Kinesin Family Member B18 Is Related to Gastric Mucin Phenotype and Contributes to Gastric Cancer Progression by Regulating Epithelial-Mesenchymal Transition.

INTRODUCTION: Gastric cancer (GC) remains a common health concern worldwide and is the third leading cause of death in Japan. It can be broadly classified into gastric and intestinal mucin phenotypes using immunohistochemistry. We previously reported numerous associations of kinesin family member (KIF) genes and mucin phenotypes with GC. However, no previous studies have reported on the importance of KIF18B in GC using immunostaining. Thus, in this study, we investigated the expression and functions of KIF18B, which is highly expressed in gastric mucin phenotype GC. METHODS: We performed RNA-seq of gastric and intestinal mucin type GCs, and clinicopathological studies of the KIF18B we found were performed using 96 GC cases. We also performed functional analysis using GC-derived cell lines. RESULT: RNA-seq showed the upregulation of matrisome-associated genes in gastric mucin phenotype GC and a high expression of KIF18B. KIF18B was detected in 52 of the 96 GC cases (54%) through immunohistochemistry. Low KIF18B expression was significantly associated with poor overall survival (p < 0.01). Other molecules that were significantly associated with KIF18B were MUC5AC and claudin 18; these were also significantly associated with the gastric mucin phenotype. KIF18B small interfering RNA (siRNA)-transfected GC cells showed greater growth and spheroid colony formation than the negative control siRNA-transfected cells. Furthermore, expression of snail family transcriptional repressor 1 and cadherin 2 was significantly increased and that of cadherin 1 was significantly decreased in KIF18B siRNA-transfected GC cells. CONCLUSION: These findings not only suggest that KIF18B may be a useful prognostic marker, but also provide insight into the pathogenesis of the GC phenotype.

Gastric mucin phenotype indicates aggressive biological behaviour in early differentiated gastric adenocarcinomas following endoscopic treatment.

BACKGROUND: The distribution of mucin phenotypes and their relationship with clinicopathological features in early differentiated gastric adenocarcinomas in a Chinese cohort are unknown. We aimed to investigate mucin phenotypes and analyse the relationship between mucin phenotypes and clinicopathological features, especially biological behaviours, in early differentiated gastric adenocarcinomas from endoscopic specimens in a Chinese cohort. METHODS: Immunohistochemical staining of CD10, MUC2, MUC5AC, and MUC6 was performed in 257 tissue samples from patients with early differentiated gastric adenocarcinomas. The tumour location, gross type, tumour size, histological type, depth of invasion, lymphovascular invasion, mucosal background and other clinicopathological parameters were evaluated. The relationship between mucin phenotypes and clinicopathological features was analysed with the chi-square test. RESULTS: The incidences of gastric, gastrointestinal, intestinal and null phenotypes were 21 %, 56 %, 20 and 3 %, respectively. The mucin phenotypes were related to histology classification (P < 0.05). The proportion of the gastric phenotype became greater during the transition from differentiated to undifferentiated (P < 0.05). Complete intestinal metaplasia was higher in the gastric and intestinal phenotypes than in the gastrointestinal phenotype (P < 0.05). Tumours with poorly differentiated adenocarcinoma were mainly of the gastric phenotype, which was significantly higher than that of purely differentiated tubular adenocarcinoma (P < 0.05), and the depth of invasion in the mixed type was deeper (P < 0.05). Neither recurrence nor metastasis was detected. CONCLUSIONS: The mucin phenotype of early-differentiated gastric adenocarcinoma has clinical implications, and the gastric phenotype has aggressive biological behaviour in early differentiated gastric cancers, especially in those with poorly differentiated adenocarcinoma or papillary adenocarcinoma components.

Early Gastric Cancer: identification of molecular markers able to distinguish submucosa-penetrating lesions with different prognosis.

