csi-miR-96-5p delivered by Clonorchis sinensis extracellular vesicles promotes intrahepatic cholangiocarcinoma proliferation and migration via the ferroptosis-related PTEN/SLC7A11/GPX4 axis.

BACKGROUND: Clonorchis sinensis (CS) is classified as a group 1 carcinogen and can cause intrahepatic cholangiocarcinoma (ICC). CS extracellular vesicles (CsEVs) play important roles in mediating communication between parasitic helminths and humans. Ferroptosis is a novel cell death mechanism that is mainly induced by lipid peroxidation and iron overload. However, the role of CsEVs in the regulation of ferroptosis in ICC remains unclear. This study aimed to explore the role of CS-secreted miR-96-5p (csi-miR-96-5p) delivered by CsEVs in ICC progression and ferroptosis. METHODS: Tissue samples were collected from ICC patients with CS infection (CS-ICC) or without CS infection (NC-ICC). The levels of csi-miR-96-5p and PTEN gene were determined by quantitative polymerase chain reaction (qPCR) and western blotting, and survival analysis was performed. CsEVs were isolated and identified by ultracentrifugation and transmission electron microscopy. Lentiviruses were used to establish stable cell lines with csi-miR-96-5p mimic expression, PTEN overexpression (PTEN-EXO) and PTEN CRISPR/Cas9-based knockout (PTEN-KO) and their respective negative controls. Cell proliferation was assessed by performing Cell Counting Kit-8 assays in vitro and in a tumor xenograft model in vivo, and cell migration was assessed by performing Transwell assays. Erastin is used to induce ferroptosis. Ferroptosis levels were evaluated using biomarkers. RESULTS: High csi-miR-96-5p and low PTEN expression was observed in CS-ICC tissues and was associated with poor overall survival. csi-miR-96-5p was highly enriched in CsEVs and was taken up by ICC cells. csi-miR-96-5p mimics or PTEN-KO significantly promoted the growth and migration of ICC cells in vitro and in vivo, whereas PTEN-EXO exerted the opposite effect. Mechanistically, csi-miR-96-5p mimics or PTEN-KO inhibited erastin-induced ferroptosis, including reducing the accumulation of Fe2+, lipid reactive oxygen species, and malondialdehyde, increasing the GSH/GSSG ratio and levels of SLC7A11 and GPX4, whereas PTEN-EXOs exerted the opposite effect. CONCLUSIONS: csi-miR-96-5p delivered by CsEVs reduced ferroptosis by regulating the expression of the PTEN/SLC7A11/GPX4 axis, thereby promoting ICC proliferation and migration. For the first time to our knowledge, we found that CS miRNAs could promote tumor development through ferroptosis.

Efficacy and Safety of Digital Single-Operator Cholangioscopy in the Diagnosis of Indeterminate Biliary Strictures by Targeted Biopsies: A Systematic Review and Meta-Analysis.

BACKGROUND: Biliary strictures are frequently encountered in clinical practice. The determination of their nature is often difficult. This study aims to systematically evaluate the efficacy and safety of the second generation of digital single-operator cholangioscopy (SpyGlass DS, DSOC) in indeterminate biliary strictures (IBDS) through biopsies. METHODS: All relative studies published in Medline, the Cochrane Library, Web of Science, and EMBASE were included. The diagnostic tests for IBDS were compared to the surgical histology, autopsy, or long-term clinical follow-up. The methodological quality of the included studies was evaluated by the Quality Assessment of Studies of Diagnostic Accuracy Included in Systematic Reviews (QUADAS-2). RESULTS: A total of 11 studies, which involved 356 patients diagnosed through biopsies, were included. The pooled sensitivity, specificity, positive likelihood ratio, negative likelihood ratio and diagnostic odds ratio were 0.74 (95% CI: 0.67-0.80), 0.98 (95% CI: 0.95-1.00), 10.52 (95% CI: 5.45-20.32), 0.31 (95% CI: 0.23-0.41), and 65.18 (95% CI: 26.79-158.61), respectively. The area under the curve (AUC) was 0.9479, and the pooled adverse event rate was 7%. The sensitivity and specificity in the heterogeneity analysis were I2 = 48.1% and I2 = 25.4%, respectively. CONCLUSION: SpyGlass DS is a safe and effective technique for IBDS. However, future randomized trials are needed to determine optimal number of biopsies.

