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Primary eosinophilic gastrointestinal disorders (EGIDs) constitute chronic allergic inflammation. The number of eosinophils is one of the diagnostic criteria; more than 20 eosinophils per high-power field (HPF) in the gastrointestinal (GI) tract are considered abnormal in Japan. However, the quantity of eosinophils considered normal varies according to anatomical location and geographical region; such values have not been reported in Japanese pediatric patients, nor have the numbers of lymphocytes in the normal pediatric stomach. To establish a reference for defining diagnostic criteria for EGIDs, we evaluated the number of eosinophils in the normal Japanese pediatric GI tract.We examined 131 biopsy cases without significant clinical history, endoscopic abnormality, or histological abnormality. Immunohistochemical analysis of CD3 and CD20 was performed.The mean eosinophil density was highest in the cecum (49.5 ± 22.4 per HPF). Counts of more than 20 eosinophils per HPF were observed in the duodenum [bulb (20.0 ± 9.6) and second portion (30.0 ± 15.8)], terminal ileum (38.3 ± 22.7), cecum (49.5 ± 22.4), ascending colon (42.3 ± 25.3), transverse colon (29.4 ± 17.0), and descending colon (32.2 ± 17.9). Counts of fewer than 10 eosinophils per HPF were observed in the stomach and rectum; a count of fewer than one eosinophil per HPF was observed in the esophagus. More than 100 CD3-positive T cells per HPF were observed in the stomach.The mean numbers of eosinophils in the bowel were greater than 20 per HPF. For Japanese pediatrics, the current threshold eosinophil count should be revised.

Most gastric neuroendocrine tumors (NETs) develop from enterochromaffin-like (ECL) cells. ECL-cell NETs are classically categorized into three types according to their etiology. A 50-year-old woman presented with submucosal tumor-like lesions in the stomach, which were identified via esophagogastroduodenoscopy. Although esophagogastroduodenoscopy and pathological findings of biopsy specimens showed an absence of mucosal atrophy in the body of the stomach, sticky, adherent, dense mucus was observed. All lesions were diagnosed as ECL-cell NETs based on histological examination findings; however, ECL-cell NETs did not apply to any of the classic types I-III categorization based on laboratory, computed tomography, and 24-h intragastric pH monitoring test findings. Endoscopic submucosal dissection of the tumor was performed. Pathological findings of the excised specimen indicated that parietal cell hyperplasia with a protrusion, dilated fundic glands, and endocrine cell hyperplasia in the background mucosa, and parietal cells were not immunostained for the α-subunits of H

Background ADP-ribosylation factor-like protein 4 C (ARL4C) is a member of the ARF small GTP-binding protein subfamily. The ARL4C gene is highly expressed in colorectal cancer (CRC). ARL4C protein promotes cell motility, invasion, and proliferation. Methods We investigated the characteristics of ARL4C by comparing its expression at the invasion front and relationships with clinicopathological data using RNAscope, a highly sensitive RNA in situ method. Results In all cases, ARL4C expression was observed in cancer stromal cells and cancer cells. ARL4C expression in cancer cells was localized at the invasion front. In cancer stromal cells, ARL4C expression was significantly stronger in cases with high-grade tumor budding than in cases with low-grade tumor budding (P = 0.0002). Additionally, ARL4C expression was significantly increased in patients with high histological grade compared with those with low histological grade (P = 0.0227). Furthermore, ARL4C expression was significantly stronger in lesions with the epithelial-to-mesenchymal transition (EMT) phenotype compared with the non-EMT phenotype (P = 0.0289). In CRC cells, ARL4C expression was significantly stronger in cells that had the EMT phenotype compared with those with a non-EMT phenotype (P = 0.0366). ARL4C expression was significantly higher in cancer stromal cells than in CRC cells (P Conclusion Our analysis reinforces the possibility that ARL4C expression worsens the prognosis of patients with CRC. Further elucidation of the function of ARL4C is desired.

In 2020, the WHO published a new system for classifying invasive endocervical adenocarcinoma based on histological features and high-risk human papillomavirus (HPV) infection. However, immunophenotypes of each histological subtype require further investigation. We immunohistochemically analyzed 66 invasive endocervical adenocarcinomas using three cell-lineage–specific markers: claudin 18 (CLDN18) for gastric, cadherin 17 (CDH17) for intestinal, and PAX8 for Mullerian epithelial cells. We identified five immunophenotypes of endocervical adenocarcinoma: gastric (21%); intestinal (14%); gastrointestinal (11%); Mullerian (35%); and not otherwise specified (NOS) (20%). Adenocarcinomas with gastric immunophenotype, characterized by aging (p = 0.0050), infrequent HPV infection (p

SUMMARYHelicobacter suishosted by hogs is the most prevalent gastric non-Helicobacter pylori Helicobacterspecies found in humans. Recent studies suggest that theH. suisinfection has already induced many cases of gastric disease. However, the infection period and route ofH. suisfrom hogs remain unclear. Because diagnostic methods based on the urease activity ofH. suisoften yield negative judgments, there is no reliable method for diagnosingH. suisinfection in clinical practice without gastric biopsy specimens. We developed the world’s first ELISA to simultaneously diagnoseH. suisandH. pyloriinfection in a single test. The area under the ROC curve was 0.9648 or 0.9200 for identifyingH. suisorH. pyloriinfection, respectively. The sensitivity, specificity, and positive and negative predictive values for identifyingH. suisinfection were 100%, 92.6%, 76.9%, and 100%, and those for identifyingH. pyloriinfection were 88.2%, 87.5%, 65.2%, and 96.6%, respectively. (150 words)

Background: Tumor budding (TB) is a poor prognostic factor in colorectal adenocarcinoma (CA), but the underlying mechanism remains unclear. Interleukin-6 (IL6) is one of the main cytokines produced by cancer-associated fibroblasts. IL6 is linked with cancer progression and poor prognosis by activating cancer cells and modifying the cancer microenvironment. However, little is known about the expression of IL6 in TB and its association with TB in CA. Methods: The clinicopathological and prognostic significance of IL6 in TB was examined using a tissue microarray consisting of 36 patient samples of TB in CA. IL6 mRNA was detected by RNAscope. Patients were stratified into negative and positive IL6 expression groups. Results: IL6 expression was overwhelmingly observed in cancer stroma but was negligible in cancer cells. In the IL6-positive group in cancer stroma, TB grade was higher than in the IL6-negative group (P=0.0161). The IL6-positive group significantly exhibited the epithelial–mesenchymal transition (EMT) phenotype compared with the IL6-negative group in cancer stroma (P=0.0301). There was no significant difference in overall survival between CA cases in the IL6-positive and -negative groups in cancer stroma. Conclusions: TB may be affected by IL6 expression, and IL6expression in cancer stroma at TB may be an important prognostic marker.

