The apelin­apelin receptor signaling pathway in fibroblasts is involved in tumor growth via p53 expression of cancer cells.

Cancer­associated fibroblasts (CAFs) are pivotal in tumor progression. TP53­deficiency in cancer cells is associated with robust stromal activation. The apelin­apelin receptor (APJ) system has been implicated in suppressing fibroblast­to­myofibroblast transition in non­neoplastic organ fibrosis. The present study aimed to elucidate the oncogenic role of the apelin­APJ system in tumor fibroblasts. APJ expression and the effect of APJ suppression in fibroblasts were investigated for p53 status in cancer cells using human cell lines (TP53­wild colon cancer, HCT116, and Caco­2; TP53­mutant colon cancer, SW480, and DLD­1; and colon fibroblasts, CCD­18Co), resected human tissue samples of colorectal cancers, and immune­deficient nude mouse xenograft models. The role of exosomes collected by ultracentrifugation were also analyzed as mediators of p53 expression in cancer cells and APJ expression in fibroblasts. APJ expression in fibroblasts co­cultured with p53­suppressed colon cancer cells (HCT116sh p53 cells) was significantly lower than in control colon cancer cells (HCT116sh control cells). APJ­suppressed fibroblasts treated with an antagonist or small interfering RNA showed myofibroblast­like properties, including increased proliferation and migratory abilities, via accelerated phosphorylation of Sma­ and Mad­related protein 2/3 (Smad2/3). In addition, xenografts of HCT116 cells with APJ­suppressed fibroblasts showed accelerated tumor growth. By contrast, apelin suppressed the upregulation of phosphorylated Smad2/3 in fibroblasts. MicroRNA 5703 enriched in exosomes derived from HCT116sh p53 cells inhibited APJ expression, and inhibition of miR­5703 diminished APJ suppression in fibroblasts caused by cancer cells. APJ suppression from a specific microRNA in cancer cell­derived exosomes induced CAF­like properties in fibroblasts. Thus, the APJ system in fibroblasts in the tumor microenvironment may be a promising therapeutic target.

A novel role for Helicobacter pylori cytotoxin-associated gene A in negative regulation of autophagy in human gastric cells.

BACKGROUND: Autophagy plays an important role in carcinogenesis and tumor progression in many cancers, including gastric cancer. Cytotoxin-associated gene A (CagA) is a well-known virulent factor in Helicobacter pylori (H. pylori) infection that plays a critical role in gastric inflammation and gastric cancer development. However, its role in autophagy during these processes remains unclear. Therefore, we aimed to clarify the role of CagA in autophagy in CagA-related inflammation. METHODS: We evaluated the autophagic index of AGS cells infected with wild-type cagA-positive H. pylori (Hp-WT) and cagA-knockout H. pylori (Hp-ΔcagA) and rat gastric mucosal (RGM1) cells transfected with CagA genes. To identify the mechanisms underlying the down regulation of autophagy in AGS cells infected with H. pylori, we evaluated protein and mRNA expression levels of autophagy core proteins using western blotting and quantitative reverse transcription-polymerase chain reaction (RT-PCR). To determine whether autophagy induced the expression of the pro-inflammatory mediator, cyclooxygenase-2 (COX-2), we evaluated COX-2 expression in AGS cells treated with an autophagy inducer and inhibitor and infected with H. pylori. In addition, we evaluated whether COX-2 protein expression in AGS cells influenced beclin-1 (BECN1) expression with si-RNA transfection when infected with H. pylori. RESULTS: Autophagic flux assay using chloroquine showed that autophagy in AGS cells was significantly suppressed after H. pylori infection. The autophagic index of AGS cells infected with Hp-WT was decreased significantly when compared with that in AGS cells infected with Hp-ΔcagA. The autophagic index of RGM1 cells transfected with CagA was lower, suggesting that CagA inhibits autophagy. In addition, BECN1 expression levels in AGS cells infected with Hp-WT were reduced compared to those in AGS cells infected with Hp-ΔcagA. Furthermore, COX-2 expression in AGS cells infected with H. pylori was controlled in an autophagy-dependent manner. When AGS cells were transfected with small interfering RNA specific for BECN1 and infected with Hp-WT and Hp-ΔcagA, COX-2 was upregulated significantly in cells infected with Hp-ΔcagA. CONCLUSIONS: In conclusion, the H. pylori CagA protein negatively regulated autophagy by downregulating BECN1. CagA-induced autophagy inhibition may be a causative factor in promoting pro-inflammatory mediator production in human gastric epithelial cells.

