Influences of germacrone on proliferation,apoptosis and invasion of esophageal cancer cells by inhibiting JAK2/STAT3 signaling pathway
Objective To investigate the influences of germacrone on the proliferation,apoptosis and invasion of esophageal cancer cells by inhibiting Janus kinase 2(JAK2)/signal transducer and activator of transcription 3(STAT3)signaling pathway.Methods Meth-yl thiazolyl tetrazolium(MTT)assay was used to detect the survival rates of esophageal cancer cells Eca109 and KYSE450 and human esophageal epithelial cells Het-1A treated with germacrone at different concentrations including 0,10,20,40,80,160,and 320 μmol/L,respectively.Eca109 cells were divided into control group(conventional culture),low-dose germacrone group(40 μmol/L germacrone inter-vention for 24 h),medium-dose germacrone group(80 μmol/L germacrone intervention for 24 h),high-dose germacrone group(160 μmol/L germacrone intervention for 24 h),Coumermycin A1 group(10 μmol/L JAK2 activator coumermycin A1+160 μmol/L germacrone intervention for 24 h),and AG490 group(50 μmol/L JAK2 inhibitor AG490+160 μmol/L germacrone intervention for 24 h).Transwell assay was used to detect the migration and invasion of cells.Plate cloning assay was used to test cell cloning ability.Flow cytometry was applied to detect apop-tosis and cell cycle.Western blot was applied to detect the expression of JAK2,phos-JAK2(p-JAK2),STAT3,p-STAT3,Ki-67,Bax,Bcl-2,and p53 proteins.Results Compared with 0 μmol/L,the survival rates of Eca109 and KYSE450 cells decreased in a concentration-de-pendent manner under treatment with 10,20,40,80,160,and 320 μmol/L germacrone(all P＜0.05),and the half-inhibitory concentrations(IC50)were(98.18±2.12)μmol/L and(154.77±2.32)μmol/L,respectively,while there was no difference in the survival rate of Het-1 A cells treated with different concentrations of germacrone(all P＞0.05).Due to the more significant effect of germacrone on Eca1 09 cells,sub-sequent studies on the mechanism were conducted on Eca109 cells with germacrone treatment at concentrations of 40,80,and 160 μmol/L.For Eca109 cells,compared with the control group,the migration and invasion of cells,number of clones,and the protein expressions of p-JAK2,p-STAT3,Ki-67 and Bel-2 in the low-,medium-and high-dose germacrone groups all decreased(all P＜0.05),while the pro-portion of cells in the G0/G1 phase,early apoptosis rate,total apoptosis rate,and the protein expression of Bax and p53 all increased(all P＜0.05).Compared with the high-dose germacrone group,the migration and invasion of cells,number of clones,and the protein expression of p-JAK2,p-STAT3,Ki-67 and Bcl-2 all increased in the Coumermycin A1 group,while the proportion of cells in the G0/G1 phase,early apoptosis rate,total apoptosis rate,and the protein expression of Bax and p53 all decreased(all P＜0.05).Compared with the high-dose germacrone group,the migration and invasion of Eca109 cells,number of clones,and the protein expression of p-JAK2,p-STAT3,Ki-67 and Bcl-2 in the AG490 group reduced,while the proportion of cells in the G0/G1 phase,early apoptosis rate,total apoptosis rate,the pro-tein expression of Bax and p53 all increased(all P＜0.05).Conclusions Germacrone can inhibit the proliferation and invasion of esopha-geal cancer cells and promote cell apoptosis by inhibiting the JAK2/STAT3 pathway.
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