关键词: O⁶-甲基鸟嘌呤DNA甲基转移酶, 甲基化, 表达, 视网膜母细胞瘤

Abstract: Objective To detect the methylation status of the promoter of O6 methylguanine DNA meghyltransferase (MGMT) gene and the expression level of MGMT mRNA and protein. Design Experimental Study. Participants 20 paraffin embedded RB tissues, 4 RB cell lines (Y79, SO-Rb50, SO-Rb50/VCR, WERI-RB1). Methods A series of 20 paraffin embedded RB tissue samples and 4 RB cell line were subjected to mehtylation-specific PCR (MSP) analysis to evaluate the methylation status of the MGMT promoter. Further, the expression of MGMT mRNA and protein were studied by Realtime PCR and Western Blot. The clinical data (e.g. gender, age of onset) and pathological features (e.g. the realm of tumor encroachment, neovascularization in iris, NVI) were also collected. Main Outcome Measures Methylation status of the MGMT promoter, mRNA and protein expression level of MGMT, data of clinical and pathological features. Results Among the 20 paraffin embedded RB tissue samples, 12 cases (60%) were partially/completely methylated, the remaining 8 cases (40%) showed MGMT promoter unmethylated. As to the 4 RB cell lines, Y79, SO-Rb50, SO-Rb50/VCR were partially methylated, while WERI-RB1 was unmethylated. MGMT mRNA and protein were expressed in all 4 RB cell lines. The mRNA expression level of WERI-RB1（1.000±0.040） was higher than the other 3 cell lines（Y79，0.617±0.026；SO-Rb50，0.356±0.020；SO-Rb50/VCR，0.389±0.017）, the differences among which were statistically significant (P<0.05). The protein expression of WERI-RB1（1.506±0.493） was slightly higher than the other 3 cells lines（Y79，1.388±0.304；SO-Rb50，1.495±0.212； SO-Rb50/VCR，1.406±0.547）, the differences among which were not statistically significant (P>0.05). Conclusions There was a comparatively high MGMT promoter methylation rate in RB pathologic samples and cell lines, which had a corresponding effect to the expression level of MGMT mRNA and protein. (Ophthalmol CHN, 2012, 21: 121-126)

Key words: MGMT, methylation, expression, retinoblastoma