Nucleotide-Binding Oligomerization Domain-Like Receptor 3 Deficiency Attenuated Isoproterenol-Induced Cardiac Fibrosis via Reactive Oxygen Species/High Mobility Group Box 1 Protein Axis.

Chen Liu,Qizhu Tang,Qingwen Xie,Jinhua Zhao,Saiyang Xie,Tongtong Hu,Qi Yao,Yuan Yuan,Qing Qing Wu,Zhulan Cai,Ning Li

doi:10.3389/fcell.2020.00713

Abstract

Nucleotide-binding oligomerization domain-like receptor 3 (NLRP3) is involved in fibrosis of multiple organs, such as kidney, liver, lung, and the like. However, the role of NLRP3 in cardiac fibrosis is still controversial and remains unclear. The study aims to investigate the role of NLRP3 on cardiac fibrosis induced by isoproterenol (ISO). In vivo, NLRP3 knockout and wild-type mice were subcutaneously injected with ISO to induce the cardiac fibrosis model. The results showed that NLRP3 deficiency alleviated the cardiac fibrosis and inflammation induced by ISO. In vitro, neonatal rat ventricular myocytes (NRVMs) and primary adult mouse cardiac fibroblasts of NLRP3 knockout and wild-type mice were isolated and challenged with ISO. Adenovirus (Ad-) NLRP3 and small interfering RNAs targeting NLRP3 were used to transfect NRVMs to overexpress or knockdown NLRP3. We found that NLRP3 could regulate high-mobility group box 1 protein (HMGB1) secretion via reactive oxygen species production in NRVMs and the HMGB1 secreted by NRVMs promoted the activation and proliferation of cardiac fibroblasts. Thus, we concluded that the NLRP3/reactive oxygen species/HMGB1 pathway could be the underlying mechanism of ISO-induced cardiac fibrosis.

Highlights

Cardiac fibrosis refers to the imbalance of extracellular matrix (ECM) production and degradation and excessive deposition and abnormal distribution of collagen, which is closely related to various cardiovascular diseases, such as hypertension, chronic heart failure (HF), hypertrophic cardiomyopathy, dilated cardiomyopathy, viral myocarditis, etc. and is a potential risk factor for sudden cardiac death
Picrosirius red staining was used to detect the collagen deposition of the heart; we found that interstitial and perivascular fibrosis exacerbated in the WT mice treated with ISO; nucleotide-binding oligomerization domain-like receptor 3 (NLRP3) deficiency ameliorated the fibrosis process in mice challenged with ISO (Figures 2A,D) as well as the transcription levels of fibrosis markers—transforming growth factor β, connective tissue growth factor (CTGF), collagen I, and collagen III (Figure 2G)
To further investigate the how NLRP3 regulates cardiac fibrosis, we cocultured the Cardiac fibroblasts (CFs) with Neonatal rat ventricular myocytes (NRVMs) supernatant treated with ISO and siNC or Small interfering RNAs targeting NLRP3 (siNLRP3); we found that the α-smooth muscle actin (α-SMA) and proliferating cell nuclear antigen (PCNA) expressions of the CFs were both increased after cocultured with NRVM supernatants treated with ISO + siNC and decreased after cocultured with NRVM supernatants treated with ISO + siNLRP3 (Figure 4C)

Summary

Introduction

Cardiac fibrosis refers to the imbalance of extracellular matrix (ECM) production and degradation and excessive deposition and abnormal distribution of collagen, which is closely related to various cardiovascular diseases, such as hypertension, chronic heart failure (HF), hypertrophic cardiomyopathy, dilated cardiomyopathy, viral myocarditis, etc. and is a potential risk factor for sudden cardiac death. Pattern recognition receptors, which included Toll-like receptors (TLRs), retinoic acid-inducible gene I-like receptors, nucleotide-binding oligomerization domain (NOD)-like receptors, and C-type lectin receptors, are widely expressed in myocardial tissues. They can initiate host inflammation and are essential to recognize infectious or dangerous foreign patterns, collectively termed as pathogenassociated molecular patterns (PAMPs) and danger-associated molecular patterns (DAMPs). Targeting the cardiac DAMPs and DAMPs, such as TLR2–9, advanced glycation end products receptors (RAGE), NOD1, NOD2, etc., has been reported to exert potentially controlling inflammatory and anti-fibrosis effect in the myocardial response to injury (Turner, 2016)

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