The PDRG1 Is An Oncogene In Lung Adenocarcinoma Cells,Promoting Radioresistance Via The ATM-P53 Signaling Pathway

Objective:The aim of this research is to investigate the expression level of oncogene PDRG1 in human lung cancer cell lines.The PDRG1 overexpression and interference cell lines were established.To explore the mechanism of PDRG1 about proliferation,apoptosis and radioresistance of lung cancer cells.Methods:1.The expression of PDRG1 mRNA and protein in lung cancer cell lines and normal cell lines were detected by qRT-PCR and Western blot.2.Western blot and qRT-PCR were applied to detect the different PDRG1 plasmids protein and mRNA expression in A549 and 95-D cells which structured by liposome transfection and lentiviral transfection technology.3.CCK8 and flow cytometry were used to detect the proliferation and apoptosis of PDGR1 in the A549 and 95-D cell lines including over expression group,interference group and normal group.4.Using flow cytometry to detect the effective radiation dose of lung cancer cell line to the radioresistance.5.The correlation between PDRG1,ATM,pP53 and other related genes were determined by Western blot assay.To determine the correlation with P65 pathway.6.Western blot and CCK8 were used to detect the changes of cells proliferation and apoptosis.The ATM inhibitor blot(KU55933)had anything to do with it.7.Observing the different cell vaccination in nude mouse hypodermic the situation of tumor growth to analysis the effects of PDRG1 on the proliferation.8.Estimating the relationship between PDRG1,ATM,P53 and other proteins in vitro by immunohistochemistry.Results:1.The results of qRT-PCR and Western blot showed that compared with the normal cell line of human embryonic lung fibroblast WI38,PDRG1 in five lung cancer cell lines showed high levels of mRNA and protein.However,there are still differences among lung cancer cell lines,and the expression of PDRG1 in lung adenocarcinoma cell line A549 is higher than that in 95-D.2.The results of qRT-PCR and Western blot showed that over expression of PDRG1 and interference plasmid could be expressed in A549 and 95-D cell lines,and the expression of mRNA and protein were changed accordingly.3.CCK-8 and flow cytometry showed that overexpression of PDRG1 could promote proliferation and inhibit apoptosis,and further verified the over expression of PDRG1 and the interference of transient expression of lung cancer cell line A549 and 95-D cells.4.The results of flow cytometry showed that the PDRG1 gene could induce a significant difference in the apoptosis rate of 8Gy at about 9 Co60.5.The results of Western blot showed that irradiation could promote the expression of PDRG1 gene,the expression of PDRG1 gene was directly proportional to the ATM gene,and was inversely proportional to the P53 gene,but there was no significant relationship with the P65 pathway.6.Flow cytometry and CCK8 assay showed that over expression of PDRG1 could enhance the proliferation of lung cancer cells and inhibit the apoptosis of the cells.After the addition of ATM inhibitor,the proliferation of PDRG1 overexpression cells was significantly decreased and the apoptosis was enhanced.However,the control group did not change significantly after adding ATM inhibitor.7.The results of nude mice bearing tumor showed that the ability of cell proliferation was decreased after Co60 irradiation,and the over expression of PDRG1 gene could promote the proliferation of cells after irradiation.8.Immunohistochemistry were used to detect the tumor formation in nude mice subcutaneous tumor.The results showed that the PDRG1 gene could be activated by radiation,and the expression of PDRG1 gene was directly proportional to ATM,which was inversely proportional to P53 gene.Conclusions:1.PDRG1 were found in various human lung cancer cell lines with high expression level,and the construction of over expression and transient disturbance stable cell lines in vitro and in vivo experiments show that PDRG1 is a cancer gene in lung cancer cell lines proliferation,inhibition of apoptosis.2.Radiation could promote the increase of PDRG1 expression by injuring the cell.We structure the overexpression and interference of PDRG1,then detect ATM,P53,P65 and other genes before or after irradiation.It was found that PDGRG1 gene could enhance the tumor resistance by ATM-P53 pathway.