BACKGROUND: Early Gastric Cancer (EGC) reaches 25% of the gastric cancers surgically treated in some areas of Northeastern Italy and is usually characterized by a good prognosis. However, among EGCs classified according to Kodama's criteria, Pen A subgroup is characterized by extensive submucosal invasion, lymph node metastases and worse prognosis, whereas Pen B subgroup by better prognosis. The aim of the study was to characterize the differences between Pen A, Pen B and locally advanced gastric cancer (T3N0) in order to identify biomarkers involved in aggressiveness and clinical outcome. METHODS: We selected 33 Pen A, 34 Pen B and 20 T3N0 tumors and performed immunohistochemistry of mucins, copy number variation analysis of a gene panel, microsatellite instability (MSI), TP53 mutation and loss of heterozygosity (LOH) analyses. RESULTS: Pen A subgroup was characterized by MUC6 overexpression (p = 0.021). Otherwise, the Pen B subgroup was significantly associated with the amplification of GATA6 gene (p = 0.002). The higher percentage of MSI tumors was observed in T3N0 group (p = 0.002), but no significant differences between EGC types were found. Finally, TP53 gene analysis showed that 32.8% of Pen tumors have a mutation in exons 5-8 and 50.0% presented LOH. Co-occurrence of TP53 mutation and LOH mainly characterized Pen A tumors (p = 0.022). CONCLUSIONS: Our analyses revealed that clinico-pathological parameters, microsatellite status and frequency of TP53 mutations do not seem to distinguish Pen subgroups. Conversely, the amplification of GATA6 was associated with Pen B, as well as the overexpression of MUC6 and the TP53mut/LOH significantly characterized Pen A.

Three types of metaplasia model through Kras activation, Pten deletion, or Cdh1 deletion in the gastric epithelium.

Chronic inflammation and intestinal metaplasia are strongly associated with gastric carcinogenesis. Kras activation and Pten deletion are observed in intestinal-type gastric cancer, and Cdh1 mutation is associated with diffuse-type gastric cancer. Although various mouse models of gastric carcinogenesis have been reported, few mouse lines enable gene manipulation selectively in the stomach. Here we established a Tff1-Cre bacterial artificial chromosome transgenic mouse line in an attempt to induce gene modification specifically in the gastric pit lineage. In the stomach, Tff1-Cre-mediated recombination was most evident in the pit lineage in the corpus and in entire antral glands; recombination was also observed in a few gastric chief and parietal cells. Outside the stomach, recombination was patchy throughout the intestines, and particularly frequently in the duodenum (Brunner glands), cecum, and proximal colon. In the stomachs of Tff1-Cre;LSL-KrasG12D mice, proliferating cell clusters expanded throughout the corpus glands, with foveolar cell expansion with ectopic Alcian blue-positive mucins, oxyntic atrophy, and pseudopyloric changes with spasmolytic polypeptide-expressing metaplasia; however, gastric cancer was not observed even at 12 months of age. Corpus-derived organoids from Tff1-Cre;LSL-KrasG12D mice exhibited accelerated growth and abnormal differentiation with a loss of chief and parietal cell markers. Tff1-Cre;Ptenflox/flox mice displayed similar changes to those seen in Tff1-Cre;LSL-KrasG12D mice, both with aberrant ERK activation within 3 months. In contrast, Tff1-Cre;Cdh1flox/flox mice initially showed signet ring-like cells that were rapidly lost with disruption of the mucosal surface, and later developed gastric epithelial shedding with hyperproliferation and loss of normal gastric lineages. Eventually, the glandular epithelium in Tff1-Cre;Cdh1flox/flox mice was completely replaced by squamous epithelium which expanded from the forestomach. Tff1-Cre mice offer an additional useful tool for studying gastric carcinogenesis both in vivo and in vitro. Copyright © 2018 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Loss of gastric gland mucin-specific O-glycan is associated with progression of differentiated-type adenocarcinoma of the stomach.

