Abstract:Background and Aims: Studies demonstrated that gene mutations are closely related to the occurrence of gallstones. This study was conducted to investigate the relationship between the single nucleotide polymorphisms (SNPs) of the BChE gene and the risk of gallstone disease in Han population from Guizhou province through a case-control study, so as to provide basis for studying its regulatory mechanism in gallstone formation. 
Methods: A total of 171 patients with gallstone disease and 125 health control subjects of Han nationality in Guizhou province were enrolled. The genomic DNA was extracted from whole blood samples of gallstone disease group and healthy control group. Using the risk-related SNP ABCG8 rs11887534 (D19H) as a positive control, the obesity-related genes BChE rs1126680 and rs1803274 in gallstone disease group and healthy control group were genotyped by TaqMan SNP assay. In homozygous and heterozygous models, the risk-related to SNPs was analyzed by Pearson χ2 test or Fisher’s exact test. The odds ratios were determined by Logistic regression analysis and the relationship between SNPs and gallstone susceptibility was analyzed.  
Results: The allele frequency distribution was in accordance with the Hardy- Weinberg equilibrium (χ2=0.641, P>0.05), so the selected sample had a population representativeness. The concentrations of high-density lipoprotein cholesterol and low-density lipoprotein cholesterol were significantly higher in the whole blood sample from patients with gallstones than those from healthy subjects (both P<0.05).  The genotyping results of TaqMan SNP assay showed that 3 tag SNPs were successfully genotyped, and the success rate of genotyping for each tag SNP was greater than or equal to 99.3%. The CG genotype of the positive control ABCG8 rs11887534 was a SNP mutation site related to the risk of gallstones (P=0.006), the CT genotype of the obesity-related gene BChE rs1803274 was significantly associated with gallstone disease (P=1.312×10–15), but the obesity-related gene BChE rs1126680 was an irrelevant mutation site to gallstone disease (P=0.616). There was a relatively strong correlation between the CT genotype of BChE rs1803274 and gallstone risk females (95% CI=0.237–0.772, P=0.009).
Conclusion: In Han population of Guizhou province, the SNPsof the BChE gene is closely related to the risk of gallstones. The risk of gallstones is increased in individual, especially in women with BChE variant rs1803274. 

Abstract:Background and Aims: The selection of an appropriate operation method is of great importance for the benign prognosis of patients with cholecystolithiasis and choledocholithiasis. In the past, the implementation of T-tube drainage on the basis of laparoscopic biliary exploration increases the degree of pain in patients, which is not conducive to postoperative recovery. Antegrade nasobiliary drainage has attracted more attention because of its minimally invasive concept. Therefore, this study was conducted to investigate the feasibility and safety of application of antegrade nasal bile duct in laparoscopic biliary tract exploration with primary closure of the common bile duct. 
Methods: Sixty patients with cholecystolithiasis and concomitant choledocholithiasis underwent laparoscopic biliary tract exploration from January 2017 to June 2018 were selected and randomly assigned to control group and observation group, with 30 cases in each group. Patients in observation group were given nasal biliary drainage and primary closure of the common bile duct, and those in control group underwent T-tube placement and bile drainage. The surgical variables, postoperative recovery conditions, the volume of postoperative bile drainage, the nutritional indicators at 1 week after operation, and the liver function parameters before and 1 year after operation as well as the incidence of complications during 1-year followed-up period between the two groups were compared. 
Results: The operative time and the volume of bile drainage on the first day after operation showed no significant difference between the two groups (all P>0.05). The drainage tube retention time, the time to first anal gas passage, the time to return to normal life, length of postoperative hospital stay and hospitalization costs as well as the volume of bile drainage on postoperative day 2 and 3 were all significantly reduced in observation group compared with control group (all P<0.05). The serum levels of albumin and prealbumin in observation group were significantly higher than those in control group at 1 week after operation (both P<0.05). There were no significant differences in each liver function parameter between the two groups before operation (all P>0.05), which were all significantly improved in both groups one year after operation (all P<0.05), but all showed no significant differences between the two groups (all P>0.05). The incidence of complications was significantly lower in observation group than that in control group during the 1-year follow-up period (3.33% vs. 26.67%, P<0.05). 
Conclusion: The combined use of antegrade nasobiliary drainage, laparoscopic biliary tract exploration, and primary closure of the common bile duct can shorten the hospitalization time and accelerate postoperative recovery of patients with gallstones and common bile duct stones, and also reduces the incidence of complications to a certain extent. So, it has certain feasibility and safety.

