石蒜碱对胃癌AGS细胞恶性生物学行为的影响及机制研究OA

Objective To investigate the effects of lycorine on the malignant biological behaviors of AGS gastric cancer cells,and to preliminarily explore its possible action mechanism.Methods AGS cells were intervened with different concentrations of lycorine hydrochloride(LH)(0 μmol/L,0.015 μmol/L,0.062,5 μmol/L,0.25 μmol/L,1 μmol/L,4 μmol/L,16 μmol/L)for 0 hour,24 hours,and 48 hours,and its inhibitory concentration 50(IC50)at the 48th hour of the intervention was calculated by the cell counting kit-8(CCK-8)test to determine the LH concentration in subsequent trials.AGS cells were divided into a control group(treated with 0.1％dimethyl sulfoxide),an LH-L group(treated with 0.125 μmol/L LH),an LH-M group(treated with 0.5 μmol/L LH),or an LH-H group(treated with 2 μmol/L LH),as well as a microRNA(miRNA)-675-nc group(transfected with blank lentiviral vectors),a miRNA-675-mimic group(transfected with lentiviral vectors overexpressing miRNA-675),or an LH+miRNA-675-mimic group(transfected with lentiviral vectors overexpressing miRNA-675+treated with 2 μmol/L LH).CCK-8 test,clonogenic assay in plates,cell scratch healing assay,and Transwell test were separately used to detect the proliferation,migration,and invasion abilities of cells in each group;real-time fluorogenic quantitative PCR was used to detect the relative expression levels of miRNA-675 and glycogen synthase kinase(GSK)-3β mRNA;western blotting was used to detect the relative protein expression levels of GSK-3β and β-catenin.Results In the range of 0.015-16 μmol/L,LH could inhibit the proliferation ability of AGS cells in a time-concentration dependent manner(P＜0.05),and its IC50 was(0.81±0.13)μmol/L at the 48th hour of the intervention.The number of cell colonies after 14 days of intervention,the scratch healing rates 24 hours and 48 hours after the intervention,the numbers of invasive cells and migrated cells 48 hours after the intervention,and the relative expression level of miRNA-675 48 hours after the intervention in the control group,LH-L group,LH-M group,and LH-H group decreased successively;48 hours after the intervention,the relative expression level of GSK-3β mRNA was higher in the LH-M group than in the control group,and the relative expression level of GSK-3β mRNA was higher in the LH-H group than in the control group,LH-L group,and LH-M group;48 hours after the intervention,the relative protein expression level of GSK-3β was higher in the LH-M group than in the control group,and the relative protein expression level of GSK-3β was higher in the LH-H group than in the control group and LH-L group;48 hours after the intervention,the relative protein expression level of β-catenin was lower in the LH-H group than in the control group and LH-L group(all P＜0.05).The optical density(48 hours),number of cell colonies(14 days),numbers of invasive cells and migrated cells(24 hours),and relative protein expression level of β-catenin(48 hours)were higher in the miRNA-675-mimic group than in the miRNA-675-nc group and LH+miRNA-675-mimic group;the relative protein expression level of GSK-3β(48 hours)was lower in the miRNA-675-mimic group than in the miRNA-675-nc group and LH+miRNA-675-mimic group(all P＜0.05).Conclusion Lycorine can effectively inhibit the proliferation,invasion,and migration abilities of AGS gastric cancer cells,which may be achieved by its regulation on the miRNA-675/GSK-3β/β-catenin axis.