| BackgroundLung cancer had the highest morbidity and mortality of malignant tumor in our country and worldwide at present,its morbidity and mortality had an obvious rising trend [1-2],although the molecular targeted therapy has obtained the remarkable curative effect,The subsequent emergence of drug resistance had hindered its further development [3-4].In recent years,studies had shown that the glycolysis of tumor cells had significantly enhanced which played a critical role in tumor occurrence and development [5-6],a lot of researches displayed that we could inhibit the growth of malignant tumor according to inhibit the abnormal glycolysis of tumor cells,even could kill them,3-bromopyruvate(3-Br PA)as the inhibitor of hexose kinase had a pronounced effect on inhibiting the growth of malignant tumor,so it has been expected to be as a new antitumor drug.However,there was few research about the effect of 3-BrPA on lung cancer,and its precise mechanism was not clear recently.Therefore,this study intended to investigate the anti-proliferative effect of glycolysis inhibitor 3-bromopyruvate(3-Br PA)on lung cancer A549 cells in vitro and its possible mechanism.ObjectiveTo investigate the anti-proliferative effect of glycolysis inhibitor 3-Br PA or/and cisplatin on lung cancer A549 cells in vitro and its possible mechanism.Methods1.CCK-8(cell counting kit-8 CCK-8)was used to observe the proliferation of A549 and MRC-5 cells while different concentrations of cisplatin and/or 3-BrPA treated after 24 h,48h and 72 hours.2.Below the IC50(half maximal inhibitory concentration)of 3-Br PA was selected to combine with cisplatin(1mg / L)to deal with A549 cells.48 hours after dealt with 3-Br PA,the morphological changes of A549 cells were observed by the inverted phase contrast microscope,and flow cytometry was used to detect the apoptosis of A549 cells of each group;48 hours after different concentrations of 3-BrPA dealt with A549 cells,the cell cycle distribution of each group were analysised by flow cytometry 3.Hexokinase(HK)activity of each group were detected 48 hours after different concentrations of 3-Br PA dealt with A549 cells by HK test kits.Results1.Effect of 3-BrPA on the proliferation of A549 cells: compared with the negative control group that when the concentrations of 3-Br PA were higher than 20?mol / L,the anti-proliferation effect of 3-Br PA was significantly enhanced(P < 0.01),and the anti-proliferation effect was significantly enhanced with the increasing concentration of 3-BrPA and the prolonging action time.And compared with monotherapy groups,the proliferation of A549 cells in the combination groups were significantly enhanced(P<0.01).Effect of 3-Br PA on the proliferation of MRC-5 cells: CCK-8 assay results suggested that compared with the negative control group,the 3-BrPA had no obvious proliferation inhibitory effect on MRC-5 cells while the concentration ranged from 0 ~ 80?mol /L(P > 0.05),only when the concentration ranged from 80-160?mol / L had a slight inhibition,and the inhibitory rate were significantly lower than those of the A549 cells under the same conditions(P < 0.01).2.Inverted microscope showed that 48 hours after the drug dealt with A549 cells,the cells of the 3-BrPA group showed the morphological changes of apoptosis,the combination group showed more apoptosis cells and the morphological changes of apoptosis were more obvious,some areas even appeared cell debris,vacuolization and other signs of cell necrosis;The flow cytometry showed that compared with the the negative control group(8.103 + 2.590)%,the apoptosis rate of 3-Br PA group(53.417 + 5.259)%,cisplatin group(20.700 + 5.172)% and the combination group(77.953 + 3.587)% were significantly higher(P <0.01),and the apoptosis rate of the combination groups were significantly higher than those of the monotherapy groups(P<0.01);The flow cytometry analysis showed that 48 h after different concentrations of 3-Br PA dealt with A549 cells the proportion of cells at S phase were obviously decreased with the increasing of 3-Br PA concentration(P < 0.01),but the percentage of the G1 phase cells were significantly increased(P < 0.01),most of the A549 cells were arrested in G1 phase,while the A549 cells which arrested in G2 / M phase and S phase had decreased significantly,and the A549 cells proliferation were gradually decreased with increasing of the 3-Br PA concentration.3.48 h after different concentrations of 3-BrPA dealt with A549 cells: hexokinase activity detection results showed that in the medication groups the hexokinase activity were significantly lower than that of the negative control group and with the increasing of the 3-BrPA concentration,the cell hexokinase activity were gradually decreased.Conclusion1.3-BrPA could inhibit the proliferation of A549 cells,and when combined with cisplatin it could significantly enhance the anti-proliferative effect of cisplatin on A549 cells,while the low concentration of 3-Br PA had no obvious anti-proliferative effect on MRC-5 cells,only when turned to high concentration had a slight inhibition,suggesting that 3-BrPA could inhibit the growth of lung cancer cell lines and had little toxic effect on normal cells,and it may cause synergistic anti-tumor effect when combined with cisplatin.2.3-BrPA could induce A549 cells to apoptosis,it also could delay the cell cycle progression and then inhibit the A549 cells' proliferation.3.3-BrPA could decrease the hexokinase activity of A549 cells,and the cell hexokinase activity decreased gradually with the increasing of its concentration.These studies indicated that 3-Br PA could inhibit the proliferation of A549 cells in vitro,induce the A549 cells to apoptosis,delay the cell cycle progression,and at the same time had sensitizing effect of chemotherapy,while it had little toxic effect on normal cells.The mechanism may be that 3-Br PA could affect the energy metabolism by inhibiting the hexokinase activity of lung cancer cells and then blocking the glycolysis process. |
PDF Full Text Request