Abstract:Background and Aims: For hilar cholangiocarcinoma (HCCA), the R0 resection rate remains unsatisfactory. At present, there are still many controversies with regard to the preoperative evaluation, preoperative biliary drainage, portal vein embolization, surgical resection scope, surgical methods, vascular resection, lymph node dissection, chemotherapy and other issues. R0 resection is considered to be the most important treatment method for HCCA patients. Here, the authors summarize the experience in treatment of HCCA, and analyze the effectiveness as well as the short- and long-term efficacy of different surgical methods. 
Methods: The clinical data of 44 patients with HCCA undergoing surgical treatment from January 2015 to January 2020 were retrospectively analyzed.
Results: Among the 44 patients, 5 cases were classified as Bismuth-Corlette type I, 7 cases were type II, 
8 cases were type IIIa, 13 cases were type IIIb and 11 cases of type IV; 29 cases underwent hemihepatectomy/extended hemihepatectomy plus caudate lobectomy (combined hemihepatectomy), and 13 cases underwent hilar hepatectomy/perihilar hepatectomy plus caudate lobectomy (perihilar hepatectomy), including partial portal vein resection plus repair in 2 cases, portal vein resection plus reconstruction in 2 cases and hepatic artery resection plus reconstruction in 2 cases, and anther 2 cases underwent T-tube drainage only due to unresectable metastases. Operation was completed in all patients, with no surgical death. The postoperative pathological findings showed that there was negative microscopic margin (R0) in 37 cases (26 cases in combined hemihepatectomy group and 11 cases in perihilar hepatectomy group), and positive microscopic margin (R1) in 5 cases (2 cases in combined hemihepatectomy group and 3 cases in perihilar hepatectomy group). Results of clinical variable analysis showed that the operative time (240.4 min vs. 358.1 min), intraoperative blood loss (705.5 mL vs. 809.9 mL) and rate of positive margin of the resected specimens (6.9% vs. 23.1%) were all significantly reduced in combined hemihepatectomy group compared with perihilar hepatectomy group (all P<0.05). The results of survival analysis showed that the recurrence-free survival time and 1-year accumulate survival rate in combined hemihepatectomy group were all superior to those in perihilar hepatectomy group (both P<0.05).
Conclusion: Radical R0 resection is the only chance of cure for HCCA patients. Compared with perihilar hepatectomy, the large scope hepatectomy hemihepatectomy plus caudate lobectomy can improve the R0 resection rate, recurrence-free survival and 1-year survival rate. Accurate preoperative evaluation, appropriate perioperative treatment and selection of individualized operation plan can improve the curative effect of HCCA.

Abstract:Background and Aims: There is no agreement on embolectomy for hepatocellular cancer (HCC) with type B4 bile duct tumor thrombus (BDTT) at present. Here, the authors report the treatment procedures of a patient with HCC and type B4 BDTT, so as to provide a reference for the treatment of this condition.
Methods: The clinical data of one patient with HCC and type B4 BDTT treated in Fujian Provincial Hospital in March 2019 were reviewed. The patient underwent surgical treatment after accurate preoperative assessment. A “q-type” biliary tumor thrombectomy was performed during operation, which was namely, that a longitudinal incision from the anterior wall of the common bile duct to the anterior wall of the right hepatobiliary duct to completely expose the BDTT, and then a circular transection was made on the confluence of the right hepatic duct to the common hepatic duct to completely remove extrahepatic BDTT. The intra- and postoperative variables as well as the follow-up results were analyzed.
Results: The patient underwent right hemihepatectomy, cholecystectomy, “q-type” biliary thrombectomy, and hepatoduodenal ligament lymph node dissection. All procedures were successfully completed. The operative time was 313 min and the intraoperative blood loss was 150 mL. Postoperative pathological results showed that the right liver tumor was poorly differentiated HCC with a size of 4 cm × 3 cm× 3 cm, and the presence of the hepatic capsular invasion, and microvascular tumor thrombus, and the tumor thrombus in common bile duct was HCC. The patient recovered smoothly without complications such as bile leakage, abdominal infection or liver failure. The patient was discharged from hospital on postoperative day 7. During the 14-month follow-up, the patient had a good quality of life, with no evidence of recurrence. 
Conclusion: For HCC type B4 BDTT, “q-type” biliary thrombectomy is a safe and effective procedure and it also adheres to the tumor-free operation principle. So, it is recommended to be applied in clinical practice.

