Metallothionein inhibits myocardial apoptosis in copper-deficient mice: Role of atrial natriuretic peptide

Dietary copper restriction causes heart hypertrophy in animal models. Several studies have indicated that this cardiomyopathy is mediated by oxidative stress. Metallothionein (MT), a low molecular weight and cysteine-rich protein, functions in protecting the heart from oxidative injury. We therefore used a cardiac-specific MT-overexpressing transgenic mouse model to test the hypothesis that MT inhibits copper deficiency-induced heart hypertrophy. Dams of both transgenic pups and non-transgenic littermates were fed a copper-adequate or copper-deficient diet, starting on the fourth day post-delivery, and the weanling mice were continued on the dams' diets until they were killed. Heart hypertrophy developed in copper-deficient pups by the fourth week of the combined pre- and post-weaning feeding and aggressively progressed until the end of the experiment (6 weeks), MT overexpression did not prevent the occurrence of heart hypertrophy, but inhibited the progression of this cardiomyopathy, which correlated with its suppression of cardiac lipid peroxidation. Corresponding to the progression of heart hypertrophy, myocardial apoptosis and atrial natriuretic peptide (ANP) production in the left ventricle were detected in non-transgenic copper-deficient mice; these effects were significantly suppressed in transgenic copper-deficient mice. Measurement of apoptosis by TUNEL assay and Annexin V-FITC confocal microscopy in primary cultures of cardiomyocytes revealed that ANP was largely responsible for the myocyte apoptosis and that MT inhibited ANP-induced apoptosis. The data clearly demonstrate that elevation of MT in the heart inhibits oxidative injury and suppresses the progression of heart hypertrophy in copper deficiency, although it does not block its initiation. The results suggest that MT inhibits the transition from heart hypertrophy to failure by suppressing apoptosis through inhibition of both cardiac ANP production and its apoptotic effect.