Liuwei Dihuang, a traditional Chinese medicinal formula, inhibits proliferation and migration of vascular smooth muscle cells via modulation of estrogen receptors

Zhang,Yayun; Qian,Xing; Sun,Xin; Lin,Chao; Jing,Yi; Yao,Yuan; Ma,Zhi; Kuai,Meiyu; Lu,Ying; Kong,Xueyun; Chen,Qi; Wu,Xiang; Zhao,Xuan; Li,Yu; Bian,Huimin

doi:10.3892/ijmm.2018.3622

Liuwei Dihuang, a traditional Chinese medicinal formula, inhibits proliferation and migration of vascular smooth muscle cells via modulation of estrogen receptors

Authors:
- Yayun Zhang
- Xing Qian
- Xin Sun
- Chao Lin
- Yi Jing
- Yuan Yao
- Zhi Ma
- Meiyu Kuai
- Ying Lu
- Xueyun Kong
- Qi Chen
- Xiang Wu
- Xuan Zhao
- Yu Li
- Huimin Bian
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Affiliations: School of Pharmacy, Nanjing University of Chinese Medicine, Nanjing, Jiangsu 210023, P.R. China
Published online on: April 16, 2018 https://doi.org/10.3892/ijmm.2018.3622
Pages: 31-40

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Abstract

The phenotypic modulation of vascular smooth muscle cells (VSMCs) serves an important role in atherosclerosis‑induced vascular alterations, including vascular remodeling. However, the precise mechanisms underlying VSMC phenotypic modulation remain to be elucidated. Our previous study demonstrated that Liuwei Dihuang formula (LWDHF) could improve menopausal atherosclerosis by upregulating the expression of estrogen receptors (ERs). The present study examined the role of ERs in the effects of LWDHF on VSMC phenotypic modulation. VSMC proliferation and cell cycle progression were examined by MTT assay and flow cytometry, respectively. The expression levels of α‑smooth muscle actin, osteopontin and ERs were determined using reverse transcription‑quantitative polymerase chain reaction (RT‑qPCR) and western blot analysis. Cell ultrastructure was observed under an electron microscope. F‑actin polymerization was detected by fluorescein isothiocyanate‑phalloidin staining using fluorescence microscopy. A modified Boyden chamber assay was employed to assess VSMCs migration. Small interfering (si)RNA technology was used to examine the role of ERα in the effects of LWDHF on phenotypic modulation. The results indicated that LWDHF (3‑12 µg/ml) inhibited proliferation and induced a cell cycle arrest in VSMCs treated with angiotensin II (Ang II; 100 nM) in a concentration‑dependent manner. In addition, Ang II‑stimulated migration of VSMCs and reorganization of actin were markedly inhibited by treatment with 12 µg/ml LWDHF. Results of RT‑qPCR and western blotting demonstrated that LWDHF markedly stimulated transcription and expression of ERα and ERβ, and inhibited VSMC synthetic phenotype. Furthermore, LWDHF‑induced inhibition of phenotypic switching was partially suppressed by tamoxifen, and transfection with ERα siRNA markedly abolished the effects of LWDHF on VSMC phenotypic switching. In conclusion, these results revealed that ERα served an important role in LWDHF‑induced regulation of the VSMC phenotype, including proliferation and migration.

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