Clinical impact of bile-derived exosomal microRNAs as novel diagnostic and prognostic biomarkers for biliary tract cancers.

Sampling of bile juice during endoscopic retrograde cholangiopancreatography (ERCP) has potential benefit of being amenable to the identification of novel biomarkers in liquid biopsy. This study reports the results of a global investigation of exosomal microRNAs (miRNAs) in bile to identify potential biomarkers for biliary tract cancers (BTCs). Eighty-eight bile samples collected during ERCP (45 BTC and 43 noncancer control samples) were enrolled in this study. Eleven BTC samples and nine control samples were assigned as the discovery set. Exosomes in bile and serum samples were collected using a glass membrane column with size-controlled macroporous glass (MPG), and exosomal miRNA expression profiles were evaluated using comprehensive miRNA microarray analysis (3D-Gene). For validation, exosomal miRNA in the bile samples of 34 BTCs and 34 controls were comprehensively evaluated using 3D-Gene. In the discovery set, eight exosomal miRNAs in bile were identified as significant aberrant expression markers, while no miRNA with aberrant expression in serum was identified. In a comparison of the discovery and validation sets, miR-451a and miR-3619-3p were identified as reproducible upregulated markers, and the combination of the two bile miRNAs showed an excellent area under the curve (0.819) value for diagnosing BTCs. In addition, high miR-3619-3p expression in bile reflects poorer prognosis of BTCs (hazard ratio = 2.89). The MPG-extracted exosomal miRNAs in bile aspirated during ERCP provide a convenient new approach for diagnosing biliary diseases. Bile-derived miRNA analysis with miR-451a and miR-3619-3p represents a potentially valuable diagnostic strategy for identifying BTCs as well as a predictive indicator of BTC prognosis.

Urolithin A targets the AKT/WNK1 axis to induce autophagy and exert anti-tumor effects in cholangiocarcinoma.

Urolithin A (UA; 3,8-dihydroxybenzo[c]chromen-6-one), a metabolite generated by intestinal bacteria during the biotransformation of ellagitannins, has gained considerable attention in treating several cancers. Cholangiocarcinoma (CCA) remains one of the most lethal cancers; it grows in a special environment constantly exposed to both blood and bile. Since UA is known to undergo enterohepatic recirculation, we hypothesized that UA might have significant antitumor effects in CCA. Here, we investigated the therapeutic potential of UA in CCA and aimed to elucidate its mechanisms, including autophagy. UA treatment inhibited cell proliferation and induced G2/M phase cell cycle arrest in CCA cells. UA also suppressed cell migration and invasion, but did not cause apoptosis. Furthermore, Western blotting and immunocytochemistry demonstrated increased LC3-II accumulation, while electron microscopy demonstrated induced autophagosomes after UA treatment, suggesting that UA upregulated autophagy in CCA cells. In xenograft mice treated with UA, tumor growth was inhibited with increased LC3-II levels. On the other hand, phospho-kinase array demonstrated downregulation of the AKT/WNK1 pathway. LC3-II expression was elevated in WNK1 knocked down cells, indicating that WNK1 is the key signal for regulating autophagy. Thus, UA exerted antitumor effects by suppressing the AKT/WNK1 signaling pathway and inducing autophagy. In conclusion, UA, a natural, well-tolerated compound, may be a promising therapeutic candidate for advanced CCA.

Subclinical hepatitis E virus (HEV) infection detected by nucleic acid amplification test on blood donation: short-term positivity for immunoglobulin G class of antibody against HEV.

