Differentiating localized autoimmune pancreatitis and pancreatic ductal adenocarcinoma using endoscopic ultrasound images with deep learning.

Objectives: Localized autoimmune pancreatitis is difficult to differentiate from pancreatic ductal adenocarcinoma on endoscopic ultrasound images. In recent years, deep learning methods have improved the diagnosis of diseases. Hence, we developed a special cross-validation framework to search for effective methodologies of deep learning in distinguishing autoimmune pancreatitis from pancreatic ductal adenocarcinoma on endoscopic ultrasound images. Methods: Data from 24 patients diagnosed with localized autoimmune pancreatitis (8751 images) and 61 patients diagnosed with pancreatic ductal adenocarcinoma (20,584 images) were collected from 2016 to 2022. We applied transfer learning to a convolutional neural network called ResNet152, together with our innovative imaging method contributing to data augmentation and temporal data process. We divided patients into five groups according to different factors for 5-fold cross-validation, where the ordered and balanced datasets were created for the performance evaluations. Results: ResNet152 surpassed the endoscopists in all evaluation metrics with almost all datasets. Interestingly, when the dataset is balanced according to the factor of the endoscopists' diagnostic accuracy, the area under the receiver operating characteristic curve and accuracy were highest at 0.85 and 0.80, respectively. Conclusions: It is deduced that image features useful for ResNet152 correlate with those used by endoscopists for their diagnoses. This finding may contribute to sample-efficient dataset preparation to train convolutional neural networks for endoscopic ultrasonography-imaging diagnosis.

Role of Bile-Derived Extracellular Vesicles in Hepatocellular Proliferation after Partial Hepatectomy in Rats.

Although liver regeneration has been extensively studied, the effects of bile-derived extracellular vesicles (bile EVs) on hepatocytes has not been elucidated. We examined the influence of bile EVs, collected from a rat model of 70% partial hepatectomy (PH), on hepatocytes. We produced bile-duct-cannulated rats. Bile was collected over time through an extracorporeal bile duct cannulation tube. Bile EVs were extracted via size exclusion chromatography. The number of EVs released into the bile per liver weight 12 h after PH significantly increased. Bile EVs collected 12 and 24 h post-PH, and after sham surgery (PH12-EVs, PH24-EVs, sham-EVs) were added to the rat hepatocyte cell line, and 24 h later, RNA was extracted and transcriptome analysis performed. The analysis revealed that more upregulated/downregulated genes were observed in the group with PH24-EVs. Moreover, the gene ontology (GO) analysis focusing on the cell cycle revealed an upregulation of 28 types of genes in the PH-24 group, including genes that promote cell cycle progression, compared to the sham group. PH24-EVs induced hepatocyte proliferation in a dose-dependent manner in vitro, whereas sham-Evs showed no significant difference compared to the controls. This study revealed that post-PH bile Evs promote the proliferation of the hepatocytes, and genes promoting cell cycles are upregulated in hepatocytes.

A case of successful removal of a migrated fish bone in the bile duct after pancreaticoduodenectomy using overtube-assisted cholangioscopy.

Fish bone migration into the bile duct in patients with surgically altered anatomy is a very rare cause of bile duct stones. Recently, balloon-assisted endoscopic retrograde cholangiopancreatography (BAERCP) is performed for biliary lesions in patients with surgically altered anatomy. We report on a 73-year-old Japanese man with a history of pancreaticoduodenectomy for intraductal papillary mucinous adenoma. A 20 mm long linear hyperattenuating structure in the left intrahepatic bile duct was noted on routine follow-up computed tomography 14 years postoperatively. The linear structure persisted until follow-up computed tomography performed 15 years postoperatively, and the left intrahepatic bile duct was shown to be dilated. We performed BAERCP for the diagnosis and treatment of the linear structure but could not visualize the linear structure in the left intrahepatic bile duct via enteroscopy and fluoroscopy. We removed the enteroscope, leaving the overtube, and inserted the cholangioscope through the overtube over the guide wire. We observed a brown rod-shaped linear structure in the left intrahepatic bile duct and removed it under direct visualization via overtube-assisted cholangioscopy. We conclude that overtube-assisted cholangioscopy was useful for assessing undiagnosed biliary lesions using conventional BAERCP and removing fish bones in the bile duct of the patient with altered gastrointestinal anatomy.

