Quantification of cyclooxygenase-2 methylation in gastric mucosa by denaturing high performance liquid chromatography assay.
- Author:
Xiao-Rui NIE
1
;
Yang ZHANG
;
Kai-Feng PAN
;
Wen-Qing LI
;
Wei-Cheng YOU
;
Lian ZHANG
Author Information
- Publication Type:Journal Article
- MeSH: Cell Line, Tumor; Chromatography, High Pressure Liquid; methods; CpG Islands; Cyclooxygenase 2; genetics; DNA Methylation; Gastric Mucosa; metabolism; pathology; Humans; Promoter Regions, Genetic; Stomach Diseases; genetics; metabolism; pathology
- From: Chinese Journal of Preventive Medicine 2010;44(1):54-57
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVETo setup a quantitative assay for detection of cyclooxygenase-2 (COX-2) methylation in human gastric mucosa samples.
METHODSA 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.
RESULTSThe 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%).
CONCLUSIONThe 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.
