| Objective: Radiotherapy is one of the main treatment methods for non-small cell lung cancer.However,radioresistance is an important factor reducing the radiotherapy efficacy of non-small cell lung cancer.Meanwhile,EMT is the main cause of radioresistance.Previous studies have shown that the EMT phenotype was changed in the radiation resistant cells of lung adenocarcinoma,and the expression of mi R-183 was increased in the cell line of lung adenocarcinoma,suggesting that mi R-183 may mediate EMT to play a role in the radioresistance of lung adenocarcinoma cell.Therefore,through deep discussion on the regulation mechanism of mi R-183,which is expected to state relationships between mi R-183 and EMT and to demonstrate effects of mi R-183 and EMT on radioresistance of lung adenocarcinoma cells,providing new thoughts for seeking targets that can improve the efficacy of NSCLC radiotherapy in clinical practice.Methods: Inverted microscopes were used to observe morphological changes of cells.By the CCK-8 cell proliferation assay,after the same dose of irradiation,differences of survival rates of cells were compared.In the clone formation assay,their radioresistance was compared.Wound healing experiments were conducted to compare changes of their migration ability.q PCR was applied to test expression of mi R-183 in cells.q PCR and Western Blot were used to test changes of expression levels of molecular markers relenting to mi R-183 and EMT respectively.Lentiviral transfection experiments were carried out to up / down regulate mi R-183 gene in H1299 and H1299 R cells.Results: 1.After continuous X-ray irradiation,cellular morphology of lung adenocarcinoma changed from original closely-connected epithelial cells to long and narrow cells with extended pseudopodia that were similar with fibroblast.In previous studies,clone formation assays showed that compared with H1299 cells,H1299 R cells had a wider shoulder region of survival with stronger radioresistance(P<0.05).CCK-8 cell proliferation assays indicated that compared with H1299 cells,H1299 R cells showed a higher survival rate after receiving a sub-lethal dose(6Gy)of X-ray irradiation(P<0.05).According to q PCR tests,it was found that mi R-183,ZEB1 and Vimentin showed high expression in H1299 R cells,while expression of E-cadherin reduced(P<0.05).2.After successful construction of mi R-183-expressed lentiviral vectors,q PCR was applied to examine expression reduction of mi R-183 in H1299R-sh RNA-mi R183 cells(P<0.05).Further clone formation assays showed that compared with negative control H1299R-sh RNA-NC cells,after interference with expression of mi R-183,the survival shoulder region of H1299R-sh RNA-mi R183 cells narrowed down with reduced radioresistance(P<0.05).CCK-8 cell proliferation assays indicated that compared with H1299R-sh RNA-NC cells,H1299R-sh RNA-mi R183 cells showed a decreased survival rate after receiving 4Gy of X-ray irradiation(P<0.05).By wound healing experiments,it was found that the wound healing time of H1299R-sh RNA-mi R183 cells was extended with reduced cell migration ability(P<0.05).According to tests applying q PCR and Western Blot,it was found that in terms of gene and protein levels,expression of ZEB1 and Vimentin reduced in H1299R-sh RNA-mi R183 cells,while that of E-cadherin increased(P<0.05).3.After successful construction of mi R-183-expressed lentiviral vectors,q PCR was applied to examine expression increase of mi R-183 in H1299-EGFP-mi R183 cells(P<0.05).Further clone formation assays showed that compared with negative control H1299-EGFP-NC cells,after interference with expression of mi R-183,the survival shoulder region of H1299-EGFP-mi R183 cells broadened with increased radioresistance(P<0.05).CCK-8 cell proliferation assays indicated that compared with H1299-EGFP-NC cells,H1299-EGFP-mi R183 cells showed an increased survival rate after receiving 4Gy of X-ray irradiation(P<0.05).By wound healing experiments,it was found that the wound healing time of H1299-EGFP-mi R183 cells was shortened with enhanced cell migration ability(P<0.05).According to tests applying q PCR and Western Blot,it was found that in terms of gene and protein levels,expression of ZEB1 and Vimentin increased in H1299-EGFP-mi R183 cells,while that of E-cadherin decreased(P<0.05).Conclusion: 1.During continuous X-ray irradiation,EMT occurred in H1299 cells with enhanced proliferative activity and increased radioresistance.2.In the course of radioresistance of H1299 cells,expression of mi R-183 increased and played an important role.Overexpression of mi R-183 might promote EMT of H1299 cells and strengthen their proliferation,migration,and radioresistance.Down-regulation of expression of mi R-183 might reverse EMT of H1299 R cells and weaken their proliferation,migration,and radioresistance. |
PDF Full Text Request