[Association between A-kinase anchor proteins 12 methylation and recurrence of hepatocellular carcinoma].

Objective: To study the association between the AKAP12 promoter methylation and recurrence of hepatocellular carcinoma. Methods: A total of 142 primary liver cancer patients underwent surgery in department of Hepatobiliary surgery in Peking University Cancer Hospital from 2003 to 2009 were selected as subjects in the survey; with the inclusion criteria as hepatocellular carcinoma, no cancer cells were observed in the surgical margin(SM) samples. All patients had neither lymph nor distant metastasis at the time of surgery, and receiving complete follow-up data for at least 3 years. By the end of May 2014, a total of 75 patients had relapsed of whom 71 died and there were no lost. All samples were acquired from the frozen surgical tissues. Genomic DNA was extracted using phenol/chloroform method and performed bisulfite modification following with polymerase chain reaction (PCR). AKAP12 methylation in hepatoma and the corresponding SM samples from 142 patients was determined by denature high-performance liquid chromatography (DHPLC) and bisulfite clone sequencing. Kaplan-Meier and Cox proportion hazard regression model were used to identify the factors related to the survival time. Results: In 142 cases, 125 patients (88.0%) were male and 17 (12.0%) cases were female. The median age was 52.5 years, ranging from 34 years to 76 years. AKAP12 methylation-positive rate was significantly higher in hepatomas than SMs (54.9% vs. 10.2%, P<0.001). Patients with AKAP12 methylation-positive had less risk of the recurrence (HR=0.62, 95%CI: 0.39-0.99); with tumor diameter more than 5 cm (HR=1.53, 95%CI: 1.00-2.50),portal vein invasion(HR=4.53, 95% CI:2.69-7.64) increased the recurrence risk. Moreover, portal vein invasion had a higher risk of death (HR=2.98, 95% CI: 1.73-4.98). Conclusion: There was significant association between AKAP12 DNA methylation and low risk of recurrence and long progression-free survival of hepatocellular carcinoma patients.

Destruction of parotid glands affects nitrate and nitrite metabolism.

The role of salivary glands in nitrate and nitrite metabolism is poorly understood. The aim of the present study was to investigate the effect of parotid gland ablation on dynamic metabolism of nitrate and nitrite in miniature pigs. The parotid glands of 5 healthy miniature pigs were bilaterally ablated by methyl violet. Concentrations of nitrate and nitrite of whole saliva, serum, and urine samples were analyzed by high-performance liquid chromatography. Results showed that bilateral ablation of the parotid glands led to a significant decrease of nitrate secretion from blood to saliva (P < 0.05) and thus low nitrite levels. Dysfunction of the parotid glands temporarily increased the systemic level of nitrate in miniature pigs after nitrate loading. This study suggests that the parotid glands play an important role in the balance of nitrate and nitrite levels in both whole saliva and the body.

Confirmation of N-(nitrosomethyl) urea as a nitrosourea derived by nitrosation of fish sauce.

N-(nitrosomethyl) urea (NMU) was characterized in the carcinogenic nitrosated fish sauce (NFS) with high performance liquid chromatography (HPLC)-photohydrolysis-pyrolysis-thermal energy analysis recently. We used HPLC-electronic spray ionization-mass spectrometry and HPLC-diode array detection to confirm NMU in NFS further. It was observed that the corresponding chromatographic fraction of NMU of NFS showed the same mass spectrum (m/z 64, 102, and 145) and ultraviolet-absorbance (lambda max = 230 nm) as those of authentic NMU. These results confirmed that the component of NFS was NMU.

[Induction of gastric adenocarcinoma in Wistar rats following neonatal exposure to MNNG and neoplastic transformation assay of DNA of the induced cancer].

Adenocarcinoma, adenoma and dysplasia of the stomach of adult Wistar rats were induced following administration of MNNG for 10 days by gavage when they were new-born. The incidence of the three types of pathological lesions at the dose of 0.4 mg MNNG per rat being 39%, 50% and 100% respectively. Induction of gastric adenocarcinoma is dose-dependent. The incidence of adenocarcinoma in male rats being significantly higher then that of female rats (P < 0.02). 23 of 33 (70%) induced cancers were found in the gastric antrum. By the use of DNA cotransfection technique and the assay of carcinogenicity in the Balb/c nude mice, it was also found that the DNA of 4 of the 6 induced gastric carcinomas could transform NIH/3T3 cells into malignant cells, an indication that the induced cancer DNA contains transforming gene.

Effect of diallyl trisulfide on induction of UDS by mutagenic drugs in primary rat hepatocytes.

