Objective To explore the effect of gamma-aminobutyric acid (GABA) on the growth, apoptosis and telomerase activity of Cholangiocarcinoma cell line QBC939. Methods Cholangiocarcinoma cell line QBC939 was cultured by routine method, and then treated with different concentrations of GABA (1-1000 μmmol/L). The proliferation, apoptosis and cell cycle of QBC939 cells was investigated by MTT, Flow cytometry and transmission electron microscopy. The telomer-ase activity of QBC939 cells was examined by modified PCR-ELISA assay. Radioimmunoassay was used to measure the intracellular cyclic adenosine monophosphate(cAMP) content. Results The dif-ferent concentrations of GABA inhibited the growth of QBC939 cells and promoted the apoptosis. The apop-totic rate of QBC939 cells was increased from 4. 8% to 28. 03%, which had significant difference (P<0.05). It had no effect for distribution of cell cycle. Cell nuclear condensation and apoptotic bodies were seen by transmission electron microscopy. Telomerase activity was inhibited by GABA(0. 82±0. 048 vs 0.56±0. 054, P<0.05). The content of intracellular cAMP was increased with the increase of GABA concentration in a dose-dependent manner [(0. 59±0. 049) nmol/L vs (0. 82±0. 033)nmol/L, P<0. 05]. Conclusion GABA can inhibit the proliferation of QBC939 cells by promoting apoptosis and inhibiting telomerase activi-ty, which may be mediated by the information transmission of post-receptor.

Objective To explore the effect of GABA on the bionomics of cholangiocarcinoma cell QBC939. Methods The proliferation of QBC939 cells was investigated by MTT. The telomerase activity of QBC939 ceils was examined by modified PCR-ELISA assay. Transwell cell culture chamber assay in vitro was used to detect the ability of invasion of cholangiocarcinoma QBC939. The expression of matrix metalloproteinase (MMP) mRNA and enzymatic activity of QBC939 cells were detected by reverse transcription polymerase chain reaction(RT-PCR) and gelatin zymography, respectively. Results GABA at all tested concentration inhibited the growth and proliferation of QBC939 cells in a time-independent manner, Telomerase activity was inhibited by GABA [(0.82 ± 0.05) vs. (0.56 ± 0.05), (P<0.05)]. GABA inhibited tumor cells' trans-Matrigel ability (at 100 μmol/L GABA, cell's number decreased from 60 ± 10 to 43 ±4, P<0.05) ,and significantly inhibited the activity and expression of MMP(MMP-2、MMP- 9) in a dose dependent way. Conclusions GABA can inhibit the growth、invasion and metastasis of cholangiocarcinoma QBC939. The down-regulation of the activity of Telomerase and MMP may be involved inthis mechanism.

Objective To explore the effects of gamma-aminobutyric acid (GABA) on the proliferation, apoptosis and invasion of cholangiocarcinoma cell line QBC_(939). Methods Cholangiocarcinoma cell line QBC_(939) was cultured via routine methods, and was treated with different concentrations of GABA (1, 10,100, 1000 μmol/L, test group) or with complete culture medium (control group). The effects of GABA on the proliferation, apoptosis and invasion of cholangiocarcinoma cell line QBC_(939) were investigated by MTT assay, flow cytometry and Transwell cell culture chamber assay, respectively. The effects of GABA on the activity of matrix metalloproteinase (MMP) secreted by cholangiocarcinoma cells were analyzed by gelatin zymogram, and the content of intracellular cyclic adenosine monophosphate (cAMP) was measured by radioimmunoassay. All data were analyzed by one-way ANOVA. Results As the GABA concentration increased from 1 μmol/L to 1000 μmol/L, the inhibition rate of GABA on cholangiocarcinoma cell line QBC_(939) was increased from 2.6% to 26.8%, and the apoptosis rate also increased from 4.80%±0.04% to 28.03% ±0.01%. The number of cholangiocarcinoma cells that migrated through the Matrigel gel decreased from 60±10 to 43±4, in a dose-dependent manner (F = 7. 883, 83.765, 7. 598, P <0.05). The activity of MMP-2 and MMP-9 was decreased. The content of intracellular cAMP was increased, and there was significant difference in the content of intracellular cAMP between test and control groups (F =9.507, 9.148, 27.418, P < 0.05). Conclusions GABA can inhibit the proliferation and invasion of cholangiocarcinoma cell line QBC_(939) cells by promoting the apoptosis and inhibiting the activity of MMP-2 and MMP-9. The process may be mediated by the information transmission of post-receptor.

