CD44 regulates Epac1-mediated β-adrenergic-receptor-induced Ca2+-handling abnormalities: implication in cardiac arrhythmias

Abstract

Background: Sustained, chronic activation of β-adrenergic receptor (β-AR) signaling leads to cardiac arrhythmias, with exchange proteins directly activated by cAMP (Epac1 and Epac2) as key mediators. This study aimed to evaluate whether CD44, a transmembrane receptor mediating various cellular responses, participates in Epac-dependent arrhythmias.

Methods: The heart tissue from CD44 knockout (CD44^-/-) mice, cultured HL-1 myocytes and the tissue of human ventricle were used for western blot, co-immunoprecipitaiton and confocal studies. Line-scanning confocal imaging was used for the study of cellular Ca²⁺ sparks on myocytes. Optical mapping and intra-cardiac pacing were applied for arrhythmia studies on mice's hearts.

Results: In mice, isoproterenol, a β-AR agonist, upregulated CD44 and Epac1 and increased the association between CD44 and Epac1. Isoproterenol upregulated the expression of phospho-CaMKII (p-CaMKII), phospho-ryanodine receptor (p-RyR), and phospho-phospholamban (p-PLN) in mice and cultured myocytes; these effects were attenuated in CD44^-/- mice compared with wild-type controls. In vitro, isoproterenol, 8-CPT-cAMP (an Epac agonist), and osteopontin (a ligand of CD44) significantly upregulated the expression of p-CaMKII, p-RyR, and p-PLN; this effect was attenuated by CD44 small interfering RNA (siRNA). In myocytes, resting Ca²⁺ sparks were induced by isoproterenol and overexpressed CD44, which were prevented by inhibiting CD44. Ex vivo optical mapping and in vivo intra-cardiac pacing studies showed isoproterenol-induced triggered events and arrhythmias in ventricles were prevented in CD44^-/- mice. The inducibility of ventricular arrhythmias (VAs) was attenuated in CD44^-/- HF mice compared with wild-type HF controls. In patients, CD44 were upregulated, and the association between CD44 and Epac1 were increased in ventricles with reduced contractility.

Conclusion: CD44 regulates β-AR- and Epac1-mediated Ca²⁺-handling abnormalities and VAs. Inhibition of CD44 is effective in reducing VAs in HF, which is potentially a novel therapeutic target for preventing the arrhythmias and sudden cardiac death in patients with diseased hearts.

Keywords: CD44; Calcium leak; Cardiac arrhythmia; Exchange proteins directly activated by cAMP; Osteopontin; β-Adrenergic receptor.