Biosafety of a novel covered self-expandable metal stent coated with poly(2-methoxyethyl acrylate) in vivo.

Covered self-expandable metal stents (CSEMS) are often used for palliative endoscopic biliary drainage; however, the unobstructed period is limited because of sludge occlusion. The present study aimed to evaluate the biosafety of a novel poly(2-methoxyethyl acrylate)-coated CSEMS (PMEA-CSEMS) for sludge resistance and examine its biosafety in vivo. Using endoscopic retrograde cholangiopancreatography, we placed the PMEA-CSEMS into six normal porcine bile ducts and conventional CSEMS into three normal porcine bile ducts. We performed serological examination and undecalcified histological analysis at 1, 3, and 6 months during follow-up. In the bile ducts with PMEA-CSEMS or conventional CSEMS, we observed no increase in liver enzyme or inflammatory marker levels in the serological investigations and mild fibrosis but no inflammatory response in the histopathological analyses. Thus, we demonstrated the biosafety of PMEA-CSEMS in vivo.

A case of a smooth transition to subsequent percutaneous transjejunal biliary intervention for hepatolithiasis after biliary reconstruction by adding jejunostomy during an emergency operation for perforation due to balloon-assisted endoscopy.

Treatments for hepatolithiasis include peroral endoscopy, percutaneous cholangioscopy, and surgery. Balloon-assisted endoscopic retrograde cholangiopancreatography (BAERCP) has been widely performed in recent years for patients with hepatolithiasis after biliary reconstruction. However, accidental bowel perforation caused by BAERCP may need emergency surgery. Here, we describe a 77-year-old Japanese woman diagnosed with acute cholangitis due to hepatolithiasis after biliary reconstruction (a biliary diversion operation for pancreaticobiliary maljunction). She underwent BAERCP for treatment of hepatolithiasis, however, a small-bowel perforation occurred. She underwent an emergency operation to suture the perforation and add a catheter jejunostomy. She had no postoperative complications after surgery and was discharged 11 days after surgery. One month later, she was readmitted and underwent percutaneous transjejunal cholangioscopy-guided lithotripsy with complete removal of the calculi. Although endoscopists should be careful to avoid small-bowel perforation during BAERCP, if perforation occurs, addition of a catheter jejunostomy during emergency surgery can be easily transitioned to subsequent treatment of the hepatolithiasis.

Utility of Claudin-3 in extracellular vesicles from human bile as biomarkers of cholangiocarcinoma.

Extracellular vesicles (EVs) are released from all cells. Bile directly contacts bile duct tumor; bile-derived EVs may contain high concentrations of cancer biomarkers. We performed a proteomic analysis of human bile-derived EVs and identified a novel biomarker of cholangiocarcinoma (CCA). EVs were isolated using ultracentrifugation, and chelating agents, ethylenediaminetetraacetic acid and ethylene glycol tetraacetic acid (EDEG) and phosphate buffered saline (PBS) were used as dissolution solutions. Bile was collected from 10 CCA and 10 choledocholithiasis (stones) cases. Proteomic analysis was performed; subsequently, ELISA was performed using the candidate biomarkers in a verification cohort. The vesicles isolated from bile had a typical size and morphology. The expression of exosome markers was observed. RNA was more abundant in the EDEG group. The proportion of microRNA was higher in the EDEG group. EDEG use resulted in the removal of more contaminants. Proteomic analysis identified 166 proteins as CCA-specific. ELISA for Claudin-3 revealed statistically significant difference. The diagnostic accuracy was AUC 0.945 and sensitivity and specificity were 87.5%. We report the first use of EDEG in the isolation of EVs from human bile and the proteomic analysis of human bile-derived EV-proteins in CCA. Claudin-3 in bile-derived EVs is a useful biomarker for CCA.