Clinical and Histopathological Features of the Differentiated Type of Early Gastric Cancer with an Unclear Lateral Demarcation.

INTRODUCTION: Magnifying endoscopy with narrow-band imaging (M-NBI) is useful for determining lateral demarcation of early gastric cancers; however, this is sometimes difficult. Features related to an unclear lateral demarcation remain unknown. We evaluated the clinical and histopathological features of early gastric cancers with unclear lateral demarcation on M-NBI. METHODS: This single-center, retrospective, cohort study analyzed early gastric cancer treated by endoscopic submucosal dissection between January 2013 and August 2015. We evaluated the clinicopathological and immunohistochemical features using anti-p53, anti-Ki-67, anti-MUC5AC, anti-MUC6, anti-MUC2, and anti-CD10 antibody staining. We compared the lateral demarcation between the demarcation clear (DC) and the demarcation unclear (DU) lesions by using M-NBI. RESULTS: A total of 224 differentiated adenocarcinomas (DU group: 18 lesions; DC group: 206 lesions) were analyzed. A history of successful Helicobacter pylori eradication was significantly more frequent in the DU group (p = 0.001). We examined the tissues of 72 lesions (DU group: 18 lesions, DC group: 54 lesions [randomly selected]) immunohistochemically. The nonneoplastic superficial epithelium was observed more frequently in the DU group as compared to in the DC group (p = 0.006). Additionally, compared to the DC group, the DU group showed a significantly higher expression of the gastric phenotype markers (p = 0.023) and had lower p53 scores (p < 0.001) and Ki-67 labeling indexes (p = 0.029). Multivariate analysis revealed the nonneoplastic superficial epithelium and a low p53 score as the significant independent variables associated with an unclear lateral demarcation on M-NBI. CONCLUSIONS: The nonneoplastic superficial epithelium and a low p53 score were associated with difficulties in determining the lateral demarcation in early gastric cancers on M-NBI.

Suppression of autophagy promotes fibroblast activation in p53-deficient colorectal cancer cells.

Deficiency of p53 in cancer cells activates the transformation of normal tissue fibroblasts into carcinoma-associated fibroblasts; this promotes tumor progression through a variety of mechanisms in the tumor microenvironment. The role of autophagy in carcinoma-associated fibroblasts in tumor progression has not been elucidated. We aimed to clarify the significance of autophagy in fibroblasts, focusing on the TP53 status in co-cultured human colorectal cancer cell lines (TP53-wild-type colon cancer, HCT116; TP53-mutant colon cancer, HT29; fibroblast, CCD-18Co) in vitro. Autophagy in fibroblasts was significantly suppressed in association with ACTA2, CXCL12, TGFß1, VEGFA, FGF2, and PDGFRA mRNA levels, when co-cultured with p53-deficient HCT116sh p53 cells. Exosomes isolated from the culture media of HCT116sh p53 cells significantly suppressed autophagy in fibroblasts via inhibition of ATG2B. Exosomes derived from TP53-mutant HT29 cells also suppressed autophagy in fibroblasts. miR-4534, extracted from the exosomes of HCT116sh p53 cells, suppressed ATG2B in fibroblasts. In conclusion, a loss of p53 function in colon cancer cells promotes the activation of surrounding fibroblasts through the suppression of autophagy. Exosomal miRNAs derived from cancer cells may play a pivotal role in the suppression of autophagy.

Use of a convolutional neural network for classifying microvessels of superficial esophageal squamous cell carcinomas.

BACKGROUND AND AIM: The morphological diagnosis of microvessels on the surface of superficial esophageal squamous cell carcinomas using magnifying endoscopy with narrow-band imaging is widely used in clinical practice. Nevertheless, inconsistency, even among experts, remains a problem. We constructed a convolutional neural network-based computer-aided diagnosis system to classify the microvessels of superficial esophageal squamous cell carcinomas and evaluated its diagnostic performance. METHODS: In this retrospective study, a cropped magnifying endoscopy with narrow-band images from superficial esophageal squamous cell carcinoma lesions was used as the dataset. All images were assessed by three experts, and classified into three classes, Type B1, B2, and B3, based on the Japan Esophagus Society classification. The dataset was divided into training and validation datasets. A convolutional neural network model (ResNeXt-101) was trained and tuned with the training dataset. To evaluate diagnostic accuracy, the validation dataset was assessed by the computer-aided diagnosis system and eight endoscopists. RESULTS: In total, 1777 and 747 cropped images (total, 393 lesions) were included in the training and validation datasets, respectively. The diagnosis system took 20.3 s to evaluate the 747 images in the validation dataset. The microvessel classification accuracy of the computer-aided diagnosis system was 84.2%, which was higher than the average of the eight endoscopists (77.8%, P < 0.001). The area under the receiver operating characteristic curves for diagnosing Type B1, B2, and B3 vessels were 0.969, 0.948, and 0.973, respectively. CONCLUSIONS: The computer-aided diagnosis system showed remarkable performance in the classification of microvessels on superficial esophageal squamous cell carcinomas.