Helicobacter pylori eradication has been shown to reduce the risk of gastric cancer (GC), with the number of eradication therapy cases on the rise. However, GC can still occur after successful treatment, and the histological differences prior to eradication in patients with and without GC are unclear. This study investigated the pre-treatment histological risk factors for GC development following eradication therapy.We retrospectively enrolled consecutive adult patients diagnosed as having H. pylori infection between April 2004 and December 2018. Atrophy and intestinal metaplasia (IM) were histologically assessed according to the updated Sydney System. The operative link on gastritis assessment and the operative link on gastric intestinal metaplasia (OLGIM) were evaluated as well.Of the 247 patients analyzed in this study, 11 (4.5%) experienced GC after eradication therapy. Histological IM scores in the GC group were significantly higher at all gastric biopsy sites (p .05), and the proportion of OLGIM III/IV stage was significantly greater in GC patients (81.8% vs. 31.8%, p .01). For GC prediction, the area under the receiver operating characteristic curve for IM score at the lesser curvature of the corpus was the highest among all biopsy sites and not inferior to OLGIM results.Patients with histological IM prior to H. pylori eradication, especially at the lesser curvature of the corpus, may be at elevated risk for GC development after eradication therapy and require close surveillance.

LGR5 is a promising stem cell marker in gastric pylorus, but there are few reports on its expression in human gastric corpus. To investigate the involvement of LGR5 expression in gastric corpus ulcer regeneration in humans. LGR5 expression was analyzed in five post-ESD ulcers during the healing process of regenerating epithelial cells of the gastric corpus. LGR5 expression was detected by mRNA in situ hybridization using an RNA scope kit. Immunohistochemistry of MUC6, HIK1083, and pepsinogen 1 (PG1) was performed to identify cell differentiation. We defined MUC6+/HIK1083−/PG1−, MUC6+/HIK1083+/PG1−, MUC6+/HIK1083+/PG1+, MUC6+/HIK1083−/PG1+, and MUC6-/HIK1083−/PG1+cells as pseudopyloric mucosa (PPM) phase 1 (PPM1), PPM phase 2 (PPM2), PPM phase 3 (PPM3), immature chief cells (ICC), and mature chief cells (MCC) in order from the ulcer center, respectively. In the regenerated mucosa around post-ESD ulcers, LGR5 expression was observed throughout the gland in PPM1–PPM3, but it was limited to the bottom of the gland in ICC and MCC. Furthermore, LGR5 expression was not identified in the normal gastric corpus. The H-score of PPM2 was significantly higher than that of PPM3 (P = 0.0313). The H-score of PPM3 was significantly higher than that of ICC (P = 0.0313). The LGR5 H-score was higher at the immature stage, which decreased gradually with progression of the differentiation stage. LGR5 expression appears to contribute to mucosal regeneration in the human gastric corpus. The application of LGR5 expression analysis to mucosal regeneration and fundic gland-type gastric tumors is expected.

Background: Interleukin-6 (IL6) is one of the main cytokines produced by cancer-associated fibroblasts (CAFs). IL6 is linked with cancer progression and poor prognosis by activating cancer cells and modifying the cancer microenvironment. However, little is known about the expression of IL6 in tumor budding (TB) and its association with TB in colorectal adenocarcinoma. Methods: The clinicopathological and prognostic significance of IL6 in TB was examined using a tissue microarray consisting of 36 patient samples of TB in CA. IL6 mRNA was detected by RNAscope kit. Patients were stratified into negative and positive IL6 expression groups. Results: IL6 expression was overwhelmingly observed in CAFs but was negligible in cancer cells. In the IL6-positive group in CAFs, TB grade was higher than in the IL6-negative group (P=0.0161). There was a significant difference in overall survival (OS) between CA cases in the IL6-positive group and the IL6-negative group (log rank test, P=0.0367). Cox proportional hazard regression model revealed that the IL6-negative group (OR = 0.25; 95% CI: 0.05–0.96; P=0.0440) had better OS for CA than the IL6-positive group. Conclusions: TB may be affected by IL6 expression, and IL6 expression in CAFs at TB may make IL6 an important prognostic marker.

Background. Immunoglobulin (Ig) G4-related diseases (RDs) are systemic diseases in which serum IgG4 levels are frequently elevated. They can cause diffuse or focal tumor formation, organ swelling, and tissue thickening in organs infiltrated by IgG4 + plasma cells. The diagnostic criteria for IgG4-RDs include an IgG4/IgG ratio >40%, but counting IgG+ cells can be difficult because of the weakness of IgG staining density. We hypothesized that an antibody cocktail of mixed IgG1, IgG2, IgG3, and IgG4 (AC-IgG) might give immunohistochemistry results comparable with those of IgG in IgG4-RD. Methods. We compared AC-IgG reactivity with IgG expression in type 1 autoimmune pancreatitis (AIP), a representative IgG4-RD. We compared immunohistochemistry results using AC-IgG and IgG-only in 10 cases of AIP. The coefficient of variation (Cv) was used to analyze differences between AC-IgG and IgG findings in AIP by 13 board-certified pathologists. Results. Although mean values for IgG+ cells did not significantly differ between AC-IgG (34.3; range = 27.4-37.1) and IgG (30.0; range = 23.0-45.6; P = .6254), Cv was lower for AC-IgG (33.4%) than for IgG (51.4%; regression equation; y[IgG] = 0.988 x + 0.982; correlation coefficient = 0.907). The data showed that the results of both methods were largely consistent. Conclusion. AC-IgG could replace IgG to count IgG+ cells because of its lower Cv.

