MGMT and MLH1 methylation in Helicobacter pylori-infected children and adults

AIM:To evaluate the association between Helicobacter pylori(H.pylori) infection and MLH1 and MGMT methylation and its relationship with microsatellite instability(MSI).METHODS:The methylation status of the MLH1 and MGMT promoter region was analysed by methylation specific methylation-polymerase chain reaction(MSPPCR) in gastric biopsy samples from uninfected or H.pylori-infected children(n = 50),from adults with chronic gastritis(n = 97) and from adults with gastric cancer(n = 92).MLH1 and MGMT mRNA expression were measured by real-time PCR and normalised to a constitutive gene(β actin).MSI analysis was performed by screening MSI markers at 4 loci(Bat-25,Bat-26,D17S250 and D2S123) with PCR;PCR products were analysed by single strand conformation polymorphism followed by silver staining.Statistical analyses were performed with either the χ 2 test with Yates continuity correction or Fisher's exact test,and statistical significance for expression analysis was assessed using an unpaired Student's t-test.RESULTS:Methylation was not detected in the promoter regions of MLH1 and MGMT in gastric biopsy samples from children,regardless of H.pylori infection status.The MGMT promoter was methylated in 51% of chronic gastritis adult patients and was associated with H.pylori infection(P < 0.05);this region was methylated in 66% of gastric cancer patients,and the difference in the percentage of methylated samples between these patients and those from H.pylori-infected chronic gastritis patients was statistically significant(P < 0.05).MLH1 methylation frequencies among H.pylori-infected and non-infected chronic gastritis adult patients were 13% and 7%,respectively.We observed methylation of the MLH1 promoter(39%) and increased MSI levels(68%) in samples from gastric cancer patients in comparison to samples from H.pylori-infected adult chronic gastritis patients(P < 0.001 and P < 0.01,respectively).The frequency of promoter methylation for both genes was higher in gastric cancer samples than in H.pylori-positive chronic gastritis samples(P < 0.05).The levels of MLH1 and MGMT mRNA were significantly reduced in chronic gastritis samples that were also hypermethylated(P < 0.01).CONCLUSION:In summary,MGMT and MLH1 methylation did not occur in earlier-stage H.pylori infections and thus might depend on the duration of infection.