Usefulness of a dedicated mouthpiece for the Valsalva maneuver to visualize the hypopharynx during transoral endoscopy.

Background and study aims Patients with esophageal squamous cell carcinoma (SCC) are at high risk of developing second primary SCCs in the hypopharynx. However, such second primary tumors are difficult to observe because of lumen closure. The Valsalva maneuver using a dedicated mouthpiece is a promising technique to visualize the hypopharynx during transoral endoscopy. In the current study, we investigated the utility of this method. Patients and methods The current study was a randomized, controlled, crossover trial. Patients with esophageal SCC were randomly assigned first to undergo pharyngeal observation using the dedicated mouthpiece followed by observation using a conventional mouthpiece, or vice versa. The primary endpoint was complete visualization of the hypopharynx, which was assessed blindly by three external evaluators. Results A total of 68 pharyngeal examinations were analyzed - 34 with the dedicated mouthpiece and 34 with a conventional mouthpiece. Complete visualization was achieved in 68 % of the examinations (23/34) using the dedicated mouthpiece, whereas none of the examinations using the conventional mouthpiece achieved complete visualization of the hypopharynx. Observation scores of the oropharynx were not significantly different between both types of examination ( P  = 0.50). No serious adverse events (AEs) occurred. Conclusions Endoscopic view of the hypopharynx was markedly improved by the Valsalva maneuver using the dedicated mouthpiece, with no serious AEs. This procedure should be included in the endoscopic examinations for the patients with esophageal SCCs.

Additional effect of magnifying narrow-band imaging on estimating the invasion depth of superficial esophageal cancer.

BACKGROUND AND AIM: To investigate whether assessment by magnifying narrow-band imaging (M-NBI) based on the classification of the Japan Esophageal Society provides additional value to the estimation of the invasion depth of superficial esophageal squamous cell carcinoma (SCC) compared with assessment by white light endoscopy (WLE) alone. METHODS: Endoscopic images of 211 consecutive superficial esophageal SCCs resected by endoscopic submucosal dissection were separated into WLE and M-NBI images. Depth estimation was performed independently by five evaluators using the numerical depth estimation scale (0 = epithelium (EP)/lamina propria (LPM), 1 = EP/LPM > muscularis mucosa (MM)/shallow submucosa (SM1), 2 = MM/SM1 > EP/LPM, 3 = MM/SM1, 4 = MM/SM1 > deep submucosa (SM2), 5 = SM2 > MM/SM1, 6 = SM2), using primarily WLE images (step 1), and subsequently both WLE and M-NBI images (step 2). The discordance scores, determined by the average of the five evaluators' difference between the estimated score (from 0 to 6) and pathological score (0 for histologically proven EP/LPM, 3 for MM/SM1, and 6 for SM2), were analyzed in steps 1 and 2. RESULTS: The discordance scores significantly decreased in step 2 (0.53 ± 0.06) compared with those in step 1 (0.79 ± 0.07) (P < 0.001). When the discordance score < 1.5 was regarded as a clinically correct diagnosis, the rate of the clinically correct diagnosis significantly increased in step 2 compared with that in step 1 (81% to 91%, P < 0.001). CONCLUSION: M-NBI has an additive value for estimating the invasion depth of superficial esophageal SCCs.

Exosomal microRNAs derived from colon cancer cells promote tumor progression by suppressing fibroblast TP53 expression.