A classification system for invasive endocervical adenocarcinoma (ECA) focusing on high-risk human papillomavirus (HPV) detection has been recently developed. However, precursor lesions of each ECA subtype and immunohistochemical markers that effectively subcategorize ECAs with gastric and intestinal differentiation have not been fully described. Here, we aimed to subcategorize endocervical adenocarcinoma in situ (AIS) by immunophenotype and to characterize the histopathology of each AIS subtype. We immunohistochemically analyzed 36 AIS and 25 lobular endocervical glandular hyperplasia (LEGH) samples using three cell lineage-specific markers (CLDN18, gastric epithelial cells; CDH17, intestinal epithelial cells; and PAX8, Müllerian epithelial cells). The AISs were immunophenotypically classified as gastric-type (G-AIS; n = 2), intestinal-type (I-AIS; n = 10), gastrointestinal-type (GI-AIS; n = 3), Müllerian-type (M-AIS; n = 18), and AIS, not otherwise specified (AIS-NOS; n = 3). All 25 LEGHs were categorized as gastric-type. G-AIS had pale eosinophilic or clear cytoplasm with a small amount of apical mucin and fewer mitotic bodies. I-AIS comprised various numbers of goblet cell-type tumor cells. GI-AIS showed intermediate or mixed features of G-AIS and I-AIS. M-AIS, as with the usual-type ECA, was typically characterized by mucin depletion; however, several lesions had abundant cytoplasmic mucin. High-risk HPV was detected in most AISs but was negative in 100% (2/2) of G-AIS, 10% (1/10) of I-AIS, and 6% (1/18) of M-AIS lesions. In summary, the AIS subtypes defined by immunophenotype had distinct histopathological and etiological characteristics. Thus, immunophenotyping with CLDN18, CDH17, and PAX8 might improve the diagnostic accuracy of histopathological classifications of ECAs.

Background Leucine-rich repeat-containing G-protein-coupled receptor 5 (LGR5) is a strong cancer stem cell marker in colorectal cancer; however, there are many unclear aspects of LGR5 expression in pancreatic cancer. It has been reported that the interaction between tumor cells and stroma at the fat infiltration site has a significant effect on pancreatic cancer prognosis. Therefore, we report a clinicopathological study of LGR5 expression at the fat invasion front in pancreatic cancer. Methods LGR5 expression was analyzed in 40 pancreatic ductal adenocarcinoma cases with RNAscope, which is a newly developed high-sensitivity in situ hybridization method. Epithelial-mesenchymal transition (EMT) was analyzed by the expression of E-cadherin and vimentin via immunohistochemistry. Results LGR5-positive dots were identified in all cases, especially with glandular formation. In the fat invasion front, a high histological grade showed significantly reduced LGR5 expression compared with a low histological grade (p=0.0126). LGR5 expression was significantly higher in the non-EMT phenotype group than in EMT phenotype group (p=0.0003). Additionally, LGR5 expression was significantly lower in cases with high vascular invasion than in those with low vascular invasion (p=0.0244). Conclusions These findings suggest that decreased LGR5 expression in the fat invasion front is associated with more aggressive biological behavior in pancreatic ductal adenocarcinoma, with higher tumor grade, EMT phenotype, and higher vascular invasion.

Background: Leucine-rich repeat-containing G-protein-coupled receptor 6 (LGR6) promotes carcinogenesis and progression in some cancer types. However, there are few reports of LGR6 expression in esophageal squamous cell carcinoma (ESCC). LGR6 expression and clinicopathological features in ESCC were investigated by RNAscope, a high-sensitivity RNA in situ hybridization method.Methods: Appropriate tumors were selected from 41 cases of ESCC from which tissue microarrays were generated, and LGR6 expression was identified by RNAscope. Results: Thirty-seven patients had LGR6 expression. High LGR6 expression was observed in 17 cases and low LGR6 expression in 24 cases. LGR6 expression was significantly higher in high histological grade ESCC than in low histological grade ESCC (P=0.0023). ESCC patients who received neoadjuvant chemotherapy had significantly higher LGR6 expression than those without neoadjuvant chemotherapy (P=0.0109). Furthermore, high LGR6 expression showed a poorer prognosis than low LGR6 expression (log-rank test, P=0.0365).Conclusions: LGR6 may be a prognostic factor and a potential new therapeutic target in ESCC.

Background Lobular endocervical glandular hyperplasia (LEGH) was first described by Nucci et al. in 1999 and is believed to be a precancerous lesion of minimal deviation adenocarcinoma and gastric-type adenocarcinoma in the uterine cervix. LEGH lesions do not always exhibit apparent cellular and structural atypia, so are difficult to distinguish from normal endocervical cells (EC cells) with cytological examination. Therefore, we often struggle to make a definite diagnosis of LEGH. Methods We used microscopy images of cytological specimens that were diagnosed as EC cells and LEGH cells. Signal intensity in whole nuclear area and in heterochromatin and euchromatin regions, euchromatin area ratio, and nuclear morphological features were quantified in each cell nucleus of the cases. Statistical analyses were conducted to determine statistical significance. Finally, we performed linear support vector machine (LSVM) modeling as a discriminant analysis using the quantified features. Results Signal intensity in whole nuclear area, and heterochromatin and euchromatin regions of EC cell nuclei were higher than that of the LEGH cell nuclei. Morphologically, EC cell nuclei were larger than LEGH cell nuclei, and nuclei of LEGH cells had irregular nuclear respectively membrane structure and an elongated shape. The LSVM accuracy of 10-fold cross validation and leave-one-case-out cross-validation (LOCOCV) using all measured features were 84.7% to 89.3% and 78.6% to 86.0%, respectively. Conclusions The LVSM analysis using features extracted from signal intensity and morphological analysis was useful for discrimination of EC cells vs LEGH cells. We therefore believe that this image analysis method could be used for early detection of LEGH.

Background Non-Helicobacter pylori Helicobacter (NHPH) is not widely recognized as a cause of acute gastric mucosal lesions (AGML), as only a few cases of AGML caused by NHPH have been reported. We present here one case and examine the species and eradication of NHPH together with the three previously reported cases. Case presentation A 52-year-old woman presented with a two-day history of severe epigastric pain, nausea, and vomiting. An esophagogastroduodenoscopy showed mucosal edema, multiple erosions, and ulcerations in the antrum. Biopsy specimens taken from the antrum revealed long spiral-shaped organisms, suggesting NHPH. As both serum anti-Helicobacter pylori (H. pylori) antibody and H. pylori stool antigen test were negative, this case was diagnosed as AGML caused by NHPH. After the administration of esomeprazole 20 mg for 14 days and the interval of the following 12 days, AGML was deemed to have been cured endoscopically. In addition, microscopic examination and PCR analysis confirmed the success of NHPH eradication. Conclusions NHPH should be considered a probable cause of AGML in cases that are not attributed to the other causes already recognized. Taking probability of spontaneous eradication into consideration, it is appropriate to start eradication therapy after confirming the chronicity of NHPH infection.

