Sex-specific associations between exposure to metal mixtures and mitochondrial DNA copy number: A repeated-measures...

Few repeated-measures studies with multiple time points have evaluated the sex-specific association between mitochondrial DNA copy number (mtDNAcn) and long-term exposure to metal mixtures. We conducted a repeated-measures study with three time points and 2550 observations in the manganese-exposed workers healthy cohort. Specifically, the blood concentrations of 15 metals and mtDNAcn were measured in 2012, 2017, and 2021. We employed a machine learning approach (GLMMLASSO) to select the metals most associated with mtDNAcn and used Bayesian kernel machine regression to examine their joint effects. In cross-sectional analyses (2550 visits), statistical methods consistently showed Calcium (Ca) was positively associated with mtDNAcn among overall visits (β = 0.292 in the linear mixed-effects model (LMM)) and dominated the positive overall effects of magnesium, Ca, titanium, iron, nickel, rubidium on mtDNAcn. An inverted "U"-shaped exposure-response curve between Ca and mtDNAcn appeared in males but not in females. Interaction analysis showed the association between Ca and mtDNAcn was significantly modified by gender. In repeated-measures analyses (807 visits), we explored the tendency for metals and mtDNAcn to change over three time points; the results confirmed the cross-sectional analyses. In overall visits, the positive association between Ca and mtDNAcn remained significant (β = 0.207 in LMM). For metal selection, Ca was identified as the predictor for mtDNAcn, and an inverted "U"-shaped exposure-response curve was found with mtDNAcn in males. Our findings reveal a consistent positive association between Ca and mtDNAcn with sex-dependent heterogeneity and suggest Ca may mitigate mitochondrial dysfunction induced by other metals.