OBJECTIVETo investigate the effect of photodynamic therapy (PDT) mediated by hematoporphyrin derivative (HPD) on apoptosis and invasion of cholangiocarcinoma QBC939 cell lines.

METHODSIn vitro cultured cholangiocarcinoma QBC939 cell line was exposed to 2, 4, 6, 8, 10, 12, and 14 μg/ml HPD with 5, 10, and 15 J/cm2 light intensity, respectively. The optical density at 450 nm of the QBC939 cells was measured by CCK8 assay and its growth inhibition ratio was calculated. Flow cytometry and transwell migration assay were applied to detect cell apoptosis and invasion respectively. RT-PCR and immunocytochemistry analyses were used to detect expressions of vascular endothelial growth factor-C (VEGF-C), cyclooxygenase-2 (COX-2), and proliferating cell nuclear antigen (PCNA). Enzyme-linked immunosorbent assay (ELISA) was carried out to examine the secretion of VEGF-C and COX-2 in QBC939 cells.

RESULTSExposure to HPD-PDT can significantly suppress the growth of QBC939 cells (all P<0.05). HPD-PDT can promote apoptosis of QBC939 cells at the early stage. When the concentration of HPD was 2 μg/ml and light irradiation was 5 J/cm2, HPD-PDT had no obvious inhibitory effect on QBC939 cell growth, but can obviously inhibit cell invasion, and significant difference was observed between the HPD-PDT and control groups (P<0.01). The HPD-PDT can reduce the mRNA and protein expressions of VEGF-C, COX-2, and PCNA, and decrease the secretion of VEGF-C and COX-2 in QBC939 cells.

CONCLUSIONPDT could promote apoptosis and inhibit growth and invasion of cholangiocarcinoma cells QBC939 in vitro.

Apoptosis ; drug effects ; Bile Duct Neoplasms ; drug therapy ; pathology ; Bile Ducts, Intrahepatic ; Cell Line, Tumor ; Cell Movement ; drug effects ; Cholangiocarcinoma ; drug therapy ; pathology ; Humans ; Neoplasm Invasiveness ; Photochemotherapy ; Proliferating Cell Nuclear Antigen ; analysis

OBJECTIVETo explore the molecular mechanism of γ-aminobutyric acid (GABA) inhibitory on the growth of cholangiocarcinoma cell line (QBC939).

METHODSQBC939 cells were cultured in different groups and treated with GABA, GABA + bicuculine (A receptor antagonist), GABA + phaclofen (B receptor antagonist) for 48 hours. MTT assay was used to determine the proliferation of QBC939 cells. Annexin V-FITC/PI binding assay was used to detect apoptosis in the QBC939 cells. Western blot was applied to check the expression of signal transducer and activator of transcription 3 (STAT3) and phosphorylation-STAT3 (p-STAT3) proteins in different groups of QBC939 cells. Animal models of cholangiocarcinoma bearing nude mice were established by subcutaneous injection of QBC939 cells and randomized into 2 groups: control and GABA-treated groups. The effect of GABA was evaluated after 5 weeks, including the body weight and tumor volume. The expression of p-STAT3 was detected by immunohistochemistry and Western blot in xenograft tumors.

RESULTSMTT and FCM assays both showed that the effect of GABA inhibitory on the proliferation (15.30% ± 0.80% vs. 2.66% ± 0.74%, t = 23.15, P = 0.00) and induced apoptosis (23.15% ± 0.21% vs. 4.30% ± 0.69%, t = 52.40, P = 0.00) of QBC939 cells could be antagonized by phaclofen, but not bicuculine. The expression of STAT3 and p-STAT3 proteins were all observed in the QBC939 cells and GABA significantly down-regulated p-STAT3 protein expression (0.77 ± 0.00 vs. 0.45 ± 0.01, t = 63.14, P = 0.00), this action was also antagonized by phaclofen (0.45 ± 0.01 vs. 0.76 ± 0.01, t = 56.25, P = 0.00). Xenograft tumor volume ((0.62 ± 0.03) cm³ vs. (0.34 ± 0.03) cm³, t = 13.45, P = 0.00) and the expression of p-STAT3 protein were significantly decreased in GABA-treated group as compared with control group.

CONCLUSIONSGABA may inhibit the growth of cholangiocarcinoma cells QBC939 through GABA(B) receptor, and down-regulation of the p-STAT3 expression perhaps is one of its anti-tumor mechanisms.

Animals ; Apoptosis ; drug effects ; Bile Duct Neoplasms ; pathology ; Bile Ducts, Intrahepatic ; pathology ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; Cholangiocarcinoma ; pathology ; Humans ; Mice ; Mice, Nude ; STAT3 Transcription Factor ; metabolism ; Tumor Burden ; gamma-Aminobutyric Acid ; pharmacology

OBJECTIVETo investigate the effects of mutant exogenous P27(kip1) gene on chemosensitivity of human cholangiocarcinoma cell line.

