[Typing and surgical treatment choice for pancreatic ductal stone].

OBJECTIVE: To explore the improvement of typing and reasonable surgical treatment for pancreatic ductal stone (PDS). METHODS: Totally 89 patients with pancreatic ductul stone treated underwent surgeries from January 2000 to December 2012 were involved into this study. There were 57 male and 32 female patients, the average age was (52 ± 23) years. According to the magnetic resonance cholangiopancreatography imaging and finding during surgery, pancreatolithiasis was classified into three types: type I, the stones were located in the main pancreatic duct; type II, the stones were located both in main and branch pancreatic duct; type III, the stones were diffusely scattered in the branch pancreatic duct; the position of PDS within pancreatic parenchyma were subtitled. In this group, 43 type I PDS were extracted with endoscopic papillotomy or endoscopic pancreatic sphincterotomy, or pancreatolithotomy plus pancreato-jejunal lateral anastomosis with wide anastomotic stoma; 39 type II cases were treated by pancreatolithotomy plus pancreato-jejunal lateral anastomosis or/and resection of pancreatic section; 7 type III PDS were managed with resection of pancreatic section. RESULTS: All surgeries were performed successfully. Among complications, 6 cases (6.7%) were pancreatic leakage which recovered after systematic non-surgical treatment, 2 cases (2.2%) were anastomotic bleeding which led to 1 death, 6 cases (6.7%) were residual pancreatolithiasis in branch pancreatic duct type. Seventy-eight patients were followed up for 6 to 131 months, 57 cases were still alive so far. Five cases were intermittent abdominal pain, 7 cases were diabetes resulted from 2 subtotal pancreatectomy and 5 distal pancreatectomy, 5 cases occurred pancreatolithiasis recurrence and 3 underwent secondary surgeries. CONCLUSIONS: The basis of this modified typing of pancreatolithiasis is the position of stone in pancreatic duct rather than pancreas parenchyma. It is more important and valuable for surgical principle of taking stones out completely and maintaining pancreatic function.

Efficacy of gemcitabine plus platinum agents for biliary tract cancers: a meta-analysis.

The objective of this study was to carry out a meta-analysis of the efficacy of gemcitabine+platinum agent regimens in the treatment of advanced biliary tract cancer (BTC). PubMed and Google Scholar were searched using the following combination of search terms: gemcitabine, oxaliplatin, cholangiocarcinoma, biliary, gallbladder, bile duct. Studies were eligible for inclusion in the meta-analysis if they were randomized trials on the use of gemcitabine plus a platinum agent for the treatment of advanced (unresectable or metastatic cancer) BTC. Outcomes of interest were response rate, overall survival, and progression-free survival. Pooled odds ratios/differences in median survival and 95% confidence intervals (CIs) were determined for each outcome. A total of 47 records were identified in the initial search. Ultimately, three open-label randomized trials (two phase 2 and one phase 3) met the eligibility criteria and were included in the meta-analysis. Two studies compared gemcitabine plus cisplatin with gemcitabine alone, whereas the other study compared gemcitabine plus oxaliplatin with fluorouracil-folinic acid. The total number of patients in the studies ranged from 54 to 410. The overall analyses revealed that all survival outcomes assessed were significantly more favorable for patients treated with gemcitabine plus platinum agents than for patients not treated with this combination. Response rates: odds ratio=2.639, 95% CI=1.210-5.757, Z=2.439, P=0.015; pooled difference in median overall survival=3.822 months, 95% CI=1.798-5.845 months, Z=3.702, P<0.001; pooled difference in median progression-free survival=3.268 months, 95% CI=1.996-4.541 months, Z=5.035, P<0.001. Patients with advanced BTC who are treated with gemcitabine plus platinum agents may experience better survival outcomes compared with patients who are not treated with this combination of chemotherapy.

Emodin regulating excision repair cross-complementation group 1 through fibroblast growth factor receptor 2 signaling.

