO promoters. Thus, CTCF/cohesin‐mediated insulators maintain the chromatin loop formation and the localization of transcriptional apparatus at the promoters, suggesting an essential role of chromatin insulation in controlling the expression of clustered genes.

Bile acids directly augment caudal related homeobox gene Cdx2 expression in oesophageal keratinocytes in Barrett’s epithelium

Background and aims: The mechanism of transformation to intestinal metaplasia in Barrett’s oesophagus has not been clarified. We investigated the effects of various bile acids on expression of the caudal related homeobox gene Cdx2 in cultured oesophageal squamous epithelial cells. In addition, morphological and histochemical changes in squamous cells to intestinal epithelial cells were studied in response to bile acid induced expression of Cdx2. Methods: A rat model of Barrett’s oesophagus was created by anastomosing the oesophagus and jejunum, and Cdx2 expression was investigated by immunohistochemistry. Also, the response of various bile acids on Cdx2 gene expression was studied in the human colon epithelial cell lines Caco-2 and HT-29, as well as in cultured rat oesophageal squamous epithelial cells using a Cdx2 promoter luciferase assay. In addition, primary cultured oesophageal squamous epithelial cells were transfected with Cdx2 expression vectors and their possible transformation to intestinal-type epithelial cells was investigated. Results: Oesophagojejunal anastomoses formed intestinal goblet cell metaplasia in rat oesophagus specimens and metaplastic epithelia strongly expressed Cdx2. When the effects of 11 types of bile acids on Cdx2 gene expression were examined, only cholic acid (CA) and dehydrocholic acid dose dependently increased Cdx2 promoter activity and Cdx2 protein production in Caco-2 and HT-29 cells, and cultured rat oesophageal keratinocytes. Results from mutation analysis of Cdx2 promoter suggested that two nuclear factor κB (NFκB) binding sites were responsible for the bile acid induced activation of the Cdx2 promoter. When bile acids were measured in oesophageal refluxate of rats with experimental Barrett’s oesophagus, the concentration of CA was found to be consistent with the experimental dose that augmented Cdx2 expression in vitro. Furthermore, transfection of the Cdx2 expression vector in cultured rat oesophageal keratinocytes induced production of intestinal-type mucin, MUC2, in cells that expressed Cdx2. Conclusions: We found that CA activates Cdx2 promoter via NFκB and stimulates production of Cdx2 protein in oesophageal keratinocytes with production of intestinal-type mucin. This may be one of the mechanisms of metaplasia in Barrett’s oesophagus.

Regenerating gene protein may mediate gastric mucosal proliferation induced by hypergastrinemia in rats.

BACKGROUND & AIMS Regenerating gene (Reg) has been isolated from rat regenerating pancreatic islets, and Reg protein is mitogenic to islet cells. We have recently shown that Reg gene and Reg protein are expressed in gastric enterochromaffin-like (ECL) cells. This study aimed to clarify whether gastrin enhances Reg protein production in ECL cells and whether Reg protein is mitogenic to gastric mucosal cells. METHODS Reg gene expression in response to acute and chronic hypergastrinemia was investigated in rats. Immunohistochemical studies, Northern blotting, and in situ hybridization were performed to investigate the expression of Reg protein and Reg gene. The direct effect of gastrin on Reg gene expression was investigated using isolated ECL cells, and the trophic effect of Reg protein on cultured gastric epithelial cells was assessed by [3H]thymidine uptake. RESULTS Both chronic hypergastrinemia and short-term gastrin administration stimulated Reg gene expression and Reg protein production in fundic mucosa. Reg gene expression was also augmented in isolated ECL cells after incubation with rat gastrin. Reg protein was mitogenic to cultured rat gastric epithelial cells. CONCLUSIONS Gastrin stimulates the production of Reg protein in gastric ECL cells, which may be involved in the gastrin-induced gastric mucosal cell growth.

CYP2C19 genotype status and intragastric pH during dosing with lansoprazole or rabeprazole

CYP2C19 has an important role in the catabolism of several proton pump inhibitors. However, the relative contribution of CYP2C19‐mediated metabolism varies among the different proton pump inhibitors.

Essential Role of MD-2 in TLR4-Dependent Signaling during Helicobacter pylori-Associated Gastritis

TLR4, a member of pattern recognition receptors, is the main receptor of LPS. MD-2 physically associates with TLR4 on the cell surface and confers LPS responsiveness. Helicobacter pylori LPS is one of the major virulence factors for induction of gastritis. We demonstrated in this study the role of MD-2 in TLR4-dependent signaling in H. pylori-associated gastritis. Gastric biopsy samples collected from patients with and without H. pylori infection and four gastric cancer cell lines were used for this study. TLR-4 and MD-2 expression in biopsy specimens and the cell lines was examined by using RT-PCR. Localization of TLR-4 in histological sections was evaluated by immunohistochemistry. For in vitro functional assays, we established stable transfectants of AGS cells expressing TLR4 and MD-2. Cellular distribution of TLR4 was examined by flow cytometry. NF-κB activation and activation of IL-8 and MD-2 promoters were assessed by reporter gene assay. H. pylori infection up-regulated the TLR4 and MD-2 expression in gastric mucosa. TLR4 staining was observed predominantly in epithelial cells, located in both the cytoplasm and at the apical surface. MD-2 transfection in AGS cells markedly increased cell surface expression of TLR4 and augmented the activation of NF-κB and IL-8 promoter upon stimulation with H. pylori LPS. Live H. pylori also stimulated transcriptional activation of MD-2. This study revealed that MD-2 expression is elevated in gastric epithelial cells during H. pylori infection, suggesting that the TLR4/MD-2 system is a potent receptor complex involved in the response to H. pylori LPS in the stomach.