Gastric gland mucin secreted from the lower portion of the gastric mucosa contains unique O-linked oligosaccharides having terminal α1,4-linked N-acetylglucosamine (αGlcNAc) residues largely attached to a MUC6 scaffold. Previously, we generated A4gnt-deficient mice, which totally lack αGlcNAc, and showed that αGlcNAc functions as a tumor suppressor for gastric cancer. Here, to determine the clinicopathological significance of αGlcNAc in gastric carcinomas, we examined immunohistochemical expression of αGlcNAc and mucin phenotypic markers including MUC5AC, MUC6, MUC2, and CD10 in 214 gastric adenocarcinomas and compared those expression patterns with clinicopathological parameters and cancer-specific survival. The αGlcNAc loss was evaluated in MUC6-positive gastric carcinoma. Thirty-three (61.1%) of 54 differentiated-type gastric adenocarcinomas exhibiting MUC6 in cancer cells lacked αGlcNAc expression. Loss of αGlcNAc was significantly correlated with depth of invasion, stage, and venous invasion by differentiated-type adenocarcinoma. Loss of αGlcNAc was also significantly associated with poorer patient prognosis in MUC6-positive differentiated-type adenocarcinoma. By contrast, no significant correlation between αGlcNAc loss and any clinicopathologic variable was observed in undifferentiated-type adenocarcinoma. Expression of MUC6 was also significantly correlated with several clinicopathological variables in differentiated-type adenocarcinoma. However, unlike the case with αGlcNAc, its expression showed no correlation with cancer-specific survival in patients. In undifferentiated-type adenocarcinoma, we observed no significant correlation between mucin phenotypic marker expression, including MUC6, and any clinicopathologic variable. These results together indicate that loss of αGlcNAc in MUC6-positive cancer cells is associated with progression and poor prognosis in differentiated, but not undifferentiated, types of gastric adenocarcinoma.

Filiform serrated adenomatous polyposis arising in a diverted rectum of an inflammatory bowel disease patient.

A 54-year-old man, previously colectomized for inflammatory bowel disease, developed carcinoma in the inflamed rectum stump. The malignant growth was surrounded by a filiform polyposis, grossly considered as pseudopolyps. The histology disclosed, however, a morphology corresponding to the recently described filiform subset of serrated adenoma (FSA). The clustering of the FSA amounted to a filiform serrated adenomatous polyposis, a hitherto unreported observation. It is speculated that neoplastic transformation of pre-existing pseudopolyps and prolaps-related events lead to this peculiar morphology. Minor zones with a villous structure were admixed as were small areas of traditional serrated adenoma and patches of flat dysplasia. Although a combined gastric and intestinal (positivity for MUC5AC, MUC2, MUC6, CDX2) immunoprofile characterized the adenomatous component, a downregulation of the gastric mucin along with a loss of the serrated attribute accompanied the malignant transformation. An added dynamic shift during the adenoma carcinoma sequence included the acquisition of CK7 expression in the malignant portion. Gastric mucin may play a role in the initial step of the neoplastic evolution and CK7 may denote neoplastic progression. This case confirms the notion of a widely variegated morphology of precursor lesions of colorectal carcinoma arising in a chronically inflamed bowel as opposed to the generally more monotonous appearance of adenomas in a sporadic context.

Role of mucin Lewis status in resistance to Helicobacter pylori infection in pediatric patients.

BACKGROUND: Helicobacter pylori causes gastritis, peptic ulcer and is a risk factor for adenocarcinoma and lymphoma of the stomach. Gastric mucins, carrying highly diverse carbohydrate structures, present functional binding sites for H. pylori and may play a role in pathogenesis. However, little information is available regarding gastric mucin in children with and without stomach diseases. MATERIALS AND METHODS: Expression of mucins and glycosylation was studied by immunohistochemistry on gastric biopsies from 51 children with and without H. pylori infection and/or peptic ulcer disease. RESULTS: In all children, MUC5AC was present in the surface epithelium and MUC6 in the glands. No MUC6 in the surface epithelium or MUC2 was detected in any section. The Le(b) and Le(a) blood group antigens were present in the surface epithelium of 80% and 29% of children, respectively. H. pylori load was higher in Le(b) negative children than in Le(b) positive individuals (mean +/- SEM 17.8 +/- 3.5 vs 10.8 +/- 1.5; p < 0.05), but there was no correlation between Le(a) or Le(b) status and gastritis, nodularity, and gastric or duodenal ulcer (DU). Expression of sialyl-Le(x) was associated with H. pylori infection, and DU. CONCLUSIONS: Mucin expression and glycosylation is similar in children and adults. However, in contrast to adults, pediatric H. pylori infection is not accompanied by aberrant expression of MUC6 or MUC2. Furthermore, the lower H. pylori density in Le(b) positive children indicates that H. pylori is suppressed in the presence of gastric mucins decorated with Le(b), the binding site of the H. pylori BabA adhesin.