Abstract:Background and Aims: The incidence of gallbladder cholesterol stone (GCS) in Tibet and other plateau areas is several times higher than that in plain regions, which brings serious harm to the residents in the plateau areas. Previous studies have shown that the disorder of lipid metabolism is the initial factor for the formation of GCS. Flavin monooxygenase 3 (FMO3) and its metabolite trimethylamine-N-oxide (TMAO) are involved in the formation of GCS (plain region). However, the roles of FMO3 and TMAO in the pathogenesis of GCS in plateau areas is not clear. This study was conducted to observe the changes in FMO3 and TMAO levels in mice GCS model created in a plateau region and then analyze the potential clinical significance. 
Methods: Adult male C57BL/6J mice were reared in Naqu, Tibet (4 500 m above sea level) and randomly divided into GCS model group and control group. Mice in model group were fed high-fat diet (basic diet +15% fat + 1% cholesterol + 0.5% cholic acid) to induce GCS, and those in control group were given normal diet. Eight weeks later, the mice were sacrificed, and the samples were harvested (the presence of cholesterol stones in the gallbladder in model group was a standard for successful model creation, and 20 mice were used in each group). Blood lipids and bile biochemical indexes were measured by automatic biochemical analyzer, plasma TMAO level was measured by high performance liquid chromatography (HPLC), the expression of FMO3 mRNA in liver was detected by qRT-PCR. Spearman correlation coefficient was used to analyze the correlation among the observed variables, and multivariate Logistic regression analysis was used to analyze the risk factors.
Results: The serum total cholesterol (TC), triglyceride (TG), low density lipoprotein (LDL), bile TC, bile acid, phospholipid and cholesterol saturation index (CSI) in model group were significantly higher than those in the control group, while the high-density lipoprotein (HDL) in model group was significantly lower than that in the control group (all P<0.05). The plasma level of TMAO and FMO3 mRNA expression level in the liver in model group were significantly higher than those in control group (both P<0.05). Spearman correlation coefficient analysis showed that FMO3 mRNA and TMAO were positively correlated with plasma TC, TG, LDL, bile TC, bile acid, phospholipid and CSI, and negatively correlated with HDL (all P<0.05). Multivariate Logistic regression showed that the increase of FMO3 mRNA and TMAO were the risk factors for GCS in plateau areas.
Conclusion: The high FMO3 and TMAO levels are closely related to the occurrence GCS in plateau areas, and the mechanism may be probably associated with the lipid metabolism disorders caused by their increase. This provides a new strategy and direction for the prevention and treatment of GCS in plateau areas.

Abstract:Background and Aims: The underlying mechanism for the occurrence of gallbladder carcinoma (GBC) is still unclear at present. The available data at the genomic and transcriptomic levels provide the basic data source for investigation of the molecular biological mechanisms of GBC. Therefore, this study was conducted to to analyze the differentially expressed genes in and normal gallbladder tissues and key protein regulatory molecules in GBC by bioinformatics approaches, so as to explore the potential molecular biological mechanism of GBC. 
Methods: The differentially expressed genes were screened based on two GBC transcriptional datasets from GEO database, and the three GO functional annotations were performed on these genes. The STRING database was applied to construct a protein interaction network, and perform module mining in the investigation of key protein regulatory genes. Finally, the expression and predictive efficacy of the identified key protein regulatory genes were comprehensively evaluated.
Results: A total of 140 repeatable differentially expressed genes (20 up-regulated genes and 120 down-regulated genes) in GBC were screened, which are mainly related to the forebrain development and positive regulation of neurogenesis, and participate in the composition of the postsynaptic membrane and transverse tubules. Meanwhile, the SFRP1, a key protein regulatory molecule, had a certain ability in predicting the occurrence of GBC.
Conclusion: The information expressed by transcription spectrum of GBC obtained in this study can provide framework and thinking structure for studying the molecular mechanism of GBC. The key protein regulatory molecule SFRP1 probably plays a pivotal role in the occurrence and development of GBC. 

