Nuclear receptor Nur77 protects against oxidative stress by maintaining mitochondrial homeostasis via regulating mitochondrial fission and mitophagy in smooth muscle cell

https://doi.org/10.1016/j.yjmcc.2022.05.007Get rights and content

Highlights

  • Nur77 protected against AngII-induced oxidative stress and mitochondrial damage in VSMCs.

  • Nur77 alleviated mitochondrial fission and restored mitophagic flux in VSMCs upon AngII stimulation.

  • Nur77 maintained mitochondrial homeostasis via regulating mitochondrial fission and mitophagy.

Abstract

Angiotensin II (AngII) induces disruption of mitochondrial homeostasis and oxidative stress. Nuclear receptor NR4A1 (Nur77) plays an important role in vascular smooth muscle cells (VSMCs) function. However, the role of Nur77 in AngII-induced mitochondrial dynamics and oxidative stress in VSMCs remains unknown. In an in vitro model of AngII-treated cells, we discovered that Nur77 knockout aggravated AngII-induced oxidative stress in VSMCs, whereas activation of Nur77 by celastrol diminished them. Concomitantly, disturbance of mitochondrial dynamics induced by AngII was further exacerbated in Nur77 deficient VSMCs compared to wild-type (WT) VSMCs. Interestingly, Nur77 deletion increased mitochondrial fission but not fusion as evidenced by upregulated fission related genes (Fis1 and Drp1) but not fusion (Opa1 and Mfn2) under AngII stimulation in VSMCs. Mechanically, Nur77 could directly bind to the promoter regions of Fis1 and Drp1 and repress their transcription. Furthermore, we observed that Nur77 additionally promoted mitochondrial homeostasis by increasing mitophagic flux in a transcription-independent manner upon AngII challenge. By using an in vivo model of AngII-induced abdominal aortic aneurysm (AAA), we finally validated the protective role of Nur77 involved in the mitochondrial fission process and mitophagic flux in aortas, which was correlated with the occurrence and development of AAA in AngII-infused mice. Our data defines an essential role of Nur77 in regulating oxidative stress by maintaining mitochondrial homeostasis in VSMCs via both transcription-dependent and transcription-independent manner, supporting the therapeutic potential of Nur77 targeting in vascular diseases.

Introduction

The renin-angiotensin-aldosterone system (RAAS) is the major hormonal system that involved in the regulation of cardiovascular function [1,2]. AngII, a potent vasoconstrictive peptide hormone of RAAS, has gained notable attention as a therapeutic target of various cardiovascular diseases such as abdominal aortic aneurysm (AAA) [3,4], hypertension [5], and heart failure [6]. Abnormal activation of RAAS results in an augmented burden of vascular walls, which is partially linked to sustained responses of VSMCs to high levels of AngII, leading to VSMC dysfunction and vascular diseases [7].

Oxidative stress is a primary factor leading to vascular damage under AngII stimulation [8]. Generation of reactive oxygen species (ROS) by damaged mitochondria contributes to numerous physiologic [9] and pathologic processes including metabolic process [10] and cell death [11,12]. Maintenance of mitochondrial homeostasis depends on the restricted quality control accomplished by several molecular machineries that related to mitochondrial biogenesis, mitochondrial dynamics (fission and fusion), and mitophagy [[13], [14], [15]]. Recent studies show that disrupted mitochondrial dynamics are implicated in AAA [16] and hypertension [17], which are particularly associated with VSMCs dysfunction. However, the underlying mechanisms referred to mitochondrial damage in VSMCs under AngII challenge remain elusive.

Nur77/NR4A1 is an orphan nuclear receptor that belongs to NR4A family. These receptors sense changes of microenvironment and control physiological and pathological processes including cellular proliferation, angiogenesis, and cell death through genomic and non-genomic actions [18]. Apart from transcriptional regulation, Nur77 could translocate to mitochondria and inhibit inflammation and oxidative stress through activating autophagy [19]. However, the role of Nur77 in mitochondrial homeostasis and oxidative stress in the setting of AngII challenge is still poorly understood. In this study, we unveil that Nur77 regulates oxidative stress by sustaining mitochondrial homeostasis via attenuating mitochondria fission process and promoting mitophagy under AngII challenge both in vitro and in vivo, supporting the therapeutic potential of Nur77 targeting in vascular diseases.

Section snippets

Animal experiment protocols

Animal experiments were approved by the Animal Ethics Committee of Renji Hospital (RJ2018-1012) and followed the National Institutes of Health (NIH) Guidelines for the Care and Use of Laboratory Animals. Nur77 global knockout mice were generated using CRISPR/Cas9 technology as described previously [20]. The efficiency of Nur77 knockout was shown in Fig. S1. ApoE−/− mice were obtained from the Shanghai Model Organisms Center. ApoE−/-Nur77−/− mice were generated by crossing Nur77−/− mice with AopE

Nur77 deficiency aggravates AngII-induced oxidative stress in VSMCs

We first investigated whether Nur77 regulated AngII-induced oxidative stress in VSMCs isolated from aortas. As shown in Fig. 1A, AngII remarkably induced the ROS production in VSMCs as revealed by DHE staining. Of note, Nur77 knockout further increased ROS generation in AngII-treated VSMCs (Fig. 1A), indicating a protective role of Nur77 against oxidative stress. Similarly, knockdown of Nur77 by transfected with specific siRNA in human VSMCs (hASMCs) also increased ROS generation compared with

Discussion

In the current study, we provided new insights into the protective role of Nur77 in regulation of mitochondrial homeostasis and oxidative stress in the setting of AngII stimulation. First, Nur77 deficiency exacerbated oxidative stress in AngII-treated VSMCs, which was accompanied by disrupted mitochondrial dynamics. Second, Nur77 specifically inhibited mitochondrial fission but not fusion process by transcriptionally repressed the expressions of mitochondrial fission related genes Fis1 and Drp1

Author contributions

Qin Shao designed the study; Na Geng, Taiwei Chen and Long Chen conducted the experiments, analyzed the data, and wrote the paper; Hengyuan Zhang, Qingqing Xiao and Zhenyu Tao performed experiments; Lingyue Sun, Yuyan Lyu and Xinyu Che participated in the animal models; Qin Shao reviewed and edited the manuscript.

Disclosures

None.

Funding

This study was supported by Grants from the National Natural Science Foundation of China (81870338, 81570390).

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    1

    Na Geng, Taiwei Chen, Long Chen contributed equally to this work.

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