Valproic Acid Inhibits RhoA-Mediated Vascular Smooth Muscle Cell Contraction

Vascular smooth muscle cells (VSMCs) play an important role in regulating vessel diameter and blood pressure. Dysregulation of VSMC contraction contributes to the development of coronary and post-subarachnoid hemorrhagic (SAH) vasospasms. We investigated the molecular mechanisms by which valproic acid (VPA) inhibits Ras homolog family member A (RhoA)-mediated VSMC contraction in rat VSMCs and isolated aortas.

In rat VSMCs, western blot analyses, quantitative real-time reverse transcription-polymerase chain reaction, ectopic expression of the constitutively active (CA)-RhoA gene or wild-type (WT)-histone deacetylase (HDAC) 5 gene, and inhibitor studies were performed. Active RhoA-GTP levels and Rho-associated protein kinase activity in VSMCs were also measured. We performed a phenylephrine (PE)-induced aortic contraction assay using isolated rat aortas, as well as post hoc analyses of an endothelium-dependent aortic relaxation assay using aortas from VPA-administered mice.

VPA decreased the phosphorylation of the myosin light chain at Ser19 (p-MLC-Ser¹⁹) in a dose- and time-dependent manner. Interestingly, VPA significantly decreased RhoA mRNA and protein expression, as well as the active RhoA-GTP level. Furthermore, ectopic expression of CA-RhoA gene almost completely reversed VPA-inhibited p-MLC-Ser¹⁹. VPA markedly increased the acetylation levels of histone 3 (H3K9ac/K14ac). VPA and sodium butyrate but not valpromide significantly decreased the expression levels of both RhoA and p-MLC-Ser¹⁹ in VSMCs. However, this decrease was not reversed by overexpression of the WT-HDAC5 gene, indicating that HDAC5 was not responsible for this decrease. Consistent with the in vitro results, VPA attenuated PE-induced aortic contraction, decreased RhoA and p-MLC-Ser¹⁹ expression, and increased H3K9ac/K14ac levels in isolated rat aortas. The post hoc analysis revealed that the VPA-inhibited RhoA pathway accounted for 30% of the total aortic relaxation induced by VPA.

This study showed that VPA inhibits RhoA-mediated VSMC and vessel contraction by decreasing RhoA expression, which is mediated by the inhibitory action of VPA on HDACs. These results suggest that VPA may be useful in the treatment and prevention of spastic vascular diseases, including coronary and post-SAH vasospasms.