Expression of B7-H3 and B7-H4 in Gastric Gastrointestinal Stromal Tumors.

Gastric gastrointestinal stromal tumors (GISTs) are mesenchymal neoplasms with variable behavior characterized by differentiation toward the interstitial cells of Cajal occurring anywhere in the gastrointestinal stromal tract. The management of GIST was revolutionized by the introduction of imatinib, a KIT inhibitor, which has become the standard first-line treatment for metastatic GIST. However, despite a clinical benefit rate of 80%, the majority of patients with GIST experience disease progression after 2 to 3 years of imatinib therapy. This shows the need for novel treatment approaches for imatinib refractory GISTs. The checkpoint proteins B7-H3 and B7-H4 inhibit the activation and function of T cells by potently suppressing the proliferation, cytokine production, and cytotoxicity of activated T cells, which is a mechanism for immune escape. This study aims to clarify B7-H3 and B7-H4 expression in gastric GISTs using immunohistochemistry and reverse transcription polymerase chain reaction (RT-PCR). We confirmed B7-H3 expression (H-score ≥50 points) in 92% and B7-H4 expression in 0% of GIST samples. We examined B7-H3 mRNA expression in 3 representative GIST samples, each having their respective immunostained areas detected by RT-PCR. B7-H3 is expressed at a particularly high rate in GISTs. This suggests that B7-H3 might operate as part of an immune checkpoint in GISTs.

Virtual multi-staining in a single-section view for renal pathology using generative adversarial networks.

Sections stained in periodic acid-Schiff (PAS), periodic acid-methenamine silver (PAM), hematoxylin and eosin (H&E), and Masson's trichrome (MT) stain with minimal morphological discordance are helpful for pathological diagnosis in renal biopsy. Here, we propose an artificial intelligence-based re-stainer called PPHM-GAN (PAS, PAM, H&E, and MT-generative adversarial networks) with multi-stain to multi-stain transformation capability. We trained three GAN models on 512 × 512-pixel patches from 26 training cases. The model with the best transformation quality was selected for each pair of stain transformations by human evaluation. Frechet inception distances, peak signal-to-noise ratio, structural similarity index measure, contrast structural similarity, and newly introduced domain shift inception score were calculated as auxiliary quality metrics. We validated the diagnostic utility using 5120 × 5120 patches of ten validation cases for major glomerular and interstitial abnormalities. Transformed stains were sometimes superior to original stains for the recognition of crescent formation, mesangial hypercellularity, glomerular sclerosis, interstitial lesions, or arteriosclerosis. 23 of 24 glomeruli (95.83 %) from 9 additional validation cases transformed to PAM, PAS, or MT facilitated recognition of crescent formation. Stain transformations to PAM (p = 4.0E-11) and transformations from H&E (p = 4.8E-9) most improved crescent formation recognition. PPHM-GAN maximizes information from a given section by providing several stains in a virtual single-section view, and may change the staining and diagnostic strategy.

GATA4 is diagnostically useful for distinguishing primary ovarian mucinous carcinomas from metastatic colorectal adenocarcinomas to the ovary.

AIM: Determining the primary origin of an ovarian mucin-producing carcinoma can be challenging at times because some metastases of primary colorectal origin may exhibit gross, microscopic, and/or immunohistochemical features that overlap with those of primary ovarian mucinous carcinomas (OMCs). We hypothesized that GATA binding protein 4 (GATA4) might be a novel, useful marker for differentiating primary OMCs from metastatic colorectal adenocarcinomas to the ovary. METHODOLOGY: For comparison with the usefulness of other markers (special AT-rich sequence-binding protein 2 (SATB2) and caudal type homeobox 2 (CDX2)), we elucidated the expression profiles of GATA4 in OMCs, colorectal non-mucinous adenocarcinomas (CNMACs), and colorectal mucinous adenocarcinomas (CMACs) using immunohistochemistry. RESULTS: We confirmed GATA4 expression (H-score ≥50 points) in 93%, SATB2 in 0%, and CDX2 in 64% of 14 OMCs. GATA4 was expressed in 13%, SATB2 in 90%, and CDX2 in 93% of 30 CNMACs. GATA4 was expressed in 20%, SATB2 in 73%, and CDX2 in 100% of 30 CMACs. CONCLUSION: The expression of GATA4 in a mucus-producing ovarian tumor strongly supports it being a primary OMC rather than a metastatic colorectal carcinoma: GATA4 expression indicates OMC and SATB2 expression indicates colorectal adenocarcinoma. However, three cases of colorectal adenocarcinoma were GATA4-positive and SATB2-negative, so the GATA4/SATB2 marker combination is not absolute for determining the primary site. Further research for more markers is necessary to find the ideal combination.

