Over-expression of microRNA-145 drives alterations in β-adrenergic signaling and attenuates cardiac remodeling in heart failure post myocardial infarction

Figures

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  Figure 1. miR-145 attenuated structural remodeling after HF. (A) relative expression of miR-145 (n=3); (B) quantification of infarcted size (n=5); (C) statistical results of LV fibrosis (n=5); (D) representative imagine of HE and masson’s stain; (E) representative western blots; (F–H) quantitative analysis of the immunoreactive band displayed by bar graph (n=3). (a) sham group; (B) HF group; (C) AD-EGFP; (D) AD-miR-145 group; (E) M+C group; IS, infarcted size. Data are presented as mean ± SD.

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  Figure 2. Characters of ECG parameters. (A), typical ECG recording after HF; (B–G), statistical results of P duration, RR interval, PR interval, QRS interval, QT interval and QTc interval (n=6). Data are presented as mean ± SD.

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  Figure 3. MiR-145 depressed the electrophysiological susceptibility of HF. (A) representative MAP recording at a PCL=150ms, APD alternans and PES; (B–D) statistical results of APD90, APD alternans threshold and arrhythmia score (n≥5); PCL, pacing cycle length; L, longer APD; S, shorter APD. Data are presented as mean ± SD.

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  Figure 4. Representative images of M-mode echocardiography at baseline and 5min after ISO injection, respectively.

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  Figure 5. Role of miR-145 in alteration of β-adrenergic signaling. (A) Representative western blots. (B–G) quantitative analysis of the immunoreactive band displayed by bar graph (n=3). Data are presented as mean ± SD.

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  Figure 6. miR-145 increased ICa density and reduced the sensitivity of HF to β-adrenergic stimulation. (A) representative ICa traces recording; (B) ICa-voltage relationship of cardiomyocytes (n≥7); (C) summary of peak ICa (n≥7); (D) the steady-state ICa activation curves (n≥7); (E) the steady-state ICa inactivation curves (n≥7); (F) the recovery curves following inactivation (n≥7); (G) representative western blots of Cav1.2, p-CaMKII and CaMKII; (H–J) quantitative analysis of the immunoreactive band displayed by bar graph (n=3); (K) ICa-voltage relationship in presence of ISO for each group (n≥7); (L) summary of peak ICa in the presence of ISO (n≥7); (M) changes of peak ICa for each group (n≥7); (A) sham group; (B) HF group; (C) AD-EGFP; (D) AD-miR-145 group; (E) M+C group. Data are presented as mean ± SD for C, H, I, J, L and M. While for B, D, E, F and K, data are presented as mean ± SEM.

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  Figure 7. miR-145 ameliorated HF-induced defective in Ca2+ homeostasis. (A) representative imagines of resting fluorescence at baseline (The arrow marks cardiomyocyte stained with Rhod-2AM); (B) statistical analysis of F0 (n≥9); (C) representative imagines of cardiomyocytes sarcomere shortening from isolated HF hearts; (D) representative recording (F/F0) of steady-state Ca2+ transients followed by Ca2+ waves (The arrow marks Ca2+ wave); (E–H) statistical analysis of Ca2+ transients in systole and diastole (Baseline and in presence of ISO, n≥9); (I–J) statistical analysis of cell shortening (Baseline and in presence of ISO, n≥9). Data are presented as mean ± SD.

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  Figure 8. miR-145 ameliorated HF-induced defective in Ca2+ homeostasis. (A–B) statistical analysis of Ca2+ waves (Baseline and in presence of ISO, n≥9); (C) Representative western blots; (D–H) quantitative analysis of the immunoreactive band displayed by bar graph (n=3). (a) sham group; (B) HF group; (C) AD-EGFP; (D) AD-miR-145 group; (E) M+C group. Data are presented as mean ± SD.

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  Figure 9. Mechanisms of miR-145-mediated cardioprotective effects against MI. miR-145 promotes the expression of β2AR via activation of Akt/CREB cascades and enhances β2AR-Gi activity, which in turn restricts β1/2AR-Gs signaling and leads to reduced response to β-adrenergic stimuli. Furthermore, MI-induced hyperactivation of CaMKII promote RyR2-mediated Ca2+ release and result in activation of NCX1, which finally leads to delayed depolarization. At the meantime, CaMKII suppressed SERCA2a and Cav1.2 expression via HDAC4 and NF-κB (p65) pathway respectively, thus impair Ca2+ re-uptake, excitation-contraction coupling and cardiac performance. However, miR-145-mediated inhibition on CaMKII expression partially reversed the related disadvantages.