Activation of protein kinase B rescues against thapsigargin-elicited cardiac dysfunction through regulation of NADPH oxidase and ferroptosis.

Endoplasmic reticulum (ER) stress is a known contributor to cardiac remodeling and contractile dysfunction. Although NADPH oxidase has been implicated in ER stress-induced organ damage, its specific role in myocardial complications resulting from ER stress remains unclear. This study aimed to investigate the possible involvement of NADPH oxidase in ER stress-induced myocardial abnormalities and to evaluate the impact of Akt constitutive activation on these myocardial defects. Mice with cardiac-specific overexpression of active mutant of Akt (Myr-Akt) and their wild-type (WT) littermates were treated with ER stress instigator thapsigargin (1 mg/kg, i. p. 72 hrs) before evaluating myocardial morphology and function. Our results noted that thapsigargin significantly impaired echocardiographic parameters and cell shortening indices, including elevated LVESD, decreased ejection fraction, fractional shortening, peak shortening, electrically-stimulated intracellular Ca release, and cardiomyocyte survival. These functional deteriorations were accompanied by upregulation of NADPH oxidase, O production, mitochondrial damage, carbonyl formation, lipid peroxidation, apoptosis, and interstitial fibrosis, with unchanged myocardial size. Constitutive Akt hyperactivation did not generate any response on myocardial morphology and function, although it greatly suppressed or nullified thapsigargin-induced myocardial remodeling and dysfunction. Thapsigargin also triggered dephosphorylation of Akt and its downstream signal GSK3β, along with development of ferroptosis, all of which were nullified by Akt hyperactivation. In vitro studies further revealed that thapsigargin provoked cardiomyocyte mechanical anomalies and lipid peroxidation, similar to in vivo results. These effects were reverted by inhibitors of NADPH oxidase and ferroptosis (apocynin and LIP1). Collectively, our data denote an important protective role for Akt hyperactivation in thapsigargin-evoked myocardial anomalies, likely through NADPH oxidase-mediated regulation of ferroptosis.