Abstract:Background and Aims: Hepatocellular carcinoma (HCC) is one of the most common malignant tumors in clinical practice. Invasion, metastasis and postoperative recurrence are the main causes of death for HCC patients. At present, the dysregulation of long non-coding RNAs (lncRNA) is considered to be related to the occurrence and metastasis of various cancers. Studies have showed that lncRNA SNHG12 expression is increased in HCC tissue, but its specific function is still unclear. Therefore, this study was conducted to investigate the action of lncRNA SNHG12 in HCC cells and the mechanism. 
Methods: The gene expressions of SNHG12 as well as the predicted targeted miRNA and gene (miR-199a-5p and FZD6) in HCC cell lines HepG2 and human intrahepatic bile duct cell line HIBEC were detected by qRT-PCR method. The effects of down-regulation of SNHG12 on the proliferation, invasion and the expressions of proteins associated with epithelial-mesenchymal transition (EMT) in HepG2 cells were observed. The association between SNHG12 and miR-199a-5p was analyzed by miRanda and Dual luciferase reporter gene assay, and then, the effects of down-regulation of miR-199a-5p on the proliferation, invasion and EMT in HepG2 cells as well as the influences of SNHG12 exerted by miR-199a-5p were analyzed. The relationship between miR-199a-5p and FZD6 was analyzed by the TargetScan and Dual luciferase reporter gene assay, and then, the effects of up-regulation and down-regulation of FZD6 on the proliferation, invasion and EMT in HepG2 cells as well as the effects of SNHG12 and miR-199a-5p on FZD6 expression were detected.
Results: The expression level of SNHG12 was significantly increased, miR-199a-5p was significantly decreased and FZD6 was significantly increased in HepG2 cells compared with those in HIBEC cells (all P<0.01). In HepG2 cells, the proliferation, invasion and EMT were significantly inhibited after down-regulation of SNHG12, while its overexpression resulted in opposite effects (all P<0.05). The 3'UTR of SNHG12 was targeted by miR-199a-5p. The proliferation, invasion and EMT of HepG2 cells were augmented by down-regulation of miR-199a-5p, and were suppressed by its overexpression (all P<0.05). The effects of miR-199a-5p overexpression on HepG2 cells were partially reversed by the simultaneous overexpression of SNHG12 (all P<0.05). The miR-199a-5p were targeted by FZD6. The proliferation, invasion and EMT of HepG2 cells were inhibited by down-regulation of FZD6, while were increased by up-regulation of FZD6 (all P<0.05). Down-regulation of SNHG12 inhibited FZD6 gene and protein expressions, while down-regulation of miR-199a-5p promoted FZD6 expression (all P<0.01). Compared with down-regulation of miR-199a-5p alone, the FZD6 expression was significantly decreased by both SNHG12 and miR-199a-5p down-regulations (P<0.01).
Conclusion: Upregulation of lncRNA SNHG12 in HCC cells can promote the proliferation, invasion and EMT process of HCC cells through influencing the miR-199a-5p/FZD6 axis.