AIM: To investigate the molecular mechanisms underlying the reversal effect of emodin on platinum resistance in hepatocellular carcinoma. METHODS: After the addition of 10 µmol/L emodin to HepG2/oxaliplatin (OXA) cells, the inhibition rate (IR), 50% inhibitory concentration (IC50) and reversal index (IC50 in experimental group/IC50 in control group) were calculated. For HepG2, HepG2/OXA, HepG2/OXA/T, each cell line was divided into a control group, OXA group, OXA + fibroblast growth factor 7 (FGF7) group and OXA + emodin group, and the final concentrations of FGF7, emodin and OXA in each group were 5 ng/mL, 10 µg/mL and 10 µmol/L, respectively. Single-cell gel electrophoresis was conducted to detect DNA damage, and the fibroblast growth factor receptor 2 (FGFR2), phosphorylated extracellular signal-regulated kinase 1/2 (p-ERK1/2) and excision repair cross-complementing gene 1 (ERCC1) protein expression levels in each group were examined by Western blotting. RESULTS: Compared with the IC50 of 120.78 µmol/L in HepG2/OXA cells, the IC50 decreased to 39.65 µmol/L after treatment with 10 µmol/L emodin; thus, the reversal index was 3.05. Compared with the control group, the tail length and Olive tail length in the OXA group, OXA + FGF7 group and OXA + emodin group were significantly increased, and the differences were statistically significant (P < 0.01). The tail length and Olive tail length were lower in the OXA + FGF7 group than in the OXA group, and this difference was also statistically significant. Compared with the OXA + FGF7 group, the tail extent, the Olive tail moment and the percentage of tail DNA were significantly increased in the OXA + emodin group, and these differences were statistically significant (P < 0.01). In comparison with its parental cell line HepG2, the HepG2/OXA cells demonstrated significantly increased FGFR2, p-ERK1/2 and ERCC1 expression levels, whereas the expression of all three molecules was significantly inhibited in HepG2/OXA/T cells, in which FGFR2 was silenced by FGFR2 shRNA. In the examined HepG2 cells, the FGFR2, p-ERK1/2 and ERCC1 expression levels demonstrated increasing trends in the OXA group and OXA + FGF7 group. Compared with the OXA group and OXA + FGF7 group, the FGFR2, p-ERK1/2, and ERCC1 expression levels were significantly lower in the OXA + emodin group, and these differences were statistically significant. In the HepG2/OXA/T cell line that was transfected with FGFR2 shRNA, the FGFR2, p-ERK1/2 and ERCC1 expression levels were significantly inhibited, but there were no significant differences in these expression levels among the OXA, OXA + FGF7 and OXA + emodin groups. CONCLUSION: Emodin markedly reversed OXA resistance by enhancing OXA DNA damage in HepG2/OXA cells, and the molecular mechanism was related to the inhibitory effect on ERCC1 expression being mediated by the FGFR2/ERK1/2 signaling pathway.

Mutual regulation between microRNA-373 and methyl-CpG-binding domain protein 2 in hilar cholangiocarcinoma.

AIM: To investigate the reciprocal modulation between microRNA (miRNA) and DNA methylation via exploring the correlation between miR-373 and methyl-CpG-binding domain protein (MBD)2. METHODS: MiR-373 expression was examined using the TaqMan miRNA assay. Methylation of miR-373 was investigated using methylation-specific polymerase chain reaction, and recruitment of methyl binding proteins was studied using the chromatin immunoprecipitation assay. Mutation analysis was conducted using the QuikChange™ Site-Directed Mutagenesis kit. The activity of miR-373 gene promoter constructs and targeting at MBD2-three prime untranslated region (3'UTR) by miR-373 were evaluated by a dual-luciferase reporter gene assay. RESULTS: In hilar cholangiocarcinoma, miR-373 decreased and was closely associated with poor cell differentiation, advanced clinical stage, and shorter survival. The promoter-associated CpG island of miR-373 gene was hypermethylated and inhibited expression of miR-373. MBD2 was up-regulated and enriched at the promoter-associated CpG island of miR-373. Methylation-mediated suppression of miR-373 required MBD2 enrichment at the promoter-associated CpG island, and miR-373 negatively regulated MBD2 expression through targeting the 3'UTR. CONCLUSION: MiR-373 behaves as a direct transcriptional target and negative regulator of MBD2 activity through a feedback loop of CpG island methylation.

Identification of methylation profile of HOX genes in extrahepatic cholangiocarcinoma.