Objective To investigate the expressions of microRNA-21 in cholangiocarcinoma tissues and the relationship between epithelial-mesenchymal transition (EMT) and the prognosis of patients.Methods Forty-one samples of cholangiocarcinoma and 10 samples of adjacent tissues from 10 patients who received radical resection of cholangiocarcinoma at the Provincial Hospital of Anhui Medical University from January 2005 to January 2010 were collected.The expressions of microRNA-21,E-cadherin and N-cadherin were detected by in situ hybridization and immunohistochemistry,and effect of their expressions on the prognosis was analyzed.Enumeration data were analyzed using chi-square test.The correlation between microRNA-21 and EMT markers was analyzed using the Spearman correlation coefficient.The survival curve was drawn by Kaplan-Meier method,and the survival rate was analyzed using the Log-rank test.Results The expression rate of microRNA-21 in the cholangiocarcinoma tissues was 63％,which was significantly higher than 30％ of that in the adjacent tissues (x2 =0.324,P ＜ 0.05).The expression of microRNA-21 was closely related with the tumor differentiation degree,lymph node metastasis,perineural invasion (x2 =6.365,0.552,11.896,P ＜ 0.05),but not with gender,age,tunor location and tumor type (x2 =0.322,0.588,0.510,0.256,P ＞ 0.05).The expressions of E-cadherin and N-cadherin were related with lymph node metastasis and perineural invasion (x2 =4.630,5.512;6.600,7.152,P ＜0.05),but not with gender,age,tumor location,tumor differentiation degree and tumor type (x2 =0.266,0.013,0.067,0.666,0.003; 1.036,0.997,1.808,2.997,0.812,P ＞0.05).A positive correlation between the expression of microRNA-21 and EMT related markers E-cadherin and N-cadherin was detected (r =0.373,0.614,P ＜0.05).The results of survival analysis showed that the overall survival rate and tumor-free survival rate of patients with low expression of microRNA-21 were significantly higher than those of high expression of microRNA-21 (x2 =3.999,4.376,P ＜ 0.05).Conclusion Over expression of microRNA-21 in cholangiocarcinoma and metastatic lymph nodes may accelerate the invasion and metastasis of cholangiocarcinoma through inducing EMT,microRNA-21 might predict the prognosis of patients.