Gastric mucus alterations associated with murine Helicobacter infection.

The C57BL/6 mouse has been shown to develop gastric adenocarcinoma after Helicobacter felis infection. This model was used to determine whether mucin and trefoil factor (TFF) expression after infection was altered in a similar fashion to the changes seen in the protective gastric mucus layer of the human stomach after H. pylori infection. Our results indicate that this mouse model mimics many of the changes seen after human H. pylori infection, including increased expression of muc4 and muc5b and loss of muc5ac. These alterations in mucin expression occurred as early as 4 weeks postinfection, before the development of significant mucous metaplasia or gastric dysplasia. The decrease in muc5ac expression occurred only in the body of the stomach and was not secondary to the adaptive immune response to infection, because a similar decrease in expression was seen after infection of B6.Rag-1(-/-) mice, which lack B and T cells. Intriguingly, the increased expression of Muc4 and Muc5b in infected C57BL/6 mice was not seen in the infected B6.Rag-1(-/-) mice. Because B6.Rag-1(-/-) mice do not develop gastric pathology after H. felis infection, these findings point to the potential role of Muc4 and Muc5b in disease progression. This manuscript contains online supplemental material at http://www.jhc.org. Please visit this article online to view these materials.

The MUC gene family: their role in the diagnosis and prognosis of gastric cancer.

Early diagnosis of gastric cancer and its differential diagnosis from other non-malignant gastric diseases like gastritis is still a major clinical problem. Most patients are asymptomatic in the early stages of gastric cancer, and there is no reliable marker available for the early and specific diagnosis of gastric cancer. Many attempts have been made to define the biological profile of gastric cancer to improve the chances of its early diagnosis, prognosis and treatment. Several studies have shown the aberrant expression profile of mucins in different malignancies, suggesting that mucins have a great potential to be used as a diagnostic and prognostic marker in gastric cancer. In this review, we have briefly described the different types of gastric adenocarcinomas and the progression of gastric cancer. Furthermore, the role of mucins and their related carbohydrate epitope is discussed in the normal stomach and in the diagnosis and prognosis of gastric adenocarcinomas.

Morphogenesis of pancreatic cancer: role of pancreatic intraepithelial neoplasia (PanINs).

INTRODUCTION: Pancreatic ductal adenocarcinoma (i.e., pancreatic cancer) is an almost universally lethal disease. The identification of precursor lesions of pancreatic cancer provides an opportunity for early detection and potential therapeutic intervention before the development of invasive cancer. DISCUSSION: It is now established that pancreatic cancers do not arise de novo but rather exhibit a sequential histological and genetic progression of precursor lesions culminating in frank, invasive neoplasia. Pancreatic intraepithelial neoplasia (PanIN) is the most common non-invasive precursor lesion of pancreatic cancer. The development of a consensus nomenclature scheme for PanINs has facilitated research into pancreatic cancer precursors and enabled standardization of results across institutions. CONCLUSION: PanINs harbor many of the molecular alterations observed in invasive pancreatic cancer, confirming their status as true non-invasive precursor lesions. Recently developed genetically engineered mouse models of pancreatic cancer also demonstrate the stepwise PanIN progression model, underscoring the commonalities in pancreatic neoplasia between mouse and man.

Characteristics of gastric and intestinal mucin phenotypes of gastric carcinoma.