Expression of TGF-ß1 in Gastrointestinal Stromal Tumor (GIST) and the Occurrence of Frequent Desmoplasia.

Gastrointestinal stromal tumors (GISTs) are mesenchymal neoplasms with variable behavior characterized by differentiation toward the interstitial cells of Cajal occurring anywhere in the gastrointestinal stromal tract. Frequently, GISTs have fibrous stroma within tumor cell proliferation areas, which is unlike other types of malignant tumors. If this desmoplasia is active, there is a possibility that some sort of transmitter exists between GIST cells and cells related to fibrosis in the tumor cell proliferation areas. Transforming growth factor (TGF)-ß isoforms, particularly TGF-ß1, are critical for fibrosis pathogenesis. TGF-ß1 regulation of myofibroblasts and fibroblasts during fibrosis is well described. The induced fibroblast activation resulting in myofibroblast differentiation has been reported as an important source of collagen, glycoproteins, proteoglycans, and matrix metallopeptidases in wound healing and fibrosis. However, there are a few reports on the relationship between TGF-ß1 and GISTs. This study aims to clarify TGF-ß1 expression in 30 gastric GISTs using immunohistochemistry and reverse transcription polymerase chain reaction (RT-PCR). For comparison, we also enrolled 30 samples of gastric tubular adenocarcinoma (GTAC). We confirmed TGF-ß1 expression (H-score ≥50 points) in 57% of GIST and 13% of GTAC samples, a significant difference between the 2 tumor types ( P =0.001). We examined the TGF-ß1 mRNA expression of 3 representative GIST samples, each having their respective immunostained areas detected by RT-PCR. Finding TGF-ß1 expression may indicate that this cytokine plays a part in the formation of desmoplasia within GIST cell proliferative areas.

The Expression and Role of Aquaporin 4 in Colon Cancer.

BACKGROUND/AIM: Aquaporins (AQPs) were initially discovered as water channel proteins that facilitate transcellular water movements. Recent studies have shown that AQPs are expressed and play an oncogenic role in various cancers. However, the expression and role of Aquaporin 4 (AQP4) in colon cancer have not been investigated. This study aimed to examine the clinical and pathophysiologic significance of AQP4 in colon cancer. PATIENTS AND METHODS: Immunohistochemistry (IHC) of AQP4 for 145 primary tumor samples obtained from patients with stage II or III colon cancer was performed, and the relationship between AQP4 expression and patients' prognoses was analyzed. Knockdown experiments with AQP4 small interfering RNA using human colon cancer cells were conducted to analyze the effects on cell invasiveness. RESULTS: IHC revealed that AQP4 was scarcely expressed in the noncancerous colonic mucosa. Of the 145 patients who enrolled in this study, 109 (75.2%) and 36 (24.8%) patients were classified as negative and positive for AQP4 expression, respectively. A high level of AQP4 expression is significantly associated with deeper tumors with lymph node metastasis and venous invasion. A 5-year progression-free survival rate of AQP4-positive patients was significantly worse than that of AQP-4 negative patients (70.7% vs. 87.0%, p=0.049). Furthermore, AQP4 knockdown significantly inhibited cell migration and invasion in HCT116 cells. CONCLUSION: AQP4 may be a novel biomarker and therapeutic target for colon cancer.

Immunohistochemical detection of cancer genetic abnormalities.