Most of anticancer drugs are mutagenic. A possible exception is diallyl trisulfide (DAT), a component of garlic. It is an antimutagenic anticancer chemical although it is mainly used as antibiotic. Its modifying effect on induction of UDS by mutagenic mitomycin C (MMC), cyclophosphamide (CP) and cis-diamine dichloroplatin (DDP) was investigated with the UDS assay in the primary cultures of Wistar rat hepatocytes (hpc) using the autoradiographic technique. Results showed that 1.0-4.0 nmol/ml of DAT did not induce UDS and that MMC, CP and DDP resulted in a significant induction of dose-dependent UDS. DAT enhanced induction of UDS by these drugs. A dose-effect relationship was observed between dose of DAT and enhancement of induction of UDS. However, the mechanism of the enhancement is not clear.

[Modulation of mutagenic drug-induced unscheduled DNA synthesis (UDS) in primary rat hepatocytes by diallyl trisulfide].

Most anticancer drugs are mutagenic/carcinogenic. A possible exception is diallyl trisulfide (DAT), a component of garlic, which inhibits growth of transplantable tumors in vitro and in vivo and mutagenicity and carcinogenicity of genotoxic agents. It is an antimutagenic anticancer chemical. Its modulating effect on induction of UDS by mutagenic mitomycin C (MMC), cyclophosphamide (CP), and cis-diamine dichloroplatin (DDP) was investigated with the assay in primary cultures of Wistar rat hepatocytes by autoradiographic technique. Results showed that MMC (1-10 mumol/L), CP (0.316-3.16mmol/L), and DDP(3.16-31.6mumol/L) resulted in a significant induction of dose-dependent UDS and that DAT (0.5-4.0 mumol/L) significantly enhanced induction of UDS by MMC, CP and DDP while DAT itself did not. A dose-response relation was also observed between the dosage of DAT and the enhancement of induction of UDS. Hepatocellular enzymes for metabolic activation of indirect mutagens may not be involved in the enhancement of UDS-induction since DAT also increased UDS level induced by direct mutagen DDP. DAT promotes UDS induction probably by increasing repair of damaged 4-DNA. DAT, an anti-infection antibiotic, may be used in cancer chemotherapy to alleviate the adverse side effects of chemotherapeutic agents with mutagenic/carcinogenic activities.

Role of nitrosamides in the high risk for gastric cancer in China.

Gastric cancer is the leading cause of death from cancer in China. Samples of fish sauce, a traditional seasoning, were collected in the high-risk area for gastric cancer in the Fuzhou area, Fujian Province. When fish sauce samples were nitrosated at pH 2.0, direct mutagenicity and high contents of N-nitrosamide were detected (30.9-78.0 microM); the N-nitrosamide content of three samples of fish sauce made in Guangdong and purchased from a market outside Fujian were low (2.1-6.0 microns). When the nitrosated fish sauce extract was given to newborn rats by gavage, dysplasia and adenocarcinoma were induced in the glandular stomach in the 4th and 16th experimental week, respectively. N-Nitrosamides were also found in fasting gastric juice from patients with chronic gastritis in the high-risk area of Putian. The mean concentration of total N-nitrosamides in the extracts correlated with the severity of gastritis in the stomach. These findings indicate that N-nitrosamides may play an important role in causing gastric cancer in China.

[Effect of beta-carotene on sister chromatid exchanges induced by MNNG and aflatoxin B1 in V79 cells].

In order to elucidate the mechanism of tumor prevention of beta-carotene, its effect on sister chromatid exchanges (SCE) induced by MNNG in cultured V79 cells, under condition free of the enzyme system to convert beta-carotene into vitamin A, was studied. It was found that SCE level was significantly increased by high doses beta-carotene (10(-5)-10(-4) M) and the enhancement of SCE was restored to its original level by addition of alpha-tocopherol (final concentration 2 micrograms/ml). This may be due to the latter inhibiting the oxidation of beta-carotene and reducing the amount of oxidated carotene, which is toxic for cultured cells. Combination of beta-carotene and alpha-tocopherol at low doses inhibited SCE induced by MNNG (P less than 0.05) but no protective activity was observed when used separately. It was also found that beta-carotene (2 x 10(-7) M) and retinol (16 micrograms/ml) inhibited SCE induced by aflatoxin B1, which is activated by S-9 mixture. The present data clearly show that the antitumor activity of beta-carotene may be attributed to both itself and its degraded compound vitamin A, and may take part in the initiation of carcinogenesis. Combination of beta-carotene and other cancer preventive drugs is more effective and safer than it used individually.