BACKGROUND/AIMS: To better understand the carcinogenesis pathway in gastric cancer, we investigated the relationship between mucin phenotype and clinicopathologic features. METHODOLOGY: The study was conducted in 203 consecutive patients with newly diagnosed gastric cancer. Sections from representative paraffin blocks of each case were immunostained with human gastric mucin, MUC2, CD10, and paradoxical concanavalin A class III. Gastric cancers were divided into three phenotypes; gastric phenotype (G-type), intestinal phenotype (I-type), and the Null phenotype (N-type). Surrounding non-cancerous mucosa was also phenotyped as G- or I-type. RESULTS: In G-type surrounding non-cancerous mucosa, I-type cancers were more frequent among the differentiated type gastric cancers whereas G-type cancers were more frequent among the undifferentiated type gastric cancers (p=0.004). As for surrounding non-cancerous mucosa, the G-type mucosa was more frequently seen than the I-type mucosa in incomplete intestinal metaplasia (p<0.001). I-type gastric cancers were more frequent in non-cancerous surrounding mucosa with incomplete intestinal metaplasia, and G-type cancers were more frequent in non-cancerous surrounding mucosa with no intestinal metaplasia (p=0.03). CONCLUSIONS: G-type gastric cancers may develop in G-type gastric mucosa with incomplete intestinal metaplasia and progress to the G-type undifferentiated carcinomas or the I-type differentiated carcinomas.

Tumor differentiation phenotype in gastric differentiated-type tumors and its relation to tumor invasion and genetic alterations.

AIM: To clarify the relations between tumor differentiation phenotype and tumor invasion or genetic alterations in gastric differentiated-type tumors. METHODS: We examined the tumor differentiation phenotype, the presence of mutations in APC and p53, and the microsatellite instability (MSI) status in 48 gastric adenomas and 171 differentiated-type carcinomas. The tumor differentiation phenotype was determined by examining the expression of human gastric mucin (HGM), MUC6, MUC2 and CD10. The tumors were then classified into gastric- (G-), gastric and intestinal mixed- (GI-), or intestinal- (I-) phenotypes, according to the immunopositivity of the above markers. The presence of mutations in APC and p53 and the MSI status were also investigated in all the tumors. RESULTS: Gastric adenomas were significantly associated with CD10 expression, I-phenotype tumors and the presence of APC mutations, compared with carcinomas (66.7% vs 25.1%, P < 0.0001; 56.3% vs 14.6%, P < 0.0001; 39.6% vs 14.0%, P < 0.0001, respectively) and inversely associated with expressions of HGM and MUC6 and the presence of p53 mutations (10.4% vs 62.6%, P < 0.0001; 39.6% vs 64.3%, P = 0.003; 2.0% vs 26.3%, P = 0.001, respectively). The frequency of APC mutations was significantly higher in HGM-negative tumors, MUC6-negative tumors, CD10-positive tumors and I-phenotype tumors than in HGM-positive tumors, MUC6-positive tumors, CD10-negative tumors and G-phenotype tumors (32.7% vs 7.1%, P < 0.0001; 27.8% vs 14.0%, P = 0.0182; 37.3% vs 10.4%, P < 0.0001; and 38.5% vs 9.5%, P = 0.0017, respectively). The frequency of MSI was significantly higher in MUC6-positive tumors, CD10-negative tumors and G-phenotype tumors than in MUC6-negative tumors, CD10-positive tumors and I-phenotype tumors (24.8% vs 6.7%, P = 0.0009; 22.2% vs 8.0%, P = 0.0143; and 28.6% vs 9.6%, P = 0.0353, respectively). CONCLUSION: The tumor differentiation phenotype is closely related to tumor invasion and genetic alterations in gastric differentiated-type tumors.

Hath1 up-regulates gastric mucin gene expression in gastric cells.

The Notch signaling pathway is known to mediate the differentiation and fate specification of cells in embryonic stage and adult tissues. Several tumors exhibit aberrant expression of Notch signaling component genes, such as Notch1/2/3 and Hath1. In this study, we investigated the mRNA expression of seven Notch-related genes, Notch1/2/3, Hes1/2/3, and Hath1, and then compared it with the expression of gastric mucin genes, MUC5AC and MUC6, in eight gastric cancer (GC) cell lines. Notch1/2/3 and Hes1 were expressed in most GC cell lines as well as normal gastric mucosae, while Hes2/3 were expressed in neither these cell lines nor the normal stomach. As for Hath1, five GC cell lines exhibited undetectable levels, while normal gastric mucosa expressed Hath1. The expression patterns of Hath1 and MUC6 were closely related in most GC cell lines. Many MUC5AC-positive cases also tended to show Hath1 expression. Over-expression of Math1, a mouse Hath1 homolog, in the GC cells strongly enhanced both the MUC6 and MUC5AC mRNA levels. Moreover, knockdown of Hath1 by means of RNA interference significantly decreased the expression of both mucin genes. These data indicate that Hath1 is one of the transcriptional regulators for MUC6 and MUC5AC in GC cells. It is also possible that loss of Hath1 expression may play a role in gastric carcinogenesis.

