Levosimendan Reverses Cardiac Malfunction and Cardiomyocyte Ferroptosis During Heart Failure with Preserved Ejection Fraction via Connexin 43 Signaling Activation

In recent decades, the occurrence of heart failure with preserved ejection fraction (HFpEF) has outweighed that of heart failure with reduced ejection fraction by degrees, but few drugs have been demonstrated to improve long-term clinical outcomes in patients with HFpEF. Levosimendan, a calcium-sensitizing cardiotonic agent, improves decompensated heart failure clinically. However, the anti-HFpEF activities of levosimendan and underlying molecular mechanisms are unclear. In this study, a double-hit HFpEF C57BL/6N mouse model was established, and levosimendan (3 mg/kg/week) was administered to HFpEF mice aged 13 to 17 weeks. Different biological experimental techniques were used to verify the protective effects of levosimendan against HFpEF. After four weeks of drug treatment, left ventricular diastolic dysfunction, cardiac hypertrophy, pulmonary congestion, and exercise exhaustion were significantly alleviated. Junction proteins in the endothelial barrier and between cardiomyocytes were also improved by levosimendan. Among the gap junction channel proteins, connexin 43, which was especially highly expressed in cardiomyocytes, mediated mitochondrial protection. Furthermore, levosimendan reversed mitochondrial malfunction in HFpEF mice, as evidenced by increased mitofilin and decreased ROS, superoxide anion, NOX4, and cytochrome C levels. Interestingly, after levosimendan administration, myocardial tissue from HFpEF mice showed restricted ferroptosis, indicated by an increased GSH/GSSG ratio; upregulated GPX4, xCT, and FSP-1 expression; and reduced intracellular ferrous ion, MDA, and 4-HNE levels. Regular long-term levosimendan administration can benefit cardiac function in a mouse model of HFpEF with metabolic syndromes (namely, obesity and hypertension) by activating connexin 43-mediated mitochondrial protection and sequential ferroptosis inhibition in cardiomyocytes.