Objective To systematically evaluate the safety and effectiveness of combined portal vein resection and reconstruction in the resection of hilar cholangiocarcinoma.Methods Literatures were researched using Cochrane Library,PubMed,Embase,China Biology Medicine disc,China National Knowledge Infrastructure,Wanfang database,VIP database from January 31,2006 to January 31,2016 with the key words including “hilar cholangiocarcinoma”“Klatskin tumor”“Bile duct neoplasm”“Vascular resection”“portal vein resection”“肝门部胆管癌”“血管切除”“门静脉切除”.The clinical studies of resection of hilar cholangiocarcinoma with portal vein resection and construction and without vascular resection and construction were received and enrolled.Two reviewers independently screened literatures,extracted data and assessed the risk of bias.Patients who underwent resection of hilar cholangiocarcinoma combined with portal vein resection and reconstruction were allocated into the portal vein resection group and patients who didn't undergo vascular resection were allocated into the no vascular resection group.Analysis indicators included (1) results of literature retrieval;(2) results of Meta-analysis:① incidence of postoperative complications (hepatic failure,biliary fistula,intra-abdominal hemorrhage),② postoperative mortality,③ patients' prognosis,④ related indicators of postoperative pathology (lymph node metastasis rate,moderate-and low-differentiated rate,nerve invasion rate,negative rate of resection margin).The heterogeneity of the studies was analyzed using the I2 test.The hazard ratio (HR) and 95％ confidence interval (CI) were used for assessing the prognostic indicators.The incidence of complications,mortality and pathological indicators were evaluated by the odds ratio (OR) and 95％ CI.Results (1) Results of literature retrieval:13 retrospective studies were eurolled in the meta-analysis,and the total sample size was 1 668 cases including 437 in the portal vein resection group and 1 231 in the no vascular resection group.(2) Results of Meta-analysis:① incidence of postoperative complications was respectively 39.86％ in the portal vein resection group and 35.27％ in the no vascular resection group,with no statistically significant difference between the 2 groups (OR =1.12,95％ CI:0.82-1.53,P ＞0.05).The results of subgroup analysis showed that hepatic failure,biliary fistula and intra-abdominal hemorrhage were postoperative main complications,and the incidences were 17.09％,8.79％,6.25％ in the portal vein resection group and 10.62％,9.69％,2.51％ in the no vascular resection group,respectively,with no statistically significant difference between the 2 groups (OR =0.48,1.13,0.82,95％ CI:0.23-1.02,0.45-2.83,0.21-3.12,P ＞ 0.05).② Postoperative mortality was respectively 5.38％ in the portal vein resection group and 3.88％ in the no vascular resection group,with no statistically significant difference between the 2 groups (OR =1.16,95％ CI:0.62-2.14,P ＞ 0.05).③ There was statistically significant difference in patients' prognosis between the 2 groups (HR =1.81,95％ CI:1.52-2.16,P ＜ 0.05).④ The related indicators of postoperative pathology:lymph node metastasis rate,moderate-and low-differentiated rate and negative rate of resection margin were 41.55％,76.42％,63.74％ in the portal vein resection group and 33.42％,66.75％,64.29％ in the no vascular resection group,respectively,with no statistically significant difference between the 2 groups (OR =1.45,1.59,0.67,95％ CI:0.95-2.21,0.97-2.61,0.37-1.20,P ＞ 0.05).The nerve invasion rate was 83.47％ in the portal vein resection group and 64.90％ in the no vascular resection group,with a statistically significant difference between the 2 groups (OR =2.61,95 ％ CI:1.45-4.70,P ＜ 0.05).Conclusion Combined portal vein resection and reconstruction is safe and feasible in the treatment of hilar cholangiocarcinoma,and the prognosis of patients with portal vein invasion is worse than that without portal vein invasion.

Objective To compare the safety and effectiveness of duodenum-preserving pancreatic head resection (DPPHR) with pylorus-preserving pancreaticoduodenectomy (PPPD) in the treatment of chronic pancreatitis with a pancreatic head mass.Methods Medline,Biosis,Cochrane Library,Science Citation Index Database,CBM Database,Wan Fang and CNKI were searched systematically.The bias risk of the included trials was assessed according to the assessing tools as suggested by the Cochrane Handbook.Review Manage 5.2 was used to perform the statistical analysis.Results 7 RCTs with 226 patients were included in the meta-analysis which showed that there were no significant differences between PPPD and DPPHR in overall postoperative morbidity,postoperative hospital stay,complete pain relief,pancreatic fistula,exocrine insufficiency,symptom score at 5 to 7-year follow-up,and quality of life score at 14 to 15-year follow-up (P ＞ 0.05).While DPPHR had significant superiorities in operation time,blood replacement,delayed gastric emptying,occupational rehabilitation after the operations,weight gain,quality of life score at 1 to 2-year follow-up,symptom score at 5 to 7-year follow-up,and physical functioning score at 14 to 15-year follow-up.Conclusions DPPHR is more favourable than PPPD in reducing the use of blood replacement,shortening operation time,delayed gastric emptying,occupational rehabilitation after the operations,weight gain,physical functioning,and in improving quality of life of patients.

