Predictors of global methylation levels in blood DNA of healthy subjects : a combined analysis

Background: Estimates of global DNA methylation from repetitive DNA elements, such as Alu and LINE-1, have been increasingly used in epidemiological investigations because of their relative low-cost, high-throughput and quantitative results. Nevertheless, determinants of these methylation measures in healthy individuals are still largely unknown. The aim of this study was to examine whether age, gender, smoking habits, alcohol drinking and body mass index (BMI) are associated with Alu or LINE-1 methylation levels in blood leucocyte DNA of healthy individuals. Methods: Individual data from five studies including a total of 1465 healthy subjects were combined. DNA methylation was quantified by PCR-pyrosequencing. Results: Age [beta = -0.011% of 5-methyl-cytosine (% 5mC)/year, 95% confidence interval (CI) -0.020 to -0.001% 5mC/year] and alcohol drinking (beta = -0.214, 95% CI -0.415 to -0.013) were inversely associated with Alu methylation. Compared with females, males had lower Alu methylation (beta = -0.385, 95% CI -0.665 to -0.104) and higher LINE-1 methylation (-0.796, 95% CI 0.261 to 1.330). No associations were found with smoking or BMI. Percent neutrophils and lymphocytes in blood counts exhibited a positive (beta=0.036, 95% CI 0.010 to 0.061) and negative (beta = -0.038, 95% CI -0.065 to -0.012) association with LINE-1 methylation, respectively. Conclusions: Global methylation measures in blood DNA vary in relation with certain host and lifestyle characteristics, including age, gender, alcohol drinking and white blood cell counts. These findings need to be considered in designing epidemiological investigations aimed at identifying associations between DNA methylation and health outcomes