Study On Effects And Mechanisms Of Long Non-coding RNAs On Proliferation, Migration And Radiosensitivity In Pulmonary Epithelial Cells

Aim: To screen differentially expressed m RNAs in X53654-overexpressing Beas-2B cell line, and to investigate its effects and mechanisms on proliferatin, migration and radiosensitivityMethods:(1) Lnc RNA-X53654-overexpressing Beas-2B cell line was constructed by lipofectamine-mediated X53654 recombinant plasmid transfection and G418 selective culture.(2) The total RNAs were extracted and microarray-hybridized for m RNAs profiling. The raw data was collected and analised using Gene Ontology(GO) and Kyoto Encyclopedia of Genes and Genomes( KEGG).(3) MTT assay, flow cytometry, wound healing, transwell migratory assay and gelatin zymography were used to examine the effects of X53654 overexpression on Beas-2B cell line, whose relating proteins were examined by western blotting.(4) RT-PCR was used to test lnc RNA-X53654 changed expression levels of post-irradiation. Data of cloniogenic assay was fitted using multi-target-single-hitting model. Flow cytometry was used to examine modulation of cell circle after irradiation, and the formation of post-irradiated γH2AX foci was tested by immunefluorescence and observed through confocal microscopy. The DNA double strands breakage was examined using comet assay. And DNA damage repairing proteins were tested using western blotting.Results: The overexpression of X53654 enhanced the proliferation of Beas-2B cell line by cell circle facilitation. The X53654-overexpression reduced cell migration, but enhanced invasion. The inhibiton of tight-junction-relating proteins(i.e. Occludin and Claudin 1) was observed in the reconstructed cell line, and invasion-relating proteins(i.e. MMP-2 and MMP-9) were up regulated. This phenomenon may be attributed to the activation of TGF-β/Smad3 pathway. And the strengthening of this pathway can explain the enhanced radiation tolerance of the X53654-overexpressed Beas-2B cell line.