Diallyl Disulfide Suppresses Aerobic Glycolysis By Down-regul Ating PKM2 In Gastric Cancer Cells

Aim:To identify the mechanisms that by which diallyl disulfide(DADS)regulates aerobic glycolysis by down-regulating PKM2 in gastric cancer cells.Materials and Methods: In this study,HGC-27 gastric cancer cells were cultured in vitro.With different concentrations of DADS(0mg/L?10mg/L?20mg/L?40mg/L)treatment of gastric cancer cells,the levels of glucose concentration and lactate production level in the medium were respectively examined with using a Glucose Colorimetric Assay Kit II and a Lactate Assay Kit,respectively.Cell lysates were analyzed by western blot using antibodies specific for PKM2 and LDHA,and the mRNA levels of PKM2 were determined by real-time PCR using specific primers.The optimal DADS concentration was selected for the next experiment.The best selection of PKM2 fragment,using RNA interference technology to silence the expression of PKM2 gene in HGC-27 cells,the experiment was divided into five groups(mock group,scramble group,sh-PKM2 group,sh-PKM2+DADS group and DADS group),and we detected the concentration of glucose and lactic acid in HGC-27 cells PKM2 silenced treated with DADS for 48 h,and the changes of PKM2 mRNA,PKM2 and LDHA protein expression at the same time.Results: 1.DADS inhibits glucose uptake and lactate production in gastric cancer cells.Inhibition in of anaerobic glycolysis was observed in diallyl disulfide-treated HGC-27 cells,with including an increased glucose concentration in the medium(P <0.05),decreased lactate production in the medium(P <0.05)and down-regulation of PKM2 and LDHA expression and showed a dosed-effect dependence.2.DADS regulates the expression of PKM2 and LDHA protein in gastric cancer cells.Compared with the blank group,the expression of PKM2 mRNA was down regulated(P < 0.05),and the expression level of PKM2 and LDHA protein decreased in a concentration dependent manner.3.PKM2 is the target gene of DADS.The results showed that the levels of glucose concentration in medium of the sh-PKM2+DADS group were up-regulated and it was not significantly regulated compared with sh-PKM2 group(p > 0.05),and the levels of lactate were up-regulated and it was not significantly regulated compared with sh-PKM2 group(p > 0.05).Futhermore,the PKM2 mRNA expression in the sh-PKM2+DADS group was of no significantly difference compared with the sh-PKM2 group(P >0.05),but were down-regulated compared with scrambled oligonucleotide-treated cells(P <0.05).Futhermore,compared with the mock group,the glucose concentration increased,the lactate concentration decreased in medium and PKM2 mRNA decreased of the DADS group,the difference was statistically significant(P<0.05).Compared with sh-PKM2 group,the expression of PKM2 and LDHA protein in sh-PKM2+DADS group was not significantly different.Compared with mock group,the expression of PKM2 and LDHA protein in sh-PKM2 group and sh-PKM2+DADS group were significantly decreased.Conclusion: DADS suppressed aerobic glycolysis in HGC-27 human gastric cancer cells by down-regulation of PKM2.