METHODSThe recombinant vector was constructed and the mutant P27(kip1) gene was transfected into human cholangiocarcinoma cell line (QBC(939)). RT-PCR and Western blot were used to determine the expression of target genes. The effects of 5-fluorouracil (5-FU), mitomycin C (MMC) and cyclophosphamide (CTX) on the transfected cells were detected by assaying the apoptotic rate and growth inhibition by methabenzthiazuron (MTT) assay and flow cytometry (FCM).

RESULTSThe mutant exogenous P27(kip1) gene was expressed effectively in the cells, and the expression enhanced the apoptosis and growth inhibition of QBC(939) inducted by 5-FU, MMC and CTX. The ratio of growth inhibiting increased significantly from 41.89% (5-FU), 45.59% (MMC), 38.91% (CTX) to 56.15% (5-FU), 55.65% (MMC), 51.69% (CTX), and apoptosis index from 13.76% +/- 3.03% (5-FU), 11.76% +/- 3.99% (MMC), 10.46% +/- 2.10% (CTX) to 41.39% +/- 4.32% (5-FU), 35.94% +/- 2.71% (MMC), 34.46% +/- 2.32% (CTX) (P < 0.05).

CONCLUSIONSThe exogenous P27(kip1) gene transfer can remarkably increase the drug sensibility of the cholangiocarcinoma cells. The strategy targeted to control the cell cycle may be more effective in cancer treatment by combination of P27(kip1) gene therapy.

Adenoviridae ; genetics ; Antineoplastic Agents ; pharmacology ; Apoptosis ; drug effects ; Bile Duct Neoplasms ; pathology ; Bile Ducts, Intrahepatic ; Cell Division ; drug effects ; Cell Line, Tumor ; Cholangiocarcinoma ; pathology ; Cyclin-Dependent Kinase Inhibitor p27 ; genetics ; pharmacology ; Drug Synergism ; Genetic Vectors ; Humans ; Transfection

Allopurinol-induced hypersensitivity syndrome is characterized by an idiosyncratic reaction involving multiple-organs, which usually begins 2 to 6 weeks after starting allopurinol. In rare cases, the adverse reactions to allopurinol are accompanied by a variety of liver injury, such as reactive hepatitis, granulomatous hepatitis, vanishing bile duct syndrome, or fulminant hepatic failure. Here we report a case with granulomatous hepatitis and ductopenia. A 69-year-old man with chronic renal failure, hyperuricemia, and previously normal liver function presented with jaundice, skin rash, and fever 2 weeks after taking allopurinol (200 mg/day). In histopathology, a liver biopsy specimen showed mild spotty necrosis of hepatocytes, marked cholestasis in parenchyma, and some granulomas in the portal area. There were vacuolar degeneration in the interlobular bile ducts and ductopenia in the portal tracts. Pathologic criteria strongly suggested the presence of allopurinol-induced granulomatous hepatitis with ductopenia and cholestasis. The patient fully recovered following the early administration of systemic corticosteroid therapy.
Aged ; Allopurinol/*adverse effects/therapeutic use ; Antimetabolites/*adverse effects/therapeutic use ; Bile Duct Diseases/*chemically induced/diagnosis/pathology ; Bile Ducts, Intrahepatic/*drug effects/pathology ; Cholestasis/*chemically induced/diagnosis/pathology ; Drug Eruptions/pathology ; Granuloma/*chemically induced/pathology ; Hepatitis, Toxic/*pathology ; Humans ; Kidney Failure, Chronic/complications/drug therapy ; Male

By way of gastrogavage, we administered CANELIM capsules to rat for prepering the drug-contained serums. And then the serums obtained were used to plant cholangiocarcinoma cells. Lastly, using the micropipette aspiration technique, we investigated the effects which the drug-contained serums of different doses have on the viscoelasticity and adhesive mechanical properties of cholangiocarcinoma cells. The results showed that cholangiocarcinoma cells presented a characteristic of high elastic coefficient and low viscous coefficient. After being treated by the high dose and middle dose drug-contained serums, the viscoelastical properties of cholangiocarcinoma cells K1, K2 and micro evidently decreased (P < 0.01). But the properties of low dose did not evidently change. The adhesive force between cholangiocarcinoma cells and CD44v6 protein significantly reduced with the increasing of the dose of CANELIM capsules (P < 0.01). It is suggested that CANELIM capsules would destroy the cytoskeleton of cholangiocarcinoma cells, restrain the adhesion molecule CD44v6 on membrane from expressing, reduce the adhesion probability between cholangiocarcinoma cells and vasal endothelial cells, and finally, prevent the metastasis of cholangiocarcinoma cells.
Animals ; Antineoplastic Agents, Phytogenic ; pharmacology ; Bile Duct Neoplasms ; pathology ; Bile Ducts, Intrahepatic ; pathology ; Capsules ; Cell Adhesion ; drug effects ; Cholangiocarcinoma ; pathology ; Drugs, Chinese Herbal ; pharmacology ; Elasticity ; drug effects ; Endothelial Cells ; cytology ; Female ; Humans ; Male ; Rats ; Serum ; Tumor Cells, Cultured ; Viscosity ; drug effects