AIM: To identify methylation profile and novel tumor marker of extrahepatic cholangiocarcinoma (CCA) with high throughout microarray. METHODS: Differential methylation profile was compared between normal bile duct epithelial cell lines and CCA cell lines by methyl-DNA immunoprecipitation (MeDIP) microarray. Bisulfite-polymerase chain reaction (BSP) was performed to identify the methylated allels of target genes. Expression of target genes was investigated before and after the treatment with DNA demethylating agent. Expression of candidate genes was also evaluated by immunofluorescence in 30 specimens of CCA tissues and 9 normal bile duct tissues. RESULTS: Methylation profile of CCA was identified with MeDIP microarray in the respects of different gene functions and signaling pathways. Interestingly, 97 genes with hypermethylated CpG islands in the promoter region were homeobox genes. The top 5 hypermethylated homeobox genes validated by BSP were HOXA2 (94.29%), HOXA5 (95.38%), HOXA11 (91.67%), HOXB4 (90.56%) and HOXD13 (94.38%). Expression of these genes was reactivated with 5'-aza-2'-deoxycytidine. Significant expression differences were found between normal bile duct and extrahepatic CCA tissues (66.67%-100% vs 3.33%-10%). CONCLUSION: HOXA2, HOXA5, HOXA11, HOXB4 and HOXD13 may work as differential epigenetic biomarkers between malignant and benign biliary tissues.

[Effect of protecting parathyroid in situ in the operation of total thyroidectomy].

OBJECTIVE: To evaluate the effect of protecting parathyroid glands in situ in the operation of total thyroidectomy by detecting parathyroid hormone after the operation. METHODS: In the surgical team, 1019 consecutive patients with thyroid diseases were treated with total thyroidectomy. During the operation, parathyroid glands were protected in situ with correctly identifying the parathyroid glands, precisely dissecting its envelope and protecting its blood supply. Serum calcium level and parathyroid hormone were measured before and 24 hours after operation. The patients who had symptomatic hypocalcemia or hypoparathyroidism were given supportive treatment and followed-up. RESULTS: At least one of the parathyroid glands was preserved and remained in situ in all cases. Eighty-nine cases (8.7%) had decreased parathyroid hormone levels and 42 cases (4.1%) had complicated symptomatic hypocalcemia. The symptoms of hypocalcemia in all these cases could be controlled by supportive treatment, and serum calcium level and parathyroid hormone had all recovered 1 - 6 months later. If 3 and 4 parathyroid were conserved in situ, the postoperative complication rate was significantly lower than those with 1 and 2 parathyroid conserved (decreased PTH 69/999 vs 20/20, symptoms of hypocalcemia 25/999 vs 17/20, all P < 0.01). CONCLUSION: The techniques to protect parathyroid glands in situ are effective measure to prevent the postoperative hypoparathyroidism in total thyroidectomy.

[Conservative therapy in the treatment of cervical chylous leakage].

OBJECTIVE: To explore and evaluate the combined conservative managements in the treatment of cervical chylous leakage. METHODS: Thirty nine cases of cervical chylous leakage from June 1992 to June 2008 were retrospectively analyzed in this hospital. All of the 39 cases were cured by treating with conservative individualized therapy, including the applying of diet with high calorie, high protein and low fat and fatty food should only contains medium-chain triglycerides, total parenteral nutrition, keep the balance of hydrogen and electrolyte and correct hypoproteinemia, local pressure dressing, high persistent vacuum drainage (-50 approximately -80 kPa) and/or somatostatin analogue. RESULTS: All the cases of chylous leakage happened 2nd to 5th days after the operation. Among the 39 cases, 7 were high flow (drainage>or=500 ml/d) chylous leakage, the amount of drainage reached as high as 1440 ml per day. The time of chylous leakage closure was 3 approximately 12 days, and the mean time was 7 days. No one experienced re-operation, wound hydrops or wound infection. CONCLUSIONS: The conservative individualized therapy may play a key role in the treatment of cervical chylous leakage.

Expression of P-aPKC-iota, E-cadherin, and beta-catenin related to invasion and metastasis in hepatocellular carcinoma.

