Ken Nishikura

Infrequent K- ras codon 12 mutation in serrated adenomas of human colorectum

Background—Serrated adenoma is a new morphological subtype of colorectal adenoma. The lesion provides a distinct morphological route to carcinoma, but the underlying genetic changes have not yet been investigated. Aims—To determine the frequency of K-ras mutation in serrated adenoma. Methods—The frequency of K-ras codon 12 point mutation in 20 serrated adenomas, five atypical hyperplastic polyps, and 58 sporadic polypoid adenomas was investigated by nested polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) methods. Results—Although most of the serrated adenomas were large (average size 11.4 mm) and polypoid, K-rascodon 12 point mutation was detected in only one of the 20 (5%), which is a significantly lower frequency than that in sporadic polypoid adenomas (18/60; 30%) (p = 0.017). No mutation was detected in the atypical hyperplastic polyps. Three of 20 (15%) serrated adenomas contained a focus of carcinoma in situ, indicating their malignant potential and the existence of a serrated adenoma-carcinoma sequence, but no mutation was detected in the foci of carcinoma in situ. Conclusions—K-ras mutation is uncommon in serrated adenomas, indicating a different spectrum of genetic alterations in these lesions from those in typical polypoid sporadic adenomas. This subtype of colorectal adenoma represents a new genetic pathway in the histogenesis of colorectal carcinoma.

APC, K-ras codon 12 mutations and p53 gene expression in carcinoma and adenoma of the gall-bladder suggest two genetic pathways in gall-bladder carcinogenesis

Currant histopathoiogical evidence suggests that gall‐bladder cancer has two main morphological pathways for Its development: de novo (ab Inltlo) origin and adenoma‐carcinoma sequence. In order to investigate the genetic difference between them, APC mutations were examined by RNase protection analysis, K‐ras mutations by nested porymerase chain reaction‐restriction fragment length polymorphism analysis, and p53 gene overexpression by immunoriisto‐chemical analysis in both tumors and benign lesions of the gall‐bladder. Overexpression of the p53 gene was detected in 105 of 164 (64%) de novo carcinomas regardless of size and depth of Invasion, but not in 16 tumors of carcinoma‐ln‐pyloric‐gland‐type adenoma, or In 51 adenomas (47 pyloric gland‐type and 4 Intestinal‐type). K‐ras codon 12 mutation was detected in 4 of 40 (10%) de novo carcinomas, ail four being associated with p53 gene overexpression, but not in 12 tumors of carcinoma in adenoma or 16 adenomas (14 pyloric gland‐type and 2 intestinal‐type). APC mutation was not found in 16 de novo carcinomas or the one pyloric gland‐type adenoma examined. These results suggest that there are two distinct genetic pathways in gall‐bladder carcinogenesis; that is, de novo carcinoma develops from a predominant p53 alteration with low K‐ras mutation, and carclnoma‐in‐pylortc‐gland‐type adenoma develops from p53‐, K‐ras‐, and APC‐gene‐unrelated, as yet unknown, alteration.

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 ≤5 mm were examined using human gastric mucin (HGM) and MUC5AC, MUC6 and M‐GGMC‐1 (or paradoxical concanavalin A type III mucin (Con A)), MUC2 and CD10 stains, and a new method to separate the previous intestinal type into intestinal type and small intestinal type. The phenotypes of carcinomas were classified into gastric, gastrointestinal, intestinal, small intestinal, and unclassified types. MUC5AC or HGM, MUC6, MUC2, and CD10 stains were all necessary to define gastric cancer phenotypes. The incidence of gastric, gastrointestinal, intestinal, small intestinal, and unclassified type was 6%, 49%, 0%, 45%, and 0%, respectively, when the percentage of positive mucin phenotype was set at >0%, and was 33%, 33%, 3%, 30%, and 0%, respe‐ctively, when the percentage of positive mucin phenotype was set at ≥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.

Carcinogenesis of gastric endocrine cell carcinoma: analysis of histopathology and p53 gene alteration

BackgroundEndocrine cell carcinoma of the stomach is characterized by endocrine differentiation and aggressive biological behavior, and is frequently accompanied by an adenocarcinoma component. Because the carcinogenic pathway and genetic alterations remain unclear, we investigated the histogenesis of this tumor by histopathological and p53 gene analysis.MethodsThe materials were 68 gastric endocrine cell carcinomas and 30 carcinoid tumors, which were resected from 93 Japanese patients for histopathological and immunohistochemical investigation. We also analyzed the concordance of p53 mutational status between the associated adenocarcinoma and endocrine cell carcinoma components, using microdissection and direct sequencing techniques.ResultsAn adenocarcinoma component was associated with 70.6% (48/68) of endocrine cell carcinomas, of which 42 (87.5%) were of well- to moderately differentiated type, while 36 of these 42 (85.7%) demonstrated histological continuity with the endocrine cell carcinoma components. Overexpression of p53 protein was observed in 58.8% (20/34) of cases. Common p53 mutational status between the two components was revealed in 73.3% (11/15) of cases analyzed. In contrast, carcinoid tumors did not exhibit p53 protein overexpression (0/15) or gene mutation (0/5).ConclusionsThese data suggest that gastric endocrine cell carcinomas predominantly arise from endocrine precursor cell clones occurring in preceding adenocarcinoma components (particularly the differentiated type), transforming into endocrine cell carcinoma during rapid clonal expansion under the influence of p53 gene alteration.

