Abstract B35: Altered DNA methylation at imprinted insulin-like growth factor-2 (IGF2) resulting from heavy metal exposure in vitro and in vivo in infants from the Newborn Epigenetic STudy (NEST).

Background: Early life exposure to heavy metals, including lead, has adverse consequences on health and development but the mechanism(s) underlying these effects are unclear. Epigenetic deregulation resulting from lead exposure may underlie these effects, and imprinted genes may be particularly vulnerable since imprinting is established and maintained in large part through DNA methylation. The best studied imprinted regulatory region is the insulin-like growth factor 2-H19 (IGF2) domain that is regulated by at least two differentially methylated regions (DMRs). Altered methylation at the IGF2 DMRs has been associated with obesity, neurological development disorders, and cancer. Data from the Newborn Epigenetics STudy (NEST), which is focused on the influence of early life environmental exposures on children's health. This study has indicated that epigenetic alterations resulting from these environmental exposures are often sex-specific and exacerbated in African Americans as compared to whites. Objective: We sought to determine if in vitro exposure to lead alters DNA methylation at two regulatory DMRs is important to imprinting and expression of IGF2 and to assess how these results compared to the effects of lead exposure in the NEST cohort. Methods: Immortalized normal human embryonic kidney cells (HEK293) were exposed to lead acetate (0-25 ug/dL) for 48-72 hours followed by purification of genomic DNA. Bisulfite modification of HEK293 and NEST cohort DNA specimens was followed by pyrosequencing to quantify DNA methylation. The North Carolina State Laboratory for Public Health performed the blood lead level testing as part of a routine check-up for children between the ages of 2-3 years old. The NEST cohort participants were born between 2005-2008 at obstetrics-care facilities in Durham County, North Carolina. For this study we used a sub-population of participants whose mother lived at the same address since conception in order to more accurately determine early and consistent environmental lead exposure. Analysis of variance (ANOVA) tests were done on the in vitro data to examine the difference between mean values of the lead treatment conditions. For the NEST samples we used generalized linear models in a sample of n=182 participants to examine aberrant methylation at the IGF2 DMRs, in children exposed to environmental lead from conception to age 2-3 years old. Results: In our in vitro model of early environmental lead exposure we saw a 3-5% decrease (p= 0.006) in IGF2 DMRs methylation profiles with increasing lead acetate concentration. Similar to the in vitro studies, in the NEST cohort we observed 4% decrease (p <0.0001) in methylation at the H19 DMRs of IGF2/H19 gene in female children with blood lead levels between 2-4 ug/dL and an 8% decrease (p<0.0001) in methylation for female infants with blood lead levels equal to or higher than 5 ug/dL of lead. Conclusions: Lead exposure was associated with aberrant methylation at the IGF2/H19 DMR in vitro and in a cohort study of young children exposed to lead in their home environment pre and post-natal. The implications of these findings in vitro and in vivo suggest a link between lead exposure and altered methylation at the IGF2/H19 DMRs. Larger studies are required. Citation Format: Monica D. Nye, Adriana C. Vidal, Marilie D. Gammon, Cathrine Hoyo. Altered DNA methylation at imprinted insulin-like growth factor-2 (IGF2) resulting from heavy metal exposure in vitro and in vivo in infants from the Newborn Epigenetic STudy (NEST). [abstract]. In: Proceedings of the Fifth AACR Conference on the Science of Cancer Health Disparities in Racial/Ethnic Minorities and the Medically Underserved; 2012 Oct 27-30; San Diego, CA. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2012;21(10 Suppl):Abstract nr B35.