OBJECTIVES: Atypical protein kinase C iota (aPKC-iota) and its associated intracellular molecules, E-cadherin and beta-catenin, are important for cell polarization in tumorigenesis and progression. Expression of aPKC-iota, P-aPKC-iota (activated aPKC-iota), E-cadherin, and beta-catenin in hepatocellular carcinoma (HCC) was measured, and correlation with clinicopathological characteristics of HCC was analyzed. METHODS: Paraffin-embedded tumor tissue was obtained from patients with HCC after resection without preoperative radiotherapy or chemotherapy. Gene expression was detected by polymerase chain reaction (PCR), and protein expression was detected by immunohistochemistry and Western blot analysis. Expressions of aPKC-iota, P-aPKC-iota, E-cadherin, and beta-catenin were analyzed with relation to the clinicopathological data. RESULTS: The gene and protein expression of aPKC-iota are obviously higher in HCC tissues than that in peritumoral tissues and normal tissues by semiquantitative PCR and immunohistochemistry methods. Accumulation of aPKC-iota in HCC cytoplasm and nucleolus inhibited the later formation of belt-like adherens junctions (AJs) and/or tight junctions (TJs) in cell-cell contact. E-cadherin was reduced and accumulation of cytoplasm beta-catenin was increased in HCC. The expression of aPKC-iota was closely related to pathological differentiation, tumor size, invasion, and metastasis of HCC. CONCLUSION: Accumulation of cytoplasm aPKC-iota may reflect pathological differentiation, invasion, and metastasis potential of HCC. In this regard, our study on HCC revealed the potential usefulness of aPKC-iota, E-cadherin, and beta-catenin as a prognostic marker, closely related to pathological differentiation, invasion, metastasis, and prognosis of HCC.

Significance and expression of atypical protein kinase C-iota in human hepatocellular carcinoma.

BACKGROUND AND OBJECTIVE: Previous studies have indicated that abnormal expression of atypical protein kinase C (aPKC-iota) plays a critical role in occurrence and progression of malignant tumor. This study analyzed the correlation of aPKC-iota with clinicopathology in hepatocarcinoma and Cyclin E and investigated molecular mechanisms of invasion and metastasis of hepatocellular carcinoma. MATERIALS AND METHODS: The expression of the aPKC-iota gene was examined by reverse transcription-polymerase chain reaction in 7 specimens of normal liver tissues and 43 of hepatoma and adjacent tissues. Expression of aPKC-iota and Cyclin E protein was detected using immunohistochemistry and Western blot. Finally, we analyzed the correlation of aPKC-iota with clinicopathologic characteristics and invasion of hepatoma. RESULTS: The expression value (0.844 +/- 0.315) of aPKC-iota gene is obviously higher in hepatoma than the value (0.530 +/- 0.217) in adjacent tissues and the value (0.372 +/- 0.130) in normal tissue (P = 0.009). The positive expression rate (58.1%) of aPKC-iota protein in hepatoma is remarkably higher than the rate (23.3%) of adjacent tissues. The expression of aPKC-iota has a positive correlation with the expression of Cyclin E, differentiation degree, and invasion of tumor (P < 0.05). CONCLUSIONS: Differentiation degree and invasion of hepatoma are related to the expression of aPKC-iota, which plays an important role in invasion and metastasis of hepatoma.

[Prevention and cure of the complications after radical pancreatoduodenectomy].

OBJECTIVE: To investigate the causes and the measures of prevention and cure of the dangerous complications (bleeding, pancreatic fistula, biliary fistula and death) after radical pancreatoduodenectomy (RPD) for periampullary malignant tumor. METHODS: The rate and management of dangerous complications of 156 cases with RPD which were continuous performed by Department of Biliary-Pancreatic Surgery, Affiliated Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology between January 2006 and June 2008 were analyzed retrospectively, including 97 males and 59 females with 37 - 79 years old, the mean age was 56.9 years old. RESULTS: Among the 156 cases with RPD, four patients had massive hemorrhage of gastrointestinal tract due to stress ulcer, two patients had bleeding in the pancreas-intestinal anastomosis after the operation, the rate of postoperative bleeding was 3.9% (6/156). One patient with massive hemorrhage of gastrointestinal tract due to stress ulcer had severe pulmonary infection and ARDS, and died of respiratory failure finally (the overall mortality rate was 0.7%) after ICU for two months. One patients with bleeding in the pancreas-intestinal anastomosis had pancreatic fistula (the rate of pancreatic fistula was 0.7%) 3 days after the second laparotomy to open the jejunum of the pancreas-intestinal anastomosis and make a transfixion of the bleeding points in the stump. Another patient who had the tumor located in the inferior segment of the bile common duct had biliary fistula 11 days after the operation (the rate of biliary fistula was 0.7%). Two patients with fistula had good recovery by expectant treatment of ultrasound-guided puncture and drainage. CONCLUSIONS: Prompt and effective treatment of the complications of bleeding, pancreatic fistula, biliary fistula could maximally decrease the perioperative death rate.

