Quantification of cyclooxygenase-2 methylation in gastric mucosa by denaturing high performance liquid chromatography assay
Nie, X.-R.; Zhang, Y.; Pan, K.-F.; Li, W.-Q.; You, W.-C.; Zhang, L.
Zhonghua Yu Fang Yi Xue Za Zhi 44(1): 54-57
2010
ISSN/ISBN: 0253-9624 PMID: 20388365 Document Number: 647345
To setup a quantitative assay for detection of cyclooxygenase-2 (COX-2) methylation in human gastric mucosa samples. A standard analysis system was established by denaturing high performance liquid chromatography (DHPLC) under the condition of 55 degrees C oven temperature and a linear acetonitrile gradient (4.0/min). While, a total of 10 cases of gastric biopsy samples were detected for methylation status of COX-2. The complete methylated human promyelocytic leukemia cells (HL-60) and unmethylated gastric cancer cell line (MGC803) were used as positive and negative control. The proportion of the methylated copies of COX-2 was calculated according to the peak heights of methylated (M) and unmethylated (U) COX-2 in same PCR amplicon. The formula was Y = 1.0608 x M/(M + U), R(2) = 0.9894. Among 10 biopsy samples, the proportions of methylated copies of COX-2 in 2 cases of dysplasia were higher than superficial gastritis and chronic atrophy gastritis (24.5%, 18.4% vs 7.6%, 9.6%). The methylation of COX-2 promoter CpG islands can be detected in human gastric mucosa samples by quantitative DHPLC assay, which could be used in the population-based study of precancerous gastric lesions.