Dysregulation of WNT5A/ROR2 Signaling Characterizes the Progression of Barrett-Associated Esophageal Adenocarcinoma.

UNLABELLED: The mechanism underlying the progression of normal esophageal mucosa to esophageal adenocarcinoma remains elusive. WNT5A is a noncanonical WNT, which mainly functions via the receptor tyrosine kinase-like orphan receptor 2 (ROR2), and has an unclear role in carcinogenesis. In this study, we aimed to determine the role of WNT5A/ROR2 signaling in esophageal adenocarcinoma. Analysis of WNT5A and ROR2 expression patterns in healthy controls, Barrett and esophageal adenocarcinoma patients' esophageal clinical specimens as well as in various esophageal cell lines demonstrated a ROR2 overexpression in esophageal adenocarcinoma tissues compared with Barrett and healthy mucosa, whereas WNT5A expression was found significantly downregulated toward esophageal adenocarcinoma formation. Treatment of esophageal adenocarcinoma OE33 cells with human recombinant WNT5A (rhWNT5A) significantly suppressed proliferation, survival, and migration in a dose-dependent fashion. rhWNT5A was found to inhibit TOPflash activity in ROR2 wild-type cells, whereas increased TOPflash activity in ROR2-knockdown OE33 cells. In addition, ROR2 knockdown alone abolished cell proliferation and weakened the migration properties of OE33 cells. These findings support an early dysregulation of the noncanonical WNT5A/ROR2 pathway in the pathogenesis of esophageal adenocarcinoma, with the loss of WNT5A expression together with the ROR2 overexpression to be consistent with tumor promotion. IMPLICATIONS: The dysregulation of WNT5A/ROR2 noncanonical WNT signaling in Barrett-associated esophageal adenocarcinoma introduces possible prognostic markers and novel targets for tailored therapy of this malignancy. Mol Cancer Res; 14(7); 647-59. ©2016 AACR.

Wnt/ß-Catenin Signaling Activation beyond Robust Nuclear ß-Catenin Accumulation in Nondysplastic Barrett's Esophagus: Regulation via Dickkopf-1.

INTRODUCTION: Wnt/ß-catenin signaling activation has been reported only during the late steps of Barrett's esophagus (BE) neoplastic progression, but not in BE metaplasia, based on the absence of nuclear ß-catenin. However, ß-catenin transcriptional activity has been recorded in absence of robust nuclear accumulation. Thus, we aimed to investigate the Wnt/ß-catenin signaling in nondysplastic BE. METHODS: Esophageal tissues from healthy and BE patients without dysplasia were analyzed for Wnt target gene expression by quantitative reverse transcription polymerase chain reaction (qRT-PCR) and immunohistochemistry. Esophageal squamous (EPC1-& EPC2-hTERT), BE metaplastic (CP-A), and adenocarcinoma (OE33) cell lines were characterized for Wnt activation by qRT-PCR, Western blot, and luciferase assay. Wnt activity regulation was examined by using recombinant Wnt3a and Dickkopf-1 (Dkk1) as well as Dkk1 short interfering RNA. RESULTS: Wnt target genes (AXIN2, c-MYC, Cyclin D1, Dkk1) and Wnt3a were significantly upregulated in nondysplastic BE compared with squamous mucosa. Elevated levels of dephosphorylated ß-catenin were detected in nondysplastic BE. Nuclear active ß-catenin and TOPflash activity were increased in CP-A and OE33 cells compared with squamous cells. Wnt3a-mediated ß-catenin signaling activation was abolished by Dkk1 in CP-A cells. TOPFlash activity was elevated following Dkk1 silencing in CP-A but not in OE33 cells. Dysplastic and esophageal adenocarcinoma tissues demonstrated further Dkk1 and AXIN2 overexpression. CONCLUSIONS: Despite the absence of robust nuclear accumulation, ß-catenin is transcriptionally active in nondysplastic BE. Dkk1 overexpression regulates ß-catenin signaling in BE metaplastic but not in adenocarcinoma cells, suggesting that early perturbation of Dkk1-mediated signaling suppression may contribute to BE malignant transformation.

Endothelial-mesenchymal transition in normal human esophageal endothelial cells cocultured with esophageal adenocarcinoma cells: role of IL-1ß and TGF-ß2.