A case of subclinical hepatitis E virus (HEV) infection was detected by nucleic acid amplification test on blood donation. The patient was followed-up until day 220 after the blood donation but showed no symptoms throughout the observation period. Aspartate aminotransferase and alanine aminotransferase levels reached the maximum values on day 37 with a slight increase but remained in normal ranges from day 67 to 220. The quantity of HEV RNA at the initial examination on day 13 was 1.1 × 102 copies/mL, which increased to 2.8 × 103 copies/mL by day 37. It was not detected from day 67 to 220. Immunoglobulin G class antibody to HEV (anti-HEV IgG) was below the cut-off value until day 37 and exceeded the cut-off value to positive on day 67, accompanied by normalization of liver function and negative conversion of HEV RNA. Thereafter, the titer decreased gradually, falling below the cut-off value on day 163, and continuing negative until day 220. Although the persistent duration of anti-HEV IgG positive is believed to be generally long, it was within only 126 days for this subclinical case. Further investigation is needed to determine whether short-term positivity for anti-HEV IgG is typical in subclinical HEV infection.

Relevance of gene mutations and methylation to the growth of pancreatic intraductal papillary mucinous neoplasms based on pyrosequencing.

We aimed to assess some of the potential genetic pathways for cancer development from non-malignant intraductal papillary mucinous neoplasm (IPMN) by evaluating genetic mutations and methylation. In total, 46 dissected regions in 33 IPMN cases were analyzed and compared between malignant-potential and benign cases, or between malignant-potential and benign tissue dissected regions including low-grade IPMN dissected regions accompanied by malignant-potential regions. Several gene mutations, gene methylations, and proteins were assessed by pyrosequencing and immunohistochemical analysis. RASSF1A methylation was more frequent in malignant-potential dissected regions (p = 0.0329). LINE-1 methylation was inversely correlated with GNAS mutation (r = - 0.3739, p = 0.0105). In cases with malignant-potential dissected regions, GNAS mutation was associated with less frequent perivascular invasion (p = 0.0128), perineural invasion (p = 0.0377), and lymph node metastasis (p = 0.0377) but significantly longer overall survival, compared to malignant-potential cases without GNAS mutation (p = 0.0419). The presence of concordant KRAS and GNAS mutations in the malignant-potential and benign dissected regions were more frequent among branch-duct IPMN cases than among the other types (p = 0.0319). Methylation of RASSF1A, CDKN2A, and LINE-1 and GNAS mutation may be relevant to cancer development, IPMN subtypes, and cancer prognosis.

Impact of physiologically shaped pancreatic stent for chronic pancreatitis.

Endoscopic pancreatic stenting is used to prevent main pancreatic duct obstruction and relieve painful symptoms of chronic pancreatitis. However, the stent typically needs to be exchanged and the rate of adverse events is high. Few studies have evaluated the effect of stent shape on those outcomes. We evaluated the adverse events, stent patency, and total medical cost within 90 days of patients who received an 8.5 French (Fr) physiologically shaped pancreatic stent by comparing these features with those associated with a conventional straight-type stent for ≥ 90 days. The total stent-related adverse event rate was significantly lower for the physiologically shaped pancreatic stent (physiologically shaped, 6.7% [2/30]; straight-type, 50.6% [44/87]; P < 0.001). Stent occlusion was significantly less frequent (P < 0.001) and the total medical costs were significantly lower (P = 0.002) for the physiologically shaped stent. The stent-related adverse event rate was significantly higher for the 10 Fr straight type stent than for the 8.5 Fr physiologically shaped stent (10 Fr, straight-type vs. 8.5 Fr, physiologically shaped: 36.1% [13/36] vs. 6.7% [2/30]; P = 0.007). In conclusion, a physiologically shaped pancreatic stent was superior to a straight-type stent in terms of the patency rate and medical costs.

Four-Step Classification of Endoscopic Transpapillary Gallbladder Drainage and the Practical Efficacy of Cholangioscopic Assistance.

