The Study On The Ferroptosis Induced By Artesunate Via System Xc-/ GSH/ GPX4 Pathway In Colon Cancer Cells And Its Relationship With The Prognosis Of Colon Cancer

Objective: To explore the ferroptosis induced by artesunate in colon cancer cells,and to preliminarily elucidate the mechanism of ferroptosis.To further investigate the expression of glutathione peroxidase 4(GPX4)in colon cancer tissue and its relationship with clinicopathological parameters,and to expound its potential clinical significance of GPX4 in the diagnosis and treatment of colon cancer.Methods: Colon cancer cells HCT116 were treated with artesunate,and Ferrostatin-1,an inhibitor of ferroptosis,and the specific inhibitor of programmed necrosis,necrostatin-1 and the apoptosis inhibitor z-vad-fmk were used to intervene the colon cancer HCT116 cells,CCK8 method(Cell Counting Kit-8,CCK-8)to detect the changes of cell vitality;the aggregation of reactive oxygen species(ROS)in cells was detected by flow cytometry;the changes of mitochondria in cells were observed by transmission electron microscope,and the changes of iron content were detected by iron detection kit.Western-blot was used to detect the changes of expression of solute carrier family seven members 11(SLC7A11)、glutathione peroxidase 4(GPX4)and glutathione(GSH).The colon cancer transcriptome data from The Cancer Genome Atlas(TCGA)database and the clinical data of 398 colon cancer patients and 39 normal subjects were downloaded,and the expression of GPX4 in colon cancer and the clinical information of colon cancer patients(gender,age,tumor grade,lymph node metastasis,tumor distant metastasis and clinical stage);immunohistochemical detection of GPX4 expression in colon cancer and adjacent tissues;gene enrichment analysis(KEGG and GO)to explore the possible pathways of GPX4 regulating colon cancer.Results: The cell viability of HCT116 cells decreased after artesunate treatment(P<0.05),and this effect could be partially reversed by Ferrostatin-1,necrostatin-1 and z-vad-fmk(P<0.05).After artesunate treated HCT116 cells,a large amount of intracellular ROS accumulated,the number and morphology of mitochondria were changed,and the iron content increased.The expression of SLC7A11,GSH and GPX4 in HCT116 cells were decreased after artesunate treatment.Bioinformatics results showed that GPX4 was highly expressed in colon cancer tissues and lowly expressed in paracancerous tissues(P<0.01);among colon cancer patients,the difference between the expression of GPX4 and the survival time of patients was statistically significant(P<0.05).However,there was no significant difference between the expression of GPX4 with age,lymph node metastasis,gender,T,N,M stage and disease stage(P>0.05).Immunohistochemical results showed high expression of GPX4 in colon cancer tissues.Conclusions: Artesunate may be through the classical ferroptosis pathway: that is,when the expression of SLC7A11 is downregulated,the intracellular GSH synthesis is reduced and the activity of GPX4 is ultimately decreased,lipid oxides cannot be metabolized by the reduction reaction catalyzed by GPX4,and then oxidized by divalent iron.A large number of reactive oxygen species are produced in colon cancer cells,which promotes ferroptosis in colon cancer HCT116 cells.The high expression of GPX4 is related to the poor prognosis in patients.The detection of GPX4 has certain clinical value in the diagnosis of colon cancer,which is helpful for the prognosis monitoring of colon cancer.