Experimental Study Of 3-bromopyruvate Inhibiting The Glycolysis In Apoptosis Of Lung Adenocarcinoma Cancer Cells

Objectives: To investigate the inhibitory effect of 3-bromopyruvate(3-BP)on the proliferation and apoptosis of human lung adenocarcinoma cells(m A549 and H1975)and to explore the changes of glycolysis and energy metabolism in the process of 3-BP acting on human lung adenocarcinoma cells.Methods: The lung adenocarcinoma cell line m A549 and H1975 were treated with different concentration of 3-BP,the cell viability was detected by MTT assay.Morphological changes of the two lung adenocarcinoma cells were observed under microscope.Cell colony formation assay was used to check the growth inhibition induced by 3-BP in m A549 and H1975 cells.Scratch and Transwell assays were used to detect the effect of different concentration of 3-BP on the migration ability of m A549 cells and H1975 cells.Annexin V-FITC/PI double staining assay was used to examine the cell apoptosis rate induced by different concentration of 3-BP on lung adenocarcinoma cells m A549 and H1975.ATP and active oxygen test kits were used to detect the changes of ATP level and ROS level in lung adenocarcinoma cells m A549 and H1975.Western blot assay was used to detect the expression of apoptosis and glycolysis related proteins by different concentrations of 3-BP on lung adenocarcinoma cells m A549 and H1975.Heterotopic transplanted tumor model of lung adenocarcinoma cancer was established to observe the antitumor effect of 3-BP in vivo.Results: MTT results showed that 3-BP could inhibit the proliferation of lung adenocarcinoma cells m A549 and H1975.The cytoplasm of lung adenocarcinoma cells was contracted and the gap between cells became larger with 3-BP treatment.The number of colony formation of lung adenocarcinoma cells was reduced by 3-BP.The results of the migration assay showed that compared with the negative group,the scratch healing rate of the groups with 3-BP treatment were decreased significantly.With the increased of the concentration of 3-BP,the migration ability of the cells was gradually decreased.Compared with the negative group,the number of the cell penetrating membrane of the lung adenocarcinoma cells decreased significantly after 3-BP treatment.The results of double staining assay showed that 3-BP could induce apoptosis of two kinds of lung adenocarcinoma cells.The level of ATP in the group with 3-BP treatment was significantly lower than the negative group.The level of active oxygen in the group with 3-BP treatment was significantly higher than the negative group on lung adenocarcinoma cells m A549 and H1975 so that 3-BP induced cell apoptosis may be related to the increase of active oxygen level.Compared with the negative group,the expression of HK-Ⅱ,MCT1 and PDK protein were significantly decreased and the expression of Bax protein was significantly increased,and the expression of Bcl-2 protein was significantly decreased in the group with 3-BP treatment.The protein level also showed that 3-BP could induce cell apoptosis of lung adenocarcinoma cells by inhibiting glycolysis.The nude mice model of heterotopic lung adenocarcinoma transplantation tumor with double fluorescence labeling was established.The nude mice were randomly divided into three groups: PBS group,DDP group and 3-BP group.The results showed that 3-BP could inhibit the growth of lung adenocarcinoma transplanted tumor in nude mice.HE and immunohistochemistry results showed that 3-BP could inhibit the proliferation of tumor cells and induce its apoptosis.At the same time,the results of liver and kidney function assays showed that compared with PBS group,3-BP was slightly toxic to liver and kidney.Conclusion: 3-BP can inhibit the proliferation of human lung adenocarcinoma cells m A549 and H1975 by inhibiting glycolysis and induce the apoptosis of lung adenocarcinoma cells.3-BP has good antitumor effect and low hepatorenal toxicity on lung adenocarcinoma in vivo.