Signet-ring cell carcinoma of the ampulla of Vater: a case diagnosed via repeated biopsies.

Signet-ring cell carcinoma of the ampulla of Vater is a rare tumor. A 74-year-old woman presented with epigastric pain and was diagnosed with cholangitis. Her liver enzyme levels were elevated. Computed tomography showed an enhanced area in the periampullary region and marked common bile duct dilatation. On endoscopic retrograde cholangiopancreatography (ERCP), the ampulla exhibited a normal appearance without ulcer or mass. Histological biopsy confirmed the absence of malignancy. During follow-up, the patient again presented with acute cholangitis multiple times and underwent ERCP each time. The ampulla had the appearance of a reddish and erosive mucosa. Although biopsy was repeated, histological examination did not show any malignancy. After a total of 13 biopsies, the patient was diagnosed with ampullary carcinoma of non-exposed protruded type following the third ERC-guided biopsy. Careful follow-up and frequent endoscopic biopsies are important in cases of papillary carcinoma of non-exposed protruded type with normal ampullary mucosa on initial endoscopy because this condition is challenging to diagnose with a single biopsy.

Usefulness of rapid on-site evaluation specimens from endoscopic ultrasound-guided fine-needle aspiration for cancer gene panel testing: A retrospective study.

Pancreatic cancer (PC) is a highly lethal malignancy, with a 5-year survival rate of 6%. Cancer gene panel testing is expected to allow selection of suitable therapeutic drugs in individual patients with PC and improve their prognosis. Although somatic mutations can be identified in formalin-fixed, paraffin-embedded samples derived from surgical specimen, the rate of surgical indication among patients with PC is only 20%. To acquire genome information with a less invasive method, we used rapid on-site evaluation (ROSE) specimens from endoscopic ultrasound-guided fine-needle aspiration. The present study aimed to retrospectively evaluate the utility of comprehensive cancer gene panel testing with ROSE specimens. DNA was extracted from preserved ROSE specimens of 26 patients diagnosed with PC between 2011 and 2017. DNA sequences of oncogenes and cancer-related genes were determined using the Ion AmpliSeq Comprehensive Caner Panel. We compared KRAS mutations between cancer gene panel testing by next-generation sequencing (NGS) and KRAS mutation analysis by polymerase chain reaction. The mean yield of DNA per extraction from ROSE specimens was 171 ng (range, 34-478 ng). On cancer gene panel testing, we noted KRAS mutations (92%), TP53 mutations (50%), CDKN2A mutations (15%), and SMAD4 mutations (31%). The concordance rate of KRAS mutations between cancer gene panel testing by NGS using ROSE specimens and KRAS mutation analysis by the companion diagnostics using residual materials was 81%. Among five cases of KRAS discordance, three showed KRAS mutations in cancer gene panel testing but not in KRAS mutation analysis. Cancer gene panel testing with ROSE specimens can help stratify unresectable PC patients without additional invasive approaches, and it can be used for therapeutic drug selection.

A novel pathogenic variant of ATP-binding cassette subfamily B member 4 causing gallstones in a young adult.

The low phospholipid-associated cholelithiasis (LPAC) syndrome was reported in European adults with cholelithiasis and a mutation of the ATP-binding cassette subfamily B member 4 (ABCB4). The ABCB4 encodes multidrug resistance 3, which is a phospholipid translocator. Reduced phospholipid transport can lead to the formation of biliary cholesterol stones. Here, we describe a 31-year-old Japanese man diagnosed with recurrent biliary colic. Although he recovered quickly after endoscopic treatment for the most recent presentation, he had a family history of similar problems. His mother had required endoscopic treatment for choledocholithiasis and his maternal aunt had died at age 29 years because of liver failure (etiology unknown). We, therefore, performed genetic analysis, which revealed a heterozygous ABCB4C717S. LPAC syndrome was diagnosed and the patient has received ursodeoxycholic acid for 2 years with no recurrence. The same variant was identified in the patient's mother, who was subsequently found to have a left intrahepatic calculus requiring left-sided lobectomy. She has received ursodeoxycholic acid for 1 year with no recurrence. ABCB4C717S is a novel pathogenic variant, and this is the first patient diagnosed with LPAC syndrome in Japan. We should consider LPAC syndrome in young adults with recurrent cholesterol gallstones to ensure early therapy.

