ABSTRACT
Objective: To investigate the effects of maslinic acid (MA) on the proliferation and autophagy in nasopharyngeal carcinoma CNE2 cells and to elucidate the regulatory role of PI3K/Akt/mTOR pathway in this process. Methods: The effect of MA on the proliferation of CNE2 cells was assessed by CCK-8. MDC staining of autophagic vacuoles was performed for autophagy analysis. Additionally, the levels of autophagy-and PI3K/Akt/mTOR-associated proteins were examined using western blot analysis. Results: MA significantly inhibited the proliferation of CNE2 cells in a dose-and time-dependent manner. MA displayed autophagy-inducing effect, as shown by the increased MDC-labeled vacuoles, up-regulated LC3-II/LC3-I ratio and Atg5, as well as the down-regulated p62 level after MA treatment. Moreover, we observed that MA inhibited the expression of PI3K-p110α and the phosphorylation of Akt and mTOR. Conclusion: MA inhibits the proliferation and induces the autophagy of CNE2 cells, the mechanism may be related to the PI3K/Akt/mTOR signaling pathway. These results imply that MA may be a potential anti-cancer agent for use in the treatment of nasopharyngeal carcinoma.
ABSTRACT
To study the effect of aqueous extracts of Yiqi Jiedu formula (YQ) on the proliferation of CNE2 cells in human nasopharyngeal carcinoma, and investigate its mechanism to provide a new theoretical basis for the clinical application of YQ. CNE2 cells were treated with different concentrations (0.125, 0.25, 0.5, 0.25 g·L⁻¹) of YQ, positive control medicine (cisplatin 4.0 mg·L⁻¹), inhibitor PD98059 (50 μmol·L⁻¹), activator isoproterenol hydrochloride (20 μmol·L⁻¹), activator isoproterenol hydrochloride (ISO)+YQ 0.5 g·L⁻¹. Then cell labeling by using real-time analyzer (RTCA) and CCK 8 method were used to detect cell proliferation activity, and the half inhibitory concentration (IC₅₀) was calculated. The cell cycle distribution was detected by fluorescence double dye flow cytometry PI staining, and Western blot method was used to detect the expression levels of related protein and MAPK/ERK signaling pathway. The results of RTCA and CCK-8 test showed that as compared with the control group, YQ group could effectively inhibit the proliferation of CNE2 cells (<0.01), with a dose and time dependence, and 48 h IC₅₀ value was 0.5 g·L⁻¹. The results of cell cycle showed that after 48 h of water extract treatment, the cell cycle was significantly changed, the proportion of G₀/G₁ was reduced, the ratio of G₂/M increased, and the cell cycle was in G₂/M period (<0.01). Western blot results showed that after 48 h treatment with different concentrations of aqueous extract, cell cycle-related proteins cyclinD1, cyclinD3 and CDK2 expression levels were down-regulated; MAPK/ERK signaling pathway related protein p-c-Raf, p-MEK, p-ERK1/2 expression level significantly lower as compared with the control group (<0.05). After adding activator and inhibitor in MAPK/ERK signaling pathway on this basis, the results showed that after adding activator ISO, cell proliferation was significantly higher than that in the Control group; the cycle related proteins cyclinD1, cyclinD3, and CDK2 expression levels were increased; at the same time, key protein p-c-Raf, p-MEK, p-ERK1/2 expression levels in the signal pathways were relatively increased. While after the addition of inhibitor PD98059, the cell proliferation was significantly lower than that in the Control group, and the expression level of corresponding protein was decreased, which was significantly different from the Control group (<0.05). So YQ could block cell cycle and inhibit the proliferation of CNE2 cells mainly by reducing the expression of MAPK/ERK signaling pathway key protein p-c-Raf, p-MEK and p-ERK1/2.