Research Low-Level Environmental Cadmium Exposure Is Associated with DNA Hypomethylation in Argentinean Women Mohammad Bakhtiar Hossain,1,2 Marie Vahter,3 Gabriela Concha,4 and Karin Broberg 1 1Department of Laboratory Medicine, Division of Occupational and Environmental Medicine, Lund University, Lund, Sweden; 2International Centre for Diarrhoeal Disease Research, Bangladesh, Dhaka, Bangladesh; 3Institute of Environmental Medicine, Karolinska Institutet, Stockholm, Sweden; 4Risk and Benefit Assessment Department, National Food Agency, Uppsala, Sweden Background : Cadmium, a common food pollutant, alters DNA methylation in vitro. Epigenetic effects might therefore partly explain cadmium’s toxicity, including its carcinogenicity; however, human data on epigenetic effects are lacking. o Bjectiv : We evaluated the effects of dietary cadmium exposure on DNA methylation, consider­ ing other environmental exposures, genetic predisposition, and gene expression. Methods : Concentrations of cadmium, arsenic, selenium, and zinc in blood and urine of non­ smoking women (n = 202) from the northern Argentinean Andes were measured by inductively coupled mass spectrometry. Methylation in CpG islands of LINE-1 (long interspersed nuclear element­1; a proxy for global DNA methylation) and promoter regions of p16 [cyclin­dependent kinase inhibitor 2A (CDKN2A)] and MLH1 (mutL homolog 1) in peripheral blood were measured by bisulfite polymerase chain reaction pyrosequencing. Genotyping (n = 172) for the DNA (cytosine­ 5)­m­ethyltransferase 1 gene ( DNMT1 rs10854076 and rs2228611) and DNA (cytosine5­)­m­ethyl­ transferase 3 beta gene (DNMT3B rs2424913 and rs2424932) was performed with Sequenom iPLEX GOLD SNP genotyping; and gene expression (n = 90), with DirectHyb HumanHT­12 (version 3.0). r esult : Cadmium exposure was low: median concentrations in blood and urine were 0.36 and 0.23 µg/L, respectively. Urinary cadmium (natural log transformed) was inversely associated with LINE-1 methylation (β = –0.50, p = 0.0070; β = –0.44, p = 0.026, adjusted for age and coca chew­ ing) but not with p16 or MLH1 methylation. Both DNMT1 rs10854076 and DNMT1 rs2228611 polymorphisms modified associations between urinary cadmium and LINE-1 (pv­alues for interaction in adjusted models were 0.045 and 0.064, respectively). The rare genotypes demonstrated stronger hypomethylation with increasing urinary cadmium concentrations. Cadmium was inversely associated with DNMT3B (rS = –0.28, p = 0.0086) but not with DNMT1 expression (rS = –0.075, p = 0.48). c onclusi : Environmental cadmium exposure was associated with DNA hypomethylation in peripheral blood, and DNMT1 genotypes modified this association. The role of epigenetic modifica­ tions in cadmiuma­ssociated diseases needs clarification. k ey w ords : cadmium, DNMT1, DNMT3B, epigenetic, genotype, LINE-1, MLH1, p16, pyro­ sequencing. Environ Health Perspect 120:879–884 (2012). http://dx.doi.org/10.1289/ehp.1104600 [Online 1 March 2012] Cadmium is a widespread environmental pollutant associated with adverse health effects, including cancer, kidney damage, low bone-mineral density, bone fractures, and probably impaired early-life development (Engström A et al. 2011; Järup and Åkesson 2009; Kippler et al. 2011). The general popu- lation is exposed to cadmium mainly through the diet, in particular, through cereals and vegetables, and through smoking. Potential mechanisms of cadmium toxicity described so far include interactions with sulfhydryl groups and endocrine disruption, as well as oxidative stress (Hartwig 2010; Johnson et al. 2003; Monroe and Halvorsen 2006; Xu et al. 2011; Yu et al. 2010). DNA methylation is essential for embryo- genesis and for maintaining cell-lineage– specific gene expression throughout life. Dysregulation of epigenetic processes, such as DNA methylation, may lead to impaired child- hood development or chronic diseases later in life, including cancer (Faulk and Dolinoy 2011; Feinberg 2007; Robins et al. 2011). In carcinogenesis, global DNA methylation is generally decreased, whereas specific regions of DNA that contain tumor suppressor genes often show hypermethylation (Chalitchagorn et al. 2004; Deng et al. 2006; Gaudet et al. 2003; Robertson and Jones 2000). In vitro studies have indicated that cadmium may interfere with epigenetic processes: Both gene- specific DNA hypermethylation with gene silencing of p16INK4a [CDKN2A (cyclin- dependent kinase inhibitor 2A)], RASSF1 [Ras association (RalGDS/AF-6) domain family member 1], and MT1 (metallothionein 1) genes, and global DNA hypomethylation have been reported (Benbrahim-Tallaa et al. 2007; Huang et al. 2008; Jiang et al. 2008; Martinez-Zamudio and Ha 2011; Takiguchi et al. 2003). However, there are no reports on the epigenetic effects of cadmium exposure in humans. The aim of this study was to evaluate whether environmental cadmium exposure, mainly through the normal diet, may affect DNA methylation, both of repetitive genetic Environmental Health Perspectives • volume 120 | number 6 | June 2012 elements (as a proxy for global methylation) and of specific genes. Materials and Methods Participants. We studied 202 women living in the Andean plateau (3,800 m above sea level) in northern Argentina, an area that has minimal industrial or vehicle-derived pollu- tion. Most of the study participants (161) were from the village of San Antonio de los Cobres, which has about 5,000 inhabitants. The 41 remaining participants were from small surrounding villages. The study area is known to have elevated concentrations of arsenic (Vahter et al. 1995), but not cadmium (Concha et al. 2010), in the drinking water. The study individuals were recruited with the assistance of local medical personnel, except in the small mining village of Tolar Grande, where we went from house to house to explain the study and invited the adults to participate. We included only women because men were often away from home for longer periods for work and were therefore likely to be exposed to a different variety of environmental hazards. Furthermore, women may be more susceptible to cadmium exposure than are men because of their increased gastrointestinal cadmium absorption when iron stores are low (Berglund et al. 1994), as is common in women of child- bearing age. Only four study participants smoked cigarettes, and one reported drinking alcohol, but almost half (46.5%) reported that they chewed coca leaves. As reported before, interviews with the study participants revealed that almost all women drank exclusively public water and that their diets consisted mainly of corn, beans, chicken, and pork (Broberg et al. 2011). Only three women reported taking Address correspondence to K. Broberg, Department of Laboratory Medicine, Division of Occupational and Environmental Medicine, Lund University, Klinikgatan 21, SE-22185 Lund, Sweden. Telephone: 46046173819. Fax: 46046143207. E-mail: karin. broberg_palmgren@med.lu.se Supplemental Material is available online (http:// dx.doi.org/10.1289/ehp.1104600). We thank the Department of Oncology, Lund University, for technical assistance. This research was supported by the Swedish Council for Working Life and Social Research, Karolinska Institutet, and Kungliga fysiografiska sällskapet, European Union (FP6; “PHIME” FOOD-CT-2006- 016253). The authors declare they have no actual or potential competing financial interests. Received 10 October 2011; accepted 1 March 2012. 879