New applications of immunohistochemistry (IHC) expand rapidly due to the development of molecular analyses and an increased understanding of molecular biology. IHC becomes much more important as a screening or even a confirmatory test for molecular changes in cancer. The past decades have witnessed the release of many immunohistochemical markers of the new generation. The novel markers have extensively high specificity and sensitivity for the detection of genetic abnormalities. In addition to diagnostic utility, IHC has been validated to be a practical tool in terms of treatments, especially molecular targeted therapy. In this review, we first describe the common alterations of protein IHC staining in human cancer: overexpression, underexpression, or loss of expression and altered staining pattern. Next, we examine the relationship between staining patterns and genetic aberrations regarding both conventional and novel IHC markers. We also mention current mutant-specific and fusion-specific antibodies and their concordance with molecular techniques. We then describe the basic molecular mechanisms from genetic events to corresponding protein expression patterns (membranous, cytoplasmic, or nuclear patterns). Finally, we shortly discuss the applications of immunohistochemistry in molecular targeted therapy. IHC markers can serve as a complementary or companion diagnostic test to provide valuable information for targeted therapy. Moreover, immunohistochemistry is also crucial as a companion diagnostic test in immunotherapy. The increased number of IHC novel antibodies is broadening its application in anti-cancer therapies.

Molecular and clinicopathological features of granzyme B-negative extranodal NK/T-cell lymphoma.

Extranodal NK/T-cell lymphoma (ENKTL) generally expresses cytotoxic molecules, including granzyme B (GZMB), T-cell-restricted intracellular antigen-1 (TIA-1), and perforin; however, the expression of these molecules varies across cases. We performed gene expression profiling and identified unique biological and clinicopathological features of GZMB-negative ENKTL. We reviewed the clinicopathological characteristics of 71 ENKTL samples. Gene expression profiling on nine ENKTLs using multiplexed, direct, and digital mRNA quantification divided ENKTLs into Groups A (n = 7) and B (n = 2) through hierarchical clustering and t-distributed stochastic neighbor embedding. Group B was characterized by downregulation of genes associated with IL6-JAK-STAT3 signaling and inflammatory responses. GZMB mRNA expression was significantly downregulated in Group B. GZMB protein expression was evaluated with immunohistochemistry in all 71 ENKTLs, and expression data of Tyr705-phosphorylated STAT3 (pSTAT3) and MYC from our previous study was utilized. T-cell receptor gamma (TRG) gene rearrangement in the selected samples was also assessed using PCR. GZMB expression was higher in pSTAT3-positive (p = 0.028) and MYC-positive (p = 0.014) ENKTLs. Eighteen percent (13/71) of all ENKTLs were negative for GZMB (defined by positivity <10 %); patients with GZMB-negative ENKTLs were often in a higher clinical stage (p = 0.016). We observed no other correlations with clinical parameters or TRG rearrangement and no significant association between GZMB expression and survival. In conclusion, GZMB expression is highly heterogeneous in ENKTLs and is associated with the activation of the JAK-STAT3 pathway and higher MYC expression. GZMB-negative ENKTLs correlate with an advanced clinical stage, suggesting the potential utility of GZMB immunohistochemistry as a biomarker of ENKTL.

Endometrioid Carcinoma of the Uterine Corpus With a Micropapillary Component: A Novel Prognostic Factor For Metastasis.

BACKGROUND/AIM: Micropapillary pattern is a morphologically distinctive form of carcinoma composed of small, hollow, or morula-like clusters of cancer cells surrounded by clear stromal spaces. The neoplastic cells characteristically display a reverse polarity, also known as an ''inside-out'' growth pattern, that is linked to higher frequencies of lymphovascular invasion and lymph nodal metastasis. To the best of our knowledge, it has not been previously recognized in uterine corpus. CASE REPORT: We report 2 cases of endometrioid carcinoma of the uterine corpus with a micropapillary component. In these cases, histological examination identified an endometrioid carcinoma that had invaded the myometrial layer. The carcinoma cells that constructed the micropapillary components were immunohistochemically positive for EMA. They lined the stromal facing surface of the cell membrane, confirming the inside-out growth pattern, and D2-40 immunohistochemistry confirmed lymphovascular invasion of carcinoma cells. CONCLUSION: We believe that the micropapillary pattern linked to higher frequencies of lymphovascular invasion and lymph nodal metastasis may be one of the most important invasive patterns in endometrioid carcinomas of the uterine corpus for predicting aggressive malignant potential, prognosis, and recurrence, although further, larger studies are required to evaluate its clinical significance.