Gastric adenocarcinoma (GAC) can be divided immunophenotypically into gastric, intestinal, or mixed gastric and intestinal phenotypes. Cadherin 17 (CDH17) and CD10 have been used as comprehensive markers for intestinal epithelial cells and for small intestinal absorptive cells in GACs, respectively. Sucrase-isomaltase (SI) and CD10 are expressed in small intestinal absorptive cells and SI is more frequently expressed than CD10 in gastric intestinal metaplasia (IM). The aim of this study was to evaluate the potential of SI as a marker for intestinal absorptive cells compared to CD10 in differentiated-type (DT) GACs. We compared the immunohistochemical expression of CDH17, SI, and CD10 in IMs and tissue microarrays of 40 samples of DTGACs. In IMs and DTGACs, CDH17 showed a diffuse lateral cytoplasmic membrane staining both in columnar and goblet cells. SI and CD10 were expressed on the luminal surfaces of the columnar cells. In IMs, SI was positive both in both complete-type IMs and in incomplete-type IMs. CD10 was positive only in complete-type IMs. In DTGACs, CDH17, SI, and CD 10 were positive in 37 (92.5%), 22 (55%), and 11 (27.5%) cases, respectively. In SI-positive cases, the degrees of expression of SI were equal to (7 cases) or less than (15 cases) those of CDH17; the degrees of expression of SI were equal to (5 cases), more than (16 cases), or less than (1 case) those of CD10. In conclusion, SI is a more sensitive immunohistochemical marker for intestinal absorptive cells than CD10 in DTGACs.

Background and aim There are only a few reports of non-Helicobacter pylori Helicobacter (NHPH) gastritis in Japanese patients. We aimed to examine its prevalence, clinical features, and esophagogastroduodenoscopy (EGD) findings based on 50 patients encountered in one facility. Materials and methods Subjects were all patients who had undergone gastric mucosal biopsy endoscopically at Kenwakai Hospital for approximately 10 years. NHPH infection was diagnosed by microscopic findings of Giemsa staining performed on all specimens. PCR analysis of urease genes was performed to detect and identify NHPH, when informed consent was obtained. Helicobacter pylori-diagnostic tests were also performed. NHPH-infected patients were questioned about symptoms and animal contact. Results NHPH gastritis was found in 50 of 3847 patients (1.30%). The percentage increased to 3.35% (30 of 896 patients) in the latter 2 years and 4 months with increasing recognition of its characteristic endoscopic findings by endoscopists. PCR analysis, performed in 30 patients, detected NHPH in 28 patients: 26 as Helicobacter suis and 2 as Helicobacter heilmanii/Helicobacter ailurogastricus. Helicobacter pylori-diagnostic tests were almost negative. However, anti-H. pylori antibody showed high-negative titer (3.0-9.9 U/ml) in 12. Of 50 patients (consisting of 49 men and 1 woman), almost all were asymptomatic, and 25 were keeping pets. Regarding EGD findings, in all 50 patients, "crack-like mucosa" and/or nodular gastritis was noted in gastric antrum, and regular arrangement of collecting venules (RAC) was noted in gastric corpus. None of the patients infected with NHPH were co-infected with H. pylori. Conclusions The prevalence was finally estimated to be approximately 3.35%. Helicobacter suis was the most common NHPH species. "Crack-like mucosa" and/or nodular gastritis in gastric antrum, RAC in gastric corpus, and H. pylori-negativity by H. pylori-diagnostic tests especially containing a high-negative titer of anti-H. pylori antibody may indicate NHPH infection.

Background It is difficult to distinguish between multicentric Castleman’s disease (MCD) and IgG4-related lung disease (IgG4-LD), an IgG4-related disease (IgG4-RD) in the lung. Methods We focused on IL-6, which is elevated in MCD, to distinguish between MCD and IgG4-LD by RNAscope, a highly sensitive RNA in situ method. Six cases of MCD and four cases of IgG4-LD were selected. Results In all cases of MCD and IgG4-LD, 10 or more IgG4-positive cells were found in one high-power field. All MCD cases were inconsistent with the pathological IgG4-related comprehensive diagnostic criteria, but 2 of 6 cases had an IgG4/IgG ratio greater than 40%. In all IgG4-LD cases, histological features were consistent with the pathological IgG4-RD comprehensive diagnostic criteria. IL-6 expression was observed in all MCD and IgG4-LD cases except for one IgG4-LD biopsy. IL-6-expressing cells were mainly identified in the stroma. Sites of IL-6 expression were not characteristic and were sparse. IL-6 expression tended to be higher in MCD compared with IgG4-LD. A positive correlation was found between the IL-6 H-score and serum IL-6 level. Conclusion Differences in IL-6 expression may help distinguish between MCD and IgG4-LD. In addition, the presence of high IL-6 levels may help elucidate the pathological mechanisms of IgG4-LD.

AIMS Claudin 18 (CLDN18) is a member of the claudin family of cell surface proteins, which are widely expressed in epithelial cells and play a role in cell-cell adhesion. CLDN18 isoform 2 (CLDN18.2) is specifically expressed in gastric epithelial cells, and is frequently expressed at high levels in gastric adenocarcinoma. On the basis of this, zolbetuximab, a targeted monoclonal antibody, has been developed for patients with CLDN18.2-positive gastro-oesophageal adenocarcinoma. Colitis-associated colorectal adenocarcinomas (CACs) tend to lose intestinal markers and show aberrant gastric mucin expression. Furthermore, clinical trials of human epidermal growth factor receptor 2 (HER2) inhibitor therapy for colorectal carcinoma are ongoing. However, the expression profile of CLDN18.2 and HER2 has not been described in a series of human CACs. METHODS AND RESULTS We performed immunohistochemistry for CLDN18 and HER2 on 56 consecutive CACs from 55 inflammatory bowel disease patients, and compared the expression profile with that of a control group of 56 sporadic colorectal adenocarcinomas (CRCs). CLDN18.1 expression and CLDN18.2 expression were validated by reverse transcription polymerase chain reaction (PCR) in paraffin-embedded CRC tissues. CLDN18 was positive in 27% (15/56) of CACs and in 5% (3/56) of sporadic CRCs (P = 0.004), and CLDN18-positive CACs were more likely to have lymph node metastasis than CLDN18-negative CACs (67% versus 36%; P = 0.017). CLDN18 expression was significantly associated with MUC5AC expression (P

