Salvianolic acid B inhibits Ang II-induced VSMC proliferation in vitro and intimal hyperplasia in vivo by downregulating miR-146a expression.

BACKGROUND: Salvianolic acid B (Sal B), a water-soluble compound extracted from Salvia miltiorrhiza that has been widely used to treat cardiovascular diseases for hundreds of years in China, exerts cardiovascular protection by multiple mechanisms. miR-146a is involved in vascular smooth muscle cell (VSMC) phenotypic modulation and proliferation. However, it has yet to be investigated whether the cardiovascular protective effect of Sal B is mediated by miR-146a. PURPOSE: To determine the relationship among the cardiovascular protective effect of Sal B, miR-146a expression, and VSMC proliferation. METHODS: MTS assay and cell counting were performed to evaluate the effect of Ang II, Sal B and miR-146a on VSMC proliferation. The neointima hyperplasia was assessed by hematoxylin/eosin staining. qRT-PCR was used to detect the expression of miR-146a, KLF5, cyclin D1 and PCNA. Western blot analysis was used to detect the expressions of KLF5, cyclin D1 and PCNA after miR-20b-5p was knocked down or overexpressed in VSMC. RESULTS: Sal B suppressed intimal hyperplasia induced by carotid artery ligation and decreased Ang II-induced VSMC proliferation by down-regulating the positive cell-cycle regulators KLF5 and cyclin D1. Further experiments showed that VSMC proliferation and upregulation of KLF5 and cyclin D1 induced by Ang II were accompanied by elevated miR-146a level. Furthermore, overexpression of miR-146a promoted and knockdown of miR-146a reduced Ang II-induced VSMC proliferation and ameliorated intimal hyperplasia induced by carotid artery ligation. Sal B inhibited Ang II-induced VSMC proliferation by suppressing miR-146a expression. CONCLUSION: Sal B inhibited Ang II-induced VSMC proliferation in vitro and intimal hyperplasia in vivo by downregulating miR-146a expression.

Application of galangin, an active component of Alpinia officinarum Hance (Zingiberaceae), for use in drug-eluting stents.

In clinical pathology, stent interposition is used to treat vascular disease but can lead to restenosis. Drug-eluting stents (DES) are most commonly used to suppress restenosis but can also have side effects. Therefore, we investigated the anti-proliferative effect and its possible target in vitro and in vivo. We found that Alpinia officinarum Hance (AO) extract efficiently inhibited VSMC proliferation by arresting the transition from the G0/G1 to the S phase via the up-regulation of p27KIP1 expression. Galangin (GA) was determined to be a significant component of this extract, with the same anti-proliferative activity as the raw extract. Immunoblotting and immunofluorescence staining showed that both the AO extract and GA targeted the up-regulation of p27KIP1 expression. Therefore, we next examined the effect of these compounds in a cuff-injured neointimal hyperplasia model in vivo. In this animal model, both the AO extract and GA completely suppressed the neointima formation, and this inhibitory effect was also demonstrated to target the up-regulation of p27KIP1, including the suppression of proliferating cell nuclear antigen expression. Our findings indicate that AO extract and GA have a potent anti-proliferative activity, targeting the up-regulation of p27 expression. Thus, GA may represent an alternative medicine for use in DES.

NFAT regulates the expression of AIF-1 and IRT-1: yin and yang splice variants of neointima formation and atherosclerosis.

AIMS: Alternative transcription and splicing of the allograft inflammatory factor-1 (AIF-1) gene results in the expression of two different proteins: AIF-1 and interferon responsive transcript-1 (IRT-1).  Here, we explore the impact of AIF-1 and IRT-1 on vascular smooth muscle cell (VSMC) activation and neointima formation, the mechanisms underlying their alternative splicing, and associations of AIF-1 and IRT-1 mRNA with parameters defining human atherosclerotic plaque phenotype. METHODS AND RESULTS: Translation of AIF-1 and IRT-1 results in different products with contrasting cellular distribution and functions. Overexpression of AIF-1 stimulates migration and proliferation of human VSMCs, whereas IRT-1 exerts opposite effects. Adenoviral infection of angioplasty-injured rat carotid arteries with AdAIF-1 exacerbates intima hyperplasia, whereas infection with AdIRT-1 reduces neointima. Expression of these variants is modulated by changes in nuclear factor of activated T-cells (NFAT) activity.  Pharmacological inhibition of NFAT or targeting of NFATc3 with small interfering RNA (siRNA) lowers the AIF-1/IRT-1 ratio and favours an anti-proliferative outcome.  NFAT acts as a repressor on the IRT-1 transcriptional start site, which is also sensitive to interferon-γ stimulation. Expression of AIF-1 mRNA in human carotid plaques associates with less extracellular matrix and a more pro-inflammatory plaque and plasma profile, features that may predispose to plaque rupture. In contrast, expression of IRT-1 mRNA associates with a less aggressive phenotype and less VSMCs at the most stenotic region of the plaque. CONCLUSION: Inhibition of NFAT signalling, by shifting the AIF-1/IRT-1 ratio, may be an attractive target to regulate the VSMC response to injury and manipulate plaque stability in atherosclerosis.