The tumor microenvironment offers favorable conditions for tumor progression, and activated fibroblasts, known as cancer-associated fibroblasts, play a pivotal role. TP53-deficient cancer cells are known to induce strong fibroblast activation. We aimed to elucidate the oncogenic role of exosomes derived from TP53-deficient colon cancer cells in fibroblast proliferation and tumor growth. Cancer cell-derived exosomes (CDEs) were isolated from the conditioned media of cancer cells using a sequential ultracentrifugation method. The effects of exosomes on tumor growth were evaluated using human cell lines (TP53-WT colon cancer, HCT116; TP53-mutant colon cancer, HT29; and fibroblasts, CCD-18Co and WI-38) and an immune-deficient nude mouse xenograft model. HCT116 (HCT116sh p53 ) cells deficient in TP53 accelerated cocultured fibroblast proliferation compared to TP53-WT HCT116 (HCT116sh control ) cells in vitro. Exosomes from HCT116sh p53 cells suppressed TP53 expression of fibroblasts and promoted their proliferation. Xenografts of HCT116sh p53 cells grew significantly faster than those of HCT116sh control cells in the presence of co-injected fibroblasts, but this difference was diminished by CDE inhibition. Microarray analysis identified upregulation of several microRNAs (miR-1249-5p, miR-6737-5p, and miR-6819-5p) in TP53-deficient CDEs, which were functionally proven to suppress TP53 expression in fibroblasts. Exosomes derived from TP53-mutant HT29 cells also suppressed TP53 expression in fibroblasts and accelerated their growth. The proliferative effect of HT29 on cocultured fibroblasts was diminished by inhibition of these miRNAs in fibroblasts. Our results suggest that CDEs play a pivotal role in tumor progression by fibroblast modification. Cancer cell-derived exosomes might, therefore, represent a novel therapeutic target in colon cancer.

The heart's exposure to radiation increases the risk of cardiac toxicity after chemoradiotherapy for superficial esophageal cancer: a retrospective cohort study.

BACKGROUND: Chemoradiotherapy effectively treats superficial esophageal cancer and is optimal to preserve organs. However, late toxicity, particularly in cardiac diseases, obstructs clinical outcomes. We revealed the risk factors for cardiac event development post-chemoradiotherapy. METHODS: Data from 80 patients who were diagnosed with submucosal invasive esophageal cancer without metastasis (confirmed using multiple modalities) and who underwent chemoradiotherapy between 2006 and 2014 were analyzed. Patients were 11% (9/80) female, and the median age and follow-up were 66.5 y and 73 mo, respectively. We calculated the individual radiation dose to the heart and analyzed relationships between the cardiac event occurrence rate and each clinical factor. RESULTS: The 5-y overall and recurrence-free survival rates were 74.6 and 62.4%, respectively. Among the total number of deaths, 34.6% was caused by esophageal cancer. During the follow-up, 13 patients developed severe cardiac events (ischemic heart diseases, n = 7; pericardial effusion, n = 3, atrial fibrillation, n = 1; and sudden death, n = 2). The significant risk factor for cardiac events post-chemoradiotherapy was the level of the heart's exposure to radiation, with higher exposure associated with greater occurrence. History of smoking, obesity, comorbidity, and history of cardiac disease were unrelated to cardiac event occurrence post-chemoradiotherapy. CONCLUSIONS: Chemoradiotherapy is a favorable intervention for superficial esophageal cancer. Reducing the radiation dose to the heart likely contributes to preventing cardiac toxicity post-chemoradiotherapy.

Adenoma of colorectal laterally spreading tumor nongranular type with biological phenotypic features similar to cancer.

BACKGROUND AND AIM: Colorectal laterally spreading tumors (LSTs) are morphologically subdivided into granular (LST-G) and nongranular (LST-NG) categories. We aimed to elucidate the differences in oncogenic characteristics between the two types. METHODS: Laterally spreading tumors resected by endoscopic submucosal dissection and surgery from March 2009 to May 2017 were examined for p53 positivity, Ki-67 labeling index (LI), microvessel density, degree of fibrosis, intensities of inducible nitric oxide synthase (iNOS) and nitrotyrosine (NT), and expression of acid mucins. We compared these factors between adenomas, noninvasive cancers, and invasive cancers, both LST-G and LST-NG. RESULTS: Ninety-three LST-G (53 adenomas [LST-GA] and 40 cancers [LST-GC]) and 55 LST-NG (24 adenomas [LST-NGA] and 31 cancers [LST-NGC]) were evaluated. Although p53 positivity was lower in LST-GA than in LST-NGA (P < 0.001), there was no difference between LST-GC and LST-NGC. Ki-67 LI was higher in LST-NGA than in LST-GA (P < 0.001) and higher in LST-NGC than in LST-GC of noninvasive cancers (P < 0.001). Microvessel density and degree of fibrosis were higher in LST-NGA than in LST-GA (P < 0.001), and intensities of iNOS and NT were also higher in LST-NGA than in LST-GA (P < 0.001). Expression of acid mucins was lower in LST-NGA than in LST-GA (P < 0.001). Although there were significant differences in p53 positivity, Ki-67 LI, microvessel density, degree of fibrosis, intensities of iNOS and NT, and expression of acid mucins between LST-GA and LST-NGA, these factors were only slightly different between LST-GC and LST-NGC of invasive cancers. CONCLUSIONS: Unlike LST-GA, LST-NGA possessed phenotypic features similar to cancer.

