Abstract 5648: Epigenetic Basis of Nitrate Tolerance: Fundamental Role of Protein Acetylation

Abstract

INTRODUCTION : We recently demonstrated that nitric oxide (NO) plays a key role regulating epigenetic enzymes, including histone acetylases (HAT) and deacetylases (HDAC). In this context, we hypothesized that during nitrate tolerance the HDAC/HAT balance could be altered and that drugs capable of modulate HDAC and/or HAT function could have an impact on this alteration. METHODS & RESULTS : Adult male rats (n=6, each group) were treated for 72 hours with subcutaneous nitroglycerin (NTG) alone (50 mg/kg) or in combination with different HDAC inhibitors (DI) including trichostatin A (TSA, 0.6 mg/kg), suberoylanilide hydroxamic acid (SAHA, 0.6 mg/kg), MS-27–275 (0.6 mg/kg) and valproic acid (VPA, 5 mg/kg). Other animals received the HAT inhibitor anacardic acid (ANAC, 2.5 mg/kg) or equivalent amounts of solvent. Ex vivo experiments performed on phenylephrine-contracted aortic rings (AR) showed that the induction of tolerance (-logEC=5.9±0.2 in tolerant vs -logEC=7.6±0.1 in control) was reversed by TSA, SAHA (-logEC=7.6±0.1) and VPA (-logEC=7.4±0.2). ANAC treatment displayed value similar to nitrotolerant (-logEC=6.2±0.1). Moreover, our data revealed that in AR from controls animals a single NTG administration induced relaxation paralleled by a rapid increase in HAT activity. Nitrate tolerant rings, however, exhibited a higher HDAC activity and failed to up-regulate HATs’ function. Remarkably, in AR from animals treated with DI in combination with NTG, nitrate tolerance was prevented while animals treated with ANAC developed resistance to both arterial relaxing or contracting stimuli suggesting that the intracellular level of HAT activity is important for arterial adaptive responses. Mechanistically, in isolated smooth muscle cells the NTG increased myosin light chain (MLC) and smooth muscle actin (SMA) acetylation paralleled by a decreased MLC phosphorylation and contractile protein association. These effects were rescued to normal by DI treatment. CONCLUSIONS : an alteration in HAT/HDAC functional balance occurs during nitrate tolerance in which the content of acetylated contractile proteins is markedly reduce. DIs, preventing this effect, may be envisaged as novel tools for the treatment of nitrate tolerance.