Yoshifumi Higashimoto

STAT3 mediates the survival signal in oncogenic ras-transfected intestinal epithelial cells.

The oncogenic ras mutation is a common and critical step in gastrointestinal carcinogenesis. In a previous study, we demonstrated that oncogenic ras activated the EGF‐related peptide autocrine loop and that the apoptosis resistance observed in the oncogenic ras‐stimulated cell (IEC‐ras cell) was dependent on this activated EGF‐related peptide autocrine loop. STATs (signal transducers and activators of transcription), first identified as intracellular signal transducers stimulated by cytokines, are known to also be activated by EGF. However, the role of STATs in the survival signal of IEC‐ras cells is not clear. In the present study, we demonstrate that STAT3 is constitutively activated in ras‐stimulated cells and that STAT3 activation is considerably suppressed by the EGF‐specific receptor kinase inhibitor AG1478. We also show that disruption of the STAT3 pathway by introduction of a dominant‐negative STAT3 mutant abolishes the apoptosis resistance against UVC and MMC treatment observed in IEC‐ras cells without affecting proliferation. Moreover, the expression of Bcl‐2 and Bcl‐xL, apoptosis‐suppressive proteins, is reduced in dominant‐negative STAT3‐transfected cells. Thus, STAT3 appears to be an important mediator of the anti‐apoptotic signal in IEC‐ras cells. Int. J. Cancer 78:326–330, 1998.© 1998 Wiley‐Liss, Inc.

Gastrin Induces Heparin-Binding Epidermal Growth Factor-like Growth Factor in Rat Gastric Epithelial Cells Transfected With Gastrin Receptor

BACKGROUND & AIMS Parietal cells express heparin-binding epidermal growth factor (EGF)-like growth factor (HB-EGF). However, it is unknown whether HB-EGF mediates the trophic action of gastrin. The purpose of this study was to determine whether gastrin modulates the expression of HB-EGF, which mediates the proliferative effects of gastrin on gastric epithelial cells. METHODS RGM1 cells, a rat gastric epithelial cell line, were transfected with a human gastrin receptor complementary DNA. Gastrin induction of messenger RNAs (mRNAs) for EGF-related polypeptides was assayed by Northern blotting. Processing of cell surface-associated proHB-EGF and secretion of HB-EGF were determined by flow cytometry and Western blotting, respectively. Tyrosine phosphorylation of the EGF receptor was assayed by immunoprecipitation and Western blotting with an antiphosphotyrosine antibody. Cell growth was evaluated by [3H]thymidine incorporation. RESULTS Gastrin induced expression of HB-EGF mRNA, processing of proHB-EGF, release of HB-EGF into the medium, and tyrosine phosphorylation of the EGF receptor. The growth-stimulatory effects of gastrin were partly inhibited by anti-rat HB-EGF serum and completely blocked by AG1478, an EGF receptor-specific tyrphostin. CONCLUSIONS The findings suggest that HB-EGF at least partially mediates the proliferative effects of gastrin on gastric epithelial cells.

Increased production of interleukin 1 beta and hepatocyte growth factor may contribute to foveolar hyperplasia in enlarged fold gastritis.

BACKGROUND AND AIMS: It has been reported that eradication of Helicobacter pylori improves fold width in H pylori associated enlarged fold gastritis. The aim of this study was to clarify the mechanism of fold thickening in this condition. PATIENTS AND METHODS: In eight patients with enlarged fold gastritis and 13 patients without enlarged folds, the presence of H pylori infection, inflammatory infiltrates, mucosal plasia, and epithelial cell proliferation in the body mucosa were investigated, and production of transforming growth factor alpha (TGF alpha), hepatocyte growth factor (HGF), and interleukin 1 beta (IL 1 beta) was determined by a competitive reverse transcription/polymerase chain reaction method and in vitro short-term culture of biopsy specimens. RESULTS: In the patients with enlarged fold gastritis, inflammatory infiltrates including macrophages increased with H pylori colonisation in the body. Foveolar thickness and proliferating cell nuclear antigen (PCNA) labelling index were increased. Messenger RNA levels of HGF, but not TGF alpha, were increased, and release of HGF and IL 1 beta was increased. HGF release, which was positively correlated with IL 1 beta release and foveolar thickness, decreased in the presence of IL 1 receptor antagonist. After eradication of H pylori, inflammatory infiltrates, IL 1 beta and HGF release decreased with concomitant decreases in PCNA labelling index, foveolar thickness and fold width. CONCLUSIONS: Increased IL 1 beta and HGF production caused by H pylori infection may contribute to fold thickening of the stomach by stimulating epithelial cell proliferation and foveolar hyperplasia in patients with enlarged fold gastritis.

