Investigation On Bioactivities And Mechanism Of Crocetin On SGC7901 Cells Of Gastric Cancer

Objective:Gastric cancer SGC7901 cells were used as an in vitro experimental model of gastric cancer to investigate the efficacy and mechanism of crocin-1,crocetin,and geniposide on gastric cancer SGC7901 cells.Methods:CCK8 method was used to detect the inhibitory effect of drugs on the proliferation of gastric cancer cell line SGC7901.12.5μmol/L、25μmol/L、50μmol/L doses of crocetin acted on SGC7901 cells for 48 h.Flow cytometry was used to detect the cells.Apoptosis and changes in mitochondrial membrane potential of cells;Detection of Caspase-3 activity by microplate reader;Real-time fluorescent quantitative PCR to detect Bcl-2 and Bax gene expression;Western blot to detect cleaved Caspase-3,Bcl-2,and Bax proteins expression.Results:1 The inhibitory effect of drugs on the proliferation of SGC7901 cellsCrocetin has a significant proliferation inhibitory effect on SGC7901 cells,and has a time-concentration effect relationship.After 24 hours of SGC7901 cells treated with crocin-1,crocetin,and geniposide,IC₅₀0 was 2229.90μmol/L,44.57μmol/L and 623.37μmol/L.Data comparison shows that the IC₅₀0 of crocetin is significantly lower than that of crocin-1 and geniposide,and the anti-stomach effect of crocetin is significantly better than crocin-1 and geniposide.2 The apoptotic effect of crocetin on SGC7901 cellsUnder ordinary light microscope,the SGC7901 cells in the control group had good adherence,full body and regular shape.In the crocetin group,the cell growth was significantly inhibited,the cell density was significantly lower than that in the control group,and the cell morphology changed,including:the cells became smaller and shrunk,rounded and floating particulate matter appeared in the culture medium.In addition,crocetin induced apoptosis in SGC7901 cells in a concentration-dependent manner.Low,medium,and high dose crocetin treatment of SGC7901 cells for 48 h resulted in apoptotic rates of 21.41±1.13%,28.28±2.42%,39.83±2.36%,which were statistically significant compared with the control group（p<0.01）.3 Effects of crocetin on mitochondrial membrane potential of SGC7901cellThe Mitochondrial Membrane Potential Kit（JC-1）was used to detect the effects of crocetin on the mitochondrial membrane potential of gastric cancer SGC7901 cells.The results showed that the mitochondrial membrane potential of SGC7901 cells treated with12.5μmol/L、25μmol/L and 50μmol/L crocetin for 48 h decreased significantly compared with the control group（p<0.05,p<0.01）.4 Effects of crocetin on Caspase-3 activity in SGC7901 cells12.5μmol/L,25μmol/L,50μmol/L crocetin treated SGC7901 cells for 48 h,and Caspase-3 activity increased significantly.The low,medium,and high doses of crocetin Caspase-3 activity were 4.88±0.34,7.23±0.31,and 11.66±0.57,which were statistically significant compared with the control group（p<0.01）.5 Effects of crocetin on Bcl-2 and Bax gene expression in SGC7901 cellsThe results of real-time quantitative PCR showed that the expression of Bcl-2 gene in crocetin group was significantly reduced,and the expression of Bax gene was significantly increased.6 Effects of crocetin on the expression of SGC7901 cell-related proteinsWestern blot results showed that Bax and cleaved Caspase-3 protein expression levels were significantly increased in the crocetin group,while Bcl-2 protein expression levels were significantly decreased.Conclusion:1.Gardenia has inhibitory effect on gastric cancer cell SGC7901.Among them,crocin-1and geniposide have no obvious anti-gastric pharmacological activity,while crocetin has significant anti-gastric cancer effect.The main ingredient crocin may be metabolized to crocetin to exert anti-gastric cancer effect,crocetin may be one of the important active molecules of crocin in anti-gastric cancer in vivo.2.The crocetin induced apoptosis of SGC7901 cells may be through activating Caspase-3,up-regulating the expression of cleaved Caspase-3 and Bax proteins,and down-regulating Bcl-2 protein,thereby activating the mitochondrial-mediated apoptosis pathway and reducing cell mitochondrial membrane potential.Eventually induce apoptosis to exert anti-gastric cancer efficacy.