Exploring Carotenoid-Methylation Associations

Concentrations of plasma carotenoids are available in Supplementary Table S1. Plasma total carotenoid concentrations (µmol/L of plasma) were defined as the sum of α-carotene, β-carotene, β-cryptoxanthin, lutein, lycopene, and zeaxanthin concentrations. Regressions between normalized DNA methylation levels of all 472,245 CpG sites and total carotenoid concentrations adjusted for the family ID were computed using R software v2.14.1 (R Foundation for Statistical Computing; http://www.r-project.org) [36 ]. Plasma total carotenoid concentrations (independent variable) were used to predict DNA methylation levels (dependent variable). A p-value ≤ 0.05 was used to identify significant associations. Regressions were adjusted for family ID (fixed effect) to account for the familial structure. In order to maintain a more exploratory approach, the regressions were not adjusted for other confounding factors. Choices of linear model and confounding factors were made for comparison purposes with similar study by our group [37 (link)]. Weighted gene correlation network analysis (WGCNA) was performed with methylation levels of 20,687 CpG sites showing a significant association (p-value ≤ 0.05) with total carotenoid concentrations. This allowed evaluating co-methylation similarities only in CpG sites that are associated with carotenoids.