Inhibition of DNA Methylation Attenuates Low-Dose Cadmium-Induced Cardiac Contractile and Intracellular Ca2+ Anomalies

Cadmium is a human carcinogen with unfavorable health impact probably associated with its DNA methylation property. Recent data suggest that environmental cadmium exposure is associated with incidence of myocardial infarction and peripheral arterial disease. Nonetheless, the effect of chronic cadmium exposure on cardiac contractile function remains elusive. This study was designed to examine the impact of low-dose cadmium exposure on cardiac contractile function and intracellular Ca2+ homeostasis. Adult male mice were exposed to cadmium for 4 weeks with or without the DNA methylation inhibitor (5-aza-2’-deoxyctidene, 5-AZA). Cardiac contractile and intracellular Ca2+ properties were analyzed including echocardiographic left ventricular parameters, fractional shortening (FS), peak shortening amplitude (PS), maximal velocity of shortening/relengthening (± dL/dt), time-to-PS (TPS), time-to-90% relengthening (TR90), electrically-stimulated increase of intracellular Ca2+ and intracellular Ca2+ decay. Our results revealed that cadmium exposure depressed FS, PS, ± dL/dt and electrically-stimulated rise in intracellular Ca2+ without affecting TPS, TR90, intracellular Ca2+ level and decay rate, the effects of which were significantly attenuated or nullified by 5-AZA. Cadmium exposure led to overt interstitial fibrosis (collagen deposition), the effect of which was mitigated by 5-AZA. Western blot analysis showed unchanged expression of ICAM-1, TNF-α and Cleaved caspase-3 in response to cadmium exposure and/or 5-AZA treatment, suggesting a relatively minor role of pro-inflammatory cytokines and apoptosis in cadmium- and 5-AZA-induced cardiac responses. Taken together, our data demonstrated for the first time direct cardiac depressant effect following cadmium exposure, which may be rescued by DNA methylation inhibition. Keywords: Cadmium, cardiac, contraction, intracellular Ca2+ INTRODUCTION Cadmium is a ubiquitous environmental toxin with the major known sources of exposure encompassing emissions from industrial activity and waste management, intake of foods (e. g. leafy vegetables, grains, organ meats and crustaceans) and exposure to cigarette smoke.1–3 Accumulation of cadmium leads to tissue toxicity exerting a broad spectrum of biological actions possibly through induction of inflammation and apoptosis. Cadmium toxicity and carcinogenesis has been extensively examined in organs such as testis, liver, lung and kidney.4 For instance, our laboratory recently demonstrated persistent liver and renal damage at more than 40 weeks after a 4-week exposure to low-dose cadmium in rodents. 5–7. Nonetheless, cardiac toxicity, if any, and the underlying mechanisms remain poorly elucidated. It has been reported that cadmium exerts its adverse cardiovascular effects by promoting atherosclerosis as well as disadvantageous functional and metabolic changes in the heart.1 In particular, high susceptibility of heart to cadmium was reported in a human 15-year following up study,8 in line with the fact that heart is a relatively sensitive organ due to its low antioxidant capacity. Similarly, high susceptibility of the heart to cadmium was also found in experimental animals.9, 10 Prenatal exposure to cadmium is known to lead to change of cardiovascular function during adulthood.1, 11, 12 Environmental exposure to cadmium has been found to be associated with an increased prevalence of heart failure1 although the precise toxic response of cadmium exposure on myocardial function remains elusive. More recent data from the National Health and Nutrition Examination Survey have strongly suggested that cadmium, at substantially low levels of exposure, remains a rather important determinant of all-cause cardiovascular mortality in certain populations of U.S. adults.13 It is thus pertinent to examine the toxic effect of cadmium on myocardial function, if any, and the mechanism of action involved. Traditional concepts for the mechanisms for cadmium toxicity include its generation of reactive oxygen or nitrogen species (ROS and RNS), inhibition of antioxidants, and impairment of DNA repair enzymes.14–16 However, a recent study has shown that even though a soy-based diet was capable of ameliorating cadmium-induced oxidative stress and damage, it failed to prevent against cadmium-elicited cardiac damage,17 suggesting possible involvement of other mechanisms including epigenetic modulating properties in cadmium-elicited toxicity.18–24 Epigenetics refers to the heritable regulation of gene expression via modification of chromosomal components without an alteration in the nucleotide sequence of the genome. Such modifications include methylation of genomic DNA as well as acetylation, methylation, phosphorylation, ubiquitination, and SUMOylation of core histone proteins. Epigenetic changes in the heart following environmental stress and life style changes have been implicated in the pathogenesis of cardiac remodeling and dysfunction.25–29 Recently we have demonstrated the induction of global DNA hypermethylation in the liver of rats and mice exposed to low-dose cadmium.7 Accordingly, we hypothesize that exposure to low-dose cadmium may cause cardiac global DNA methylation to modify certain key gene expression, leading to cardiac dysfunction. Therefore, the present study was designed to address the following novel and important questions: (1) whether exposure of low-dose cadmium elicits cardiac functional toxicity and involvement of DNA methylation in such pathological process; (2) the effect of DNA methylation inhibition on cadmium exposure-induced unfavorable cardiac responses, if any.