In recent years, the roles of microRNAs (miRNAs) in pulmonary diseases have been 16 widely studied and researched. However, the molecular mechanism by which miR-214 affects 17 bronchopulmonary dysplasia (BPD) remains elusive and merits further exploration. Hence, this 18 study aims to clarify the function of miR-214 in pulmonary angiogenesis and alveolarization in 19 preterm infants with BPD. BPD neonatal rat model was induced by hyperoxia, and pulmonary 20 epithelial cells were isolated from rats and exposed to hyperoxia. Gain-or loss-of-function 21 experiments were performed in BPD neonatal rats and hyperoxic pulmonary epithelial cells. 22 MiR-214 and PlGF expression in BPD neonatal rats, and eNOS, Bcl-2, c-myc, Survivin, α-SMA 23 and E-cadherin expression in hyperoxic pulmonary epithelial cells were detected using RT-qPCR 24 and western blot analysis. The interaction between PlGF and miR-214 was identified using dual 25 luciferase reporter gene assay and RIP assay. ELISA was adopted to assess IL-1β, TNF-a, IL-6,
26ICAM-1 and Flt-1 expression in rats. Decreased miR-214 expression and elevated PlGF expression 27 were evident in the lung tissues of neonatal rats with BPD. PlGF was a target of miR-214, and 28 miR-214 downregulated PlGF to inactivate the STAT3 signaling pathway. miR-214 overexpression 29 or PlGF silencing decreased apoptosis of hyperoxic pulmonary epithelial cells and declined 30 pulmonary angiogenesis and alveolarization in BPD neonatal rats. Collectively, miR-214 can 31 protects against pulmonary angiogenesis and alveolarization in preterm infants with BPD by 32 suppressing PlGF and blocking STAT3 signaling pathway.33 34 Pulmonary angiogenesis; Alveolarization 36 37 59miR-214/PlGF/STAT3 signaling pathway may be involved in BPD. Therefore, the current study 60 was conducted with the aim to verify the expected involvement of miR-214/PlGF/STAT3 axis in 61 BPD, and to elucidate the underlying molecular mechanisms. 62 63 4 MATERIALS AND METHODS 64 Ethics statement. Animal experiment protocols were approved by the Experimental Animal 65 Ethics Committee of Affiliated Hangzhou First People's Hospital, Zhejiang University School of 66 Medicine. All animal experiments were performed in accordance with the Guide for the Care and 67 Use of Laboratory animals published by the US National Institutes of Health. Extensive efforts 68 were made to ensure minimal suffering of animals during the study. 69 70 Analysis of BPD gene expression dataset and bioinformatics prediction. The gene expression 71 dataset GSE25293 of mouse BPD models was retrieved from the annotation platform GPL1261 in 72 the Gene Expression Omnibus (GEO) database (https://www.ncbi.nlm.nih.gov/gds) and then 73 77 (https://www.exiqon.com/miRSearch) were used to analyze the intersected upstream miRNAs in 78 human body. A box plot was drawn using R language to extract key miRNA expression data from 79 the miRNA dataset GSE25293 from the annotation platform GPL11199 in the GEO database.80Protein-protein interaction (PPI) analysis was perfo...