Effect of mutant p27(kipl) gene on human cholangiocarcinoma cell line, QBC(939).

AIM: To investigate the effects of exogenously mutated p27(kipl) (p27) on proliferation and apoptosis of human cholangiocarcinoma cell line, QBC(939)in vivo. METHODS: Adenviral vectors were used to transfect mutated p27 cDNA into human QBC(939)cell line. Expression of p27 was detected by RT-PCR. Western blot. Cell growth, morphological change, cell cycle, apoptosis and cloning formation were determined by MTT assay and flow cytometry. RESULTS: The expression of p27 protein and mRNA was increased significantly in QBC(939) cell line transfected with Ad-p27mt. The transfer of Ad-p27mt could significantly inhibit the growth of QBC(939) cells, decrease the cloning formation rate and induce apoptosis. p27 over expression caused cell cycle arrest at G(0)/G(1)phase 72 h after infection with Ad-p27mt. CONCLUSION: p27 may cause cell cycle arrest at G(0)/G(1)phase and subsequently lead to apoptosis. Recombinant adenovirus expressing mutant p27 may be potentially useful in gene therapy for cholangiocarcinoma.

[The technique of radical pancreaticoduodenectomy for malignant tumor in pancreatic head with pressed superior mesenteric blood vessels or portal vein].

OBJECTIVE: To investigate the technique of radical pancreaticoduodenectomy for malignant tumor in pancreatic head with pressed superior mesenteric blood vessel or portal vein. METHODS: From March 2005 to March 2007, thin slice scan and vessel-reconstruction of 56 patients of malignant tumor in pancreatic head with pressed superior mesenteric blood vessels or portal vein were carried out using multidetector spiral CT to evaluate whether peripheral vessels of pancreatic tumor were invaded and whether the tumor was resectable. During the operation, 3 vascular blocking bands for superior mesenteric vein, portal vein and spleen vein or 4 vascular blocking bands (additional one for inferior mesenteric vein) were preset. Under the cross and traction between superior mesenteric vein and superior mesenteric artery, resected the uncinate process of pancreas thoroughly. Using those methods, radical pancreaticoduodenectomy for 56 patients above-mentioned were successfully accomplished. RESULTS: The accuracy for preoperative judging by using multidetector spiral CT whether the peripheral vessels of pancreatic cancer were invaded and whether the tumor was resectable was 98% and 100% separately. Thirty-seven of 56 patients, whose superior mesenteric blood vessels or portal veins were pressed by the tumor of pancreatic head, were operated using 3 vascular blocking bands and 2 patients using 4 vascular blocking bands, followed by suturing the bleeding points of the superior mesenteric vein with 5-0 vascular suture Proline. One patient's superior mesenteric vein was partially resected and restored. The operations cost 5-8 h each and the blood loss was 200-600 ml. There were no operative or postoperative hemorrhage or pancreatic juice leakage. According to the follow-up up to now, 2 patients died of multiple live tumor metastases 7 and 9 months separately after operation, the other 54 patients were still alive. CONCLUSIONS: Thin slice scan and vessel-reconstruction using multidetector spiral CT can accurately judge whether the blood vessels near the pancreatic tumor were invaded and whether the tumor was resectable, using 3 vascular blocking bands or 4 vascular blocking bands and cross, traction of the superior mesenteric blood vessels, operator can easily accomplish the radical pancreaticoduodenectomy of malignant tumor in pancreatic head with pressed superior mesenteric blood vessels and portal vein, which was not resectable or need combined resection of the blood vessels in the traditional opinion.

Effect of histone deacetylase inhibitor on proliferation of biliary tract cancer cell lines.

