Biased activation of β₂-AR/Gi/GRK2 signal pathway attenuated β₁-AR sustained activation induced by β₁-adrenergic...

Heart failure is the terminal stage of many cardiac diseases, in which β₁-adrenoceptor (β₁-AR) autoantibody (β₁-AA) has a causative role. By continuously activating β₁-AR, β₁-AA can induce cytotoxicity, leading to cardiomyocyte apoptosis and heart dysfunction. However, the mechanism underlying the persistent activation of β₁-AR by β₁-AA is not fully understood. Receptor endocytosis has a critical role in terminating signals over time. β₂-adrenoceptor (β₂-AR) is involved in the regulation of β₁-AR signaling. This research aimed to clarify the mechanism of the β₁-AA-induced sustained activation of β₁-AR and explore the role of the β₂-AR/Gi-signaling pathway in this process. The beating frequency of neonatal rat cardiomyocytes, cyclic adenosine monophosphate content, and intracellular Ca²⁺ levels were examined to detect the activation of β₁-AA. Total internal reflection fluorescence microscopy was used to detect the endocytosis of β₁-AR. ICI118551 was used to assess β₂-AR/Gi function in β₁-AR sustained activation induced by β₁-AA in vitro and in vivo. Monoclonal β₁-AA derived from a mouse hybridoma could continuously activate β₁-AR. β₁-AA-restricted β₁-AR endocytosis, which was reversed by overexpressing the endocytosis scaffold protein β-arrestin1/2, resulting in the cessation of β₁-AR signaling. β₂-AR could promote β₁-AR endocytosis, as demonstrated by overexpressing/interfering with β₂-AR in HL-1 cells, whereas β₁-AA inhibited the binding of β₂-AR to β₁-AR, as determined by surface plasmon resonance. ICI118551 biasedly activated the β₂-AR/Gi/G protein-coupled receptor kinase 2 (GRK2) pathway, leading to the arrest of limited endocytosis and continuous activation of β₁-AR by β₁-AA in vitro. In vivo, ICI118551 treatment attenuated myocardial fiber rupture and left ventricular dysfunction in β₁-AA-positive mice. This study showed that β₁-AA continuously activated β₁-AR by inhibiting receptor endocytosis. Biased activation of the β₂-AR/Gi/GRK2 signaling pathway could promote β₁-AR endocytosis restricted by β₁-AA, terminate signal transduction, and alleviate heart damage.