Background/Aims: Although endoscopic transpapillary gallbladder drainage (ETGBD) has been reported as an alternative procedure for acute cholecystitis, it requires advanced endoscopic techniques. In terms of the certainty of achieving drainage, it remains a challenging procedure. The aim of the current study was to elucidate the practical efficacy of cholangioscopic assistance and to develop a new classification that could be used to evaluate the technical difficulty of ETGBD and provide a theoretical strategy to apply cholangioscopy appropriately for difficult ETGBD. Methods: A total of 101 patients undergoing ETGBD were retrospectively studied. The characteristics and technical outcomes of ETGBD with conventional ETGBD (C-ETGBD) and SpyGlass DS-assisted ETGBD (SG-ETGBD) were evaluated. The characteristics and technique-dependent factors of unsuccessful C-ETGBD/SG-ETGBD were evaluated using the classification based on the steps of the procedure. The predictive factors of successful C-ETGBD/SG-ETGBD were examined. Results: C-ETGBD was successful in 73 patients (72.3%). SG-ETGBD was successful in 11 of 13 patients (84.6%) who had C-ETGBD failure. Optional SG-ETGBD significantly increased the final success rate (94.1%) compared to C-ETGBD alone (p=0.003). ETGBD procedures could be classified into four steps. SG-assistance worked as an excellent troubleshooter in step 1 (failure to identify the cystic duct orifice) and step 2 (failure of guidewire advancement across the downturned angle of cystic duct takeoff). Magnetic resonance cholangiopancreatography could provide predictive information based on the classification. Conclusions: Optional SG-ETGBD achieved a significantly higher success rate than C-ETGBD alone. Step classification is helpful for determining the technical difficulty of ETGBD and developing a theoretical strategy to apply cholangioscopy in a coordinated manner.

Autotaxin in ascites promotes peritoneal dissemination in pancreatic cancer.

Peritoneal dissemination and malignant ascites in pancreatic ductal adenocarcinoma (PDAC) patients represent a major clinical issue. Lysophosphatidic acid (LPA) is a lipid mediator that modulates the progression of various cancers. Based on the increasing evidence showing that LPA is abundant in malignant ascites, we focused on autotaxin (ATX), which is a secreted enzyme that is important for the production of LPA. This study aimed to elucidate the importance of the ATX-LPA axis in malignant ascites in PDAC and to determine whether ATX works as a molecular target for treating peritoneal dissemination. In a PDAC peritoneal dissemination mouse model, the amount of ATX was significantly higher in ascites than in serum. An in vitro study using two PDAC cell lines, AsPC-1 and PANC-1, showed that ATX-LPA signaling promoted cancer cell migration via the activation of the downstream signaling, and this increased cell migration was suppressed by an ATX inhibitor, PF-8380. An in vivo study showed that PF-8380 suppressed peritoneal dissemination and decreased malignant ascites, and these results were validated by the biological analysis as well as the in vitro study. Moreover, there was a positive correlation between the amount of ATX in ascites and the degree of disseminated cancer progression. These findings demonstrated that ATX in ascites works as a promotor of peritoneal dissemination, and the targeting of ATX must represent a useful and novel therapy for peritoneal dissemination of PDAC.

Early Detection of High-grade Biliary Intraepithelial Neoplasia (BilIN-3) in the Cystic Duct Visualized by SpyGlass DS Cholangioscopy.

An 84-year-old man was admitted with epigastralgia. Computed tomography showed contrast-enhanced wall thickness in the cystic duct. An endoscopic examination revealed short irregular stricture in the cystic duct, and per-oral cholangioscopy revealed a reddish papillary tumor at the stricture site. Surgical resection revealed high-grade biliary intraepithelial neoplasia (BilIN) at the stricture site of the cystic duct. To our knowledge, this is the first case of a solitary high-grade BilIN epithelium in the cystic duct detected by per-oral cholangioscopy.

Metachronous Pancreatic Ductal Adenocarcinoma with Adjacent Serous Cystadenoma that Was Preoperatively Diagnosed by EUS-FNA: A Case Report and Review of the Literature.