Endothelial-mesenchymal transition (EndoMT) has been recognized as a key determinant of tumor microenvironment in cancer progression and metastasis. Endothelial cells undergoing EndoMT lose their endothelial markers, acquire the mesenchymal phenotype, and become more invasive with increased migratory abilities. Early stages of esophageal adenocarcinoma (EAC) are characterized by strong microvasculature whose impact in tumor progression remains undefined. Our aim was to determine the role of EndoMT in EAC by investigating the impact of tumor cells on normal primary human esophageal microvascular endothelial cells (HEMEC). HEMEC were either cocultured with OE33 adenocarcinoma cells or treated with IL-1ß and transforming growth factor-ß2 (TGF-ß2) for indicated periods and analyzed for EndoMT-associated changes by real-time PCR, Western blotting, immunofluorescence staining, and functional assays. Additionally, human EAC tissues were investigated for detection of EndoMT-like cells. Our results demonstrate an increased expression of mesenchymal markers [fibroblast-specific protein 1 (FSP1), collagen1α2, vimentin, α-smooth muscle actin (α-SMA), and Snail], decreased expression of endothelial markers [CD31, von Willebrand factor VIII (vWF), and VE-cadherin], and elevated migration ability in HEMEC following coculture with OE33 cells. The EndoMT-related changes were inhibited by IL-1ß and TGF-ß2 gene silencing in OE33 cells. Recombinant IL-1ß and TGF-ß2 induced EndoMT in HEMEC. Although the level of VEGF expression was elevated in EndoMT cells, the angiogenic property of these cells was diminished. In vivo, by immunostaining EndoMT-like cells were detected at the invasive front of EAC. Our findings underscore a significant role for EndoMT in EAC and provide new insights into the mechanisms and significance of EndoMT in the context of tumor progression.

Reclassification of serous ovarian carcinoma by a 2-tier system: a Gynecologic Oncology Group Study.

BACKGROUND: A study was undertaken to use the 2-tier system to reclassify the grade of serous ovarian tumors previously classified using the International Federation of Gynecology and Obstetrics (FIGO) 3-tier system and determine the progression-free survival (PFS) and overall survival (OS) of patients treated on Gynecologic Oncology Group (GOG) Protocol 158. METHODS: The authors retrospectively reviewed demographic, pathologic, and survival data of 290 patients with stage III serous ovarian carcinoma treated with surgery and chemotherapy on GOG Protocol 158, a cooperative multicenter group trial. A blinded pathology review was performed by a panel of 6 gynecologic pathologists to verify histology and regrade tumors using the 2-tier system. The association of tumor grade with PFS and OS was assessed. RESULTS: Of 241 cases, both systems demonstrated substantial agreement when combining FIGO grades 2 and 3 (overall agreement, 95%; kappa statistic, 0.68). By using the 2-tier system, patients with low-grade versus high-grade tumors had significantly longer PFS (45.0 vs 19.8 months, respectively; P = .01). By using FIGO criteria, median PFS for patients with grade 1, 2, and 3 tumors was 37.5, 19.8, and 20.1 months, respectively (P = .07). There was no difference in clinical outcome in patients with grade 2 or 3 tumors in multivariate analysis. Woman with high-grade versus low-grade tumors demonstrated significantly higher risk of death (hazard ratio, 2.43; 95% confidence interval, 1.17-5.04; P = .02). CONCLUSIONS: Women with high-grade versus low-grade serous carcinoma of the ovary are 2 distinct patient populations. Adoption of the 2-tier grading system provides a simple yet precise framework for predicting clinical outcomes.

Multifocal vulvar smooth muscle tumor with an unusual intravascular growth pattern and multiple local recurrences in a 10-year-old child: a diagnostic dilemma.

Smooth muscle tumors of the vulva are rare entities with variable morphologic features and clinical behavior. Our report summarizes the case of a multifocal vulvar smooth muscle tumor of undetermined malignant potential with an unusual intravascular growth pattern and subsequent local recurrence in a 10-year-old girl, the youngest case reported to date, and reviews the specific pathologic findings of that category. Vulvar smooth muscle tumors of undetermined malignant potential represent a diagnostic, prognostic, and therapeutic challenge. The histologic features of this case were predictive of local recurrence, although the significance of the intravascular growth pattern is still uncertain. The relationship of this multifocal, recurrent tumor to the clinical entity, vulvar leiomyomatosis, is unknown at this time.

Modulatory effect of curcumin on survival of irradiated human intestinal microvascular endothelial cells: role of Akt/mTOR and NF-{kappa}B.