[Hypopituitarism associated with autoimmune pancreatitis: a case report].

We herein report the case of a 64-year-old male patient with hypopituitarism associated with autoimmune pancreatitis (AIP). The patient was previously diagnosed with AIP based on the presence of a swollen pancreas, elevated serum immunoglobulin G4, and narrowing of the pancreatic duct by imaging. Magnetic resonance imaging revealed a pituitary stem tumor, and loading test showed a decrease in the function of the anterior lobe suggesting severe failure of growth hormone secretion. Treatment with steroids was effective in reducing the pituitary lesion and improving the function of the anterior lobe. The present case illustrates the importance of pituitary function evaluation before steroid treatment in patients with AIP.

A case of pancreatic intraepithelial neoplasia-3 with autoimmune pancreatitis.

We report a case of pancreatic intraepithelial neoplasia-3 (PanIN-3) with autoimmune pancreatitis (AIP). The patient, a 75-year-old man, had been diagnosed to have AIP with stenosis of the main pancreatic duct. After six years, computed tomography demonstrated dilatation of the main pancreatic duct in the mid-pancreas. Although we could not confirm the presence of any pancreatic tumor on the basis of imaging modalities alone, cytological examination of the pancreatic juice obtained by endoscopic retrograde pancreatography revealed atypical cells. Therefore, we performed pancreatoduodenectomy and obtained a pathologic diagnosis of PanIN-3 with AIP. The present case is informative in the context of pancreatic carcinogenesis in AIP.

High Glucose Accelerates Cell Proliferation and Increases the Secretion and mRNA Expression of Osteopontin in Human Pancreatic Duct Epithelial Cells.

BACKGROUND: The incidence of pancreatic cancer is increasing year-by-year in Japan. Among the diseases that complicate pancreatic cancer, diabetes is the most common. Recently, it has become evident that patients suffering from diabetes and obesity show increased expression of osteopontin (OPN). The purpose of this study was to investigate the effect of high glucose and high insulin culture conditions on a human pancreatic duct epithelial cell line (HPDE-6), focusing particularly on OPN expression. METHODS: HPDE-6 were cultured under various conditions, employing several combinations of glucose (normal, 6 mM high, 30 mM, and 60 mM) and insulin (0.1 nM, 1 nM) concentration. RESULTS: HPDE-6 cell proliferation was significantly accelerated under high glucose culture conditions in comparison to samples in 6 mM glucose, and was more prominent under high insulin conditions. At the same time, the expression of OPN mRNA was also increased significantly. In comparison with 6 mM glucose, the expression of 8-OHdG DNA was increased in high glucose culture. CONCLUSION: HPDE-6 cells show accelerated proliferation and increased OPN expression when cultured under high glucose and high insulin conditions. Furthermore, the cells show increased oxidative stress in the presence of high glucose.

Specific MAPK-Associated MicroRNAs in Serum Differentiate Pancreatic Cancer from Autoimmune Pancreatitis.