Pancreatic serous cystic neoplasms (SCNs), such as serous cystadenoma (SCA), are generally recognized as benign because malignant counterparts of SCNs have been extremely rare. In clinical practice, pancreatic cystic neoplasms diagnosed as SCNs have been managed by conservative observation, as long as the patients remained asymptomatic. We herein report a case of metachronous ductal adenocarcinoma that was discovered during long-term follow-up of SCN and review the related literature. To our knowledge, this was the first reported case of the local presence of ductal adenocarcinoma adjacent to SCA that was preoperatively diagnosed by endoscopic ultrasound-guided fine-needle aspiration.

Predictive factors for the mortality of acute pancreatitis on admission.

BACKGROUND AND AIMS: The revised Atlanta classification is widely used for the evaluation of acute pancreatitis (AP) severity. However, this classification cannot be used within 48 hours of AP onset. The aim of this study was to investigate the predictive factors of mortality in patients with AP on admission. METHODS: We evaluated the association between AP mortality and clinical parameters at the time of admission in patients with AP from April 2013 to December 2017 at one university hospital and one tertiary care referral center. RESULTS: A total of 203 consecutive patients were enrolled. Nine patients (4.4%) died despite multidisciplinary treatment. In a multivariable analysis, hematocrit ≥ 40% (odds ratio [OR], 1.07; 95% confidence interval [CI], 1.01-1.13; P = 0.021), blood urea nitrogen (BUN) ≥ 40 mg/dL (OR, 1.26; 95% CI, 1.11-1.42; P < 0.001), base excess < -3.0 mmol/L (OR, 1.15; 95% CI, 1.04-1.26; P = 0.004), and inflammation extending to the rectovesical excavation (OR, 1.19; 95% CI, 1.10-1.30; P < 0.001) on admission were significantly associated with mortality. CONCLUSION: Among the imaging findings, inflammation extending to the rectovesical excavation was the only independent predictive factor for mortality in AP. This simple finding, obtained on computed tomography without contrast agent on admission, might be a promising prognostic factor for AP.

Laterally Spreading Adenocarcinoma Involving the Lower Bile Duct and Duodenum Expressing Heterogeneous Immunohistochemical Phenotypes.

A 70-year-old man was admitted to our hospital due to elevated levels of hepatobiliary and pancreatic enzymes. Computed tomography showed contrast-enhanced mucosal hypertrophy from the duodenal papilla to the distal bile duct. Endoscopic examinations revealed a laterally spreading granular tumor and ampullary swelling. After surgical resection, an examination revealed well-differentiated adenocarcinoma of the ampulla with tubular adenoma spreading from the distal common bile duct to the second part of the duodenum showing both bile duct and duodenal phenotypes. To our knowledge, this is the first case of a tumor spreading from the bile duct to the duodenum that exhibited multiple phenotypes.

Feasibility and safety of duodenal covered self-expandable metallic stent fixation: an experimental study.

BACKGROUND AND AIMS: Migration of duodenal covered self-expandable metallic stents (C-SEMS) is the main cause of stent dysfunction in patients with malignant gastric outlet obstruction. However, the ideal method to prevent migration has not been clarified. We aimed to evaluate the feasibility and safety of duodenal C-SEMS fixation in this experimental study. METHODS: We used the over-the-scope clip (OTSC), suture, and clip methods to fix duodenal C-SEMS and evaluated the gripping force of each device and invasion depth based on pathological findings. RESULTS: The OTSC and suturing systems had a significantly higher mean gripping force compared with the clipping system (OTSC vs. clip: 13.2 vs. 1.0 Newtons [N], P < 0.001; suture vs. clip: 8.5 vs. 1.0 N, P < 0.001). OTSC compression was stronger compared with suturing (OTSC vs. suture: 13.2 vs. 8.5 N, P = 0.006). The submucosal layer, but not the muscle layer, was compressed more widely and deeply by OTSC compared with clips based on pathological findings by hematoxylin and eosin staining. CONCLUSION: Both OTSC and suturing methods used for duodenal C-SEMS fixation were feasible compared with the clipping method. The pathological evaluation of invasion depth indicated that OTSC may be safe even for preventive use. This study suggested that these methods can be applied clinically for duodenal C-SEMS fixation.

