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Objective O6-methylguanine-DNA methyltransferase(MGMT)is an effective DNA repair enzyme. The aberrant methylation of its promoter region CpG islands may be one of the important mechanisms of carcinogenesis. We studied the methylation status of MGMT gene and clinical significance in cholangiocarcinoma. Methods Target DNA was modified by sodium bisulfite, coverting all unmethylated, but not methylated, cytosines to uracil. Then, the promoter hypermethylation of MGMT gene and the expression of MGMT protein were detected by methylation-specific PCR and immunohistochemical staining. Results Aberrant methylation of MGMT gene was detected in 17 of 36(47.2%) in cholangiocarcinoma tissues. 21 cases showed negative immunoreactivities of MGMT protein. Among them, there were 14 cases with aberrant methylation of MGMT gene .15 cases showed positive immunoreactivities of MGMT protein, but only 3 cases with aberrant methylation of MGMT gene. There were statistically differences in the exent of differentiation, invasion and the MGMT protein expression between the methylated tissues and those unmethylated but not in pathologic type and lymph node metastasis. Conclusion Hypermethylation of the MGMT promoter is a frequency molecular event in cholangiocarcinoma and might be involved in carcinogenesis. The methylation status of MGMT gene may be helpful to predict prognosis in patients with cholangiocarcinoma. [ABSTRACT FROM AUTHOR]

Background: Death-associated protein kinase (DAPK) is a Ca2+/calmodulin-regulated Ser/Thr kinase which is involved in apoptosis. The aberrant methylation of its promoter region CpG islands may be one of the important mechanisms of carcinogenesis. We studied the relationship of methylation status and expression of the DAPK gene with the clinical findings in cholangiocarcinoma., Methods: Target DNA was modified by sodium bisulfite, coverting all unmethylated, but not methylated, cytosines to uracil, and subsequently detected by methylation-specific PCR. Moreover, mRNA expression of the DAPK gene was assessed by RT-PCR., Results: Aberrant methylation of the DAPK gene was detected in 11 (30.6%) of 36 tissue specimens of cholangiocarcinoma, and in 2 (5.6%) of 36 specimens of adjacent normal tissues. DAPK mRNA was not expressed in tumor and adjacent tissues with hypermethylation of the DAPK promoter. There were no statistical differences in the extent of differentiation and invasion, lymph node metastasis or pathologic type between the methylated and unmethylated tissues., Conclusions: The frequency of DAPK gene methylation in cholangiocarcinoma is high and it may offer an effective means for earlier auxiliary diagnosis of the malignancy. The DAPK gene is probably suppressed by methylation, and it could become resistant to apoptosis and immunological surveillance. The DAPK gene epigenetically affected by methylation may be associated with the carcinogenesis of cholangiocarcinoma.

Aim: To characterize the tumor suppressor gene p53 mutations and study the correlation of p53 gene mutation and the expression of P53 protein in cholangiocarcinoma., Methods: A total of 36 unselected, frozen samples of cholangiocarcinoma were collected. p53 gene status (exon 5-8) and P53 protein were examined by automated sequencing and immunohistochemical staining, combined with the clinical parameters of patients., Results: p53 gene mutations were found in 22 of 36 (61.1%) patients. Nineteen of 36 (52.8%) patients were positive for P53 protein expression. There were significant differences in extent of differentiation and invasion between the positive and negative expression of P53 protein. However, there were no significant differences in pathologic parameters between the mutations and non-mutations., Conclusion: The alterations of the p53 gene evaluated by DNA sequence analysis is relatively accurate. Expression of P53 protein could not act as an independent index to estimate the prognosis of cholangiocarcinoma.