The Expression and Role of NADPH Oxidase 2 in Colon Cancer.

BACKGROUND/AIM: Recent studies have reported that nicotinamide adenine dinucleotide phosphate oxidases (NOXs) are expressed in various cancers and play important roles in tumor progression. However, no studies have examined the expression and role of NOX2 in colon cancer. The aim of this study is to investigate the pathophysiological roles of NOX2 in colon cancer patients and cell lines. PATIENTS AND METHODS: One-hundred and sixteen primary colon cancer samples of patients who underwent radical resection for locally advanced colon cancer were used for immunohistochemistry of NOX2 protein. The relationship between NOX2 expression and clinicopathological factors was assessed and the prognostic significance of NOX2 expression was evaluated in colon cancer patients. NOX2 siRNA transfection experiments were performed using two colon cancer cell lines (HCT116 and RKO) to analyze the impact of NOX2 expression on cellular physiological functions. RESULTS: The expression of NOX2 protein in noncancerous tissue was scarcely observed, and 45 samples (38.8%) showed positively stained NOX2 expression in cancer tissue. There were no clinicopathological factors significantly associated with NOX2 expression. The 5-year recurrence-free survival rate of the NOX2 positive group was significantly lower than that of the NOX2 negative group (61.1% vs. 79.3%, p=0.029). NOX2 depletion significantly inhibited cell proliferation with G1 arrest, and motility in the two cell lines. CONCLUSION: NOX2 expression level has a close association with the prognosis of colon cancer patients and physiological functions of colon cancer cells. NOX2 may be a useful prognostic biomarker for colon cancer patients.

Three-dimensional structural analysis of papillary thyroid carcinoma nuclei with serial block-face scanning electron microscopy (SBF-SEM).

Nuclear morphology of carcinoma cells is critical for the pathological diagnosis of papillary thyroid carcinoma (PTC). However, three-dimensional architecture of PTC nuclei is still elusive. In this study, we analyzed the three-dimensional ultrastructure of PTC nuclei using serial block-face scanning electron microscopy which takes advantage of the high-throughput acquisition of serial electron microscopic images and three-dimensional reconstruction of subcellular structures. En bloc-stained and resin-embedded specimens were prepared from surgically removed PTCs and normal thyroid tissues. We acquired two-dimensional images from serial block-face scanning electron microscopy and reconstructed three-dimensional nuclear structures. Quantitative comparisons showed that the nuclei of carcinoma cells were larger and more complex than those of normal follicular cells. The three-dimensional reconstruction of carcinoma nuclei divided intranuclear cytoplasmic inclusions into "open intranuclear cytoplasmic inclusions" connecting to cytoplasm outside the nucleus and "closed intranuclear cytoplasmic inclusions" without that connection. Cytoplasm with abundant organelles was observed in open inclusions, but closed inclusions contained fewer organelles with or without degeneration. Granules with a dense core were only observed in closed inclusions. Our observations suggested that open inclusions originate from nuclear invaginations, and disconnection from cytoplasm leads to closed inclusions.

Carcinoembryonic antigen levels in pancreatic juice are associated with histological subtypes of intraductal papillary mucinous neoplasm of the pancreas.

Background: The present study aimed to examine the correlation between preoperative carcinoembryonic antigen levels in pancreatic juice (PJ-CEA) and the histological subtype of intraductal papillary mucinous neoplasm (IPMN). Methods: We enrolled IPMN patients who underwent endoscopic retrograde pancreatography between March 2002 and March 2018. Clinical factors associated with IPMN histological subtypes of 67 patients who underwent surgery were analyzed. Furthermore, the relationship between CEA immunohistochemistry findings and histological subtypes was investigated. Results: Median PJ-CEA were 15 ng/ml in the gastric type, 150 ng/ml in the intestinal type, and 175 ng/ml in the pancreatobiliary type. Both intestinal and pancreatobiliary types had significantly higher PJ-CEA than the gastric type (p = 0.001). In the analysis of histological subtype predictors, high PJ-CEA (≥63 ng/ml) only showed a significant difference in multivariate analyses (95% confidence interval 4.8-70.2; p < 0.001). Immunohistochemistry findings revealed significantly higher CEA expression in the non-gastric type than in the gastric type (p < 0.001). The non-gastric type showed a significantly worse prognosis than the gastric type (p = 0.017). Conclusion: PJ-CEA was an independent predictor of IPMN histological subtypes in a preoperative setting. High PJ-CEA predict the non-gastric type, while low PJ-CEA predict the gastric type.

