Multi-components from Pulsatilla Chinensis saponins synergistically inhibit proliferation of NCI-H460 cells based on tumor glycolysis mechanism / 中草药

ABSTRACT

Objective: The study was designed to investigate the antitumor effect of Pulsatilla chinensis saponins components and the synergistic effect of its effect components on tumor glycolysis. Methods: NCI-H460 cell lines were cultured in vitro, MTT assay was used to detect the inhibitory rate of various components of Pulsatilla chinensis saponins. The compatibility and synergistic antitumor effect of different effect components were studied by Calcusyn 3.0 software. Biochemical assay and Elisa assay were used to detect the glycolysis related metabolites (pyruvate, lactic acid, and glucose) and enzymes (GLUT1, HK, PKM2, and LDHA). Energy metabolism related gene members from KEGG pathway database and key protein (ERK1/2, Ras, GLUT1, MCT4) were detected by qRT-PCR and Western blotting. Results: The antitumor results in vitro showed that PSD, R13 and PSA in Pulsatilla chinensis saponins had the strongest antitumor activity for human lung cancer NCI-H460 cells and the IC50 were 5.2, 4.6, and 7.9 μg/mL, respectively. Using Calcusyn 3.0 software to confirm the distribution ratio of three effective monomers in Pulsatilla chinensis saponins and determine the synergistic antitumor effect after compatibility. The results of biochemical and Elisa assay showed that the content of pyruvate, lacate, glucose, HK, PKM2 and LDHA were significantly decreased and the GLUT1 content was significantly increased compared with the blank group (P < 0.05),; Compared with each monomer group, the content of pyruvic acid, lactic acid, glucose, HK, PKM2 and LDHA in the combined group were significantly decreased and the content of GLUT1 was significantly increased (P < 0.05). Real-time PCR results show that the combinations group has the most nodes in the network diagram, and its target was more than each monomer group. Western blotting results showed that compared with the blank group, the combinations group significantly decreased the expression of ERK1/2, Ras, GLUT1, and MCT4 protein (P < 0.05). Conclusion: The combination of effect components of Pulsatilla chinensis saponins has synergistic antitumor effect on NCI-H460 cells and the antitumor mechanism may be associated with the regulation of glycolysis.