AIM: To explore the effect of histone deacetylase inhibitor, trichostatin A (TSA) on the growth of biliary tract cancer cell lines (gallbladder carcinoma cell line and cholangiocarcinoma cell line) in vivo and in vitro, and to investigate the perspective of histone deacetylase inhibitor in its clinical application. METHODS: The survival rates of gallbladder carcinoma cell line (Mz-ChA-l cell line) and cholangiocarcinoma cell lines (QBC939, KMBC and OZ cell lines) treated with various doses of TSA were detected by methylthiazoy tetrazolium (MTT) assay. A nude mouse model of transplanted gallbladder carcinoma (Mz-ChA-l cell line) was successfully established, and changes in the growth of transplanted tumor after treated with TSA were measured. RESULTS: TSA could inhibit the proliferation of gallbladder carcinoma cell line (Mz-ChA-l cell line) and cholangiocarcinoma cell lines (QBC939, KMBC and OZ cell lines) in a dose-dependent manner. After the nude mouse model of transplanted gallbladder carcinoma (Mz-ChA-l cell line) was successfully established, the growth of cancer was inhibited in the model after treated with TSA. CONCLUSION: TSA can inhibit the growth of cholangiocarcinoma and gallbladder carcinoma cell lines in vitro and in vivo.

Clinical analysis of solid-pseudopapillary tumor of the pancreas: report of 15 cases.

BACKGROUND: Solid-pseudopapillary tumor of the pancreas (SPTP) is an uncommon and enigmatic pancreatic neoplasm that occurs mainly in young women. Although more and more cases have been reported in recent years, misdiagnosis and incorrect treatment still frequently take place. This study was designed to stimulate consideration of this tumor. METHODS: We retrospectively reviewed the experience of diagnosis and treatment of 15 patients with SPTP and compared them with 516 patients with pancreatic cancer from January 1997 to March 2007. RESULTS: Most of the SPTP cases were asymptomatic except for one palpable mass. Almost all SPTPs demonstrated a solid structure with hypo- or iso-attenuation, cystic structure with hypo-attenuation on pre-contrast CT scan, and enhancement of solid portions on post-contrast CT scan. By contrast, most cases of pancreatic carcinoma had multiple symptoms and abnormal blood results. The tumors showed hypo-attenuation on both pre-contrast and post-contrast CT scan, and only a few showed iso-attenuation on post-contrast CT scan. All cases of SPTP in our group were cured by surgical resection, while only 16.86% of patients with pancreatic carcinoma could undergo a radical resection. CONCLUSIONS: Clinical features and CT scans were helpful to differentiate SPTP from pancreatic carcinoma. Radical surgical resection was the most effective and safe method for the treatment of SPTP.

Correlation of aPKC-iota and E-cadherin expression with invasion and prognosis of cholangiocarcinoma.

BACKGROUND: The abnormal expression of atypical protein kinase C-iota (aPKC-iota) subtype and E-cadherin play important roles in tumor occurrence and progression. This study was designed to investigate the correlation of expression of aPKC-iota and E-cadherin with the clinicopathological characteristics and prognosis of cholangiocarcinoma, and to analyze the molecular mechanisms of invasion and metastasis of the tumor. METHODS: EnVision immunohistochemistry was used to detect the expression of aPKC-iota and E-cadherin in 9 specimens of benign bile duct tissues, 35 specimens of cholangiocarcinoma and 6 specimens of metastatic cholangiocarcinoma. The relationship of the expression with clinicopathological characteristics, invasion and prognosis of cholangiocarcinoma was analyzed. A multivariate regression analysis was made of these data by the Cox proportional hazard model. RESULTS: The positive expression level of aPKC-iota in cholangiocarcinoma was remarkably higher than that in benign bile duct tissues (68.6% vs. 11.1%, P=0.006), but the expression level of E-cadherin was lower in cholangiocarcinoma than in benign bile duct tissues (37.1% vs. 88.9%, P=0.016). Correlation analysis revealed that the expression of aPKC-iota was positively related to tumor differentiation and invasion, whereas that of E-cadherin was entirely the contrary. Moreover, there was a negative relationship between the expression of aPKC-iota and that of E-cadherin (r=-0.287, P<0.05). Univariate analysis showed that the overall survival rate of the group with a higher expression of aPKC-iota in cholangiocarcinoma was remarkably lower than that of the group with a lower expression (P<0.01); multivariate analysis revealed that the expressions of aPKC-iota and E-cadherin are important prognostic factors for cholangiocarcinoma (P<0.05). CONCLUSIONS: The expressions of aPKC-iota and E-cadherin may reflect the differentiation and invasive potential of cholangiocarcinoma. aPKC-iota and E-cadherin may be independent prognostic factors and, when used in combination with clinicopathological characteristics, may increase the accuracy in predicting the prognosis of patients with cholangiocarcinoma. As a polar regulative associated protein, aPKC-iota may play an important role in the invasion and metastasis of cholangiocarcinoma.