Utility of multiple endoscopic techniques in differential diagnosis of gallbladder adenomyomatosis from gallbladder malignancy with bile duct invasion: A case report.

BACKGROUND: Gallbladder adenomyomatosis (GAM) is a benign lesion, characterized by thickening of the gallbladder wall and a focal mass, which overlap with the features of gallbladder malignancy. Consequently, differential diagnosis of GAM from gallbladder cancer is difficult and approximately 20% of suspected malignant biliary strictures are postoperatively confirmed as benign lesions. Herein, we report a case in which a preoperative diagnosis of GAM was made by a combination of endoscopic and imaging techniques. CASE SUMMARY: A 40-year-old man was referred to our hospital chiefly for a fever and right upper abdominal pain with dark urine. Enhanced computed tomography showed thickening of the gallbladder wall and a mass in the gallbladder neck with involvement of the hepatic bile ducts, which was suspected to be malignant. Gallbladder malignancy with bile duct invasion was ruled out by subsequent endoscopic examinations, including endoscopic retrograde cholangio-pancreatography, intraductal ultrasound, and SpyGlass. Endoscopic examinations showed a homogeneous hyperechoic lesion with smooth margins of benign bile duct stricture suggestive of inflammatory stenosis of the bile duct. The patient underwent laparoscopic cholecystectomy. GAM was postoperatively diagnosed and confirmed based on the histopathology results, which are consistent with the preoperative diagnosis. Notably, no malignant event occurred in the patient during a 12-mo follow-up period. CONCLUSION: A combination of endoscopic techniques may help in the differential diagnosis of GAM from gallbladder cancer.

The clinical profiles, management, and prognostic factors of biliary mixed neuroendocrine nonneuroendocrine neoplasms: A systematic review of the literature.

BACKGROUND: Mixed neuroendocrine nonneuroendocrine neoplasms (MiNENs) originating from the biliary system (gallbladder, biliary tract, or ampulla of Vater) are extremely rare and have not been discussed in detail or systematically. We aimed to present the demographics, clinicopathological characteristics, management, and prognostic factors of biliary MiNENs. METHODS: A systematic search of electronic biomedical databases (Web of Science, PUBMED, and Embase) was performed to identify eligible studies. Survival was analyzed with the Kaplan-Meier method. Log-rank tests were used to evaluate the differences between groups, and the effects of various clinical and histopathological features on prognosis were analyzed by univariate and multivariate Cox regression. RESULTS: Fifty-three publications (patients, n = 67) were included. The median overall survival time was 21.0 months. Fifty-one patients (76.1%) underwent radical surgery and median survival for 41 months (P < .001). Twenty-two patients who received adjuvant radiochemotherapy treatment after radical surgery had a median survival for 43 months (P = .076). Radical resection (P < .001), Ki-67 index (P = .011), tumor stage (P < .001), neuroendocrine (NEC) grade (P = .011), and non-NEC grade (P = .017) were independent statistically significant prognostic factors according to univariate analysis; radical resection (P = .010) and small morphological subtype (P = .036) were independent statistically significant prognostic factors associated with higher overall survival according to multivariate analysis, and radical resection (P = .005) and age < 65 years (P = .026) were associated with higher recurrence free survival time. CONCLUSION: Radical resection is essential for long-term survival. Aggressive multimodality therapy with adjuvant radiochemotherapy and biotherapy may improve survival of biliary MiNENs. Further randomized controlled trials are needed to determine the standard treatment.