Background Leucine-rich repeat-containing G-protein-coupled receptor 5 (LGR5), a component of the Wnt receptor complex, is thought to lineage label gastric and intestinal stem cells. LGR5 expression is increased in colorectal carcinoma (CRC) compared to normal tissue. Colitis associated colorectal adenocarcinoma (CAC) often shows distinct morphologic and molecular phenotypes compared to sporadic cases. However, the expression profile of LGR5, and by extension the potential role of an intestinal stem cell phenotype, has not been well described in a series of human CAC. Method RNA in situ hybridization (ISH) for LGR5 expression on 30 CACs (12 cases with conventional morphology and 18 cases with non-conventional type morphology) from 29 inflammatory bowel disease (IBD) patients was performed and compared the expression profile to a control group of 10 sporadic CRCs. Immunohistochemistry for beta-catenin and SATB2 was performed on the 30 CACs. Result LGR5 was positive in 30% (9/30) of CAC cases and 90% (9/10) of sporadic CRCs (p = 0.002). A large majority (89%) of LGR5 positive CACs were of the conventional histologic type, and conventional type CAC showed a significantly higher LGR5 score (median 3.0; interquartile range 1.75–3.25) than non-conventional type CAC (median 1.5; interquartile range 1.00–2.00) (p = 0.034). CAC with conventional morphology did have a lower level of LGR5 expression than sporadic CRC. Sporadic CRCs showed a significantly higher LGR5 level score than non-conventional type CACs (p LGR5 expression (p = 0.003), however no significant association was identified between SATB2 expression and LGR5 expression status in CACs. Conclusion These findings suggest that the wider spectrum of tumor morphology in CAC may be associated with absence of a LGR5-expressing intestinal stem cell phenotype.

Colon cancer stem cells (CSCs) are closely related to tumorigenesis and treatment response, and LGR5 is currently the most robust and reliable CSC marker in colorectal cancer (CRC). However, LGR5 expression in CRC tumor budding (TB) is not well understood. We examined the clinicopathological and prognostic significance of LGR5 in CRC TB. LGR5 expression was evaluated by RNAscope, a newly developed RNA in situ hybridization technique, using a tissue microarray consisting of 55 patient samples of TB in colon adenocarcinoma (CA) selected from the medical archives at our hospital. Patients were stratified into negative and positive LGR5 expression groups. Tumor-infiltrating lymphocytes (TILs) and histological grade were lower in the LGR5-positive group compared with the LGR5-negative group (P = .0407 and P = .0436, respectively). There was no significant difference in overall survival between the LGR5-positive group and the LGR5-negative group (log-rank test, P = .6931). LGR5 expression did not remain a predictor of prognosis in univariate analysis (OR = 0.84, 95% CI: 0.33–2.02, P = .6928). LGR5 expression may be affected by TILs, which have been demonstrated to be associated with worse prognosis in the budding area of CA and is an important potential marker of prognosis.

BackgroundLeucine-rich repeat-containing G-protein-coupled receptor 5 (LGR5), a component of the Wnt receptor complex, is thought to lineage label gastric and intestinal stem cells. LGR5 expression is increased in colorectal carcinoma (CRC) compared to normal tissue. Colitis associated colorectal adenocarcinoma (CAC) often shows distinct morphologic and molecular phenotypes compared to sporadic cases. However, the expression profile of LGR5, and by extension the potential role of an intestinal stem cell phenotype, has not been well described in a series of human CAC. MethodRNA in situ hybridization (ISH) for LGR5 expression on 30 CACs (12 cases with conventional morphology and 18 cases with non-conventional type morphology) from 29 inflammatory bowel disease (IBD) patients was performed and compared the expression profile to a control group of 10 sporadic CRCs. ResultLGR5 was positive in 30% (9/30) of CAC cases and 90% (9/10) of sporadic CRCs (p=0.002). A large majority (89%) of LGR5 positive CACs were of the conventional histologic type, and conventional type CAC showed a significantly higher LGR5 score (median 3.0; interquartile range 1.75-3.25) than non-conventional type CAC (median 1.5; interquartile range 1.00-2.00) (p = 0.034). CAC with conventional morphology did have a lower level of LGR5 expression than sporadic CRC. Sporadic CRCs showed a significantly higher LGR5 level score than non-conventional type CACs (p ConclusionThese findings suggest that the wider spectrum of tumor morphology in CAC may be associated with absence of a LGR5-expressing intestinal stem cell phenotype.

Background: Interleukin-6 (IL6) is one of the main cytokines produced by cancer-associated fibroblasts (CAFs). IL6 is linked with cancer progression and poor prognosis by activating cancer cells and modifying the cancer microenvironment. However, little is known about the expression of IL6 in tumor budding (TB) and its association with TB in colon adenocarcinoma (CA). Methods: The clinicopathological and prognostic significance of IL6 in TB was examined using a tissue microarray consisting of 36 patient samples of TB in CA. IL6 mRNA was detected by RNAscope kit. Patients were stratified into negative and positive IL6 expression groups.Results: IL6 expression was overwhelmingly observed in CAFs but was negligible in cancer cells. In the IL6-positive group in CAFs, TB grade was higher than in the IL6-negative group (P=0.0161). There was a significant difference in overall survival (OS) between CA cases in the IL6-positive group and the IL6-negative group (log rank test, P=0.0367). Cox proportional hazard regression model revealed that the IL6-negative group (OR = 0.25; 95% CI: 0.05–0.96; P=0.0440) had better OS for CA than the IL6-positive group. Conclusions: TB may be affected by IL6 expression, and IL6 expression in CAFs at TB may make IL6 an important prognostic marker.