Over-expression of bcl-xL gene in human gastric adenomas and carcinomas

The present study was designed to clarify whether bel‐xL is involved in the development of carcinoma in the stomach. Levels of bel‐xL and bc1‐2 mRNA were determined by a reverse‐transcription/polymerase‐chain reaction in endoscopic gastric biopsy specimens from 10 control subjects, 11 patients with adenomas and 14 patients with carcinomas. In 6 of 11 adenomas, 5 of 8 early carcinomas and 3 of 6 advanced carcinomas, the bel‐xL gene was over‐expressed. In carcinomas, overexpression of the bel‐xL gene was observed in 6 of 9 intestinaltype carcinomas and 2 of 5 diffuse‐type carcinomas. No correlation was observed between bel‐xL and bc1‐2 gene expression. In cases in which the bel‐xL gene was over‐expressed, an apparent increase in the protein level of Bcl‐xL was observed by immunoblot analysis and intense Bcl‐x immunoreactivity was detected immunohistochemically within the tumor cells. In conclusion, we showed that bel‐xL is over‐expressed in gastric carcinomas at both the RNA and protein levels, suggesting that overexpression of bel‐xL may play a role in gastric carcinogenesis.

Expression of hepatocyte growth factor and c-met in ulcerative colitis.

Abstract.Objective and Design: This study was designed to determine if the hepatocyte growth factor (HGF)-Met system is involved in the repair process of inflamed mucosa of ulcerative colitis (UC) and in the development of UC-associated colorectal cancer.¶Materials and Methods: HGF and c-met gene expressions were quantified in colonic mucosal specimens from healthy control subjects, patients with UC and patients with UC-associated colorectal cancer, using the competitive reverse transcription-polymerase chain reaction. Expression of HGF protein was determined by immunoblot analysis. Expression of c-Met protein was analyzed immunohistochemically.¶Results: HGF and c-met gene expressions were increased in inflamed mucosa of UC, compared with control subjects. Gene expression of HGF was also increased in the surrounding inflamed mucosa of UC-associated cancers. In cases in which the HGF gene expression was increased, an apparent increase in protein levels of HGF in inflamed mucosa of UC were observed by immunoblot analysis. The c-met gene was overexpressed in UC-associated cancers and a high level of immunoreactivity of the c-Met protein was immunohistochemically detected within the cancer cells.¶Conclusion: We showed that HGF and c-met expression is increased in the inflamed mucosa of UC and that c-met is overexpressed in UC-associated colorectal cancers. These observations suggest HGF-Met system is involved in the repair process of the inflamed mucosa of UC and provide further support for the view that the inappropriate expressions of both HGF and c-met genes predispose to the development of colorectal cancer in patients with UC.

Modulation of apoptosis by endogenous Bcl-xL expression in MKN-45 human gastric cancer cells

This study was designed to clarify the role of endogenous Bcl-xL expression in modulating apoptosis of malignant cells. Administration of bcl-x-antisense oligonucleotides decreased Bcl-xL protein levels in the MKN-45 human gastric cancer cell line. The decrease in Bcl-xL protein content resulted in increased cell death induced by serum deprivation or Fas-antibody administration. Flow cytometric analysis revealed that the increased apoptotic cell death was more prominent in bcl-x-antisense-treated cells as compared to control cells, bcl-x-sense-treated cells, or bcl-x-nonsense-treated cells. To inhibit the effect of intrinsic Bcl-xL protein, we overexpressed Bak, which binds Bcl-xL and inhibits the anti-apoptotic effect of Bcl-xL, by transfection into MKN-45 cells. Bak-overexpressing cells showed increased apoptotic cell death induced by Fas-antibody when compared to parent cells and MKN-neo-transfected cells. Bak-overexpressing cells also showed greater sensitization to 5-fluorouracil and cisplatin than parent cells and MKN-neo-transfected cells.  In conclusion, we demonstrated that administration of bcl-x-antisense oligonucleotides or overexpression of Bak protein induces sensitization to apoptosis in MKN-45 gastric cancer cells, suggesting that endogenous Bcl-xL expression in cancer cells is an important modulator of apoptosis.