Radiation therapy is an essential modality in the treatment of colorectal cancers. Radiation exerts an antiangiogenic effect on tumors, inhibiting endothelial proliferation and survival in the tumor microvasculature. However, damage from low levels of irradiation can induce a paradoxical effect, stimulating survival in endothelial cells. We used human intestinal microvascular endothelial cells (HIMEC) to define effects of radiation on these gut-specific endothelial cells. Low-level irradiation (1-5 Gy) activates NF-kappaB and the phosphatidylinositol 3-kinase (PI3K)/Akt pathway, which is involved in cell cycle reentry and cell survival in HIMEC. A downstream target of PI3K/Akt is mammalian target of rapamycin (mTOR), which contributes to endothelial proliferation and angiogenesis. The aim of this study was to investigate the signaling molecules involved in the radiosensitizing effects of curcumin on HIMEC subjected to low levels of irradiation. We have demonstrated that exposure of HIMEC to low levels of irradiation induced Akt and mTOR phosphorylation, which was attenuated by curcumin, rapamycin, LY294002, and mTOR small interference RNA (siRNA). Activation of NF-kappaB by low levels of irradiation was inhibited by curcumin, SN-50, and mTOR siRNA. Curcumin also induced apoptosis by induction of caspase-3 cleavage in irradiated HIMEC. In conclusion, curcumin significantly inhibited NF-kappaB and attenuated the effect of irradiation-induced prosurvival signaling through the PI3K/Akt/mTOR and NF-kappaB pathways in these gut-specific endothelial cells. Curcumin may be a potential radiosensitizing agent for enhanced antiangiogenic effect in colorectal cancer radiation therapy.

Diverse histologic appearances in pulmonary mucinous cystic neoplasia: a case report.

INTRODUCTION: Primary pulmonary mucinous cystic neoplasia comprises a group of tumors, from benign cystadenoma to mucinous cystadenocarcinoma. CASE PRESENTATION: We report a case of primary pulmonary mucinous cystadenocarcinoma in a 75-year-old woman who was found to have a right hilar mass on a routine chest X-ray. A lobectomy was performed and the resection specimen revealed a multicystic mucinous tumor. Microscopically, the tumor was composed of confluent mucin-filled cystic spaces lined by columnar mucin-secreting cells which ranged from cytologically bland to moderately atypical with 'bronchioloalveolar pattern' invasion into the adjacent parenchyma. Immunohistochemically, tumor cells were positive diffusely for Cytokeratin 7, and focally for Cytokeratin 20 and Thyroid Transcription Factor-1. CONCLUSION: This case highlights the continuous spectrum of pulmonary mucinous cystic neoplasia from benign mucinous cystadenoma to malignant mucinous cystadenocarcinoma, and the probable existence of a 'borderline' mucinous cystic tumor. Although molecular data are lacking to substantiate progression from benign to malignant in these neoplasms, the importance of recognizing the morphologic continuum lies in alerting pathologists to thoroughly examine specimens to rule out invasive foci in tumors with 'borderline' morphology.

Role of MutS homolog 2 (MSH2) in intestinal myofibroblast proliferation during Crohn's disease stricture formation.

Tissue remodeling and mesenchymal cell accumulation accompanies chronic inflammatory disorders involving joints, lung, vasculature, and bowel. Chronic inflammation may alter DNA-mismatch repair (MMR) systems in mesenchymal cells, but is not defined in Crohn's disease (CD) and its associated intestinal remodeling and stricture formation. We determined whether DNA-MMR alteration plays a role in the pathogenesis of CD tissue remodeling. Control and CD bowel tissues were used to generate primary cultures of muscularis mucosa myofibroblasts, which were assessed directly or following stimulation with TNF-alpha/LPS or H2O2. MutS homolog (MSH)2, MSH3, and MSH6 expression in tissues and myofibroblasts was determined. Immunohistochemical staining revealed an increased expression of MSH2 in CD muscularis mucosa and submucosal tissues compared with controls or uninvolved CD tissue, and MSH2 expression was increased in CD myofibroblasts compared with control cells. TNF-alpha/LPS and H2O2 further enhanced MSH2 expression in both control and CD cells, which were decreased by simvastatin. There were no significant changes in MSH3 and MSH6 expression. Proliferating cell nuclear antigen and Ki67 staining of CD tissue revealed increased proliferation in the muscularis mucosa and submucosa of chronically inflamed tissues, and enhanced proliferation was seen in CD myofibroblasts compared with controls. Simvastatin reversed the effects of inflammatory stress on the DNA-MMR and inhibited proliferation of control and CD myofibroblasts. Gene silencing with MSH2 siRNA selectively decreased CD myofibroblast proliferation. These data demonstrate a potential role for MSH2 in the pathogenesis of nonneoplastic mesenchymal cell accumulation and intestinal remodeling in CD chronic inflammation.

Ischemic uterine rupture and hysterectomy 3 months after uterine artery embolization.

The exact frequency and extent of complications after uterine artery embolization (UAE) have yet to be documented in the literature. Ischemic necrosis and rupture of the uterus is a theoretical concern of this procedure. Rupture of the uterus from any cause is a very serious gynecologic complication requiring immediate surgical intervention to prevent death. Ischemic necrosis and rupture of the uterus can occur months after UAE. In our patient they occurred 3 months after UAE for treatment of symptomatic uterine myomas, and required hysterectomy. To our knowledge, this is the first report of ischemic uterine rupture after UAE in the United States.