Gastric and intestinal phenotypic cell marker expressions in gastric differentiated-type carcinomas: association with E-cadherin expression and chromosomal changes.

Gastric and intestinal phenotypic cell markers are widely expressed in gastric carcinomas, irrespective of their histological type. In the present study, the relations between the phenotypic marker expression of the tumour, histological findings, expression of cell adhesion molecules, and the chromosomal changes in gastric differentiated-type carcinomas were examined. The phenotypic marker expression of the tumour was determined by the combination of the expression of the human gastric mucin (HGM), MUC6, MUC2 and CD10, and was evaluated in comparison with the expression of cell adhesion molecules, such as E-cadherin and beta-catenin, and chromosomal changes by comparative genomic hybridization (CGH) in 34 gastric differentiated-type carcinomas. Tumours were classified into the gastric- (G-), gastric and intestinal mixed- (GI-), intestinal- (I-), or unclassified- (UC-) phenotype according to the immunopositivity of staining for HGM, MUC6, MUC2, and CD10. G-phenotype tumours were significantly associated with a higher incidence of differentiated-type tumours mixed with undifferentiated-type component, compared with GI- and I-phenotype tumours (88.9 vs 33.3%, P=0.0498 and 88.9 vs 42.9%, P=0.0397; respectively). HGM-positive tumours were significantly associated with a higher incidence of tumours with abnormal expression of E-cadherin, compared with HGM-negative tumours (66.7 vs 21.1%, P=0.0135). GI-phenotype tumours were significantly associated with a higher incidence of tumours with abnormal expression of E-cadherin, compared with I-phenotype tumours (77.8 vs 21.4%, P=0.0131). HGM-negative tumours were significantly associated with higher frequencies of the gains of 19q13.2 and 19q13.3, compared with HGM-positive tumours (57.9 vs 20.0%, P=0.0382 and 63.2 vs 13.3%, P=0.0051; respectively). MUC6-positive tumours were significantly associated with higher frequencies of the gains of 20q13.2, compared with MUC6-negative tumours (71.4 vs 30.0%, P=0.0349). MUC2-positive tumours were significantly associated with the gain of 19p13.3, compared with MUC2-negative tumours (41.2 vs 5.9%, P=0.0391). I-phenotype tumours were significantly associated with higher frequencies of gains of 5p15.2 and 13q33-34, compared with G-phenotype tumours (66.7 vs 0%, P=0.0481, each) and also associated with higher frequencies of gain of 7p21, compared with GI-phenotype tumours (66.7 vs 0%, P=0.0481). Our present results show that gastric differentiated-type carcinomas have different characteristics according to the phenotypic marker expression of the tumour in terms of histological findings, E-cadherin expression and pattern of chromosomal changes.

A novel in vitro infection model of Helicobacter pylori using mucin-producing murine gastric surface mucous cells.

