Abstract: OBJECTIVE： This study aimed to screen for methylation changes at MGMT CpG sites and their biological significance in blood lymphocytes from healthy adults who were exposed to short-term PM_2.5. METHODS： Thirty healthy male undergraduates from Hebei Medical University participated in a 35-day panel study， during which daily PM_2.5 exposure was assessed based on individual time-activity patterns. MGMT methylation was quantified by pyrophosphate sequencing of DNA extracted from peripheral blood lymphocytes at low and high PM_2.5 exposure time points. Additionally， urinary 1-hydroxypyrene (1-OHP)， 8-hydroxy- 2'-deoxyguanosine (8-OHdG)， and plasma tumor necrosis factor-alpha (TNF-α) levels were analyzed. RESULTS： The average ambient PM_2.5 concentration for the three days before the high exposure (110.09 μg/m³) was 2.32 times higher than at low exposure (47.37 μg/m³)，while urinary 1-OHP levels，indicating internal exposure，were 1.73 times higher (P<0.01). Urinary 8-OHdG and plasma TNF-α levels were 1.18-fold and 1.66-fold higher，respectively，than those under low exposure (all P<0.01). Increase in oxidative DNA damage and inflammatory response was accompanied by a decrease in the methylation levels of MGMT CpG sites 1， 3，4，5，6，and 10 by 37.31%，42.60%，19.69%，15.29%，34.95% and 32.12%，respectively (all P<0.05). Hypomethylation at CpG sites 1，3，and 10 was negatively correlated with urinary 1-OHP (β≥-0.40，all P<0.05)，8-OHdG (β≥-0.46，all P<0.05)，and TNF-α (β≥-0.26，all P<0.05). Altered methylation at these hot CpG sites was negatively correlated with the moving average concentrations of PM_2.5 (lag0-2) (β≥-0.53，all P< 0.05). CONCLUSION： MGMT hypomethylation modifications at hot CpG sites 1，3 and 10 were found to be associated with both external and internal PM_2.5 exposure as well as biological effects，indicating their potential as biomarkers for PM_2.5 exposure in the population.

Key words: short-term PM_2.5 exposure, panel study, MGMT gene, DNA methylation, specific CpG sites