A novel murine model of atrial fibrillation by diphtheria toxin-induced injury

Abstract

The treatment of atrial fibrillation (AF) continues to be a significant clinical challenge. While genome-wide association studies (GWAS) are beginning to identify AF susceptibility genes (Gudbjartsson et al., Nature, 2007, 448, 353-357; Choi et al., Circ. Res., 2020, 126, 200-209; van Ouwerkerk et al., Circ. Res., 2022, 127, 229-243), non-genetic risk factors including physical, chemical, and biological environments remain the major contributors to the development of AF. However, little is known regarding how non-genetic risk factors promote the pathogenesis of AF (Weiss et al., Heart Rhythm, 2016, 13, 1868-1877; Chakraborty et al., Heart Rhythm, 2020, 17, 1,398-1,404; Nattel et al., Circ. Res., 2020, 127, 51-72). This is, in part, due to the lack of a robust and reliable animal model induced by non-genetic factors. The currently available models using rapid pacing protocols fail to generate a stable AF phenotype in rodent models, often requiring additional genetic modifications that introduce potential sources of bias (Schüttler et al., Circ. Res., 2020, 127, 91-110). Here, we report a novel murine model of AF using an inducible and tissue-specific activation of diphtheria toxin (DT)-mediated cellular injury system. By the tissue-specific and inducible expression of human HB-EGF in atrial myocytes, we developed a reliable, robust and scalable murine model of AF that is triggered by a non-genetic inducer without the need for AF susceptibility gene mutations.

Keywords: amiodarone; atrial fibrillation; diphtheria toxin; non-genetic cause; sarcolipin (SLN).

FIGURE 1

Design of murine model of atrial fibrillation (A). Illustration of the generation of mouse model and experimental scheme of DT treatment. DT consists of subunits A (blue) and B (red) linked by disulfide bridges. DT binds to hHB-EGF specifically expressed on the cell surface of atrial myocytes of Sln+/Cre; R26+/DTR mice. Upon endocytosis of receptor-ligand complex, A subunit translocates into the cytosol, ribosylates host EF2, and inactivates protein synthesis, resulting in mild damage in atrial muscle. Figure drawn using BioRender software. (B). Representative ECGs of two control mice at Day 18 and 21 and three AF mice at Day 17, 21 and 21 post-DT, respectively. (C). Kaplan-Meyer curve of animals free from AF after DT injection. DT was injected at Day 0 and 14. (n = 70)

FIGURE 2

Progression of atrial fibrillation after induction (A). Representative telemetry ECG indicating time in AF versus normal sinus rhythm (n = 4). DT was injected at Day 0. This mouse started to show paroxysmal AF at Day 8. The duration of AF gradually prolonged over time. NSR, normal sinus rhythm. (B). Masson’s trichrome staining of the atrial tissue of control (left) and AF mouse (right) at Day 0 (pre-DT) to Day 540 post-DT injection. Note severe fibrosis in AF mouse at Day 540. Scale bar = 500 μm. Right panel shows the quantification of fibrotic area (n = 3, each; *p < 0.05).

FIGURE 3

Cardioversion by i. p. injection of amiodarone. Amiodarone was injected at Day 28 post-DT injection. Representative of 9 out of 11 mice injected with DT that were successfully reverted to sinus rhythm.