Structural, Pro-Inflammatory and Calcium Handling Remodeling Underlies Spontaneous Onset of Paroxysmal Atrial Fibrillation in JDP2-Overexpressing Mice

Figures

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  Figure 1. Conduction defects and atrial fibrillation (AF) detected in electrocardiogram (ECG) recordings. ECGs were recorded over 30 min in non-anesthetized mice after 1, 2, 3, 4 and 5 weeks of JDP2 overexpression. (A) After 2 weeks the PQ-interval was prolonged and QRS-complexes broadened. (n = 6–11, * p < 0.05 vs. WT). (B) Paroxysmal AF in 5-week-old JDP2 mice. ECG segments with sinus rhythm are presented (left and right white panel), and P-waves are indicated by small arrows. By contrast, other episodes (middle yellow panel) represent AF. The loss of P-waves and the irregularity of RR-intervals define an episode of AF. In derivation L1, the section in the red rectangle was magnified in order to clearly show the presence of rapid electrical activity. (C) Detection of second-degree AV Block Mobitz type II episodes in different magnifications (indicated in the red circles) after 4 and 5 weeks of JDP2 overexpression. P-waves are indicated by small arrows. Lead 1 from Einthoven is presented.

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  Figure 2. Hypertrophy and fibrosis were analyzed in atria after five weeks of JDP2 overexpression. (A) mRNA expression of the hypertrophic marker gene ANP was determined by real-time RT-PCR and compared to 18SrRNA as a housekeeping gene (n = 11–12, p < 0.05 vs. WT) and a cross-sectional area of isolated cardiomyocytes was calculated from 30–40 cells per preparation (values are means ± SD of 4 independent culture preparations). (B) mRNA expression of fibrotic marker genes was determined by real-time RT-PCR and compared to 18SrRNA as a housekeeping gene (n = 11–12, p < 0.05 vs. WT), and fibrotic areas were determined after staining atrial tissue sections with azan dye (values are means ± SD of 5–6 tissue sections). * Difference from WT, p < 0.05.

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  Figure 3. Atrial inflammation contributes to AF-associated electrical remodeling in JDP2-overexpressing mice. mRNA expression of the inflammatory marker genes in atria of mice overexpressing JDP2 for 5 weeks were determined by real-time RT-PCR and compared to 18SrRNA as a housekeeping gene (n = 7–12, * p < 0.05 vs. WT).

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  Figure 4. mRNA expression of connexins, β-adrenergic receptors and calcium-handling proteins. mRNA expression of connexins (Cx40, Cx37, Cx43), β-adrenergic receptors (Adrb1, Adrb2) and calcium-handling proteins (NCX, Cav1.2, Ryr2, PLB, Serca) in atria of mice overexpressing JDP2 for 5 weeks were determined by real-time RT-PCR and compared to 18SrRNA as a housekeeping gene (n = 11–12,* p < 0.05 vs. WT).

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  Figure 5. Expression and phosphorylation of sarcoplasmic reticulum (SR) calcium-handling proteins in atria from JDP2-overexpressing mice. Original Western blots (A,C) and summarized data (B,D) from 8–9 atria isolated from mice overexpressing JDP2 for 5 weeks vs. WT mice (p-values as indicated). Expression of SERCA2a, CSQ and RyR2 was analyzed, as well as phosphorylation of RyR2 at S2808. GAPDH was used as loading control for normalization. Note that the GAPDH signals shown in (A) for SERCA2a and in (C) for pS2808 are identical, as the proteins were derived from the same Western blot.