An Increased Chromosome 7 Copy Number in Endoscopic Bile Duct Biopsy Specimens Is Predictive of a Poor Prognosis in Cholangiocarcinoma.

BACKGROUND: The ability of fluorescence in situ hybridization (FISH) assays in endoscopic transpapillary bile duct biopsy specimens to predict the prognosis of cholangiocarcinoma (CCA) has not been elucidated. AIMS: We aimed to clarify the association between the results of UroVysion FISH assays and the prognosis of CCA. METHODS: We retrospectively reviewed 49 specimens obtained by transpapillary forceps biopsy from consecutive patients with CCA. The copy numbers of chromosomes 3, 7, and 17 were evaluated by FISH assay using UroVysion. We compared the overall survival (OS) of CCA patients with and without increased copy numbers of chromosomes 3, 7, and 17. Furthermore, we evaluated the association between OS and the clinicopathological parameters of CCA patients. RESULTS: The OS was significantly shorter in patients with than without an increased chromosome 7 copy number (log-rank p = 0.015; median OS 11.9 vs. 20.7 months). In the univariate analyses, age (p = 0.012), ECOG performance status (p = 0.046), tumor stage (p = 0.046), surgery (p = 0.006), and an increased chromosome 7 copy number (p = 0.017) were significantly associated with OS. The multivariate analysis revealed that an increased chromosome 7 copy number (hazard ratio, 2.46; 95% CI 1.15-5.27; p = 0.021) and advanced clinical stage (hazard ratio, 2.26; 95% CI 1.11-4.63; p = 0.025) were independently predictive of a poor OS. CONCLUSIONS: Detection by FISH assay of an increased chromosome 7 copy number in transpapillary forceps biopsy specimens is predictive of a poor prognosis in CCA patients.

The utility and efficacy of self-expandable metal stents for treating malignant gastric outlet obstructions in patients under best supportive care.

PURPOSE: Self-expandable metallic stents (SEMSs) may be used to effectively palliate malignant gastric outlet obstructions (GOOs), but their utility and efficacy in patients under best supportive care (BSC) have not been explored. METHOD: In this multicenter retrospective study, we reviewed data on patients under BSC who underwent endoscopic SEMS placement to treat malignant GOO without chemotherapy. We evaluated the safety and efficacy of the procedure. RESULTS: We enrolled a total of 208 patients. SEMS placement was technically successful in 207 (99.5%) and clinically successful in 164 (78.8%). The mean procedure time was 25.6 ± 2.8 min. Stent dysfunction later developed in 30 (14.4%) patients, of whom 90% (27/30) underwent reintervention. The procedure-related mortality rate was 1.44%; all deaths were due to pneumonia. Subgroup analysis by Karnofsky performance status (KPS) revealed that neither technical success, stent dysfunction, reintervention rate, procedure-related pneumonia or death, nor death within 14 days differed between patients with good and poor KPS. However, the clinical success rate and the median survival time were significantly lower and shorter, respectively, in those with poor KPS (p < 0.001). CONCLUSIONS: Duodenal SEMS placement is an effective palliative treatment for malignant GOO in BSC patients. Although the GOO score did not dramatically improve in patients with poor KPS, the procedure was safe and palliatively feasible. Procedure-related pneumonia was fatal; thus, it is essential to proceed with great caution. TRIAL REGISTRATION: Clinical trial registration number: UMIN000028367.

Case of pancreatic metastasis from colon cancer in which cell block using the Trefle® endoscopic scraper enables differential diagnosis from pancreatic cancer.