Antineoplastic Agents ; pharmacology ; Bile ; physiology ; Bile Duct Neoplasms ; metabolism ; pathology ; Bile Ducts, Intrahepatic ; Celecoxib ; Cell Division ; drug effects ; Cell Line, Tumor ; Cholangiocarcinoma ; metabolism ; pathology ; Cyclooxygenase 2 ; Dinoprostone ; metabolism ; Humans ; Isoenzymes ; biosynthesis ; genetics ; Membrane Proteins ; Prostaglandin-Endoperoxide Synthases ; biosynthesis ; genetics ; Pyrazoles ; RNA, Messenger ; genetics ; Sphincterotomy, Transduodenal ; adverse effects ; Sulfonamides ; pharmacology ; Up-Regulation

OBJECTIVETo investigate the anti-tumor immune response of dendritic cells (DCs) acquiring antigens from apoptotic cholangiocarcinoma cells and their therapeutic effects on cholangiocarcinoma cells.

METHODSDCs from human peripheral blood monocytes which acquired antigen capturing and processing capacity, characteristics of maturation, were established in vitro using granulocyte-macrophage colony-stimulating factor (GM-CSF) and interleukin-4 (IL-4). Then cholangiocarcinoma cells were induced to apoptosis with mitomycin. The three groups included (1) coculture of DCs, apoptotic cancer cells and T cells, (2) coculture of DCs, necrotic cancer cells and T cells, (3) coculture of DCs, cultured cancer cells and T cells. After 7 days, DCs and T cells were riched separately to perform anti-tumor cells test and immune response test.

RESULTSthese cells had typical dendritic cell morphology, expressed high levels of CD1a and B7, acquired antigen from apoptotic cells caused by mitomycin and could stimulate T cells to inhibit, even kill cholangiocarcinoma cells.

CONCLUSIONSThe DCs from peripheral blood monocytes induced by GM-CSF and IL-4 can efficiently present antigen derived from apoptotic cells caused by mitomycin, and stimulate T cells activity obviously. It maybe become an effective therapy for tumor.

Antigens, Neoplasm ; immunology ; Apoptosis ; drug effects ; Bile Duct Neoplasms ; immunology ; pathology ; Bile Ducts, Intrahepatic ; Cholangiocarcinoma ; immunology ; pathology ; Coculture Techniques ; Dendritic Cells ; drug effects ; immunology ; Granulocyte-Macrophage Colony-Stimulating Factor ; pharmacology ; Humans ; Interleukin-4 ; pharmacology ; Mitomycin ; pharmacology ; T-Lymphocytes ; immunology ; Tumor Cells, Cultured

OBJECTIVETo investigate the efficacy of the combination of gemcitabine with capecitabine in the chemotherapy for patients with relapsed or metastatic biliary tract carcinoma.

METHODSForty-one patients with unresectable relapsed or metastatic carcinoma of the biliary tract were treated from March 2000 to December 2004. The regimen consisted of intravenous administration of gemcitabine plus oral intake of capecitabine every 3 weeks for more than 2 cycles. The parameters including tumor response, clinical benefit rate,survival and safety were observed.

RESULTSThirty-six patients were valuable and 5 patients were excluded from this series due to various reasons. Eleven patients (30.1%) had a partial response and another 11 patients (30.1%) experieced stable disease with a clinical benefit rates of 61.1%. The median overall survival time and time to progression were 10 months and 6 months, respectively. The one-year survival rate was 40.0%. The adverse events including nausea, diarrhea and hand-foot syndrome, fatigue, neutropenia, thrombocytopenia were frequently observed, which were usually in grade I or II, rarely in grade III and none in grade IV (NCI-CTC).

CONCLUSIONOur results show that the regimen of gemcitabine combined with capecitabine is effective and well tolerated in patients with unresectable relapsed or metastatic carcinoma of the biliary tract.

