Study On The Expression And Regulation Mechanisms Of LncRNAs In Pulmonary Fibrosis

Objective:Recently, the pathogenesis of idiopathic pulmonary fibrosis (IPF) remains unclear and lack of effective treatment. Researches related to genetic changes in pulmonary fibrosis gradually increased so that to find new molecular therapeutic targets. Long non-coding RNA as a new class of non-coding RNA, its regulatory mechanism and its relationship with the disease has become a hot topic. By observing the differentially expressed lncRNA profile between lung fibrosis and normal lung tissue, analyzing lncRNA structural characteristics and the position or sequence relationship between the mRNA. Finally, we choice lncRNA-MRAK088388 and MRAK081523 for further study to explore the mechanism in pulmonary fibrosis.Methods:Sprague-Dawley rats were randomly divided into control group and bleomycin-treated group. The model of pulmonary fibrosis was induced by a single intratracheal instillation of bleomycin. All rats were killed on day 28. Part of lung tissue was removed to liquid nitrogen for frozen, part was fixed in glutaraldehyde and embedded by epoxy resin, another part was fixed conventionally and embedded by paraffin. The collagen content and number of fibers were detected by Masson’s staining, TEM, and Hyp assay. The differentially expressed lncRNA and mRNA profiles between lung fibrosis and normal lung tissue were analyzed using microarrays. Gene ontology analysis and pathway analysis were performed for further research. The bioinformatic analysis of lncRNA by NCBI database and UCSC databases. lncRNA screening differentially expressed fluorescence real-time quantitative PCR validation lncRNA and related miRNA, mRNA expression; situ hybridization observed lncRNA cellular localization. qRT-PCR was used to validate the expression of lncRNA and relatived miRNA and mRNA. In situ hybridization was used to observe cellular localization of lncRNA.Results:The results showed that the number of collagen fibers in the interstitial lung tissue significantly increased in the model group compared with the normal group. In total,210 and 358 lncRNAs were up-regulated and down-regulated, respectively, along with 415 up-regulated and 530 down-regulated mRNAs in the rats with lung fibrosis. According to the GO annotation and pathway analysis, the differentially expressed genes were principally enriched for gene ontology terms related to immune response, cell differentiation, cytoskeleton, cytokine-cytokine receptor interaction, chemokine signaling pathway, Jak/STAT signaling pathway as well as PPAR signaling pathway et al. Bioinformatics analysis indicated that there was a certain correlation between lncRNA with neighboring or homologous protein-coding genes; qRT-PCR displayed MRAK088388, MRAK081523, N4bp2, Plxna4 up-regulated in the model group, whereas miR-29 and let-7i down-regulation. MRAK088388, MRAK081523 were mainly located in the cytoplasm of pulmonary interstitial cells.Conclusion:In conclusion, the expression profile of the lncRNAs was significantly altered in lung fibrosis tissue. Bioinformatics analysis showed the relationships between lncRNA and adjacent or homologous protein-coding genes can be used to predict lncRNA significance. Preliminary results predicted between MRAK088388, N4bp2 and miR-29 as well as MRAK081523, Plxna4 and let-7i had functional interactions, providing a new direction for the study of the pathogenesis of pulmonary fibrosis.