Objective To investigate the expression of STAT3 signaling pathway genes including Survivin and COX-2 in cholangiocarcinoma,as well as the relationship between expression of these genes and prognosis of patients with cholangiocarcinoma.Methods The tumor and normal tissue samples were respectively collected from 43 patients with cholangiocarcinoma and 12 patients with intra-and extrahepatic bile duct stones or hepatic duct injury in the Affiliated Provincial Hospital of Anhui Medical University from September 2007 to July 2012.The expression of STAT3,phosphorylated-STAT3 (p-STAT3),Survivin and COX-2 were examined using immunohistochemistry,and the relationship between the expression of these genes and the clinical pathological features and prognosis of patients with cholangiocarcinoma was analyzed.Patients were followed-up through outpatient examination and telephone interview until March 2014.Categorical data were analyzed using the chi-square test.Correlation analysis was done by Spearman's method.The survival curve was generated using the Kaplan-Meier method,and the survival analysis was conducted using the log-rank test.Results The positive expression rates of STAT3,p-STAT3,Survivin and COX-2 in the tumor samples were 69.8％ (30/43),65.1％ (28/43),72.1％ (31/43),79.1％ (34/43),respectively,which were compared with 41.7％ (5/12),8.3％ (1/12),16.7％ (2/12) and 41.7％ (5/12) in the normal tissue samples,showing a significant difference for the last 3 indexes (x2=12.136,9.811,4.679,P ＜ 0.05).Overexpression of p-STAT3,Survivin and COX-2 protein was correlated with lymph node metastasis (x2 =14.700,5.959,4.075,P ＜ 0.05).Overexpression of p-STAT3 was also related to neural invasion (x2=10.384,P ＜ 0.05).Expression of Survival and COX-2 protein was not associated with lymph invasion (x2=2.718,3.024,P ＞ 0.05).Expression of p-STAT3,Survivin and COX-2 was however not associated with gender,age and tumor location,differentiation and diameter (x2=0.148,0.720,1.835,1.040,0.236 ; 0.001,0.009,0.029,1.863,0.197 ; 0.433,0.686,0.002,2.974,0.029,P ＞ 0.05).Expression of Survivin and COX-2 protein was positively correlated to p-STAT3 protein (r =0.524,0.583,P ＜ 0.05).All the 43 patients were followed up for 6-60 months.Among the 17 patients with hilar cholangiocarcinoma,the median survival time was 7,9,9 months for patients with positive expression of p-STAT3,Survivin and COX-2 protein,compared with 18,11 and 11 months for patients with negative expression of these proteins.The survival rates of the patients with positive and negative expression of p-STAT3 protein were 33.3％ and 68.6％,respectively,with a statistical significance for p-STAT3 protein (x2=12.916,P ＜ 0.05).Of the remaining 26 patients with common bile duct carcinoma,the median survival time was 9,10 and 9 months for patients with positive expression of p-STAT3,Survivin and COX-2 protein,compared with 20,20 and 20 months for patients with negative expression of these proteins.The survival rates of the patients with positive expression of p-STAT3,Survivin and COX-2 protein were 20.8％,9.4％ and 8.5％,which were lower than 37.5％,37.5％ and 50.0％ of patients with the negative expression of these proteins,with a statically significance for all the 3 proteins (x2=12.787,6.245,11.161,P ＜ 0.05).Conclusions The p-STAT3,Survivin,COX-2 proteins are highly expressed in the cholangiocarcinoma and the expression levels of these proteins are positively correlated.The survivin and COX-2 may be the downstream genes of STAT signaling pathway,which are involved in the progression and prognosis of cholangiocarcinoma.