Two-dimensional electrophoresis for comparative proteomic analysis of human bile.

BACKGROUND: Proteomic analysis of bile fluid holds promise as a method to identify biomarkers of bile tract diseases, especially for tumors. Two-dimensional electrophoresis (2-DE) is a popular and proven separation technique for proteome analysis, but using this strategy for bile fluid analysis is still not fully developed. This study was undertaken to (a) establish a reliable method for general clean-up to make bile fluid samples suitable for 2-DE; (b) obtain 2-D biliary maps with high reproducibility and resolution; and (c) identify protein patterns present in 2-D biliary maps for potential tumor biomarker discovery, with the intention of distinguishing malignant from benign causes of bile duct obstruction. METHODS: Bile fluid samples were obtained from two patients suffering from malignant and benign bile tract obstruction (one patient with cholangiocarcinoma as the experimental case, the other with cholelithiasis as control). A variety of sample preparation options, including delipidation, desalination and nucleic acid removal, were adopted to remove contaminants that affect 2-DE results. After that, each 350 microg purified sample was loaded onto nonlinear IPG strips (18 cm, pH 3-10 and pH 4-7) for first-dimension isoelectric focusing, and 12.5% SDS-PAGE electrophoresis for second dimension separation. Then 2-D maps were visualized after silver staining and analyzed with the Image Master 2-D software. RESULTS: A large number of protein spots were separated in 2-D maps from the experimental and control groups, with means of 250 and 216 spots on pH 3-10 IPG strips, and 182 and 176 spots on pH 4-7 strips, respectively. Approximately 16 and 23 spots were differentially expressed in matched pairs from the experimental and control cases using pH 3-10 and pH 4-7 strips. CONCLUSIONS: This study established a reliable sample preparation process suitable for 2-DE of bile fluid. By this method, 2-D biliary maps with high reproducibility and resolution were obtained. The differentially displayed proteomes in the 2-D biliary maps from the experimental and control groups indicated the potential application for bile fluid analysis to identify disease-associated biomarkers, especially for biliary tract tumors.

[Expression and significance of aPKC-iota and E-cadherin in cholangiocarcinoma].

BACKGROUND & OBJECTIVE: Studies showed that the abnormal expression of atypical protein kinase C iota subtype (aPKC-iota) and E-cadherin plays an important role in tumor genesis and progression. This study was to investigate the expression of aPKC-iota and E-cadherin in cholangiocarcinoma, and analyze molecular mechanisms of the invasion and metastasis of cholangiocarcinoma. METHODS: The expression of aPKC-iota and E-cadherin in 9 specimens of benign bile duct tissues and 35 specimens of cholangiocarcinoma was detected by EnVision immunohistochemistry, and their correlations to the clinicopathologic characteristics and invasion of cholangiocarcinoma were analyzed. RESULTS: The positive rate of aPKC-iota was significantly higher in cholangiocarcinoma than in benign bile duct tissues (68.6% vs. 11.1%, P = 0.006), while the positive rate of E-cadherin was significantly lower in cholangiocarcinoma than in benign bile duct tissues (37.1% vs. 88.9%, P = 0.016). aPKC-iota expression was negatively correlated to E-cadherin expression (r = -0.287, P < 0.05). aPKC-iota expression was positively and E-cadherin expression was negatively correlated to the differentiation and invasion of cholangiocarcinoma (P < 0.05). CONCLUSIONS: The expression of aPKC-iota and E-cadherin may reflect the differentiation and invasive potential of cholangiocarcinoma. As a polar regulation-associated protein, aPKC-iota may play a role in the invasion and metastasis of cholangiocarcinoma.

Effect of targeted magnetic nanoparticles containing 5-FU on expression of bcl-2, bax and caspase 3 in nude mice with transplanted human liver cancer.

