HDAC Inhibition Reverses Preexisting Diastolic Dys...

Background: Diastolic dysfunction (DD) is associated with the development of heart failure (HF) and contributes to the pathogenesis of other cardiac maladies, including atrial fibrillation (AF). Inhibition of histone deacetylases (HDACs) has been shown to prevent DD by enhancing myofibril relaxation. Here, we addressed the therapeutic potential of HDAC inhibition in a model of established DD with preserved ejection fraction (EF). Methods: Four weeks following uninephrectomy (UNX) and implantation with deoxycorticosterone acetate (DOCA) pellets, when DD was clearly evident, one cohort of mice was administered the clinical-stage HDAC inhibitor ITF2357/Givinostat. Echocardiography, blood pressure measurements, and endpoint invasive hemodynamic analyses were performed. Myofibril mechanics and intact cardiomyocyte relaxation were assessed ex vivo. Cardiac fibrosis was evaluated by picrosirius red (PSR) staining and second harmonic generation (SHG) microscopy of left ventricular (LV) sections, RNA-sequencing of LV mRNA, mass spectrometry-based evaluation of decellularized LV biopsies, and atomic force microscopy (AFM) determination of LV stiffness. Mechanistic studies were performed with primary rat and human cardiac fibroblasts. Results: HDAC inhibition normalized DD without lowering blood pressure in this model of systemic hypertension. Surprisingly, in contrast to prior models, myofibril relaxation was unimpaired in UNX/DOCA mice. Furthermore, cardiac fibrosis was not evident in any mouse cohorts based on PSR staining or SHG microscopy. However, mass spectrometry revealed induction in the expression of more than one hundred extracellular matrix (ECM) proteins in LVs of UNX/DOCA mice, which correlated with profound tissue stiffening based on AFM. Remarkably, ITF2357/Givinostat treatment blocked ECM expansion and LV stiffening. The HDAC inhibitor was subsequently shown to suppress cardiac fibroblast activation, at least in part, by blunting recruitment of the pro-fibrotic chromatin reader protein, BRD4, to key gene regulatory elements. Conclusions: These findings demonstrate the potential of HDAC inhibition as a therapeutic intervention to reverse existing DD, and establish blockade of ECM remodeling as a second mechanism by which HDAC inhibitors improve ventricular filling. Additionally, our data reveal the existence of pathophysiologically relevant 'covert' or 'hidden' cardiac fibrosis that is below the limit of detection of histochemical stains such as PSR, highlighting the need to evaluate fibrosis of the heart using diverse methodologies.