Gastric-type adenocarcinoma (GA) is an aggressive subtype of cancer of the uterine cervix. Several immunohistochemical markers for gastric mucins, such as mucin 6 (MUC6) and N-acetylglucosamine α1 → 4galactose → R (αGlcNAc-R), which is recognized by HIK1083 antibody, have been introduced for diagnosis of GA and lobular endocervical glandular hyperplasia (LEGH). However, MUC6 is also expressed in normal endocervical glands and HIK1083 antibody has limited availability. Trefoil factor family 2 protein (TFF2) is secreted by gastric, but not normal endocervical glands. Here, we evaluated TFF2 immunostaining for detection of a gastric immunophenotype in endocervical glandular lesions. We compared TFF2, αGlcNAc-R, and MUC6 expression in 103 endocervical glandular lesions: LEGH (n = 23), adenocarcinoma in situ/microinvasive adenocarcinoma (AIS-MIA) (n = 29), and invasive adenocarcinoma (usual type [UA], n = 26; GA, n = 11; intestinal type [IA], n = 2; signet ring cell type [Sig], n = 2; and mucinous adenocarcinoma not otherwise specified [NOS], n = 10). TFF2 and αGlcNAc-R expression was completely concordant in each subtype: LEGH (100%), AIS-MIA (44.8%), UA (26.9%), GA (90.9%), IA (100%), Sig (0%), and NOS (20%). TFF2 staining scores were significantly correlated with those of αGlcNAc-R in these lesions. TFF2 and αGlcNAc-R immunoreactivity was present in cytoplasmic mucins and luminal secretions. TFF2 and αGlcNAc-R were not expressed in the normal endocervical glands. MUC6 was frequently expressed in normal endocervical glands and endocervical glandular lesions. Endocervical adenocarcinomas sometimes stained only for MUC6. TFF2 is a promising immunohistochemical marker and its identification in uterine cervical secretion is a potentially useful diagnostic test for endocervical glandular lesions with gastric differentiation.

LGR5 is expressed in various tumors and has been identified as a putative intestinal stem cell marker. Here we investigated LGR5 expression in colorectal neuroendocrine neoplasms and analyzed the correlation with pathological characteristics. We evaluated the clinicopathological features of 8 neuroendocrine tumor (NET) grade 1 (NET G1), 4 NET Grade 2 (NET G2), and 8 NET Grade 3 (NET G3; also termed neuroendocrine carcinoma, or NEC) cases. We examined LGR5 expression using an RNAscope, a newly developed RNA in situ hybridization technique, with a tissue microarray of the neuroendocrine neoplasm samples. LGR5 staining in individual tumor cells was semi-quantitatively scored using an H-score scale. We also performed a combination of LGR5 RNA in situ hybridization and synaptophysin immunohistochemistry. All cases contained tumor cells with some LGR5-positive dots. For all cases, H-scores showed a positive correlation with nuclear beta-catenin expression. In the NEC group, there was a strong positive correlation between H-score and beta-catenin expression. Our findings suggest that LGR5 may serve as a stem cell marker in NEC, as is the case in colon adenocarcinoma. The positive correlation between H-score and beta-catenin expression suggests that LGR5 expression might be affected by beta-catenin expression in neuroendocrine neoplasms and especially in NEC.

The relationship between methylation abnormality and autoimmune pancreatitis (AIP)-a representative IgG4-related disease-has not yet been elucidated. We identified SKI might have a significant methylation abnormality in AIP through methylation array analysis using the Illumina Infinium Human Methylation 450K BeadChip array, and investigated the relationship of SKI with AIP clinicopathological features. The methylation rate of SKI was assessed by quantitative SYBR green methylation-specific PCR, and the degree of SKI expression in tissue specimens was assessed by immunohistochemistry in 10 AIP cases, 14 cases of obstructive pancreatitis area in pancreatic ductal adenocarcinoma (PDA) without a history of AIP, and 9 normal pancreas (NP) cases. The SKI methylation ratio was significantly lower in AIP than in PDA and NP. Additionally, the immunohistochemical staining-index (SI) score for SKI was significantly higher in AIP than NP, although there was no significant difference between AIP and PDA. There was a strong negative correlation between SI score and SKI methylation ratio, and between the serum concentrations of IgG4 and the SKI methylation ratio. There was a moderate positive correlation between the serum concentrations of IgG4 and SI. SKI is thought to be an oncogene indicating that SKI hypomethylation and carcinogenesis might be linked to AIP. Furthermore, the correlation between serum concentrations of IgG4 and SKI methylation levels suggest SKI might be involved in the pathogenesis of AIP. However, the role of SKI has not been clearly elucidated. Further studies are needed to understand further the function of SKI.

Cold snare polypectomy (CSP) has not undergone sufficient histopathological evaluation. This study aimed to clarify the histopathological features of CSP specimens, including resection depth and layer, as compared with endoscopic mucosal resection (EMR). Polyps were recruited retrospectively. Sessile, semi-pedunculated, and 0-IIa polyps of ≤ 9 mm were selected by propensity score matching and classified as either a complete resection or one with an unevaluable/positive (X/+) margin. Resection depth and layer were estimated and the risk factors for an X/+ margin were evaluated. A total of 1072 polyps were enrolled. After matching, 184 polyp pairs were selected. An X/+ margin was seen in 105/184 (57%) vs. 70/184 (38%) CSP vs. EMR specimens (p

To better understand the cellular origins and differentiation of anal canal epithelial neoplasms, the immunohistochemical profiles of the anal canal epithelium in humans and swine were evaluated. Formalin-fixed tissue sections were immunostained for mucin (MUC: MUC2, MUC5AC, MUC5B), desmoglein 3 (DGS3), p63, CDX2, SOX2, and α-smooth muscle actin (α-SMA). The anal transitional zone (ATZ) epithelium covered the anal sinus and consisted of a stratified epithelium with mucous cells interspersed within the surface lining. Anal glands opened into the anal sinus. Ducts and acini of intraepithelial or periepithelial mucous type were the main structures of human anal glands, whereas those of swine were compound tubuloacinar mixed glands. Distal to the ATZ epithelium, non-keratinized stratified squamous epithelium merged with the keratinized stratified squamous epithelium of the perianal skin. MUC5AC expression predominated over MUC5B expression in the ATZ epithelium, while MUC5B expression was higher in the anal glands. SOX2 was positive in the ATZ epithelium, anal glands, and squamous epithelium except in the perianal skin. In humans, DGS3 was expressed in the ATZ epithelium, anal gland ducts, and squamous epithelium. p63 was detected in the ATZ epithelium, anal glands, and squamous epithelium. Myoepithelial cells positive for α-SMA and p63 were present in the anal glands of swine. Colorectal columnar cells were MUC5B+ /MUC2+ /CDX2+ /MUC5AC- /SOX2- . The ATZ epithelium seems to be a distinctive epithelium, with morphological and functional features allowing smooth defecation. The MUC5AC+ /SOX2+ /MUC2- /CDX2- profile of the ATZ epithelium and anal glands is a useful feature for diagnosing adenocarcinoma arising from these regions. Anat Rec, 301:796-805, 2018. © 2017 Wiley Periodicals, Inc.

