Study On The Relationship Between Long Non-coding RNA Tug1 And Lung Development In Newborn Rats And Its Potential Mechanism Of Action

Objective: Bronchial pulmonary dysplasia(bronchopulmonary dysplasia,BPD)is one of very common complication in premature infants.With the rapid development of perinatal medicine,continuous optimization of respiratory support strategies,timely application of lung surfactant,the survival rate of extremely low and ultra-low birth weight infants gradually increased,and the incidence of BPD increased year by year.The short-term or long-term adverse outcomes not only affect the life and quality of life of the children,but also bring heavy economic burden to the children's families and even the whole society.The occurrence of new BPD is closely related to lung immaturity,and premature infants with small gestational age and low birth weight are more likely to develop BPD.Fetal lung development is divided into five periods,including the key of the alveolar period for alveolar mature period,most preterm birth lung development in phase vesicles,lung anatomical structure and physiological function is not yet mature,highly vulnerable to high oxygen pressure,mechanical ventilation,the influence of such factors as the development of alveolar process,characterized by lung development stagnation,alveolar simplification and small pulmonary vascular dysplasia.In recent years,studies have confirmed that lncRNA can interact with transcription factors to participate in the regulation of lung development and the occurrence and development of BPD.LncRNA Tug1 is a development-related indicator that has been shown to play an important role in the development of the cerebral cortex,neurons and retina.However,the relationship between Tug1 and lung development has not been reported.Through bioinformatics analysis,PCR,WB,correlation analysis and chirp-ms,this study aims to clarify the relationship between Tug1 and lung development and BPD,and lay a foundation for the study on the mechanism of "novel" BPD lung development disorder.Methods: In the GEO data,mRNA expression microchips of endothelial cells in liver,cochlea,brain,small intestine and blood were selected for the adult and end-embryonic or end-embryonic stages.The control group was the adult stage by GEO2 R,and the end-embryonic or end-embryonic stage was the experimental group.The differentially expressed genes were calculated with P<0.05 and |logFC|?1 as the standard.After the intersection of five data sets,the results were analyzed by clustering.In the GEO database to select the lung tissue and epithelial cells of the alveolar type ? adulthood and embryology 18 days mRNA expression chip,with former differential expression analysis,and with the first five data sets the result of the intersection to intersection.BPD model was prepared in newborn SD rats induced by high oxygen(oxygen concentration was85%,n=40),and the control group was newborn SD rats growing normally in air environment(oxygen concentration was 21%,n=40).10 samples of lung tissue were randomly collected from each group at 1,3,7 and 14 days after birth.Pulmonary histomorphological changes were observed by hematoxylin-eosin staining,and alveolar maturity was evaluated by radiative alveolar counting(RAC).MRNA expression levelsof long non-coding RNA Tug1 and Pdgfra in lung tissues were detected by real-time fluorescence quantitative PCR(rt-pcr).Protein expression of Pdgfra was detected by Western Blot.Fluorescence in situ hybridization was used to observe the cell localization of Tug1.A high-throughput sequencing set of mRNA expression during alveolar development was selected from GEO database,and the coexpressed Tug1 genes were calculated using cot.test function.The co-expressed genes of Tug1 during alveolar development were analyzed for functional enrichment,and the proteins directly bound to Tug1 were detected by chirp-ms technology.Results: the expression of Tug1 in liver,cochlea,brain,small intestine and blood endothelial cells was significantly higher than that in adult.The expression in lung tissue and alveolar type ii epithelial cells at 18 days of embryo was higher than that in adult.In the lung tissue,the mRNA expression of Pdgfra was significantly lower than that of the control group from 7 days(1.04±0.25 P=0.002 in the control group 1.62±0.37 model group)to 14 days.Pdgfra protein expression in the control group showed an increasing trend with the increase of daily age,and it was significantly lower in the model group at7 days than in the control group(1.62±0.09 in the control group,1.04±0.13 P=0.04 in the model group)until 14 days.Tug1 expression in the control group showed an increasing trend with age,and Tug1 expression showed a significant positive correlation with RAC value(P=0.007,r=0.648),and a significant positive correlation with Pdgfra protein expression(P=0.001,r= 0.755).In the model group,the expression of Tug1 was significantly lower than that of the control group(1.68±0.20 model group 0.78±0.20P=0.04)from 7 days,and the trend continued to 14 days.During alveolar development,a total of 338 co-expressed genes with Tug1 at FDR < 0.05 were involved in cell energy metabolism and product decomposition,and were closely related to post-embryonic development.Among them,Uba52 and Rps27 a were proved to bind directly to Tug1 in chirp-ms experiments.Conclusion: Long non-coding RNA Tug1 is closely related to alveolar development,and the expression of Tug1 in the lung tissues of newborn rats with BPD is decreased.Tug1 may directly regulate Uba52 or indirectly regulate Smarca1 and Cdkn1 c to participate in normal alveolar development and block BPD alveolar development.