**Anti-Atherosclerotic E**ff**ect of** *Hibiscus* **Leaf Polyphenols against Tumor Necrosis Factor-alpha-Induced Abnormal Vascular Smooth Muscle Cell Migration and Proliferation**

#### **Cheng-Chung Chou 1, Chi-Ping Wang 2,3, Jing-Hsien Chen 2,4,\* and Hui-Hsuan Lin 2,4,\***


Received: 2 November 2019; Accepted: 3 December 2019; Published: 5 December 2019

**Abstract:** The proliferation and migration of vascular smooth muscle cells (VSMCs) are major events in the development of atherosclerosis following stimulation with proinflammatory cytokines, especially tumor necrosis factor-alpha (TNF-α). Plant-derived polyphenols have attracted considerable attention in the prevention of atherosclerosis. *Hibiscus* leaf has been showed to inhibit endothelial cell oxidative injury, low-density lipoprotein oxidation, and foam cell formation. In this study, we examined the anti-atherosclerotic effect of *Hibiscus* leaf polyphenols (HLPs) against abnormal VSMC migration and proliferation in vitro and in vivo. Firstly, VSMC A7r5 cells pretreated with TNF-α were demonstrated to trigger abnormal proliferation and affect matrix metalloproteinase (MMP) activities. Non-cytotoxic doses of HLPs abolished the TNF-α-induced MMP-9 expression and cell migration via inhibiting the protein kinase PKB (also known as Akt)/activator protein-1 (AP-1) pathway. On the other hand, HLP-mediated cell cycle G0/G1 arrest might be exerted by inducing the expressions of p53 and its downstream factors that, in turn, suppress cyclin E/cdk2 activity, preventing retinoblastoma (Rb) phosphorylation and the subsequent dissociation of Rb/E2F complex. HLPs also attenuated reactive oxygen species (ROS) production against TNF-α stimulation. In vivo, HLPs improved atherosclerotic lesions, and abnormal VSMC migration and proliferation. Our data present the first evidence of HLPs as an inhibitor of VSMC dysfunction, and provide a new mechanism for its anti-atherosclerotic activity.

**Keywords:** proliferation; migration; vascular smooth muscle cells; atherosclerosis; tumor necrosis factor-alpha; *Hibiscus* leaf polyphenols
