Park Ji-Kwang, Kim Seo-Hyeon, Lee Hansol, Hwang Yun-Jin, Cho Du-Hyong
Department of Pharmacology, College of Medicine, Yeungnam University, Daegu, Korea.
J Korean Med Sci. 2025 Aug 25;40(33):e199. doi: 10.3346/jkms.2025.40.e199.
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 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.
血管平滑肌细胞（VSMC）在调节血管直径和血压方面发挥着重要作用。VSMC收缩失调会导致冠状动脉痉挛和蛛网膜下腔出血（SAH）后血管痉挛的发生。我们研究了丙戊酸（VPA）抑制大鼠VSMC和离体主动脉中Ras同源家族成员A（RhoA）介导的VSMC收缩的分子机制。
在大鼠VSMC中，进行了蛋白质免疫印迹分析、定量实时逆转录-聚合酶链反应、组成型活性（CA）-RhoA基因或野生型（WT）-组蛋白去乙酰化酶（HDAC）5基因的异位表达以及抑制剂研究。还测量了VSMC中活性RhoA-GTP水平和Rho相关蛋白激酶活性。我们使用离体大鼠主动脉进行了去氧肾上腺素（PE）诱导的主动脉收缩试验，并对使用VPA处理小鼠的主动脉进行的内皮依赖性主动脉舒张试验进行了事后分析。
VPA以剂量和时间依赖性方式降低了肌球蛋白轻链在Ser19位点的磷酸化（p-MLC-Ser¹⁹）。有趣的是，VPA显著降低了RhoA mRNA和蛋白表达以及活性RhoA-GTP水平。此外，CA-RhoA基因的异位表达几乎完全逆转了VPA抑制的p-MLC-Ser¹⁹。VPA显著增加了组蛋白3的乙酰化水平（H3K9ac/K14ac）。VPA和丁酸钠而非丙戊酰胺显著降低了VSMC中RhoA和p-MLC-Ser¹⁹的表达水平。然而，WT-HDAC5基因的过表达并未逆转这种降低，表明HDAC5不是导致这种降低的原因。与结果一致，VPA减弱了PE诱导的离体大鼠主动脉收缩，降低了RhoA和p-MLC-Ser¹⁹表达，并增加了H3K9ac/K14ac水平。事后分析显示，VPA抑制的RhoA途径占VPA诱导的主动脉总舒张的30%。
本研究表明，VPA通过降低RhoA表达来抑制RhoA介导的VSMC和血管收缩，这是由VPA对HDAC的抑制作用介导的。这些结果表明，VPA可能对治疗和预防包括冠状动脉痉挛和SAH后血管痉挛在内的痉挛性血管疾病有用。
