The Expression And Promoter Hypermethylation Status Of MGMT, RASSF1A In Gastric Cancer

Objective:To investigate hypermethylation of MGMT(O6一methvlguanine DNAmethyltransferase) and RASSFlA (ras-association domain family1A) genes in gastriccarcinoma tissues and paired surgical marginal normal gastric tissues,and analyzingthe dynamic change of hypermethylation from gastric carcinoma tissues and pairedsurgical marginal normal gastric tissues. We want to investigate the functions ofhypermethylation of MGMT and RASSF1A genes in human gastric cancer．Gastriccancer,one of the most common tumors worldwide．Global gastric cancer mortality inall cancer mortality in the second place．An increasing number of genes that areinactivated by CpG island hypermethylation have been reported in gastric cancer，involving tumor suppressor，cell—cycle regulator，tissue-invasion-related and DNAmismatch repair genes．Methods:（1）the Municipal Hospital of Qingdao，2012to2013surgical resection，and thepathologic examination confirmed60cases of gastric cancer specimens，pairedsurgical marginal normal gastric tissues which were concluded above5cm between gastric cancer specimens in60cases． Abstract DNA from these specimensrespectively,then modificate them by sodium sulfite, at last methylation-specific PCRwas used to amplificate the modificated DNA.Methylation status of the MGMT andRASSF1A genes in60gastric carcinoma tissues and60paired surgical marginalnormal gastric tissues were detected using methylation-specific PCR. Analyzing therelationship from MGMT、RASSF1A between gastric cancer syntheticly.(2) Reverse transcript RNA which was abstracted from gastric carcinomatissues and paired surgical marginal normal gastric tissues respectively to cDNA,thenmodificate them by PCR. Real-time RT-PCR was used to detect the expression ofmRNA in MGMT and RASSF1A.（3）The immunohistochemical method was used to detect the expression ofgastric carcinoma tissues and paired surgical marginal normal gastric tissues inMGMT and RASSF1A，and results was analysed of their relationship with gastriccancer．Experimental data were used by statistical analysis package SPSSl7．0,enumeration data applied χ2test，group applied t test,P <0．05as statisticallymeaningful criteria．Results:（1）According to methylation specific PCR,gastric cancer，the positive rates ofpromoter methylation of the MGMT in45.00%(27/60) higher than8.33%(5/60) ofnormal tissue. Gastric cancer， the positive rates of promoter methylation of theRASSF1A in64.70%(37/60) higher than6.67%(4/60) of normal tissue. The positiverates of promoter methylation of the MGMT and RASSF1A genes were significantlyhigher in gastric cancer than in normal tissue(both P<0.05).（2）The positive rates of MGMT mRNA expression in gastric cancer were38.3%(23/60) lower than those in normal tissues96.67%(58/60) by Real-timeRT-PCR．The positive rates of RASSF1A mRNA expression in gastric cancer were20.00%(12/60)lower than those in normal tissues100%(60/60) by Real-timeRT-PCR． The two were significantly different (both P<0.05). （3）Immunohistochemistry was used to detect the expression of MGMT andRASSF1A. MGMT protein expression in gastric cancer31.61%（19/60）was lower thannormal tissue90.00%（54/60），RASSF1A protein expression in gastric cancer30.00%（18/60）was lower than normal tissue91.67%（55/60），the two were significantlydifferent (P<0．05)．Conclusion:（1）Methylation-specific PCR,Real-time RT-PCR and immunohistochemistrywere used to detect the expression of MGMT and RASSF1A that hypermethylation ofCpG island and loss of the protein expression in MGMT and RASSF1A genes wereexisted in gastric cancer, which was one of significant ways in effluence of gastriccancer.（2）The mRNA expression of MGMT and RASSF1A in gastric cancer was lowerthan normal tissue,it possibly plays a key role in carcinogenesis of gastric cancer．...