Morphological and functional restoration of parietal cells in Helicobacter pylori associated enlarged fold gastritis after eradication

BACKGROUND/AIM Helicobacter pylori infections are associated with hypochlorhydria in patients with pangastritis. It has previously been shown that eradication of H pylori leads to an increase in acid secretion in H pylori associated enlarged fold gastritis, suggesting that H pylori infection affects parietal cell function in the gastric body. The aim of this study was to evaluate the effects ofH pylori infection on parietal cell morphology and function in hypochlorhydric patients. PATIENTS/METHODS The presence of H pylori infection, mucosal length, and inflammatory infiltration were investigated in six patients with enlarged fold gastritis and 12 patients without enlarged folds. Parietal cell morphology was examined by immunohistochemistry using an antibody against the α subunit of H+,K+-ATPase and electron microscopy. In addition, gastric acid secretion and fasting serum gastrin concentration were determined before and after the eradication ofH pylori. RESULTS In theH pylori positive patients with enlarged fold gastritis, fold width, foveolar length, and inflammatory infiltration were increased. In addition, the immunostaining pattern of H+, K+-ATPase was less uniform, and the percentage of altered parietal cells showing dilated canaliculi with vacuole-like structures and few short microvilli was greatly increased compared with that in H pylori positive patients without enlarged folds. After eradication, fold width, foveolar length, and inflammatory infiltrates decreased and nearly all parietal cells were restored to normal morphology. On the other hand, altered parietal cells were negligible in H pylori negative patients. In addition, the basal acid output and tetragastrin stimulated maximal acid output increased significantly from 0.5 (0.5) to 4.1 (1.5) mmol/h and from 2.5 (1.2) to 13.8 (0.7) mmol/h (p<0.01), and fasting serum gastrin concentrations decreased significantly from 213.5 (31.6) to 70.2 (7.5) pg/ml (p<0.01) after eradication in patients with enlarged fold gastritis. CONCLUSION The morphological changes in parietal cells associated withH pylori infection may be functionally associated with the inhibition of acid secretion seen in patients with enlarged fold gastritis.

Mucosal interleukin-1β production and acid secretion in enlarged fold gastritis

Background: We have previously shown that eradication of Helicobacter pylori increases acid secretion in H. pylori‐associated enlarged fold gastritis.

Plasma cholecystokinin-octapeptide like immunoreactivity in patients with hepatic cirrhosis

Molecular forms of cholecystokinin (CCK) in the peripheral circulation were studied in normal subjects and cirrhotic patients. Fractionation of plasma extract collected 20 min after intraduodenal infusion of fat revealed four major peaks by Sephadex G-50 column chromatography in normal subjects. Peak I eluted at a position similar to CCK-33, peaks II and III eluted between CCK-33 and CCK-14, and peak IV eluted between CCK-14 and CCK-8. In cirrhotic patients, there was a prominent peak (peak V) eluted at a position similar to CCK-8, in addition to those four peaks. These findings are consistent with the previous observations of hepatic elimination of CCK-8, and suggest that smaller forms of CCK similar in size to CCK-8 are not major forms of CCK in plasma in normal subjects but circulate substantially in cirrhotic patients.

Successful Embolization for Uterine Hemorrhage in a Patient with Acute Promyelocytic Leukemia

A successful embolization for life-threatening uterine hemorrhage is described in a patient with acute promyelocytic leukemia and disseminated intravascular coagulation. On admission the patient was in the 6th week of gestation with incomplete abortion and uterine hemorrhage. In addition to combination chemotherapy and anticoagulant therapy, dilatation and curettage were also performed. After the operation the uterine hemorrhage was progressively increased in amount and was not responsive to usual measures. Embolization of bilateral uterine arteries using Gelfoam was performed with a dramatic improvement of the hemorrhage.