Effect Of GPD₂ Gene Knockdown By ShRNA On Biological Function Of Lung Cancer Cells

Objectives:To clarify the role of GPD2 gene in proliferation,apoptosis,cell cycle,migration and invasion of lung cancer cells Methods:First,real-time quantitative PCR(qPCR)and Western blot were used to detect the expression of GPD2 gene in four lung cancer cells H1299,A549,H460 and H23.Secondly,construct GPD2 gene-specific RNA interference lentiviral vector(shGPD2)and Corresponding control interference vector(shCtrl)was used to infect lung cancer cells H1299 and A549 with lentiviral interference vectors shGPD2 and shCtrl,respectively.qPCR and Western blot were used to determine the knockdown efficiency of GPD2 gene in two lung cancer cells.Finally,CCK-8 method was used to detect the proliferation of two lung cancer cells for 5 consecutive days.The cell clone assay was used to detect the colony forming ability of two lung cancer cells.The cell cycle changes and apoptosis of two lung cancer cells were detected by flow cytometry.Transwell and cell scratch assays were used to detect the invasion and migration of two lung cancer cells.Results:The results of qPCR and Western blot showed that the expression of GPD2 gene was the highest in lung cancer cells H1299,followed by A549 and H460,and the lowest in H23.After qPCR detection of lentiviral vector shGPD2 and shCtrl infection of H1299 and A549 cells,mRNA expression of GPD2 gene showed that the mRNA expression of GPD2 gene in shGPD2 group was significantly inhibited compared with shCtrl group(p<0.05).The reduction efficiency reached 73.5%(H1299)and 74.5%(A549);Western Blot results confirmed that the protein content of GPD2 gene was decreased in shGPD2 cells compared with shCtrl group.The proliferation of the twolung cancer cells was detected by CCK-8 assay after the lentiviral interference vector shGPD2 and shCtrl were infected with H1299 and A549 cells.Cell clone formation experiments showed that the number of cell clones in shGPD2 group was significantly lower than that in shCtrl group(P<0.05).Cell cycle was detected by flow cytometry by PI-FACS method: results were compared with shCtrl group and shGPD2 group in G1 phase.There was a significant increase(P<0.05),indicating that the cell cycle of both lung cancer cells was arrested in G1 phase after GPD2 gene knockdown;Annexin V-APC single staining flow cytometry to detect early cell apoptosis showed: In the shCtrl group,the number of early apoptosis in the shGPD2 group was significantly increased(P<0.001).Transwell results showed that the invasion and migration rate of both lung cancer cells decreased after GPD2 gene knockdown(P<0.05);The results of the scratch healing experiment to test the migration ability of lung cancer showed that the H1299 cell scratching rate was significantly decreased at 12 hours and 24 hours(P<0.05),and the A549 cell scratching rate was also significantly decreased at 24 hours and 48 hours(P< 0.05).Conclusions:GPD2 gene knockdown significantly inhibited the proliferation and clonality of lung cancer cells,leading to cell cycle arrest in G1 phase,promoting apoptosis,and significantly reducing cell invasion and migration.