Metallothionein Alleviates Glutathione Depletion–Induced Oxidative Cardiomyopathy through CISD1-Dependent Regulation of Ferroptosis in Murine Hearts

ABSTRACT

This study was designed to discern the effect of heavy scavenger metallothionein on glutathione (GSH) deprivation-evoked cardiac anomalies and mechanisms involved with an emphasis on ferroptosis. WT and cardiac metallothionein transgenic mice received GSH synthase inhibitor buthionine sulfoximine (BSO, 30 mM in drinking water) for 14 days prior to assessment of myocardial morphology and function. BSO evoked cardiac remodeling and contractile anomalies including cardiac hypertrophy, interstitial fibrosis, enlarged left ventricular chambers, deranged ejection fraction, fraction shortening, cardiomyocyte contractile capacity, intracellular Ca²⁺ handling, sarcoplasmic reticulum Ca²⁺ reuptake, loss of mitochondrial integrity (mitochondrial swelling, loss of aconitase activity), mitochondrial energy deficit, carbonyl damage, lipid peroxidation, ferroptosis and apoptosis. Metallothionein itself did not affect myocardial morphology and function although it mitigated BSO-provoked myocardial anomalies, loss of mitochondrial integrity and energy, and ferroptosis. Immunoblotting revealed downregulated SERCA2a, GPx4, the ferroptosis suppressing iron sulfur domain 1 (CISD1) and mitochondrial regulating GSK-3β phosphorylation with elevated p53, MHC-β isozyme, IκB phosphorylation, and SLC7A11 as well as unchanged SLC39A1, SLC1A5 and FSP1 following BSO challenge, all of which except glutamine transporter SLC7A11 and p53 were abrogated by metallothionein. Inhibition of CISD1 using pioglitazone nullified GSH-offered benefit against BSO-induced cardiomyocyte ferroptosis, contractile and intracellular Ca²⁺ derangement. Taken together, these findings support a regulatory modality for CISD1 in the impedance of ferroptosis in metallothionein-offered protection against GSH depletion-evoked cardiac aberration.