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LINC00612/miR-31-5p/Notch1 Axis Regulates Apoptosis, Inflammation, and Oxidative Stress in Human Pulmonary Microvascular Endothelial Cells Induced by Cigarette Smoke Extract

Authors Luo J, Li L, Hu D, Zhang X

Received 2 April 2020

Accepted for publication 21 July 2020

Published 26 August 2020 Volume 2020:15 Pages 2049—2060


Checked for plagiarism Yes

Review by Single anonymous peer review

Peer reviewer comments 2

Editor who approved publication: Dr Richard Russell

Jun Luo,1,* Li Li,2,* Die Hu,2 Xian Zhang1

1Department of Laboratory Medicine, Chengdu Second People’s Hospital, Chengdu 610000, Sichuan, People’s Republic of China; 2Department of Respiratory and Critical Care Medicine, Dujiangyan People’s Hospital, Dujiangyan 611830, Sichuan, People’s Republic of China

*These authors contributed equally to this work

Correspondence: Li Li
Department of Respiratory and Critical Care Medicine, Dujiangyan People’s Hospital, No. 622 Baolian Road, Dujiangyan 611830, Sichuan, People’s Republic of China
Tel +86-28-87121111

Background: Long non-coding RNAs (lncRNAs) have been reported as key regulators in chronic obstructive pulmonary disease (COPD). However, the precise role of LINC00612 remains unclear.
Methods: The real-time quantitative polymerase chain reaction (RT-qPCR) was used to quantify the expression levels of LINC00612, miR-31-5p, and Notch homolog 1 (Notch1) in lung tissues and cells. Under a cigarette smoke extract (CSE) stimulation condition, the apoptosis was analyzed by flow cytometry assay. Caspase-3 activity was examined with a caspase-3 activity assay kit; besides, inflammation and oxidative stress were assessed by measuring interleukin-6, tumor necrosis factor-α, glutathione/oxidized glutathione, reactive oxygen species, malondialdehyde, and superoxide dismutase activity. The interaction relationship between miR-31-5p and LINC00612 or Notch1 was predicted by bioinformatics databases, while dual-luciferase reporter, RNA immunoprecipitation, and RNA pull-down assays were performed to confirm prediction. Eventually, the related protein expression was estimated with western blot assay.
Results: LINC00612 was downregulated in COPD tissues when compared with controls. Consistently, CSE inhibited LINC00612 expression in HPMECs with a dose/time-dependent method. Gain-of-function experiments indicated that the upregulation of LINC00612 could repress cell apoptosis, inflammation, and oxidative stress in HPMECs induced by CSE. In addition, miR-31-5p was negatively regulated by LINC00612 in HPMECs treated with CSE. The overexpression of miR-31-5p could abolish LINC00612-induced effects on HPMECs exposed to CSE. Importantly, LINC00612 could weaken CSE-induced cell apoptosis, inflammation, and oxidative stress in HPMECs by regulating the miR-31-5p/Notch1 signaling pathway.
Conclusion: Current findings suggest that CSE-mediated cell apoptosis, inflammation, and oxidative stress in HPMECs were abolished by upregulation of LINC00612. Furthermore, the LINC00612/miR-31-5p/Notch1 axis may represent a novel regulator of apoptosis, inflammation, and oxidative stress of HPMECs, which may be a potential therapeutic target for COPD in the future.

Keywords: LINC00612, miR-31-5p, Notch1, COPD, CSE

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