Early Colorectal Cancer with Special Reference to the Superficial Nonpolypoid Type from a Histopathologic Point of View

AbstractThe incidence and histopathologic characteristics of nonpolypoid (superficial type) early colorectal carcinomas were studied and compared with those of the polypoid type. The superficial type was subclassified as elevated (type IIa), type IIa with central depression (type IIa+IIc), plain (type IIb), depressed (type IIc), and IIc with marginal elevation (type IIc+IIa). The superficial type comprised 22% and 27% of intramucosal and submucosal carcinomas, respectively. Pure type IIb was not found, and there were only three pure type IIc lesions. Type IIa+IIc and IIc+IIa (and IIc) showed a significantly higher rate of submucosal invasion among the small tumors (59% and 71% less than 20 mm, respectively) compared to the polypoid type; type IIa showed no significant difference. The incidence of lymph node metastasis among submucosal carcinomas showed no significant difference between the superficial type and the polypoid type. About 64% and 52% of type IIa and IIa+IIc tumors accompanied residual adenoma, suggesting that they originated from small, flat adenomas through the adenoma—carcinoma sequence, whereas type IIc+IIa (and IIc) did not have an adenomatous component, implying that they arose de novo or originated through an adenoma—carcinoma sequence at a smaller size than the type IIa and IIa+IIc lesions. Superficial-type early colorectal carcinomas are not rare, and they are not uniform in nature. Rapid growth and invasion to the submucosa is characteristic of superficial-type lesions with a central depression, and only superficial depressed (type IIc+IIa, IIc) lesions can arise de novo. Although they grow rapidly to invade the submucosa, it cannot be said that they show more aggressive behavior than the polypoid type.

Topographic distribution of Helicobacter pylori in the resected stomach

Objectives The aim of this study was to elucidate the prevalence of Heliobacter pylori and its distribution in order to clarify the frequency of H. pylori infection and the most appropriate site of endoscopie biopsy for studies of H. pylori infection associated with different gastric diseases. Designs and methods Swiss role mucosal strips from 275 resected stomachs, which included the greater curvature, anterior wall and lesser curvature of the antrum, incisura and corpus, were stained with haematoxylin-eosin and H. pylori antibody. Results The prevalence of H. pylori infection was 97% in duodenal ulcers, 98% in gastric ulcers, 98% in intestinal-type carcinomas and 99% in diffuse-type carcinomas. H. pylori was present at a rate of 100% in any site in cases of duodenal ulcer, but was diffusely distributed in the antrum and patchily distributed in the corpus. The detection rate of H. pylori was 50–100% in gastric ulcers, 30–100% in intestinal-type adenocarcinomas and 63–100% in diffuse-type adenocarcinomas depending on the site of the stomach examined. Conclusions The prevalence of H. pylori infection was very high in peptic ulcers of the duodenum and stomach and gastric carcinomas of Japanese patients. Biopsy specimens for evaluation of H. pylori infection should be taken routinely from both the greater curvature of the antrum and corpus. Immunohistochemical staining should be used to assay for H. pylori when few organisms are present or eradication therapy has been used.

Correlation of p53 protein expression with gene mutation in gall‐bladder carcinomas

The correlation of p53 protein expression and p53 mutation of 33 gall‐bladder carclnomas was studied accordlng to the depth of invasion and grade of cytological atypia. Overexpresslon of p53 protein was detected by immunostaining in seven (70.00/) of 10 intramucosal and in 16 (69.6%) of 23 invasive carcinomas. p53 mutation was detected in five (71.4%) of the seven intramucosal carcinomas with overexpresslon and In eight (50.0%) of the 16 invasive cancers with overexpression and in one (10%) of the 10 non‐overexpressing carclnomas at exons 5–8 by nested polymerase chain reactlon‐single‐strand conformatlon polymorphism. The overexpression of p53 protein was present In nine (56.3%) of 16 low‐grade carcinomas and In 14 (82.3%) of 17 high‐grade carcinomas. In cases of overexpresslon, p53 mutatlon was detectable in four (44.4%) of nlne lowgrade and in nlne (64.3%) of 14 high‐grade carclnomas. In total, p53 mutation was verified In 56.5%0 (13123) of cases involving protein overexpression and in 10% (1/10) of cases of nonoverexpresslon. The sensltivity of p53 mutation was 56.5% (13/23), the specificity was 90.0% (9/10) and the validity was 1.47. in conclusion, our study indicates that p53 protein overexpression correlates well wlth gene mutatlon and that p53 alteration may be related to increaslng grade of cytologic atypla of carclnomas.