Background: TMS1/ASC is a bipartite protein comprising two protein-protein interactive domains: pyrin (PYD) and caspase recruitment domain (CARD). Proteins containing these domains play pivotal roles in regulating apoptosis and immune response pathways. The absence of TMS1/ASC expression in some tumors is because methylation of the TMS1/ASC gene contributes to carcinogenesis and cancer development. We studied the methylation status of the TMS1/ASC gene and its clinical significance in cholangiocarcinoma., Methods: Target DNA was modified by sodium bisulfite, coverting all unmethylated, but not methylated, cytosines to uracil, and subsequently by a nested amplification with primers specific for methylated versus unmethylated DNA. The PCR product was detected by gel electrophoresis and combined with the clinical records of patients., Results: Aberrant methylation of the TMS1/ASC gene was detected in specimens of colorectal cancer tissues from 13 (36.1%) of 36 patients, and specimens of adjacent normal tissues from 3 patients (8.3%). No statistical differences were seen in the extent of differentiation and invasion, lymph node metastasis, and pathologic type between the methylated and unmethylated tissues (P>0.05)., Conclusions: The frequency of TMS1/ASC gene methylation in cholangiocarcinoma is high, but it is not related to pathologic changes. The TMS1/ASC gene is probably suppressed by methylation, and is resistant to apoptosis and immunological surveillance. The gene epigenetically affected in methylated tissues could be associated with carcinogenesis of cholangiocarcinoma.

Background: The outcome of patients with cholangiocarcinoma is poor. To evaluate the experience in the diagnosis and surgical treatment of cholangiocarcinoma, we investigated the status quo of diagnosis and treatment of cholangiocarcinoma in China., Method: The clinical data of 680 patients with cholangiocarcinoma treated at 8 hospitals from 1995 to 2001 were retrospectively analyzed with SPSS software package., Results: The incidence of the tumor was the highest in the age group of 60-65 years. Meanwhile, the incidence was higher in aged men than in aged women, with a male to female ratio of 1.39:1. Proximal cholangiocarcinoma was the commonest (41.6%) and distant cholangiocarcinoma the second (28.7%) in the 680 patients. B-mode ultrasonography for cholangiocarcinoma was performed in 80.3% of the patients. Non-traumatic examinations such as computed tomography (CT), magnetic resonance image (MRI) and magnetic resonance cholangiopancreatography (MRCP) were more widely used than that of traumatic examinations such as percutaneous transhepatic cholangiography (PTC) and endoscopic retrograde cholangiopancreatography (ERCP). The low- and middle-differentiation cancer of the proximal bile duct accounted for about 50%. Most of the patients suffered from late-stage cholangiocarcinoma. The resection rate of the tumor was low, and the rate of radical operation was only 21.6% (147/680)., Conclusions: Cholangiocarcinoma is common in the aged men. Its diagnosis and treatment have been improved, but little. Most patients are diagnosed as having late-stage cholangiocarcinoma at the time of outpatient clinic, and the rate of radical operation is low. Therefore, it is necessary to reinforce the early diagnosis and treatment of cholangiocarcinoma to improve the outcome after operation.

Background: The inhibitory effect of antisense oligodeoxynucleotide (asODN) on the replication and expression of hepatitis C virus (HCV) in vitro is not well elucidated. This study was to assess the effect of asODN on HCV in cholangiocarcinoma., Methods: The QBC939 cells transfected by a recombinant HCV containing HCV core gene cloned in vector of PBK-CMV (PBK-HCVC) were treated by 14-mers phosphorothioate ODN complementary to the HCV core genomic region. The variation of HCVmRNA level was detected by RT-PCR. Moreover, the inhibitory effect of asODN was observed in nude mice., Results: HCVmRNA was detected in transfected-QBC939 cells. The 14-mers complementary phosphorothioate ODN showed effective inhibition on HCVmRNA and unexpression HCVmRNA at 6 micromol/L. The tumorigenicity of the transfected-QBC939 cells incubated with asODN in nude mice was greatly inhibited., Conclusion: The results suggest a potential therapy of asODN for HCV infected cholangiocarcinoma.