Effects Of CASK Overexpression On Non-small Cell Lung Cancer H1299 Cells Apoptosis And Cell Cycle And Mechanism

OBJECTIVE : Lung cancer has become the number one public enemy threatening human health,and more than 80% of them are non-small cell lung cancer(NSCLC).Targeted therapy as a new treatment,its reliable efficacy and mild adverse reactions,has pointed out a new direction for the treatment of lung cancer has pointed out a new direction for the treatment of lung cancer.Studies have shown that overexpression of CASK can significantly inhibit cell proliferation.(a atrategy for extracting..)However,CASK gene and non-small cell lung cancer research is rare.Targeted therapy as a new treatment,its reliable efficacy and mild adverse reactions,has pointed out a new direction for the treatment of lung cancer.Studies have shown that overexpression of CASK can significantly inhibit cell proliferation In this study,the H1299 cell line was stably transfected by the lentiviral vector constructed in the early stage of this research group and can express human CASK gene stably.The effect of H1299 cells on cell cycle and apoptosis after CASK overexpression was studied.METHOD : In the early stage of this research group,human calcium/calmodulin-dependent serine protein kinase(CASK)geneoverexpression lentiviral vector has been constructed.1.Cell culture and CASK overexpression Lentivirus infection of H1299 cells,photographed under a fluorescent inverted microscope to observe the cell status and infection efficiency of each group.2.The CASK gene was overexpressed with lentivirus to infect H1299 cells(experimental group,ie,OE group),CON328 negative control virus was infected with H1299 cells(negative control group,ie NC group),and a blank control group(MOCK group)was also established.3.Real-time quantitative PCR was used to detect the change of CASK gene expression in H1299 cells at the mRNA level.4.Flow cytometry Annexin-V(APC staining)was used to detect the apoptosis of cells.5.Flow cytometry PI staining was used to detect cell cycle changes.RESULT : 1.Lentivirus was infected with H1299 cells for 3 days,and H1299 cells were observed by fluorescence inverted microscope.The infection efficiency was determined according to the expression of green fluorescent protein.The expression rate of green fluorescence was over 70%.The results showed that the cells were in good condition and the infection was successful.Lentivirus was infected with H1299 cells for 3 days,and H1299 cells were observed by fluorescence inverted microscope.The infection efficiency was determined according to the expression of green fluorescent protein.The expression rate of green fluorescence was over 70%.The results showed that the cells were in good condition and the infection was successful.2.The expression of CASK in H1299 cells was significantly increased by real-time PCR.The abundance of CASK in H1299 cells in OE group was 2.852 times higher than that in NC group(P<0.01).3.The apoptotic rate of H1299 cells overexpressing CASK gene and the control cells of CON328 negative virus was 5.89% in apoptotic cells in OE group and3.72% in apoptotic cells in NC group.The apoptotic rate was significantly increased(P<0.05).This indicates that overexpression of CASK gene virus can promote apoptosis of NSCLC H1299 cells.4.Cell cycle results showed that compared with NC group and MOCK group,G1 phase cells in OE group increased significantly,while cells in S phase and G2/M phase decreased significantly,the difference was statistically significant(P<0.05).These results indicate that overexpression of CASK gene virus can affect the cell cycle progression of NSCLC H1299 cells,inhibit cell cycle G1/S phase transition,cause G1 arrest,and finally inhibit the proliferation of non-small cell lung cancer H1299 cells.CONCLUSION:Overexpression of CASK gene can significantly promote the apoptosis of non-small cell lung cancer H1299 cells,and can affect the cell cycle progression of NSCLS H1299 cells,slow cell proliferation,and thus inhibit the development of lung cancer.In this study,we investigated the function and role of CASK gene in the biological characteristics of human lung cancer cells,and initially confirmed that CASK is a candidate tumor suppressor gene for non-small cell lung cancer,providing a theoretical basis for clinical molecular targeted therapy for lung cancer.Overexpression of CASK gene can significantly promote the apoptosis of non-small cell lung cancer H1299 cells,and can affect the cell cycle progression of NSCLS H1299 cells,slow cell proliferation,and thus inhibit the development of lung cancer.In this study,we investigated the function and role of CASK gene in the biological characteristics of human lung cancer cells,and initially confirmed that CASK is a candidate tumor suppressor gene for non-small cell lung cancer,providing a theoretical basis for clinical molecular targeted therapy for lung cancer.