Adult ; Aged ; Antineoplastic Combined Chemotherapy Protocols ; adverse effects ; therapeutic use ; Bile Duct Neoplasms ; drug therapy ; pathology ; Bile Ducts, Intrahepatic ; Capecitabine ; Cholangiocarcinoma ; drug therapy ; pathology ; Deoxycytidine ; administration & dosage ; analogs & derivatives ; Diarrhea ; chemically induced ; Female ; Fluorouracil ; administration & dosage ; analogs & derivatives ; Follow-Up Studies ; Humans ; Male ; Middle Aged ; Nausea ; chemically induced ; Neoplasm Metastasis ; Neoplasm Recurrence, Local ; Neutropenia ; chemically induced ; Remission Induction ; Survival Rate

OBJECTIVETo investigate the expression profile of microRNA-21 in human cholangiocarcinoma tissues and to validate its bona fide targets in human cholangiocarcinoma cells.

METHODSThe expression profile of microRNA-21 in human cholangiocarcinoma tissues and cholangiocarcinoma cell line, QBC939, was evaluated by using real-time PCR analysis. The bona fide targets of microRNA-21 were analyzed and confirmed by dual luciferase reporter gene assay and western blot, respectively. The expressional correlation of microRNA-21 and its targets was probed in human cholangiocarcinoma tissues by using real-time PCR, locked nucleic acid in situ hybridization (LNA-ISH), and immunohistochemistry analysis.

RESULTSReal-time PCR analysis revealed that microRNA-21 expression depicted a significant up-regulation in human cholangiocarcinoma tissues about 5.6-fold as compared to the matched normal bile duct tissues (P<0.05). The dual luciferase reporter gene assay revealed endogenous microRNA-21 in cholangiocarcinoma cell line, QBC939, inhibited the luciferase reporter activities of wild-type PTEN (P<0.01) and PDCD4 (P<0.05) and had no this effect on mutated PTEN and PDCD4. Moreover, loss of microRNA-21 function led to a significant increase of PTEN and PDCD4 protein levels in QBC939 cells. Elevated microRNA-21 levels were accompanied by marked reductions of PTEN and PDCD4 expression in the same cholangiocarcinoma tissue.

CONCLUSIONmicroRNA-21 expression is up-regulated in human cholangiocarcinoma and PTEN, PDCD4 are direct effectors of microRNA-21.

Apoptosis Regulatory Proteins ; genetics ; Bile Duct Neoplasms ; genetics ; pathology ; Bile Ducts, Intrahepatic ; metabolism ; pathology ; Cell Line, Tumor ; Cholangiocarcinoma ; genetics ; pathology ; Female ; Gene Expression Profiling ; Gene Expression Regulation, Neoplastic ; drug effects ; Humans ; Male ; MicroRNAs ; genetics ; physiology ; Middle Aged ; PTEN Phosphohydrolase ; genetics ; RNA-Binding Proteins ; genetics ; Transfection

Intrahepatic cholangiocarcinoma (ICC), a malignant tumor derived from the intrahepatic bile duct epithelium, has a poor prognosis and is refractory to conventional chemotherapy and radiation therapy. Thus, there is an urgent need to develop new effective therapeutic strategies for this disease. We previously found that L1 cell adhesion molecule (L1CAM) plays an important role in tumor progression of ICC, and we generated a murine mAb, A10-A3 (IgG1), that binds to the Ig1 domain of L1CAM. In the present study, we further characterized A10-A3, constructed a chimeric A10-A3 antibody (cA10-A3) containing the constant regions of human IgG1, and evaluated the therapeutic potential in a human ICC xenograft nude mice model. The affinities (K D) of A10-A3 and cA10-A3 for soluble L1CAM were 1.8 nM and 1.9 nM, respectively, as determined by competition ELISA. A10-A3 inhibited L1CAM homophilic binding and was slowly internalized into the tumor cells, but it did not significantly inhibit proliferation of ICC cells in vitro. cA10-A3 mediated antibody-dependent cell-mediated cytotoxicity in vitro and displayed anti-tumor activity in the ICC animal model. These results suggest that the humanized A10-A3 antibody may have potential as an anticancer agent for the treatment of ICC.
Animals ; Antibodies, Monoclonal/genetics/*immunology ; Antibody-Dependent Cell Cytotoxicity ; Bile Ducts, Intrahepatic/drug effects/immunology/pathology ; CHO Cells ; Cell Adhesion/drug effects ; Cell Proliferation/drug effects ; Cholangiocarcinoma/*drug therapy/immunology/pathology ; Cricetinae ; Disease Models, Animal ; Endocytosis/drug effects ; Humans ; Immunoglobulin G/genetics/*immunology ; Liver Neoplasms/*drug therapy/immunology/pathology ; Mice ; Mice, Nude ; Neoplasm Transplantation ; Neural Cell Adhesion Molecule L1/genetics/*immunology/metabolism ; Protein Binding ; Protein Structure, Tertiary ; Recombinant Fusion Proteins/immunology/metabolism/*therapeutic use