Cardiac metastasis from colon cancer effectively treated with 5-fluorouracil, leucovorin, and oxaliplatin (modified FOLFOX6) plus panitumumab: a case report.

BACKGROUND: Cardiac metastasis from colorectal cancer is rare. There is little evidence supporting the effectiveness of chemotherapy, and standard therapy for metastatic cardiac tumors has not been established. CASE PRESENTATION: A 76-year-old woman presented with a right ventricle tumor that was detected incidentally on screening cardiac ultrasonography. The initial computed tomography (CT) scan showed the cardiac tumor, which was approximately 40 mm in size, and multiple pulmonary nodules. Serum levels of tumor markers CEA and CA19-9 were elevated aberrantly. The suspected primary tumor, a well-differentiated adenocarcinoma of the transverse colon with wild-type KRAS was found by colonoscopy, and treatment with 5-fluorouracil, leucovorin, and oxaliplatin (modified FOLFOX6) plus panitumumab was initiated. After 4 courses of the therapy, a CT scan showed that the cardiac tumor size had markedly decreased and the pulmonary nodules had diminished. The serum levels of CEA and CA19-9 were also markedly decreased. After 12 courses of chemotherapy during 10 months of treatment, the patient continued to show a partial response, and she remained asymptomatic with continuation of the treatment through 15 courses. CONCLUSION: To the best of our knowledge, this is the first report of the efficacy of combination therapy using cytotoxic and molecular targeted agents against cardiac metastasis from colon cancer.

A case of gastric mixed adenoneuroendocrine carcinoma with difficulty in diagnosis before endoscopic submucosal dissection.

A woman in her 60s was referred to our hospital with a superficial depressed lesion measuring 8mm in diameter on the lesser curvature of the lower gastric body. Initial biopsy of the lesion indicated a moderately differentiated adenocarcinoma. Endoscopic submucosal dissection was performed, and pathological examination revealed a tumor comprised of adenocarcinoma and neuroendocrine carcinoma with submucosal infiltration, with the final pathological diagnosis being gastric mixed adenoneuroendocrine carcinoma (MANEC). Laparoscopic gastrectomy was subsequently performed. No recurrence was observed after 18 months. Most neuroendocrine carcinomas including MANEC are diagnosed at an advanced stage and require surgical resection. Here we report a case of gastric MANEC mimicking early gastric cancer that was removed en bloc via endoscopic submucosal dissection.

Pancreaticoduodenal artery aneurysm associated with coeliac artery occlusion from an aortic intramural hematoma.

Pancreaticoduodenal artery aneurysms are a rare type of visceral artery aneurysm, whose rupture is associated with high mortality. These aneurysms are of particular interest because local haemodynamic change caused by coeliac artery obstruction plays an important role in their development. However, the pathophysiological mechanism of coeliac artery obstruction is not completely understood. Pressure from the median arcuate ligament is most frequently reported cause. Although it is well-known that stenosis or occlusion of the visceral vessels may be caused by aortic syndrome, reports of pancreaticoduodenal artery aneurysm associated with coeliac artery occlusion due to aortic syndrome are extremely rare. Our case indicates a new aetiology for a pancreaticoduodenal artery aneurysm and demonstrates the rapid deterioration of the patient affected.

Protracted anaphylaxis developed after peginterferon α-2a administration for chronic hepatitis C.

Peginterferon is a key drug used to treat chronic viral hepatitis that is known for causing various side effects. Side effects occurring immediately after administration include headache, nausea, and influenza-like symptoms, such as fever and joint pain. However, reports of anaphylactic shock are extremely rare. Here we report a patient with protracted anaphylaxis who suffered shock symptoms after peginterferon α-2a administration for chronic hepatitis C. Although the patient improved temporarily with shock treatment, symptoms of anaphylaxis recurred. As peginterferon is often administered on an outpatient basis, it is important to recognize life-threatening side effects that may develop in a protracted manner.