Abstract Body: Postoperative atrial fibrillation (poAF) is self-limited AF that occurs days after cardiac surgery in one-third of patients. The degree of perioperative increase in systemic inflammation, particularly involving interleukin (IL)-6, correlates with poAF risk. However, no studies to-date have identified the cell types involved nor mechanistically linked IL-6 signaling to fundamental arrhythmia mechanisms.
To study poAF, we developed a mouse model consisting of atrial pericardiectomy and transient aortic cross-clamping followed by intracardiac burst pacing on postoperative day three to assess poAF inducibility. Single-cell RNA sequencing of atrial non-myocytes revealed macrophages to be the most prominently altered atrial cell type – increased 2.4-fold (P<0.001) in mice with versus without poAF. Pseudotime trajectory analyses revealed IL-6 to be the top cytokine altered in macrophages, which we confirmed by western blot demonstrating a 1.7-fold (P=0.012) increase in atrial IL-6 protein in mice with versus without poAF. Indeed, both macrophage depletion and conditional knockout of IL-6Ra in macrophages decreased poAF inducibility by 5.2-fold (P=0.048) and 5.5-fold (P=0.027), respectively. Downstream inhibition using an FDA-approved STAT3 inhibitor also rescued poAF (6.0-fold decrease, P=0.022). At the single atrial cardiomyocyte (ACM) level, IL-6 challenge led to a 6.3-fold (P<0.001) increase in calcium (Ca²⁺) spark frequency after normalization to sarcoplasmic reticulum Ca²⁺ load, indicative of ryanodine receptor-2 (RyR2) dysfunction. Indeed, RyR2 phosphorylation at Ser2814, the target of Ca²⁺/calmodulin-dependent protein kinase II (CaMKII), was increased in the atria of mice with poAF, and CaMKII inhibition was sufficient to rescue arrhythmogenic Ca²⁺sparks in ACMs from mice with poAF. We corroborated our findings in human ACMs and atrial tissue to demonstrate translational relevance.
Taken together, we use an integrated approach incorporating mouse and human biochemical, functional, and single-cell sequencing data to demonstrate that infiltrating atrial macrophages, specifically IL-6Ra from macrophages, are fundamentally required for poAF development (Fig. 1). Molecularly, enhanced atrial IL-6 signaling led to arrhythmogenic RyR2 dysfunction through CaMKII. Our findings portend therapeutic utility by providing mechanistic evidence that targeting the IL-6 axis may be used for poAF prevention and treatment in a clinically amenable timeframe.