灵芝酸 A 对非小细胞肺癌 PC9 细胞糖酵解及其关键 限速酶的影响.

Objective: To discuss the effects of different doses of ganoderic acid A (GAA) on the biological activities, glycolysis, and the expression of rate-limiting enzymes of non-small cell lung cancer (NSCLC) PC9 cells, and to clarify the mechanism. Methods: The NSCLC PC9 cells were cultured in vitro and divided into blank control group, low dose (25 µmol·L-1) of GAA group, medium dose (50 µmol·L-1) of GAA group, and high dose (100 µmol·L-1) of GAA group. The methylthiazolydiphenyltetrazolium (MTT) assay was used to detect the survival rates of the PC9 cells in various groups; Transwell chamber assay was used to detect the number of migration cells of the PC9 cells in various groups; the glucose uptakes of the PC9 cells in various groups were detected by glucose assay kit; the levels of ATP in the PC9 cells in various groups were detected by ATP assay kit; the levels of lactic acid in the PC9 cells in various groups were detected by lactate assay kit; the expression levels of hexokinase 2 (HK2) and pyruvate kinase M2 (PKM2) mRNA in the PC9 cells in various groups were detected by real-time fluorescence quantitative PCR (RT-qPCR) method; the expression levels of HK2 and PKM2 proteins in the PC9 cells in various groups were detected by Western blotting method. Results: The MTT assay results showed that, at 24, 48, and 72 h of culture, compared with blank control group, the survival rates of the cells in medium and high doses of GAA groups were significantly decreased (P<0. 05). At 72 h of culture, compared with low dose of GAA group, the survival rate of the cells in high dose of GAA group was significantly decreased (P<0. 05). The Transwell chamber assay results showed that compared with blank control group, the numbers of migration cells in medium and high doses of GAA groups were significantly decreased (P<0. 05); compared with low doses of GAA group, the number of migration cells in high dose of GAA group was significantly decreased (P<0. 05). Compared with blank control group, the glucose uptakes and levels of lactic acid in the cells in medium and high dose of GAA groups were significantly decreased (P<0. 05), and the level of ATP in the cells in high dose of GAA group was significantly decreased (P<0. 05). The RT-qPCR results showed that compared with blank control group, the expression levels of HK2 and PKM2 mRNA in the cells in medium and high doses of GAA groups were significantly decreased (P<0. 05). The Western blotting results showed that compared with blank control group, the expression levels of HK2 and PKM2 proteins in the cells in medium and high doses of GAA groups were significantly decreased (P<0. 05). Conclusion: Medium and high doses of GAA can inhibit the biological activities of proliferation and migration of the PC9 cells, reduce the glycolysis, and its mechanism may be related to the inhibition of the expressions of the key rate-limiting enzymes HK2 and PKM2. [ABSTRACT FROM AUTHOR]