Genetic and immunohistochemical profiling of NK/T-cell lymphomas reveals prognostically relevant BCOR-MYC association.

Extranodal NK/T-cell lymphoma, nasal type (ENKTL) is an Epstein-Barr virus-positive, aggressive lymphoma with a heterogeneous cell of origin and variable clinical course. Several clinical prognostic indices have been proposed for ENKTL; however, there are few pathological biomarkers. This multi-institutional study sought to identify histologically assessable prognostic factors. We investigated mutation profiles by targeted next-generation sequencing (NGS) and immunohistochemical assessments of expression of MYC, Tyr705-phosphorylated (p-)STAT3, and CD30 in 71 ENKTL samples. The median age of the patients was 66 years (range, 6-100). The most frequent mutations were in STAT3 (27%), JAK3 (4%), KMT2D (19%), TP53 (13%), BCOR (10%), and DDX3X (7%). Immunohistochemistry (IHC) revealed that ENKTLs with STAT3 mutations exhibited higher expression of pSTAT3 and CD30. BCOR mutations were associated with increased MYC expression. Univariate analysis in the entire cohort showed that stage (II, III, or IV), BCOR mutations, TP53 mutations, and high MYC expression (defined as ≥40% positive neoplastic cells) were associated with reduced overall survival (OS). Multivariate modeling identified stage (II, III, or IV) and high MYC expression as independent adverse prognostic factors. In a subgroup analysis of patients treated with anthracycline (AC)-free chemotherapy and/or radiotherapy (RT) with curative intent, BCOR but not high MYC expression was an independent adverse prognostic factor. In conclusion, activating STAT3 mutations are common in ENKTLs and are associated with increased CD30 expression. MYC overexpression is, at least in part, associated with deleterious BCOR mutations, and this BCOR-MYC linkage may have prognostic significance, underscoring the potential utility of IHC for MYC in risk stratification of patients with ENKTL.

Clinical detection of "extremely low-risk" follicular thyroid carcinoma: A population-based study of 7304 patients.

Background: Previous studies have not been consistent in the risk of metastasis in follicular thyroid carcinoma (FTC). Therefore, we conducted a large population study to stratify the risk of distant metastasis in FTC patients using only clinical parameters. Methods: We extracted FTC patients from The Surveillance, Epidemiology, and End Results (SEER) database and divided them into training and validation cohorts. Results: The two cohorts consisted of 4913 and 2391 patients, respectively. We developed a nomogram and risk table based on a logistic regression model using algorithm-selected variables. Receiver Operating Characteristic (ROC) analyses showed high discriminatory power in the training and validation cohorts (Area under the curve [AUC] of 0.85 and 0.84, respectively). Extremely low, low, intermediate, and high-risk groups had 0.3%, 1%, 3.5%, and 16.7% risk of distant metastasis, respectively. Conclusions: Our risk scoring table can separates patients into four risk groups and efficiently detect patients with almost no risk of metastasis.

Incidence, clinicopathological features and genetics of in-situ follicular neoplasia: a comprehensive screening study in a Japanese cohort.