Pancreatic ductal adenocarcinoma (PDAC) is difficult to distinguish from autoimmune pancreatitis (AIP) because of their clinical and radiological similarities, and therefore simple and minimally invasive surrogate markers for differential diagnosis would be useful. In our previous studies, we identified four microRNAs (miRNAs)-miR-7, miR-34a, miR-181d, and miR-193b -as MAPK-associated microRNAs whose expression was altered significantly with upregulation of the MAPK signaling pathway. Recently it has been reported that these miRNAs could be used as biomarkers in serum samples for accurate diagnosis of pancreatic lesions. The aim of the present study was to evaluate whether these MAPK-associated miRNAs in serum could be used as biomarkers for differentiating PDAC from AIP. We enrolled 69 patients with PDAC, 26 with intraductal papillary mucinous neoplasm (IPMN) and 15 with AIP. The expression of MAPK-associated miRNAs in serum was measured by quantitative real-time PCR. The 2-ΔCT method was used to quantify the expression of miRNAs, and the data were normalized using spiked-in synthetic cel-miR-39. Patients with PDAC or IPMN showed significantly higher amounts of serum MAPK-associated miRNAs than those with AIP (p<0.009 for miR-7, p<0.002 for miR-34a, p<0.001 for miR-181d, p<0.002 for miR-193b). ROC curve analysis demonstrated that these miRNAs had an area under the ROC curve (AUC) of 0.723-0.882 for differentiation between PDAC or IPMN from AIP. Furthermore, serum miR-181d was significantly associated with the presence of metastasis in patients with PDA (p = 0.014). Serum MAPK-associated miRNAs could be novel noninvasive biomarkers for differentiation between PDAC or IPMN and AIP.

Stromal Fibrosis and Expression of Matricellular Proteins Correlate With Histological Grade of Intraductal Papillary Mucinous Neoplasm of the Pancreas.

OBJECTIVE: The aim of the study was to clarify the correlation between the microenvironmental factors and histological grade in intraductal papillary mucinous neoplasm (IPMN). METHODS: We investigated 65 IPMNs resected at Yamagata University Hospital between 2000 and 2011, and all cases were categorized to low-inter (including low- and intermediate-grade dysplasia) and high-inv (including high-grade dysplasia and IPMN with an associated invasive carcinoma) groups. We compared between the 2 groups pathologically with regard to fibrosis and the expression of alpha-smooth muscle actin (α-SMA), periostin, and galectin-1 in the periductal stroma of IPMN. RESULTS: There were 41 low-inter and 24 high-inv. The subtype was categorized as 22 main duct type (MD-IPMN) and 43 branch duct type (BD-IPMN). The degree of fibrosis and the expression of α-SMA, periostin, and galectin-1 were significantly higher in high-inv than in low-inter within BD-IPMNs. Multivariate logistic regression analysis indicated that high expression of α-SMA (odds ratio, 13.802; 95% confidence interval, 1.108-171.893; P = 0.0414) was a significant independent related factor of high-inv in BD-IPMN. CONCLUSIONS: Stromal fibrosis and expression of α-SMA, periostin, and galectin-1 are more marked in high-inv than in low-inter within BD-IPMNs, and they could become new markers for determining the indications for surgery in BD-IPMN.

Pancreatic fat accumulation, fibrosis, and acinar cell injury in the Zucker diabetic fatty rat fed a chronic high-fat diet.

OBJECTIVE: The histological alteration of the exocrine pancreas in obesity has not been clarified. In the present study, we investigated biochemical and histological changes in the exocrine pancreas of obese model rats. METHODS: Zucker lean rats were fed a standard diet, and Zucker diabetic fatty (ZDF) rats were divided into 2 groups fed a standard diet and a high-fat diet, respectively. These experimental groups were fed each of the diets from 6 weeks until 12, 18, 24 weeks of age. We performed blood biochemical assays and histological analysis of the pancreas. RESULTS: In the ZDF rats fed a high-fat diet, the ratio of accumulated pancreatic fat area relative to exocrine gland area was increased significantly at 18 weeks of age in comparison with the other 2 groups (P < 0.05), and lipid droplets were observed in acinar cells. Subsequently, at 24 weeks of age in this group, pancreatic fibrosis and the serum exocrine pancreatic enzyme levels were increased significantly relative to the other 2 groups (P < 0.01). CONCLUSIONS: In ZDF rats fed a chronic high-fat diet, fat accumulates in pancreatic acinar cells, and this fatty change seems to be related to subsequent pancreatic fibrosis and acinar cell injury.