Long non‑coding RNA Rian protects against experimental bronchopulmonary dysplasia by sponging miR‑421

Tao,Xifeng; Fang,Yafei; Huo,Chen

doi:10.3892/etm.2021.10213

Long non‑coding RNA Rian protects against experimental bronchopulmonary dysplasia by sponging miR‑421

Authors:
- Xifeng Tao
- Yafei Fang
- Chen Huo
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Affiliations: Department of Pediatrics, The First People's Hospital of Lianyungang, Lianyungang, Jiangsu 222000, P.R. China
Published online on: May 19, 2021 https://doi.org/10.3892/etm.2021.10213
Article Number: 781
Copyright: © Tao et al. This is an open access article distributed under the terms of Creative Commons Attribution License.

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Abstract

Bronchopulmonary dysplasia (BPD) is a frequent complication characterized by accelerated lung alveolarization in newborns. Long non‑coding RNAs (lncRNAs) and microRNAs (miRs) are regarded as essential regulators in various diseases, including BPD. However, the detailed mechanism of the functions of RNA imprinted and accumulated in nucleus (Rian) lncRNA in the progression of BPD have remained elusive. The aim of the present study was to illustrate the interaction between miR‑421 and Rian in BPD models and MLE‑12 cells. The ability of Rian to protect neonatal lungs from hyperoxia‑induced lung damage was examined. A mouse model of BPD and a hyperoxia‑stimulated MLE‑12 cell damage model were generated and treated with specific plasmid/mimics for the overexpression of Rian/miR‑421. The interaction between miR‑421 and Rian was predicted and verified using StarBase and a dual‑luciferase reporter assay, respectively. The expression levels of miR‑421 or Rian in both tissues and the MLE‑12 alveolar epithelial cell line were assessed using reverse transcription‑quantitative (RT‑q)PCR. As parameters of alveolarization, the mean linear intercept (MLI), radial alveolar count (RAC) and the lung weight/body weight (LW/BW) ratio were measured. Furthermore, RT‑qPCR was used to measure mRNA levels of pro‑inﬂammatory cytokines (TNF‑α, IL‑6 and IL‑1β) in the lung tissue of mice, and ELISAs were performed to determine the levels of pro‑inﬂammatory cytokines (TNF‑α, IL‑6 and IL‑1β) in the supernatant of MLE‑12 cells. Cell growth and apoptosis were evaluated using an MTT assay and ﬂow cytometry, respectively. Furthermore, caspase‑3 activity was assessed using a caspase‑3 activity detection kit. Prediction with StarBase and the dual‑luciferase reporter assay revealed that miR‑421 directly targeted Rian. RT‑qPCR analysis confirmed that Rian was downregulated and miR‑421 was upregulated in lung tissues of the mouse model of BPD and in hyperoxia‑induced MLE‑12 cells. However, the expression of miR‑421 was decreased by Rian‑overexpression, an effect that was reversed by miR‑421 mimics. In addition, BPD was alleviated by Rian‑plasmid, as confirmed by the enhanced RAC and reduced MLI and LW/BW ratio. The present results also indicated that Rian‑plasmid inhibited the secretion of pro‑inflammatory cytokines (TNF‑α, IL‑6 and IL‑1β) in BPD mouse serum and hyperoxia‑induced MLE‑12 cells. In addition, Rian‑plasmid eliminated the effect of hyperoxia to inhibit cell viability and induce apoptosis in MLE‑12 cells. However, all of these effects of Rian were markedly reversed by miR‑421 mimics. The present results indicated that Rian may attenuate hyperoxic damage in neonatal lungs and may serve as a novel molecular target for BPD treatment.

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