Selective loss of Cx40 and conduction abnormalities underlie enhanced atrial fibrillation susceptibility in a HFpEF model in male mice

Figures

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  FIGURE 1. Phenotypic and echocardiographic characterization of HFpEF mice induced by HFD+L‐NAME Mice subjected to a high‐fat diet combined with L‐NAME administration (HFD+L‐NAME) exhibited heart failure with preserved ejection fraction (HFpEF) phenotype characterized by diastolic dysfunction with preserved systolic function. (a) Experimental protocol for induction of the HFpEF model. (b–e) Comparison of body weight (b), heart rate (c), systolic blood pressure (d) and diastolic blood pressure (e) between chow diet‐fed control mice and HFD+L‐NAME mice (Chow diet, n = 16; HFD+L‐NAME, n = 16). (f, g) Heart weight‐to‐tibial length (HW/TL) ratios in the ventricles (f) and left atrium (g) (Chow diet, n = 9; HFD+L‐NAME, n = 8). (h–k) Echocardiographic parameters. Left ventricular ejection fraction (LVEF) (h), left ventricular end‐diastolic diameter (LVDd) (i), left ventricular anterior wall thickness (LVAWth) (j) and left ventricular posterior wall thickness (LVPWth) (k) assessed by transthoracic echocardiography (TTE) (Chow diet, n = 8; HFD+L‐NAME, n = 8). (l) Representative TTE Doppler images. Top panels show transmitral E wave in chow diet–fed control (left) and HFD+L‐NAME mouse (right). Bottom…

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  FIGURE 2. Enhanced atrial arrhythmogenicity and increased AF inducibility in HFpEF mice. HFpEF mice induced by HFD+L‐NAME exhibited atrial electrical remodeling and markedly increased vulnerability to atrial fibrillation (AF) compared with chow diet‐fed controls. (a) Representative surface electrocardiograms from chow diet–fed controls and HFD+L‐NAME mice. (b–d) Quantitative analysis of P wave duration (b), PR interval (c), and QRS duration (d) (Chow diet, n = 9; HFD+L‐NAME, n = 9). (e, f) Representative surface electrocardiograms during transesophageal atrial pacing showing the absence of AF induction in chow diet mice (e) and successful AF induction in HFD+L‐NAME mice (f). (g, h) Comparison of AF inducibility, defined as the percentage of mice in which AF lasting ≥1 s was induced (g), and AF duration (h) between groups (left: Chow diet, n = 21; right: HFD+L‐NAME, n = 18). (h) Comparison of AF duration between groups (left: Chow diet, n = 3; right: HFD+L‐NAME, n = 11). AF induction rates were analyzed using Fisher's exact test, and AF duration was analyzed using the Mann–Whitney U test.

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  FIGURE 3. Impaired atrial conduction and increased conduction heterogeneity revealed by optical mapping. Optical mapping demonstrated slowed atrial conduction and increased conduction heterogeneity in HFpEF mice induced by HFD+L‐NAME. (a) Representative raw optical fluorescence images of the heart obtained prior to activation map reconstruction. The left panel shows a wide‐field view including the left atrium and ventricle, and the right panel shows a magnified view of the LA region. (b) Representative action potential (AP) traces recorded from the atrial myocardium in chow diet‐fed mice (blue) and HFD+L‐NAME mice (red). (c) Representative atrial action potential duration (APD) map, AP rise time map, isochronal activation map, and conduction velocity (CV) map from chow diet‐fed control mice (top) and HFD+L‐NAME mice (bottom). (d–f) Quantitative comparisons of APD at 90% repolarization (APD90), AP rise time, and CV at pacing cycle lengths (PCLs) of 60, 80, 100, and 120 ms (blue: Chow diet–fed controls; red: HFD+L‐NAME mice) (n = 7 per group). (g–i) Spatial heterogeneity of APD90, AP rise time, and CV expressed as the coefficient of variation (CoV) (blue: Chow diet–fed controls; red:…

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  FIGURE 4. Selective downregulation of connexin 40 expression in the atria of HFpEF mice. Connexin 40 (Cx40) expression was selectively and significantly reduced in the atria of HFpEF mice induced by HFD+L‐NAME. (a) Relative mRNA expression levels of inflammatory markers, fibrotic markers, and gap junction–related genes in atrial tissue assessed by quantitative PCR (n = 7 per group). (b, c) Representative western blot images and densitometric quantification of Cx40 and Cx43 protein expression in left atrial tissue from chow diet‐fed control mice and HFD+L‐NAME mice. (n = 6 per group). GAPDH was used as a normalized control. Data are presented as mean ± SD. Statistical comparisons were performed using unpaired t‐tests with Welch's correction.

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  FIGURE 5. Preserved atrial structure and reduced Cx40 immunoreactivity in HFpEF mice Histological analysis revealed preserved atrial structure with reduced connexin 40 (Cx40) immunoreactivity in HFpEF mice induced by HFD+L‐NAME. (a) Representative Masson's trichrome–stained sections of the left atrium from chow diet‐fed control (40×, top) and HFD+L‐NAME mice (40×, bottom). (b) Quantitative comparison of fibrotic area in the left atrium based on Masson's trichrome staining (n = 3 per group). (c) Representative immunohistochemical staining for Cx40 in the left atrium. Low‐magnification images (10×, left panels) and high‐magnification images (100×, right panels) are shown for each group. Red arrows indicate the lateralization of Cx40. Data are presented as mean ± SD. Statistical comparisons were performed using unpaired t‐tests with Welch's correction. p < 0.05 was considered statistically significant.