We investigated the expression of LGR5, the most robust and reliable known cancer stem cell (CSC) marker of colorectal cancer, and PD-L1 in tumor budding (TB), as well as clinicopathological features. Tissue microarrays (TMAs) were generated from TB samples from 32 stage II/III colorectal adenocarcinoma patients, and LGR5 expression in TMAs was evaluated by RNAscope, an extremely sensitive RNA in situ hybridization technique. LGR5 expression was significantly lower in the PD-L1-positive group than in the PD-L1-negative group (P = 0.0256). In the PD-L1-positive group, the tumor-infiltrating lymphocytes (TILs) score tended to be higher while the TNM stage was lower compared with the PD-L1 negative group (P = 0.0822 and P = 0.0765, respectively). There was no significant difference in Overall Survival between the PD-L1-positive and PD-L1-negative groups (log-rank test, P = 0.8218). This study showed that PD-L1-positive patients are a unique population with low LGR5 expression, and that LGR5-positive cells may be a promising therapeutic target in PD-L1-negative patients.

Helicobacter suis (H. suis), formerly called Helicobacter heilmannii type 1 (H. heilmannii), is a gram-negative bacterium of the Helicobacter species. This pathogen infects the stomach of humans and animals such as dogs, cats, pigs, and rodents, the latter giving rise to zoonotic infection. Here, we generated a H. suis-specific antibody useful for immunohistochemistry with formalin-fixed, paraffin-embedded tissue sections. To do so, we began by cloning the gene encoding H. suis cholesterol α-glucosyltransferase (αCgT). αCgT is the key enzyme responsible for biosynthesis of cholesteryl α-D-glucopyranoside (CGL), a major cell wall component of Helicobacter species including H. suis. The deduced amino acid sequence of H. suis αCgT had 56% identity with the corresponding Helicobacter pylori (H. pylori). We then developed a polyclonal antibody (anti-Hh-I205R) by immunizing rabbits with a 205 amino acid H. suis αCgT fragment. Immunohistochemistry with the anti-Hh-I205R antibody could differentiate H. suis from H. pylori in gastric mucosa sections derived from mice infected with either pathogen. We then probed formalin-fixed, paraffin-embedded sections of human gastric mucosa positive for H. suis infection with the anti-Hh-I205R antibody and detected positive staining. These results indicate that anti-Hh-I205R antibody is specific for H. suis αCgT and useful to detect H. suis in gastric specimens routinely analyzed in pathological examinations.

LGR5 is the most robust known stem cell marker for gastrointestinal tumors, but there are few reports in breast cancer. Triple negative breast cancer (TNBC) is the most malignant subtype of breast cancer, and thus identification of new cancer stem cell populations in TNBC may help to identify targeted therapies. LGR5 expression was evaluated by RNAscope, a newly developed RNA in situ hybridization technique, using a tissue microarray consisting of 43 patient samples of TNBC selected from the medical archives at our hospital. Patients were stratified into negative and positive LGR5 expression groups. Tumor necrosis was greater in the LGR5-positive group compared with the LGR5-negative group (P = .026). Mitosis tended to show a high value in the LGR5-positive group compared with the LGR5-negative group (P = .0831), while stage tended to show a high stage in the LGR5-positive group compared with the LGR5-negative group (P = .0617). Cox proportional hazards models revealed that the LGR5-positive group (overall survival (OS) = 2.12; 95% CI: 2.12–2.12; P = 0.1575) had no relationship with OS. LGR5 expression is associated with tumor necrosis of TNBC and suggested higher malignant potential.

We describe the case of a 70-year-old man with diabetic nephropathy undergoing hemodialysis. Four years following hemodialysis, he started taking lanthanum carbonate 1500 mg/day and lansoprazole 30 mg/day. Nine years following hemodialysis, he underwent screening esophagogastroduodenoscopy, which demonstrated the presence of the whitish cobblestone-like mucosa in the gastric corpus and multiple reddish depressed lesions with annular whitish mucosa in the antrum. With magnified narrow-band imaging endoscopy, a yellowish–white substance was observed in the villous structure, and subepithelial vessels were observed on the yellowish–white substance. Biopsies were taken from the whitish cobblestone-like mucosa of the upper corpus, a reddish depressed part of the antrum. Histologically, aggregates of cells containing amphophilic fine granular material were found in the mucosal interstitium. These cells stained positive for CD68 and were identified as histiocytes. Since he had been taking lanthanum carbonate for 5 years, we considered the possibility of histiocyte-mediated phagocytosis of lanthanum. Digital mapping via scanning electron microscopy with energy-dispersive X-ray spectrometry showed the presence of lanthanum and phosphorus in the interstitium and cytoplasm of histiocytes. The white, rough mucosa in the gastric body appeared 6 months following the commencement of lanthanum administration and still exists 3 years and 5 months after discontinuation of lanthanum.

Colon cancer stem cells (CSCs) are closely related to tumorigenesis and treatment response, and LGR5 is currently the most robust and reliable CSC marker in colorectal cancer (CRC). However, LGR5 expression in CRC tumor budding (TB) is not well understood. We examined the clinicopathological and prognostic significance of LGR5 in CRC TB. LGR5 expression was evaluated by RNAscope, a newly developed RNA in situ hybridization technique, using a tissue microarray consisting of 55 patient samples of TB in colon adenocarcinoma (CA) selected from the medical archives at our hospital. Patients were stratified into negative and positive LGR5 expression groups. Inflammatory cell infiltration was weaker and histological grade was lower in the LGR5-positive group compared with the LGR5-negative group (P = 0.0407 and P = 0.0436, respectively). There was a significant difference in OS between the LGR5-positive group and LGR5-negative group (log-rank test, P = 0.0088). Cox proportional hazards models revealed that the LGR5-positive group (Overall survival (OS) = 0.37, 95% CI: 0.17-0.79, P = 0.0101) had better OS. LGR5 expression may be affected by inflammatory cell infiltration in the budding area of CA and is an important potential marker of prognosis.

