Increased intracellular Cl concentration mediates neutrophil extracellular traps formation in atherosclerotic cardiovascular diseases

Hui Han1, Chang Liu1, Mei Li2, Jin Wang3, Yao-sheng Liu1, Yi Zhou3, Zi-cheng Li1, Rui Hu1, Zhi-hong Li1, Ruo-mei Wang4, Yong-yuan Guan1, Bin Zhang2, Guan-lei Wang1
1 Department of Pharmacology, Cardiac and Cerebral Vascular Research Center, Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou 510080, China
2 VIP Healthcare Center, the Third Affiliated Hospital of Sun Yat-sen University, Guangzhou 510630, China
3 Guangdong Cardiovascular Institute, Guangdong Provincial People’s Hospital, Guangdong Academy of Medical Sciences, Guangzhou 510080, China
4 School of Data and Computer Science, Sun Yat-sen University, Guangzhou 510006, China
Correspondence to: Bin Zhang:, Guan-lei Wang:,
DOI: 10.1038/s41401-022-00911-9
Received: 19 October 2022
Accepted: 7 April 2022
Advance online: 5 May 2022


Neutrophil extracellular traps (NETs) play crucial roles in atherosclerotic cardiovascular diseases such as acute coronary syndrome (ACS). Our preliminary study shows that oxidized low-density lipoprotein (oxLDL)-induced NET formation is accompanied by an elevated intracellular Cl concentration ([Cl]i) and reduced cystic fibrosis transmembrane conductance regulator (CFTR) expression in freshly isolated human blood neutrophils. Herein we investigated whether and how [Cl]i regulated NET formation in vitro and in vivo. We showed that neutrophil [Cl]i and NET levels were increased in global CFTR null (Cftr−/−) mice in the resting state, which was mimicked by intravenous injection of the CFTR inhibitor, CFTRinh-172, in wild-type mice. OxLDL-induced NET formation was aggravated by defective CFTR function. Clamping [Cl]i at high levels directly triggered NET formation. Furthermore, we demonstrated that increased [Cl]i by CFTRinh-172 or CFTR knockout increased the phosphorylation of serum- and glucocorticoid-inducible protein kinase 1 (SGK1) and generation of intracellular reactive oxygen species in neutrophils, and promoted oxLDL-induced NET formation and pro-inflammatory cytokine production. Consistently, peripheral blood samples obtained from atherosclerotic ApoE−/− mice or stable angina (SA) and ST-elevation ACS (STE-ACS) patients exhibited increased neutrophil [Cl]i and SGK1 activity, decreased CFTR expression, and elevated NET levels. VX-661, a CFTR corrector, reduced the NET formation in the peripheral blood sample obtained from oxLDL-injected mice, ApoE−/− atherosclerotic mice or patients with STE-ACS by lowering neutrophil [Cl]i. These results demonstrate that elevated neutrophil [Cl]i during the development of atherosclerosis and ACS contributes to increased NET formation via Cl-sensitive SGK1 signaling, suggesting that defective CFTR function might be a novel therapeutic target for atherosclerotic cardiovascular diseases.

Keywords: atherosclerotic cardiovascular diseases; neutrophil extracellular traps; intracellular chloride; CFTR; oxLDL; SGK1

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