Endoscopic transpapillary brush cytology and forceps biopsy during endoscopic retrograde cholangiopancreatology are generally used to obtain pathological evidence of biliary strictures. Recently, the new endoscopic scraper Trefle® has been reported and demonstrated high cancer detectability in malignant biliary strictures. This device is used to scrape the stricture over the guidewire, and, in the original method, the tissue and/or cell samples obtained are subjected to histological and/or cytological analysis separately. However, discrimination of chunks of tissue is hampered by the opacity of the surrounding fluid. We have developed a cell block technique for the Trefle® device without dividing obtained specimens into tissue and cellular components, which is the simplest method and enables immunohistochemical analysis. We present a case of obstructive jaundice diagnosed immunohistochemically as pancreatic metastasis from colon cancer using cell block sections obtained with the Trefle® device, which procedure is as easy as conventional brush cytology.

Covered duodenal self-expandable metal stents prolong biliary stent patency in double stenting: The largest series of bilioduodenal obstruction.

BACKGROUND AND AIM: Endoscopic biliary and duodenal stenting (DS; double stenting) is widely accepted as a palliation therapy for malignant bilioduodenal obstruction. The aim of the current study was to investigate the patency and adverse events of duodenal and biliary stents in patients with DS. METHODS: Patients who underwent DS from April 2004 to March 2017 were analyzed retrospectively with regard to clinical outcomes and predictive factors of recurrent biliary and duodenal obstruction (recurrent biliary obstruction [RBO] and recurrent duodenal obstruction [RDO]). RESULTS: A total of 109 consecutive patients was enrolled. Technical success of DS was achieved in 108 patients (99.1%). Symptoms due to biliary and duodenal obstruction were improved in 89 patients (81.7%). RBO occurred in 25 patients (22.9%) and RDO in 13 (11.9%). The median times to RBO and RDO from DS were 87 and 76 days, respectively. Placement of a duodenal uncovered self-expandable metal stent (U-SEMS) was significantly associated with RBO in the multivariable analysis (P = 0.007). Time to RBO was significantly longer in the duodenal covered self-expandable metal stent group than in the U-SEMS group (P = 0.003). No predictive factors of RDO were detected, and duodenal stent type was not associated with the time to RDO (P = 0.724). CONCLUSIONS: Double stenting was safe and effective for malignant bilioduodenal obstruction. Duodenal U-SEMS is a risk factor for RBO. The covered self-expandable metal stent is the preferred type of duodenal SEMS in patients with DS (Clinical trial registration number: UMIN000027606).

Autoimmune Hemolytic Anemia Obscured by the Obstructive Jaundice Associated with IgG4-related Sclerosing Cholangitis in a Patient with Type 1 Autoimmune Pancreatitis: A Case Report and Review of the Literature.

Type 1 autoimmune pancreatitis (AIP) is a pancreatic manifestation of IgG4-retated disease that is often associated with IgG4-related sclerosing cholangitis (IgG4-SC). Autoimmune hemolytic anemia (AIHA) is an immune-related disease that causes hemolytic anemia. Although type 1 AIP/IgG4-SC and AIHA have a shared etiology as a presumed autoimmune disease, they rarely overlap, and their association has not been clarified. Secondary AIHA might not be diagnosed appropriately because the obstructive jaundice observed in type 1 AIP/IgG4-SC can obscure the presence of hemolytic jaundice. We herein report a case of type 1 AIP/IgG4-SC overlapping with secondary AIHA along with a review of the literature.

Percutaneous Transhepatic Self-expanding Metallic Stent Placement for the Treatment of Malignant Afferent Loop Obstruction.

We report the case of a 71-year-old man with afferent loop obstruction (ALO) after Roux-en-Y reconstruction due to gastric cancer. Computed tomography showed a distended afferent loop and a dilatated bile duct. We could not reach the stricture site in the afferent loop using a gastroscope. We performed percutaneous transhepatic biliary drainage (PTBD) and placed a self-expanding metallic stent (SEMS) in the duodenal stricture through the PTBD route. Although an endoscopic approach is preferable, when PTBD can be performed, percutaneous transhepatic SEMS placement might be an alternative option for treating ALO in cases in which it is not possible to reach the site of stenosis with an endoscope.