Predominant nocturnal acid reflux in patients with Los Angeles grade C and D reflux esophagitis

Nocturnal gastric acid breakthrough (NAB) is defined as an intragastric pH < 4.0 lasting more than 1 h during the night in patients taking a proton pump inhibitor (PPI). Gastroesophageal reflux disease (GERD) patients with nocturnal gastroesophageal acid reflux accompanied by NAB are thought to be refractory to PPI treatment. The aim of this study was to endoscopically identify the patients with predominant nocturnal gastroesophageal acid reflux.

Guidelines for gastroenterological endoscopy in patients undergoing antithrombotic treatment

Guidelines for gastroenterological endoscopy in patients undergoing antithrombotic treatment have been produced by the Japan Gastroenterological Endoscopy Society in collaboration with the Japan Circulation Society, the Japanese Society of Neurology, the Japan Stroke Society, the Japanese Society on Thrombosis and Hemostasis and the Japan Diabetes Society. Previous guidelines from the Japan Gastroenterological Endoscopy Society have focused primarily on prevention of hemorrhage after gastroenterological endoscopy as a result of continuation ofantithrombotic therapy, without considering the associated risk of thrombosis. The new edition of the guidelines includes discussions of gastroenterological hemorrhage associated with continuation of antithrombotic therapy, as well as thromboembolism associated with withdrawal of antithrombotic therapy.

Role of hiatus hernia and gastric mucosal atrophy in the development of reflux esophagitis in the elderly

Background and Aims: The pathogenetic mechanism responsible for the increased prevalence of reflux esophagitis in the elderly remains controversial. The aim of this study was to determine if an increased occurrence of hiatus hernia (HH) and/or a lower rate of gastric mucosal atrophy (GMA) are the main causes of the increased prevalence of reflux esophagitis in elderly patients.

MFG-E8 Attenuates Intestinal Inflammation in Murine Experimental Colitis by Modulating Osteopontin-Dependent αvβ3 Integrin Signaling

MFG-E8 (milk fat globule-epidermal growth factor 8) deficiency is strongly associated with acquisition of immune-mediated disorders due to the loss of tissue homeostasis. However, comparatively little is known regarding its functions in gastrointestinal tract disorders, in which immune homeostasis is a major concern. Herein, we report altered MFG-E8 expression in inflamed colons during the acute phase of murine experimental colitis and found that treatment with recombinant MFG-E8, but not its arginine-glycine-aspartate mutant counterpart, ameliorated colitis by reducing inflammation and improving disease parameters. To reveal the MFG-E8-mediated antiinflammatory mechanism, we employed an in vitro system, which showed the down-regulation of NF-κB in an LPS-dependent manner. Additionally, MFG-E8 altered αvβ3 integrin-mediated focal adhesion kinase phosphorylation by impeding the binding of one of its potent ligands osteopontin, which becomes activated during colitis. Taken together, our results indicated that MFG-E8 has a novel therapeutic potential for treatment of colitis.

Co-infection by serologically-silent hepatitis B virus may contribute to poor interferon response in patients with chronic hepatitis C by down-regulation of type-I interferon receptor gene expression in the liver

Intrahepatic mRNA levels of type‐I interferon (IFN) receptor genes have been shown to correlate with the clinical efficacy of IFN therapy in patients with chronic hepatitis C. Recently, co‐infection by serologically‐silent hepatitis B virus (HBV) has been assumed to be associated with the poor IFN response in patients with chronic hepatitis C. The aim of this study was to investigate the relationship between the co‐infection of serologically‐silent HBV and type‐I IFN receptor gene expression in the liver of patients with chronic hepatitis C. The intrahepatic mRNA levels of IFNAR2, one of the two subunits of the type‐I IFN receptor, were quantified and compared with both the prevalence of HBV DNA and the hepatitis C virus (HCV) genotype in 45 patients with chronic hepatitis C, who were negative for hepatitis B surface antigen. Co‐infection, as evaluated by a nested polymerase chain reaction, was present in 22 patients (48.9%), with dominance of the HCV genotype 1b (65.2%) over genotype 2a (31.8%). Co‐infection was associated with lower IFNAR2 mRNA levels, higher levels of serum HCV RNA, and a poor IFN response, regardless of the HCV genotype. The findings suggest the possibility that co‐infection by serologically‐silent HBV is one of the factors that can lead to an unfavorable IFN response in chronic hepatitis C by down‐regulation of IFN receptor gene expression in the liver. J. Med. Virol. 63:220–227, 2001.