HDAC5 inhibition attenuates ventricular remodeling and cardiac dysfunction

Figures

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  Fig. 1. Upregulation of HDAC5 in in ventricular remodeling of TAC mice and H9C2 cells exposed to Ang II (A-B) Representative Western blots and quantification showing HDAC5 levels in sham or TAC mice (n = 6 in each group). (C) Representative images of immunofluorescence staining of HDAC5 (red), c-TNT (green), and DAPI (blue) in the ventricular remodeling zone of mouse hearts in the sham group and 14 days after TAC. c-TNT: cardiomyocyte marker. Scale bar, 50 μm. (D-E) Representative Western blots and quantification showing HDAC5 levels in H9C2 cells in each group (n = 6 in each group). (F-G) Representative images of immunofluorescence staining and fluorescence intensity of HDAC5 in H9C2 cells in each group. Scale bar, 50 μm. n > 10 in each group. Data are presented as means ± SD, **P < 0.01

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  Fig. 2. LMK235 ameliorates cardiac dysfunction induced by TAC in vivo (A) Experimental procedure for injecting mice with LMK235. (B) Representative images of M-mode echocardiography of mice in each group. (C, D, E, F) Echocardiographic parameters include left ventricular ejection fraction (LVEF), left ventricular shortening fraction (LVFS), left ventricular end-diastolic diameter (LVIDd), and left ventricular end-systolic diameter (LVIDs) (n = 6 for each group). (G) Representative Doppler echocardiographic images of mitral flow in each group. (H) Mitral inflow velocities (E/A). (I) Isovolumic relaxation time (IVRT) (n = 3 for each group). Data are presented as means ± SD, *P < 0.05; **P < 0.01

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  Fig. 3. LMK235 attenuates TAC-induced ventricular remodeling and Ang II-induced H9C2 cell hypertrophy (A) Representative images of cardiac appearance for each group. (B) Heart weight/body weight (HW/BW) (n = 6 for each group). (C) Hematoxylin and eosin staining in horizontal sections of heart tissue. Scale bar, 50 μm. (D, E) Representative images of Masson’s trichrome staining in each group and statistical analysis of the fibrotic area. Scale bar, 50 μm. (n = 6 for each group). (F, G) Representative images of WGA staining in each group and statistical analysis of cardiomyocytes cross-sectional area. Scale bar, 50 μm. (n > 50 for each group). (H-K) Western blot results and statistical analysis of BNP, ANP, and β-MHC expression in mice (n = 3 for each group). (L-O) Western blot results and statistical analysis of BNP, ANP, and β-MHC expression in H9C2 cells (n = 3 for each group). Data are presented as means ± SD, *P < 0.05; **P < 0.01

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  Fig. 4. LMK235 reverses TAC and Ang II induced HDAC5 expression increase and cytoplasmic translocation (A-C) Western blot results and statistical analysis of HDAC5 and MEF2A expression in mice (n = 3 for each group). (D) Representative images of immunohistochemical staining of HDAC5 in each group. Scale bar, 50 μm. (E-G) Western blot results and statistical analysis of HDAC5 and MEF2A expression in H9C2 cells (n = 3 for each group). (H) Representative images of immunofluorescence staining of HDAC5 in H9C2 cells in each group. Scale bar, 50 μm. Data are presented as means ± SD, *P < 0.05; **P < 0.01

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  Fig. 5. HDAC5 inhibition activates ERK/EGR1 signaling to regulate MEF2A expression during cardiomyocyte hypertrophy (A-C) Western blot results and statistical analysis of EGR1, ERK, and p-ERK expression in mice (n = 3 for each group). (D-F) Western blot results and statistical analysis of EGR1, ERK, and p-ERK expression in H9C2 cells (n = 3 for each group). (G-H) Western blot results and statistical analysis of HDAC5, MEF2A, EGR1, ERK, and p-ERK expression in H9C2 cells after transfection with HDAC5 siRNA (n = 3 for each group). Data are presented as means ± SD, *P < 0.05; **P < 0.01

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  Fig. 6. HDAC5 regulates MEF2A expression by influencing the ERK/EGR1 signaling pathway Ventricular remodeling induces cytoplasm translocation of HDAC5. Inhibition of ERK/EGR1 by HDAC5 results in a marked upregulation of MEF2A, which consequently contributes to the promotion of TAC-induced ventricular remodeling and Ang II-induced hypertrophy in H9C2 cells