AIMS: In-situ follicular neoplasia (ISFN) is a histologically recognizable neoplastic proliferation of follicular lymphoma (FL)-like B cells confined to the germinal centres. While some ISFNs are associated with overt FL, others are incidentally identified as isolated or pure forms in individuals without evidence of overt FL. The prevalence of incidentally found isolated ISFN is approximately 3% in Europe; however, no screening study has been conducted in Asia. To investigate the incidence and clinicopathological characteristics of ISFNs in the Japanese population, we conducted histopathological screening of the lymph nodes (LNs) resected for solid tumours or inflammatory conditions. METHODS AND RESULTS: We screened for ISFN in 5700 LNs from 340 individuals using immunohistochemistry for BCL2 and identified seven ISFNs, with an incidence of 2.1%. The median age of the individuals with ISFN was 67 years, none of whom developed overt FL, with a median follow-up of 59 months. Next-generation sequencing was performed in five ISFNs, and 10 variants in seven FL-associated genes were identified. The identified variants included HIST1H1E (n = 2), ARID1A (n = 2), KMT2D (n = 1), CARD11 (n = 1), BCL7A (n = 1), CREBBP (n = 1) and TNFRSF14 (n = 1). CONCLUSIONS: The incidence of isolated ISFN in the Japanese population is not significantly different from that in Europe, presumably reflecting the recent increase in FL in Japan. These incidentally found ISFNs have a low potential to transform into overt FL. Although mutations of FL-associated genes are already present in ISFNs, further molecular studies are needed to identify driver genes leading to the transformation of ISFN to overt FL.

A rare case of intracytoplasmic mucin-rich nasopharyngeal carcinoma.

Nasopharyngeal carcinoma (NPC) is an Epstein-Barr virus (EBV)-associated tumor with a high incidence in Asian countries. NPC is a type of squamous cell carcinoma originating from the nasopharyngeal mucosa. Although rare, NPCs show some uncommon histologic variants; these variations remain to be understood. We described the cytologic characteristics of a rare NPC variant with abundant intracytoplasmic mucin. A 37-year-old Japanese man presented to our hospital with bilateral ear discomfort and palpable lymph nodes. Nasopharyngeal biopsy showed tumor cells with abundant intracytoplasmic, Alcian blue-PAS-positive mucin. Immunohistochemical analysis demonstrated that the tumor cells were positive for p40 and p53. Epstein-Barr encoding region (EBER) in situ hybridization (ISH) showed EBV infection of the tumor cells. Fluorescence in situ hybridization (FISH) using MAML2 break-apart probes did not show split signals. Fine needle aspiration biopsy (FNAB) cytology of the metastatic lymph nodes was also performed. Smear samples had a necrotic and inflammatory background with both lymphocyte and neutrophil infiltration. Highly cellular tumor clusters and dispersed cells with naked nuclei were observed. The tumor cells showed a clear cytoplasm with distinct cell borders. Intracytoplasmic mucin pushing the nucleus to the periphery was observed in the scattered tumor cells in a liquid-based cytology sample. Given these findings, the final diagnosis was advanced nasopharyngeal carcinoma; cisplatin-based chemoradiation therapy was performed as the first-line treatment. The tumor recurred 8 months after completing the treatment. The recurrent nasopharyngeal tumor was a typical non-keratinizing NPC and lacked intracytoplasmic mucin.

Prognostic impact of lymph node micrometastasis in patients with gastric cancer.

PURPOSE: The clinical significance of lymph node micrometastasis (LNMM) remains controversial in gastric cancer (GC). In this study, we investigated the prognostic impact of LNMM in patients with GC. METHODS: A total of 624 patients with pathologically lymph node metastasis-negative (pN0) and N1 status (pN1) who underwent gastrectomy between 2004 and 2018 were enrolled in this retrospective study. The diameter of tumor cell clusters in metastatic lymph nodes was measured in 120 patients with pN1 GC. RESULTS: Patients with lymph node tumors < 1500 µm in diameter (LNMM) had a significantly better prognosis than those with tumors ≥ 1500 µm in diameter (p = 0.012; log-rank test). Cox's proportional hazards model revealed that LNMM (p = 0.016), several dissected lymph nodes (p = 0.049), and the provision of adjuvant chemotherapy (p = 0.002) were independent prognostic factors for the overall survival of patients with pN1 GC. There was no significant difference in the overall survival between patients with LNMM who received chemotherapy and those who did not (p = 0.332). CONCLUSIONS: LNMM is associated with a favorable prognosis and maybe an independent prognostic marker in patients with pN1 GC. LNMM in GC may be considered a factor preventing adjuvant chemotherapy.