ObjectiveTo discuss the effects of apoptosis and invasion of RBE cells caused by miRNA 21 suppression and further investigate the potential role of miRNA-21 plays on target mRNA regulation. MethodsThe RNAi technology was employed to suppress the expression of RBE cells.The changes in RECK mRNA and protein expressions were detected by RT-PCR and Western blotting respectively. Changes occurred in apoptosis was closely monitored by flow cytometry (FCM). The invasion of RBE cells was analyzed in vitro by invasion assay (transwell). ResultsThe expression of miRNA-21 was clearly suppressed while the RECK mRNA and protein were over-expressed. The rate of apoptosis was significantly accelerated and there was a dramatic decrease in RBE cells' ability to invade after miRNA-21 knockdown. ConclusionThrough miRNA-21 suppression, the rate of apoptosis of RBE cells was accelerated whereas their invasion ability was greatly reduced. RECK was found to be the target gene of miRNA-21 which participates in the regulation process of regulation.

Objective To investigate the short and long-term outcomes and risk factors after open surgery or and endoscopic hepatolithotomy for hepatolithiasis. Methods A retrospective study was made on cases of hepatolithiasis who underwent hepatolithotomy by open surgery or endoscopically from Jan 2001 to Dec 2008.Of 254 patients,189 were followed-up including 127 after open surgery and 62 endoscopically.The univariate and multivariate analyses were performed to determine the risk factors. Results Complete stone clearance was achieved in 85.0 ％ ( 108/127 ) of open surgery including hepatecomy,61.3 ％ ( 38/62 )of endoscopic treatment.After a median follow-up period of 6.0 years (2.5 to 10.5 years),stone recurred in 32.8％ (62/189) of patients,biliary cirrhosis in 7.4％ ( 14/189),cholangiocarcinoma in 7(3.7％ ),all cancer cases were dead with a mortality rate of 7(3.7％ ).Bile duct stricture (OR:7.522,95％ CI:2.642 -21.415),stones in both lobes (OR:11.630,95％ CI:3.989 -33.912),and endoscopic treatment ( OR:21.374,95％ CI:6.713 - 68.056 ) were independent risk factors ( P ＜ 0.05 ) for incomplete stone clearance by unconditional Logistic regression analysis.In addition,recurrent stones and/or cholangitis were associated with residual stones ( OR:3.059,95％ CI:1.307 - 7.159),stricture ( OR:3.702,95％ CI:1.567-8.745) and endoscopic therapy (OR:4.841,95％ CI:1.946 - 12.043) (P ＜ 0.05).Conclusions Stricture,stone in both lobes and endoscope therapy were independent risk factors for residual stones; Residual stones,bile duct stricture and endoscope therapy were independent risk factors for recurrent stones and/or cholangitis.

Objective To explore the methods of diagnosis and appropriate treatment of pancreatic duct stone.Methods Clinical data of 16 patients with pancreatic duct stone from March,2005 to August,2009 were analyzed retrospectively.Results 15 patients presented with varying degrees of upper abdominal pain,another one with irregular diarrhea.Serum and urine amylase level was higher than the upper limit of normal level in 3 patients;serum glucose was elevated in 4 patients.The diagnostic accuracy by Bultrasonography,CT,MRI,ERCP and KUB was 93.8%(15/16),68.8% (11/16),57.1% (4/7);100%(2/2) and 50% (3/6),respectively.2 cases underwent endoscopic pancreatic sphincterotomy + pancreatic stent drainage,14 cases were treated with surgery,including transpancreatic duct lithotomy + pancreatic jejunal anastomosis in 12 cases,pancreatoduodenectomy in 1 ease,and pancreatic body and tail resection +pancreatic jejunal Roux-en-Y anastomosis in one case.All operations were successful without mortality,and abdominal pain was significantly improved.Follow up of 14 cases showed no stone recurrence.Two patients were lost in follow up,so the follow up rate was 87.5% with the duration ranging from 1 to 53 months.Conclusions B-ultrasonography was the best imaging examination for pancreatic duet stone,but the combined application of imaging tests could significantly improve the diagnostic yield,and imaging examination provided an important basis for the choice of treatment method.With the improvement of endoscopic techniques,ERCP will be as important as surgery for the treatment of pancreatic duct stone.