17β-Estradiol Inhibits Proliferation and Oxidative Stress in Vascular Smooth Muscle Cells by Upregulating BHLHE40 Expression.

Dan-Dan Feng,Xiao Hao,Ying Ma,Man-Li Zhang,Xin-Hua Zhang,Jin-Kun Wen,Bin Zheng,Jing Yu

doi:10.3389/fcvm.2021.768662

Abstract

Background: Intimal hyperplasia is a major complication of restenosis after angioplasty. The abnormal proliferation and oxidative stress of vascular smooth muscle cells (VSMCs) are the basic pathological feature of neointimal hyperplasia. 17β-Estradiol can inhibit VSMCs proliferation and inflammation. However, it is still unclear whether and how 17β-Estradiol affects intimal hyperplasia.Methods: The neointima hyperplasia was observed by hematoxylin/eosin staining. The expression of PCNA, cyclin D1, NOX1, NOX4 and p47phox in neointima hyperplasia tissues and VSMCs was determined by qRT-PCR and Western blotting. MTS assay, cell counting and EdU staining were performed to detect cells proliferation. The oxidative stress was assessed by ROS staining.Results: 17β-Estradiol suppressed carotid artery ligation-induced intimal hyperplasia, which is accompanied by an increase of BHLHE40 level. Furthermore, loss- and gain-of-function experiments revealed that BHLHE40 knockdown promotes, whereas BHLHE40 overexpression inhibits TNF-α-induced VSMC proliferation and oxidative stress. 17β-Estradiol inhibited TNF-α-induced VSMC proliferation and oxidative stress by promoting BHLHE40 expression, thereby suppressing MAPK signaling pathways. In addition, enforcing the expression of BHLHE40 leads to amelioration of intimal hyperplasia.Conclusions: Our study demonstrates that 17β-Estradiol inhibits proliferation and oxidative stress in vivo and in vitro by promotion of BHLHE40 expression.

Highlights

Vascular smooth muscle cell (VSMC), which plays a crucial role in maintaining vascular structure and function, is mainly subsistence in the medial layer of the blood vessel wall [1]
Since it is known that ligation injuryinduced intimal hyperplasia is closely related to VSMC proliferation and oxidative stress, we investigate the effects of E2 on proliferation and oxidative stress-related genes expression in carotid arteries
Carotid artery ligation-induced these changes were reversed by E2 (10 mg ·kg−1·day−1) treatment (Figure 1D). qRT-PCR analysis of PCNA, cyclin D1, NOX1, NOX4, p47phox and KLF4 expression was consistent with their expression of protein level (Figure 1E)

Summary

Introduction

Vascular smooth muscle cell (VSMC), which plays a crucial role in maintaining vascular structure and function, is mainly subsistence in the medial layer of the blood vessel wall [1]. Abnormal VSMC proliferation, migration, inflammation or oxidative stress could lead to vascular remodeling, which contributes to the development of a series of vascular diseases, such as BHLHE40 Inhibits Intimal Hyperplasia atherosclerosis, hypertension and post-angioplasty restenosis [2, 3]. Researchers demonstrate that BHLHE40 is closely involved in many kinds of biological processes like cell proliferation, senescence, inflammation and oxidative stress [7,8,9,10]. We explored the function of BHLHE40 in ligation injury-induced intimal hyperplasia, providing causative evidence that proliferation and oxidative stress were negatively regulated by BHLHE40 protein in VSMC. The abnormal proliferation and oxidative stress of vascular smooth muscle cells (VSMCs) are the basic pathological feature of neointimal hyperplasia. It is still unclear whether and how 17β-Estradiol affects intimal hyperplasia

Methods

Results

Conclusion