Expression of CXCL12 in esophageal high grade dysplasia characterized pathologically by lymphocyte accumulation directly under the lesion.

Squamous dysplasia of the esophagus is an unequivocal neoplastic alteration of the esophageal squamous epithelium without invasion. Esophageal high grade dysplasia (EHGD) is characterized by >50% epithelial involvement or severe cytological atypia. Frequently, lymphocytes accumulate below EHGD lesions even though there is no invasion. If this lymphocytic accumulation is active, a transmitter should exist between the EHGD cells and the lymphocytes. C-X-C motif chemokine ligand (CXCL) 12, CXCL10 and C-C motif chemokine ligand 18 (CCL18) are all lymphocyte chemoattractants in vivo, but there are no reports on the relationship between these chemokines and EHGDs. In this study, we investigated these chemokines and C-X-C motif chemokine receptor 4 (CXCR4) (receptor for CXCL12) in 30 EHGDs using immunohistochemistry and reverse transcription polymerase chain reaction (RT-PCR). For comparison, we enrolled 30 samples of normal esophageal squamous epithelium (NESE). We confirmed CXCL12 expression (H-score≥50 points) in 70% of EHGD and 0% of NESE samples, CXCL10 expression in 3% of EHGD and 3% of NESE samples, CCL18 expression in 3% of EHGD and 0% of NESE samples, and CXCR4 expression in 53% of EHGD and 0% of NESE samples by immunohistochemistry. EHGD and NESE cases were significantly different in their expressions between the tissue types (CXCL12, p<0.001; CXCR4, p<0.001). We examined CXCL12 and CXCR4 mRNA expressions of 3 representative EHGD samples, each having their respective immunostained areas detected by RT-PCR. Finding CXCL12 expression may indicate that this chemokine plays a part in the lymphocyte accumulation that occurs directly under EHGDs.

Expressions of CXCL12, CXCL10 and CCL18 in Warthin tumors characterized pathologically by having a lymphoid stroma with germinal centers.

The Warthin tumor is a benign neoplasm of the salivary glands, histologically, the tumor has an oncocytic epithelial component forming uniform rows of cells surrounded by cystic spaces associated with a lymphoid stroma often showing the presence of germinal centers. The lymphoid stroma is a representative microscopic finding. If this lymphocytic accumulation is active, some sort of transmitter should exist between the Warthin tumor cells and lymphocytes. C-X-C motif chemokine ligand (CXCL12) 12, CXCL10 and C-C motif chemokine ligand 18 (CCL18) are a chemoattractant for lymphocytes in vivo. There is no report on the relationship between these chemokines and Warthin tumors. In this study, we investigated these chemokines expressions in 20 Warthin tumors using immunohistochemistry and reverse transcription polymerase chain reaction (RT-PCR). For comparison, we also enrolled samples of pleomorphic adenoma, which is another benign salivary gland tumor type without prominent lymphocytic infiltration. All Warthin tumors were immunopositive for CXCL12 and CXCL10, and these reactivities were diffuse. Meanwhile, the majority of pleomorphic adenomas were immunonegative for CXCL12 (95%), CXCL10 (80%) and CCL18 (85%). Warthin tumor and pleomorphic adenoma cases were significantly different in these immunostaining expressions (CXCL12, p<0.001; CXCL10, p<0.001; CCL18, p=0.024). We examined CXCL12, CXCL10 and CCL18 mRNA expressions of 3 representative Warthin tumor samples, each having these chemokines immunopositive areas detected by RT-PCR. Finding CXCL12 and CXCL10 expressions indicate that these chemokines may play a part in the formation of a lymphoid stroma within Warthin tumors. In regards to this phenomenon, the participation of CCL18 might be restrictive compared to CXCL12 and CXCL10.

Role of magnifying endoscopy with narrow-band imaging in the diagnosis of noninvasive gastric neoplasia.

