Abstract
BackgroundBronchopulmonary dysplasia (BPD) in premature infants is a predominantly secondary occurrence to intrauterine inflammation/infection and postpartum mechanical ventilation; in recent years, an association with epigenetics has also been found. DNA methylation, catalyzed by DNA methyl transferases (DNMTs), and tri-methylation of lysine 27 on histone H3 (H3K27me3), mediated by the methyltransferase, Enhancer of Zeste Homolog 2 (EZH2), are some of the most commonly found modifications in epigenetics. Runt-related transcription factor 3 (RUNX3) is associated with pulmonary epithelial and vascular development and regulates expression at the post-transcriptional level by DNA methylation through DNMT1 or DNMT3b. However, the involvements of these epigenetic factors in the occurrence of BPD are, as yet, unclear.MethodsNewborn rats were randomly assigned to a model, hyperoxia (85 % O2) or control, normoxia group (21 % O2). Lung tissues and alveolar type 2 (AT2) epithelial cells were collected between 1–14 days. The expression of DNMTs, and EZH2 was detected by immunohistochemistry, Western blot and real-time PCR. The percentage of DNA methylation and H3K27me3 levels in the RUNX3 promoter region was measured by bisulfite sequencing PCR and chromatin immunoprecipitation assay. RUNX3 protein and mRNA expression in AT2 cells was also measured after inhibition using the DNA methylation inhibitor, 5-Aza-2′-deoxycytidine, the H3K27me3 inhibitor, JMJD3, and the EZH2 inhibitor, DZNep.ResultsCompared with the control group, RUNX3 protein was downregulated and DNMT3b and EZH2 were highly expressed in lung tissues and AT2 cells of the model group (P < 0.05), while high DNA methylation and H3K27me3 modifications were present in the RUNX3 promoter region, in lung tissues of the model group (P < 0.05). Following hyperoxia in the model group, JMJD3 and DZNep significantly reversed the hyperoxia-induced down-regulation of RUNX3 expression in AT2 cells (P < 0.05), more so than 5-Aza-2′-deoxycytidine (P < 0.05).Conclusions1) DNA methylation and H3K27 trimethylation are present in the BPD model; 2) RUNX3 down-regulation is attributed to both DNMT3b-catalyzed DNA methylation and EZH2-catalyzed histone methylation.
Highlights
Bronchopulmonary dysplasia (BPD) in premature infants is a predominantly secondary occurrence to intrauterine inflammation/infection and postpartum mechanical ventilation; in recent years, an association with epigenetics has been found
Runt-related transcription factor 3 (RUNX3) expression in lung tissues and alveolar type 2 (AT2) cells following hyperoxia We studied RUNX3 protein localization by immunohistochemical staining of lung tissues from both control and BPD model groups exposed to hyperoxia over time; RUNX3 protein expression was found in the nucleus and cytoplasm of alveolar epithelial cells in lung sections of both the control and model groups (Fig. 1a, b)
We examined RUNX3 protein and mRNA levels in lung tissue and AT2 cells isolated from these groups
Summary
Bronchopulmonary dysplasia (BPD) in premature infants is a predominantly secondary occurrence to intrauterine inflammation/infection and postpartum mechanical ventilation; in recent years, an association with epigenetics has been found. Runt-related transcription factor 3 (RUNX3) is associated with pulmonary epithelial and vascular development and regulates expression at the post-transcriptional level by DNA methylation through DNMT1 or DNMT3b. A large number of studies have revealed that some genes, including those for vascular endothelial growth factor (VEGF) [2], interleukin 1-beta (IL-1β) [3] and mucin 1 (MUC1) [4], participate in the pulmonary developmental disorder process of BPD by regulating alveolar formation. Previous studies have shown that RUNX3 was expressed in mouse pulmonary epithelium at E15.5 [8], while RUNX3 knock-out caused pulmonary epithelial hyperplasia [8] and pulmonary vascularization disorder [7], similar to the pathological changes seen in BPD [1]. The mechanisms behind RUNX3 down-regulation and any potential regulators of abnormal RUNX3 expression in a BPD model have, as yet, to be defined
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
