Knocking Down AGPAT4 Inhibits The Development Of Human Lung Adenocarcinoma By Promoting Ferroptosis

Objective: As of now,non-small cell lung cancer(NSCLC)is still one of the cancers with the highest incidence in the population,and due to the high mortality rate of NSCLC,NSCLC is a serious threat to the health of our people.Ferroptosis is a new type of programmed death process different from apoptosis and necrosis.It is characterized by lipid metabolism disorder,iron metabolism disorder,and oxygen metabolism disorder,which leads to accumulation of lipid reactive oxygen species and attacks intracellular biological macromolecules.Previous studies have shown that the unique physiological structure of lung tissue makes lung tissue usually experience severe oxygen damage.Ferroptosis may play an important role in the occurrence and development of NSCLC,but there are few relevant research reports.Therefore,we hope to screen out related proteins that can regulate the ferroptosis process and affect the occurrence and development of NCLSC through our research.Methods: 1,Screen out the proteins involved in oxygen metabolism through bioinformatics methods.2,Screen the protein AGPAT4 related to NCLSC by prognostic analysis.3,DCFH-DA probe was used to select suitable NSCLC cell lines to establish cell model.4,Establish an over-expression and knock-down AGPAT4 cell line in a suitable NCLSC cell line,and use Western Blot to detect the expression efficiency.5,Use DCFH-DA and Liperfluo probes to detect the changes of intracellular reactive oxygen species after overexpression and knockdown of AGPAT4.6,Use CCK-8 experiment to detect changes in cell activity of over-expression and knock-down AGPAT4 cell lines.7,Use JC-1 experiment to detect the changes of cell mitochondrial membrane potential after overexpression and knockdown of AGPAT4.8,MDA assay was used to detect the difference in the degree of lipid metabolism disorder between overexpression and knockout of AGPTAT4 cells.9,Use Western Blot experiment to detect the changes of ferroptosis-related protein Maker after overexpression and knockdown of AGPAT4.10,After treating the AGPAT4knock-down cell line with apoptosis inhibitor,necrosis inhibitor,and Ferroptosis inhibitor,use DCFH-DA probe to detect the level of cell reactive oxygen species.11,After using apoptosis inhibitor,necrosis inhibitor,ferroptosis inhibitor to treat AGPAT4 knock-down cell line,CCK-8 experiment to detect the change of cell activity.12,Use the human lysophosphatidic acid(LPA)Elisa detection kit to detect the changes in LPA levels in the AGPAT4 knockdown group.13,Use the human coenzyme Q10(Co Q10)Elisa detection kit to detect the changes of Co Q10 levels in the AGPAT4 knockdown group.Results: In the research of this subject,we found that in the NSCLC cell line A549,after knocking down AGPAT4,the level of intracellular reactive oxygen species increased,cell activity decreased,mitochondrial membrane potential level decreased,and lipid metabolism disorder level increased,The expression of Ferroptosis protein Maker GPX4 and FSP1 is down-regulated;and apoptosis inhibitors and necrosis inhibitors cannot restore the increased levels of reactive oxygen species and cells produced after AGPAT4 knockdown.Conclusion: We found that AGPAT4 promotes the malignant progression of lung cancer by inhibiting the expression of iron death markers GPX4 and FSP1.