BACKGROUND AND AIM: There are no globally approved, distinguishing criteria enabling the classification of gastric adenomas and intramucosal carcinomas for differential diagnosis of noninvasive neoplasia (NIN). METHODS: Next-generation sequencing of 50 cancer-related genes was undertaken on 68 pathologically diagnosed microdissected gastric neoplasms (25 adenomas, 27 intramucosal carcinomas, and 16 submucosal carcinomas) obtained during endoscopic submucosal dissection. Findings from magnifying endoscopy with narrow-band imaging (M-NBI) of 52 NINs (the 25 adenomas and 27 intramucosal carcinomas) were compared with these data. RESULTS: Among all 68 neoplasms, the most frequently mutated genes were APC (76% in adenoma, 11.1% in intramucosal carcinoma, and 0% in submucosal carcinoma; P < 0.001) and TP53 in intramucosal and submucosal carcinomas (8% in adenoma, 48.1% in intramucosal carcinoma, and 75% in submucosal carcinoma; P < 0.001). Dividing the NIN neoplasms into five groups according to their mutational status (A1: APC mutation, A2: APC + α mutation, B: APC + TP53 mutation, C: TP53 mutation, D: no mutation in either APC or TP53) resulted in almost identical diagnoses by pathology and M-NBI for groups A1 (12/13, 92%), C (12/13, 92%), and D (16/17, 94%) but not for groups A2 (3/7, 43%) or B (0/2, 0%). This finding implies that NINs with the APC + α mutation have carcinoma-like endoscopic features despite most being judged as adenomas by pathology. CONCLUSION: A diagnosis of NINs by pathology or M-NBI in the subset of gastric tumors classified by cancer-related mutations is not completely identical, suggesting the possible additional role of M-NBI in diagnosing NINs. Further studies are needed to confirm this.

Progression of Papillary Thyroid Carcinoma to Anaplastic Carcinoma in Metastatic Lymph Nodes: Solid/Insular Growth and Hobnail Cell Change in Lymph Nodes Are Predictors of Subsequent Anaplastic Transformation.

Most anaplastic thyroid carcinomas (ATCs) arise from papillary thyroid carcinoma (PTC). This process is also called anaplastic transformation, and the morphological harbingers of this phenomenon in nodal recurrence have not been assessed systematically. For this reason, the current study focused on features of 10 PTCs with regional lymph node recurrence that was accompanied with disease progression due to anaplastic transformation in at least one of the nodal recurrences. The findings of additional 19 PTCs which recurred without anaplastic transformation after ≥ 10 years of follow-up served as the control group. There were no clinicopathological differences between the two groups at initial surgery including age, gender, tumor size, lymph node metastasis, distant metastasis, extrathyroidal extension, histologic subtype, and treatment. The median time from the initial thyroid surgery to anaplastic transformation in the nodal recurrence was 106 months (range 6 to 437 months). Mutational analyses showed recurrent PTCs with anaplastic transformation had a high prevalence of BRAFV600E mutation (8/9) and TERT promoter mutation (9/9), both of which were detected in primary tumors. PIK3CAH1047R mutation was detected in one case. No case had RAS mutation. Nineteen recurrent PTCs without anaplastic transformation harbored BRAFV600E mutation and seventeen of these had TERT promoter mutation. Unlike primary tumors with subsequent nodal anaplastic transformation, TERT promoter mutation was only present in the metastatic nodal recurrence from 4 patients without transformation. No patients had neither high-grade features (necrosis and increased mitotic activity) nor solid/insular growth or hobnail cell features in their primary tumors. In the group of patients with transformation, 3 had solid/insular growth in the lymph node metastasis at the time of primary tumor resection (one displaying nuclear features of PTC and solid growth with increased mitotic activity, one with insular component consistent with poorly differentiated carcinoma component, and one displaying nuclear features of PTC and solid growth), and additional 2 patients had solid/insular growth with no high-grade features or poorly differentiated carcinoma component at the time of subsequent nodal recurrence prior to anaplastic transformation. Hobnail cell features were exclusively seen in subsequent metastatic lymph nodes prior to anaplastic transformation. The control group lacked solid/insular growth and hobnail cell features in the metastatic nodal disease. Aberrant p53 expression and loss of TTF-1 featured tumor components with anaplastic transformation. This series identified a subset of recurrent PTCs with TERT promoter mutation was prone to undergo anaplastic transformation, and that solid/insular growth and hobnail cell features were morphological predictors of anaplastic transformation in the nodal recurrence.