BACKGROUND: Helicobacter pylori is found within the gastric surface mucous gel layer and in the epithelial surface. Gastric cancer cells have been used in experimental H. pylori infection in vitro, although cancer cells have some abnormalities in cellular properties. The aim of this study was to develop an in vitro H. pylori infection model using normal gastric surface cells that produce gastric mucin. MATERIALS AND METHODS: Normal murine gastric surface mucous cells (GSM06) were cultured by the liquid interface method using a serum-free medium and a collagen gel containing a fibroblast cell line (L929) and infected with H. pylori. Infection by H. pylori was assessed by enumerating the colony-forming units (CFU) of H. pylori adhered to GSM06 cells and by transmission electron microscopy. The production of mucin was determined by a lectin binding assay, sugar analysis, and MUC5AC gene expression. RESULTS: GSM06 cells cultured under these conditions produced mucin containing N-acetylgalactosamine and MUC5AC as the core protein. Significantly higher numbers of H. pylori adhered to GSM06 cells under mucin-producing conditions than under nonproducing conditions. Microscopic observation showed a filamentous structure resembling a type IV secretion system apparatus formed between the surface of GSM06 cells and H. pylori. CONCLUSIONS: This study demonstrates a novel in vitro H. pylori infection model using mucin-producing murine GSM06 cells for early stages of infection.

Mucin phenotypic expression and background mucosa of esophagogastric junctional adenocarcinoma.

BACKGROUND: In the West, the incidence of adenocarcinoma of the cardia or esophagogastric junction (EGJ) is increasing. This carcinoma is variously defined, however, and it contains heterogeneous tumors. In Japan, the frequency of this carcinoma is low, and little is known about it. We studied small esophagogastric junctional adenocarcinoma to determine its characteristics in Japanese patients. METHODS: Fifty-four patients with Siewert type II junctional adenocarcinoma (with tumors <<4 cm in maximum diameter) were studied. The carcinomas were classified into two types: tumors straddling the EGJ (EGJ-type) and tumors occurring entirely below the EGJ (bEGJ-type). Characteristics of the tumors and surrounding mucosae were evaluated. Phenotypic expression of mucin, p53 overexpression, and Helicobacter pylori infection were investigated histologically and immunohistochemically. RESULTS: Twenty-three (43%) patients had EGJ-type and 31 (57%) had bEGJ-type carcinomas. The ratio of advanced cancers was significantly higher in EGJ-type. Some between-group similarities and differences existed in histological type, mucin phenotype, and p53 protein overexpression, but they were not statistically significant. In the mucosa surrounding EGJ-type tumors vs bEGJ-type tumors, oxyntic glands were significantly better preserved, gastric intestinal metaplasia occurred significantly less frequently, and the H. pylori infection rate was lower. Barrett's metaplasia was seen in only 2 patients, with the EGJ-type. CONCLUSION: EGJ-type and bEGJ-type adenocarcinomas have different background mucosa of the stomach. EGJ-type has less atrophy of the oxyntic glands and less intestinal metaplasia in the stomach. This type appears to be the Western-type junctional adenocarcinoma that is still rare in Japan. Many cases of advanced junctional adenocarcinoma in Japan are of subjunctional origin.

A role for human MUC4 mucin gene, the ErbB2 ligand, as a target of TGF-beta in pancreatic carcinogenesis.

MUC4: encodes a large transmembrane mucin that is overexpressed in pancreatic adenocarcinomas. The molecular mechanisms responsible for that altered pattern of expression are unknown. TGF-beta, a pleiotropic cytokine, regulates numerous genes involved in pancreatic carcinogenesis via activation of the Smads proteins and MUC4 promoter is rich in Smad-binding elements. Our aim was to study whether the regulation of MUC4 expression by TGF-beta in pancreatic cancer cells was strictly dependent on Smad4 activity. Three pancreatic cancer cell lines, CAPAN-1 (MUC4+/Smad4-), CAPAN-2 (MUC4+/Smad4+) and PANC-1 (MUC4-/Smad4+), were used. By RT-PCR, transfection assays and immunohistochemistry, we show that (i) both MUC4 mRNA and apomucin expression are upregulated by TGF-beta, (ii) Smad2 positively cooperates with Smad4 to activate the promoter, (iii) activation of Smad4 by exogenous TGF-beta induces Smad4 binding to the promoter, (iv) Smad7 and c-ski both inhibit activation by Smad4. When Smad4 is mutated and inactive, TGF-beta activates MUC4 expression via MAPK, PI3K and PKA signaling pathways. Absence of expression in PANC-1 cells is due to histone deacetylation. Altogether, these results indicate that upregulation of MUC4 by TGF-beta is restricted to well-differentiated pancreatic cancer cells, and point out a novel mechanism for TGF-beta as a key molecule in targeting MUC4 overexpression in pancreatic adenocarcinomas.