AIM: To investigate the anti-tumor effect and mechanisms of magnetic nanoparticles targeting hepatocellular carcinoma. METHODS: Human hepatocellular carcinoma was induced in nude mice, and the mice were randomly divided into group A receiving normal saline, group B receiving magnetic nanoparticles containing 5-fluorouracil (5-FU), group C receiving 5-FU, and group D receiving magnetic nanoparticles containing 5-FU with a magnetic field built in tumor tissues. The tumor volume was measured on the day before treatment and 1, 4, 7, 10 and 13 d after treatment. Tumor tissues were isolated for examination of the expression of bcl-2, bax and caspase 3 by immunohistochemical method, reverse transcription polymerase chain reaction and Western blotting. RESULTS: The tumor volume was markedly lower in groups C and D than in groups A and B (group C or D vs group A or B, P<0.01). The volume was markedly lower in group D than in group C (P<0.05). The expression of protein and mRNA of bcl-2 was markedly lower in groups C and D than in groups A and B (group C or D vs group A or B, P<0.01), and was markedly lower in group D than in group C (P<0.01). The expression of bax and caspase 3 in groups C and D was significantly increased, compared with that in groups A and B (P<0.01). CONCLUSION: The targeted magnetic nanoparticles containing 5-FU can improve the chemotherapeutic effect of 5-FU against hepatocellular carcinoma by decreasing the expression of bcl-2 gene, and increasing the expression of bax and caspase 3 genes.

Effects of targeting magnetic drug nanoparticles on human cholangiocarcinoma xenografts in nude mice.

BACKGROUND: Targeting is a new therapeutic tool for malignant tumor as a result of combining nanotechnology with chemotherapeutics. The aim of our study was to investigate the effects of magnetic nanoparticles enveloping a chemotherapeutic drug on human cholangiocarcinoma xenografts in nude mice. METHODS: The human cholangiocarcinoma xenograft model was established in nude mice with the QBC939 cell line. The nude mice were randomly assigned to 7 groups. 0.9% saline or magnetic nanoparticles, including high (group 2), medium (group 4) and low (group 5) dosages, were given to nude mice through the tail vein 20 days after the QBC939 cell line was implanted. Calculations were made 35 days after treatment in order to compare the volumes, inhibition ratios and growth curves of the tumors in each group. Mice in each group were sacrificed randomly to collect tumor tissues and other organs for electron microscopy and pathological examination. RESULTS: The high and medium dosage groups were significantly different from the control group (P<0.05). The tumor inhibition ratios for the high, medium and low dosage groups were 39.6%, 14.6% and 7.9%, respectively. The tumor growth curve of groups 5, 4, and 2 changed slowly in turn. The high and medium groups showed cell apoptosis under an electron microscope. CONCLUSION: Magnetic nanoparticles can inhibit the growth of human cholangiocarcinoma xenografts in nude mice.

[Pharmacokinetics and distribution of 5-Fu magnetic albumin deuto-microsphere in normal and tumor-bearing mice].

To observe the pharmacokinetic and tissue-distribution characters of 5-flourouracil magnetic albumin deuto-microsphere (5-Fu-MAD) in normal and tumor-bearing mice, HPLC method for the determination of 5-Fu in plasma and tissues was established and applied to determine 5-Fu in mouse plasma and tissue samples. A Flame atomic absorption spectrometer was used to detect the iron concentration in mouse tissue. Plasma concentration-time curves of free 5-Fu, 5-Fu-MAD and 5-Fu-MAD plus the magnetic frame (MF) conformed to two compartment model of first order absorption and they had C(max) of 34.9, 7.95 and 5.97 mg x L(-1); T1/2 (Ke) of 22.26, 76.0 and 124.6 min, V(d) of 3.28, 30.7 and 66.1 L x kg; AUC(0-t), of 233.9, 78.3 and 50.2 mg x min x L(-1); AUC(0-infinity) of 237.2, 89.3 and 68.1 mg x min x L(-1), respectively. The distribution of 5-Fu and iron was the highest in the plenty blood perfusion organs like the liver, tumor, spleen and lung, while lower in the kidney and heart and lowest in brain and muscle. The tissue distribution of muscle and tumor increased significantly when a magnetic frame was inserted there. The pharmacokinetics and tissue distribution of 5-Fu-MAD exhibited sustained-release and target characteristics.