Background Since "Helicobacter pylori (H. pylori) infection" was set as the indication in the Japanese Society for Helicobacter Research (JSHR) Guidelines 2009, eradication treatment for H. pylori gastritis is covered under insurance since 2013 in Japan, and the number of H. pylori eradication has rapidly increased. Under such circumstances, JSHR has made the third revision to the "Guidelines for diagnosis and treatment of H. pylori infection" for the first time in 7 years. Methods The Guideline Committee held 10 meetings. Articles published between the establishment of the 2009 Guidelines and March 2016 were reviewed and classified according to the evidence level; the statements were revised on the basis of this review. After inviting public comments, the revised statements were finalized using the Delphi method. Results There was no change in the basic policy that H. pylori infectious disease is an indication for eradication. Other diseases presumed to be associated with H. pylori infection were added as indications. Serum pepsinogen level, endoscopic examination, and X-ray examination were added to the diagnostic methods. The effects of 1-week triple therapy consisting of potassium-competitive acid blocker (P-CAB), amoxicillin, and clarithromycin have improved, and high eradication rates can also be expected with proton pump inhibitors (PPI) or P-CAB combined with amoxicillin and metronidazole. If the susceptibility test is not performed, the triple PPI or P-CAB/amoxicillin/metronidazole therapy should be chosen, because the PPI/amoxicillin/metronidazole combination demonstrated a significantly higher eradication rate than PPI/amoxicillin/clarithromycin. In the proposal for gastric cancer prevention, we divided gastric cancer prevention measures by age from adolescent to elderly, who are at an increased gastric cancer risk, and presented measures for gastric cancer prevention primarily based on H. pylori eradication. Conclusion We expect the revised guidelines to facilitate appropriate interventions for patients with H. pylori infection and accomplish its eradication and prevention of gastric cancer.

A 71-year-old Japanese man was diagnosed as having stage I primary follicular lymphoma (FL) of the duodenum according to Lugano International Conference Classification and began receiving annual checkups. Endoscopic examination disclosed white villi swelling with depressed red mucosal lesions. Biopsy specimens from the area of white villi exhibited histopathological features that met the diagnostic criteria for low-grade FL. The depressed red lesions gradually enlarged over six years of follow-up. A biopsy of the white villi swelling revealed distinct well-circumscribed follicles with attenuated mantles in the lamina propria that were positive for CD20, bcl-2, and CD10. Histological findings from the depressed red lesions at 5.5 years after the initial diagnosis were compatible for FL. However, biopsy specimens 6 months later obtained from the same lesions showed a mixture of larger mononuclear cells. These follicular cells were positive for CD20 and bcl-2, but not for CD10, indicating the presence of diffuse large B-cell lymphoma (DLBCL). This case shows altered endoscopic findings in the course of DLBCL development from FL. When depressed red lesions are detected in the background of white villi swelling, repeated biopsies should be performed from both lesions.

Leucine-rich repeat-containing G-protein-coupled receptor 5 (Lgr5) is a putative intestinal stem cell marker that is also expressed in various tumors. To analyze its pathological characteristics in mucosal gastric signet-ring cell carcinoma (SRCC), we investigated Lgr5 expression in 35 intramucosal gastric SRCC patients using RNAscope, a newly developed RNA in situ hybridization technique. Lgr5 expression in individual tumor cells was scored semi-quantitatively from 0 to 400. Ki67 was also examined by immunohistochemistry, with a linear arrangement of Ki67-expressing cells present in 20 of 35 cases. This area of Ki67-expressing cells was topographically divided into upper, middle, and lower regions. All cases with linear Ki67 expression patterns also had Lgr5-positive cells arranged in a linear fashion in the lower area-which was distinct from the area of high Ki67 expression. The rate of Ki67 positivity in Lgr5-positive cells was significantly lower than that of Lgr5-negative cells in areas of high Ki67 expression. In intramucosal SRCC, the low mitotic activity of Lgr5-positive cells suggests that they may represent cancer stem cells as seen in other types of stomach carcinomas. Intramucosal SRCC may therefore contain stem cells expressing Lgr5 in the lower area of the lamina propria, akin to normal gastric pyloric mucosa.

LGR5 is expressed in various tumors and has been identified as a putative intestinal stem cell marker. Here we investigated LGR5 expression in colorectal neuroendocrine neoplasms and analyzed the correlation with pathological characteristics. We evaluated the clinicopathological features of 8 neuroendocrine tumor (NET) grade 1 (NET G1), 4 NET Grade 2 (NET G2), and 8 NET Grade 3 (NET G3; also termed neuroendocrine carcinoma, or NEC) cases. We examined LGR5 expression using an RNAscope, a newly developed RNA in situ hybridization technique, with a tissue microarray of the neuroendocrine neoplasm samples. LGR5 staining in individual tumor cells was semi-quantitatively scored using an H-score scale. We also performed a combination of LGR5 RNA in situ hybridization and synaptophysin immunohistochemistry. All cases contained tumor cells with some LGR5-positive dots. For all cases, H-scores showed a positive correlation with nuclear beta-catenin expression. In the NEC group, there was a strong positive correlation between H-score and beta-catenin expression. Our findings suggest that LGR5 may serve as a stem cell marker in NEC, as is the case in colon adenocarcinoma. The positive correlation between H-score and beta-catenin expression suggests that LGR5 expression might be affected by beta-catenin expression in neuroendocrine neoplasms and especially in NEC.

We report a case of vulval extramammary Paget disease (EMPD) with dermal invasion showing mucinous carcinoma (MC). An 80-year-old woman presented with vulvar itching and pain. A physical examination showed a pigmented vulvar, perianal erythematous plague, and a subcutaneous nodule in the left major labia. No internal malignancy, such as colorectal or genitourinary carcinoma, was identified in any of the clinical examinations. A histological examination of the resected specimen revealed Pagetoid tumor cells that had spread widely through the epidermis and invaded the dermis forming a solid nest with mucous lake-like MC. Immunohistochemical examination revealed that the tumor cells in the epidermis and dermis were positive for CK7, CEA, GCDFP-15, MUC5AC, and MUC2, but negative for CK20, MUC6, and CDX2. Only the invasive component showed overexpression of p53. A diagnosis of primary EMPD with dermal invasion showing MC of the vulva was made. This is an extremely rare diagnosis, and we suggest that immunohistochemical evaluations in addition to systemic work-ups are helpful in distinguishing between these cases and those involving vulvar or perianal skin invasion of underlying colorectal or genitourinary carcinomas, which are referred to as secondary EMPD.