Abstract:Background and Aims: The tumor suppressor gene HIV-1 Tat interactive protein 2 (TIP30) is down-regulated or absent in many human tumors, and is related to the poor prognosis of patients. However, there is no report about TIP30 in intrahepatic cholangiocarcinoma (ICC). Therefore, this study was performed to investigate the expression of TIP30 in ICC and its clinical significance. 
Methods: The TIP30 expressions in 51 paired specimens of ICC tissue and tumor adjacent tissue were detected by immunohistochemical staining. The relations of TIP30 expression in ICC tissues with the clinicopathologic features as well as the overall survival rate and cumulative recurrence rate of ICC patients were analyzed. The change in colony-forming ability in ICC cells (ICC-9810, SSP-25 and HuH-28) after transfection with TIP30-shRNA were determined by soft agar colony formation assay. The potential miRNAs targeting the 3’UTR of TIP30 were predicted using TargetScan and miRDB databases, and then the results were further verified by qRT-PCR.
Results: The results of immunohistochemical staining demonstrated that the expression level of TIP30 in ICC tissue was lower than that in adjacent tissue. The results of clinical data analysis indicated that the TIP30 expression was significantly associated with lymph node metastasis (P=0.008). The survival analysis showed that the 5-year overall survival rate was significantly higher (P=0.041 2), while the cumulative recurrence rate was significantly lower (P=0.037 4) in patients with high TIP30 expression than those in patients with low TIP30 expression. The number of colony-forming units in ICC cells (ICC-9810, SSP-25 and HuH-28) transfected with TIP30-shRNA were significantly increased over the negative controls (all P<0.001). The results of database prediction suggested that miR-124-3p could recognize and bind to the 3' UTR of TIP30. The results of qRT-PCR verification revealed that the relative content of miR-124-3p in ICC tissue was significantly higher than that in adjacent tissue (P<0.001); the expressions of TIP30 in ICC cells (9810, SSP-25 and HuH-28) transfected with miR-124-3p were lower than those in their negative controls (all P<0.05).
Conclusion: The TIP30 expression is down-regulated in ICC, and is closely related to the poor prognosis of ICC patients. The mechanism may be associated with the regulation mediated by miR-124-3p. The results of this study may provide information and basis for future studies on the pathogenesis and molecular target therapy of ICC.

Abstract:Background and Aims: Surgical resection is the preferred treatment for hepatocellular carcinoma (HCC) at present. However, the use of anatomic resection (AR) or non-anatomic resection (NAR) is still controversial. This study was conducted to evaluate the efficacy and safety of using AR and NAR in the treatment of patients with HCC by Meta-analysis, so as to obtain evidence-based information resources for clinical practice.  
Methods: The studies comparing the clinical efficacy of AR versus NAR for HCC were collected by a computer-based search in several national and international databases from their inception to September 2020. After literature screening, data extraction and bias risk evaluation by two reviewers independently, Meta-analysis was performed using RevMan 5.3 software.
Results: A total of 33 studies were finally included, comprising 1 randomized controlled trial (RCT) and 32 cohort studies, involving 6 132 patients, with 3 029 cases in AR group and 3 103 cases in NAR group. The results of Meta-analysis showed that in AR group compared with NAR group, the postoperative early recurrence rate (OR=0.67, 95% CI=0.49–0.92, P=0.01) and intrahepatic local recurrence rate (OR=0.35, 95% CI=0.20–0.59, P=0.000 1) were reduced, and the 1-, 2-, 3-, and 5-year disease-free survival rates (DFS) (OR=1.26, 95% CI=1.12–1.42, P=0.000 1; OR=1.31, 95% CI=1.17–1.46, P<0.000 01; OR=1.32, 95% CI=1.19–1.48, P<0.000 01; OR=1.40, 95% CI=1.23–1.58, P<0.000 01), as well as the 5-year overall survival rate (OS) (OR=1.16, 95% CI=1.03–1.31, P=0.02) were improved. However, there were no statistical difference in intrahepatic distant recurrence rate (OR=1.11, 95% CI=0.80–1.54, P=0.54), extrahepatic metastasis rate (OR=0.97, 95% CI=0.78–1.22, P=0.83), 1-, 2-, and 3-year OS (OR=1.01, 95% CI=0.85–1.22, P=0.88; OR=1.15, 95% CI=0.99–1.33, P=0.06; OR=1.13, 95% CI=1.00–1.28, P=0.06) as well as the incidence of postoperative complications (OR=0.98, 95% CI=0.81–1.17, P=0.79) between the two groups. The subgroup analysis showed that the 1-, 2-, 3-, and 5-year DFS (OR=1.36, 95% CI=1.07–1.73, P=0.01; OR=1.55, 95% CI=1.23–1.95, P=0.000 2; OR=1.78, 95% CI=1.38–2.30, P<0.000 01; OR=2.07, 95% CI=1.46–2.94, P<0.000 1) as well as the 2-, 3-, and 5-year OS (OR=1.54, 95% CI=1.07–2.21, P=0.02; OR=1.46, 95% CI=1.11–1.92, P=0.007; OR=1.52, 95% CI=1.13–2.03, P=0.005) for HCC combined with microvascular invasion (MVI) in AR group were higher than those in NAR group.
Conclusion: The current evidence indicates that AR may reduce the incidence of postoperative early recurrence and intrahepatic local recurrence, and improve the DFS and long-term OS compared with NAR. In particular, for patients combined with MVI, AR is significantly better than NAR in terms of short- and long-term efficacy, which should be recommended for clinical application. Limited by the quality and quantity of the included studies, the above conclusions need to be verified by more multicenter randomized controlled trials with large sample size.