Multiple K‐ras Mutations in Hyperplasia and Carcinoma in Cases of Human Pancreatic Carcinoma

Mucous cell hyperplasia (MCH) has been considered an important precursor of pancreatic ductal carcinoma based on histological and molecular research, although various K‐ras mutations rates are seen among cases with pancreatic carcinoma, chronic pancreatitis and normal pancreas, with a wide range of histological characters. To investigate the premalignant potential of MCH and the multicentricity of pancreatic carcinoma, we analyzed K‐ras mutation at codon 12 in carcinoma foci of 82 cases of surgically‐resected pancreatic carcinoma [67 solid‐type carcinomas (SCs) and 15 ductectatic‐type carcinomas (DCs)], as well as in both MCH and carcinoma foci in 42 cases (30 SCs and 12 DCs), using an enriched polymerase chain reaction (PCR)‐enzyme linked mini‐sequence assay (ELMA). K‐ras mutation was recognized in 85% (57/67) of SCs and 73% (11/15) of DCs, and multiple K‐ras mutations in 12% (8/67) of SCs and in 20% (3/15) of DCs. Multiple K‐ras mutations were also recognized in MCHs in 47% (14/30) of SCs and in 42% (5/12) of DCs. Moreover, the same sequence at K‐ras codon 12 in MCH and carcinoma was identified in 76% (32/42) of carcinoma cases and it was more frequently recognized in hyperplasias with histological atypia (51%, 37 of 72 foci) than those without atypia (24%, 16 of 68 foci) (P < 0.0007). These results further support the idea of multicentric carcinogenesis and premalignant potential of atypical hyperplasia in the human pancreas, although about half of the hyperplasias around carcinomas were not thought to be direct precursors.

Risk factors for nodal micrometastasis of submucosal gastric carcinoma: assessment of indications for endoscopic treatment

Background. Despite the widespread use of endoscopic mucosal resection (EMR) for intramucosal gastric carcinoma, there is no standardized therapy for those patients in whom the carcinoma is found, after EMR, to have invaded the submucosa.Our aim in this study was to examine the relationship between the clinicopathological features of submucosal invasive carcinomas and their incidence of nodal micrometastasis, as detected by anti-human cytokeratin immunohistochemistry, in order to assess the curative potential of submucosal carcinoma by EMR. Methods. Fifty surgically resected submucosal gastric carcinomas which would have satisfied the absolute indications for EMR, except for the criterion of submucosal invasion, were examined. The extent of submucosal invasion was determined by measuring its vertical and horizontal spread. Immunohistochemical analysis was performed with anti-human cytokeratin antibody (CAM5.2). Results. Three of 50 cases (6.0%) were positive for nodal metastasis by routine H&E examination. Nodal micrometastases were detected in 11 of 47 cases (23.4%). Statistical analysis revealed that both the depth and the width of carcinoma in the submucosa were significantly larger in cases with than in those without micrometastasis (P = 0.019 and P = 0.006, respectively). The group with lymphatic invasion showed more frequent micrometastasis than the group without (P = 0.014). There were no micrometastases in submucosal carcinomas whose submucosal invasion was less than 200 μm vertically and less than 320 μm horizontally. Conclusions. The present study indicates that differentiated gastric adenocarcinoma with minimal submucosal invasion (less than 200 μm vertically and less than 320 μm horizontally) and not accompanied by peptic ulcer or other risk factors, such as lymphatic invasion, can be considered as having high curative potential by EMR alone, without the necessity for further radical surgery.

Bidirectional gastric differentiation in cellular mucin phenotype (foveolar and pyloric) in serrated adenoma and hyperplastic polyp of the colorectum

This study examined whether gastric pyloric gland‐type mucin is expressed in serrated adenoma (SA) and in hyperplastic polyp (HP) of the colorectum, and whether cellular position‐based gastric differentiation is observed in these lesions as previously hypothesized. Immunostaining was performed for MUC6 and α‐linked GlcNAc residue (pyloric gland‐type mucin markers), human gastric mucin (HGM; foveolar‐type mucin marker) and Ki‐67 (proliferating cell marker) for 31 SA, 22 HP, 21 traditional tubular adenoma (TA) and 20 hyperplastic nodule (HN). MUC6 showed varying expression in SA, 22/31 (71.0%); HP, 15/22 (68.2%); TA, 2/21 (9.5%); and HN, 0/20 (0%) with significantly higher frequencies in SA and HP compared to those in TA and HN. The α‐linked GlcNAc residue was found only in SA (3/31, 9.7%) and in HP (2/22, 9.1%). In SA and HP, HGM was typically expressed in the entire crypt length, but some reduction in expression was shown in the basal crypt portion below the proliferative zone. MUC6 and α‐linked GlcNAc residues were expressed in the basal crypt portion below or below and including proliferative zone. These data demonstrate that SA and HP show bidirectional gastric (foveolar and pyloric gland) differentiation with respect to mucin cellular phenotype and the potential for cellular position‐based differentiation, which mimics the gastric antral mucosa.