Mucin phenotypic expression in early signet ring cell carcinoma of the stomach: its relationship with the clinicopathologic factors.

Gastric early-stage signet ring cell carcinoma (SIG) has been reported to have a lower rate of lymph node metastasis and a higher rate of favorable prognosis than other histological types. However, the development and progression mechanisms of early-stage SIG (early SIG) are controversial. This study examined the correlation between the mucin phenotype of early SIG and its clinicopathologic factors, particularly for the sake of less invasive surgery. Sixty-nine early SIGs were studied immunohistochemically with gastric mucin (M1 and MUC6) and intestinal mucin (MUC2). SIGs were classified into gastric (G), intestinal (I), gastrointestinal (GI), or unclassified (U) type. The intramucosal spreading patterns of SIG were investigated and then classified as either expansive or infiltrative. SIGs were classified into G-type (59.4%) and GI-type (40.6%). Neither the I- nor the U-type was observed. The GI-type expression correlated with the depth of tumor invasion in SIGs (P < 0.05). In contrast, there was no increase in GI-type expression in relation to tumor size. Intramucosal infiltrative growth correlated with intestinal metaplasia (IM) of background mucosa of SIGs (P < 0.01). There was no significant correlation between phenotypes and intramucosal spreading pattern. In conclusion, the GI-type expression of SIG is a clinically useful factor for predicting submucosal invasion. The findings of SIG surrounded with IM revealed the need to exercise great care in determining the surgical margin.

Re-evaluation of mucin phenotypes of gastric minute well-differentiated-type adenocarcinomas using a series of HGM, MUC5AC, MUC6, M-GGMC, MUC2 and CD10 stains.

We examined which, and how many, mucin markers are necessary to define the phenotypes of gastric cancers, and re-evaluated the incidence of their mucin phenotypes and whether minute gastric carcinomas arise as unclassified type. Well-differentiated-type minute gastric carcinomas (n = 33) measuring 0%, and was 33%, 33%, 3%, 30%, and 0%, respectively, when the percentage of positive mucin phenotype was set at >or=10%. Thus, a panel of MUC5AC (or HGM), MUC6, MUC2 and CD10 stains is indispensable for accurately determining the mucin phenotypes of gastric carcinomas, and the above-mentioned classification is important for studying changes in the histological types of well-differentiated-type adenocarcinomas during change to the poorly differentiated type, as well as corresponding genetic abnormalities.

Transcriptional regulation of human mucin MUC4 by bile acids in oesophageal cancer cells is promoter-dependent and involves activation of the phosphatidylinositol 3-kinase signalling pathway.

Abnormal gastro-oesophageal reflux and bile acids have been linked to the presence of Barrett's oesophageal premalignant lesion associated with an increase in mucin-producing goblet cells and MUC4 mucin gene overexpression. However, the molecular mechanisms underlying the regulation of MUC4 by bile acids are unknown. Since total bile is a complex mixture, we undertook to identify which bile acids are responsible for MUC4 up-regulation by using a wide panel of bile acids and their conjugates. MUC4 apomucin expression was studied by immunohistochemistry both in patient biopsies and OE33 oesophageal cancer cell line. MUC4 mRNA levels and promoter regulation were studied by reverse transcriptase-PCR and transient transfection assays respectively. We show that among the bile acids tested, taurocholic, taurodeoxycholic, taurochenodeoxycholic and glycocholic acids and sodium glycocholate are strong activators of MUC4 expression and that this regulation occurs at the transcriptional level. By using specific pharmacological inhibitors of mitogen-activated protein kinase, phosphatidylinositol 3-kinase, protein kinase A and protein kinase C, we demonstrate that bile acid-mediated up-regulation of MUC4 is promoter-specific and mainly involves activation of phosphatidylinositol 3-kinase. This new mechanism of regulation of MUC4 mucin gene points out an important role for bile acids as key molecules in targeting MUC4 overexpression in early stages of oesophageal carcinogenesis.