Abstract:Background and Aims: Pancreatic cancer is characterized by features such as low early detection rate, high distant metastasis risk, and poor chemotherapy response. The majority of the patients with pancreatic cancer may face a dismal prognosis. So, development of early diagnostic tools for pancreatic cancer is of great importance. This study was conducted to serum miRNAs that can be used for differential detection among samples and construct a decision tree model using bioinformatics approaches and machine learning method. 
Methods: Serum miRNA expressions datasets of GSE113486 and GSE85589 were downloaded from the Gene Expression Omnibus (GEO) database. Batch effect removal was performed using the ComBat function. The hub miRNAs for distinguishing the tumor from normal serum samples were screened by LASSO regression analyses. Decision-tree model was constructed based on the hub miRNAs by rpart software package. The diagnostic efficacy of the decision tree was evaluated with ROC curves. The expression differences of the variables of the decision tree between the tumor and normal serum samples were compared by Wilcoxon test. Finally, the target mRNAs of the hub miRNAs were predicted using miRDB, miRTarBsae and TargetScan databases, and enrichment analysis was also performed.
Results: After removing the batch effects, the serum miRNA expression profiles of 119 healthy controls and 128 pancreatic cancer samples were included in the study. Using LASSO regression with 10-fold cross validation, 33 miRNAs were screened. Next, all samples were randomly divided into a training set (60%) and a validation set (40%). In training set, 33 hub miRNAs were used for construction of decision tree model. After pruning, miR-4532 and miR-4668-5p were selected as observation variables of the decision tree. The results of ROC curve evaluation showed the area under the curve (AUC) was 0.948 1 in training set and was 0.902 4 in validation set. In addition, miR-4532 and miR-4668-5p were significantly overexpressed in the pancreatic cancer serum samples compared to the normal serum samples. Six target mRNAs were predicted for miR-4532, and 73 target mRNAs predicted for miR-4668-5p. They were mainly associated with the functions such as transcription regulator complex, nuclear chromatin, transcription repressor complex, cell-subtrate junction organization, regulation of megakaryocyte differentiation, and negative regulation of focal adhesion assembly, and mainly enriched in the pathways involving the transcriptional misregulation in cancer, FoxO, adherens junction, pancreatic cancer, hepatitis B, hepatocellular carcinoma, TGF-β and MAPK.
Conclusion: The decision tree constructed by miR-4532 and miR-4668-5p has a good efficacy in distinguishing normal and pancreatic cancer serum samples, and it has certain value in early detection of pancreatic cancer.

Abstract:Common bile duct stones often lead to acute biliary obstruction, cholangitis and pancreatitis. Endoscopic stone extraction has become the main treatment for bile duct stones. Most of the common bile duct stones can be removed by the conventional endoscopic method, but there are still 10% to 15% of the complex choledocholithiasis that need the combined treatment of endoscopic lithotripsy. The authors review and summarize the current endoscopic management options for complex choledocholithiasis, such as endoscopic balloon papillary dilatation, endoscopic guided lithotripsy and endoscopic ultrasound-guided choledocholithotomy, so as to provide endoscopic diagnosis and treatment suggestions and basis for the clinical treatment of refractory choledocholithiasis.   

Abstract:Stone disease accounts for the highest proportion among the disorders of the biliary system in our country. It is characterized by complex pathologies, high relapse rates, low cure rates and multiple complications. In its advanced stage, the end-stage biliary disease such as biliary cirrhosis, portal hypertension and intrahepatic cholangiocarcinoma may occur, so it has long been an intractable problem in the field of hepatobiliary surgery. In recent years, there are increasing reports of using various minimally invasive procedures in the treatment of cholelithiasis, and the development of minimally invasive techniques represented by laparoscopy and endoscopy has opened up a new era for the treatment of this condition. Here, the authors present a review of the systemaltization and progress of minimally invasive management of cholelithiasis guided by the “three-approach” theory in the new period based on the experience of our own team and related literature.