The Role Of Phenylmetformin In Glioma Cells And Its Related Molecular Mechanisms

Objective:Glioma has the characteristics of fast growth,high degree of malignancy,poor prognosis and easy recurrence.The treatment and prognosis of glioma are still facing great challenges.Related studies have shown that phenylmetformin has anti-tumor effects,but its role in glioma is still unclear.The purpose of this study was to investigate the role of phenylformin in glioma cells and its related molecular mechanisms.Methods:Glioma cell lines U87 and U251 were selected as research objects to determine the IC50 of phenylmetformin on glioma cells.CCK8 kit was used to detect the effects of different concentrations(0-1000um)of phenylformin on the proliferation of glioma cells at different time points(12h,24 h,48h).The effect of phenylmetformin on glioma cell migration was detected by scratch healing experiment.Transwell method was used to detect the effect of phenylmetformin on the invasion of glioma cells.The effect of phenylmetformin on cell cycle and apoptosis of glioma was detected by flow cytometry.TCGA and CGGA databases were used to analyze the expression levels of SKP2 in patients with glioma of different levels and the relationship between SKP2 and clinical prognosis.RNA and protein levels of SKP2 in glioma cells treated with phenylformin were detected by rt-pcr and Western blot.Meanwhile,the protein levels of p-ampk,p-mtor,S6 K and PPARy were detected.Results:1.The IC50 of phenylmetformin on U87 and U251 glioma cells were 564.1kb and 523.7kb,respectively.Phenylmetformin inhibited the proliferation of both types of glioma cells in a concentration-dependent and time-dependent manner.The results of cell cycle experiments showed that the cell cycle of glioma was blocked in G0/G1 phase after treatment with phenylmetformin.Moreover,the results of apoptosisexperiments showed that phenylformin increased the proportion of apoptosis.In addition,phenylmetformin can significantly inhibit the invasion and migration of U87 and U251 cells.2.The higher the grade of glioma(grade II,III,IV),the higher themRNA expression level of SKP2,and the survival period of patients in the SKP2high-expression group was significantly lower than that in the SKP2 low-expression group(P<0.05).3.The expression level of SKP2 in glioma cells significantly decreased after treatment with phenylformin(P<0.05).Compared with the control group,the expression level of p-ampk increased significantly.The expression levels of p-mtor,S6 K and PPARy decreased significantly,and the difference was statistically significant(p <0.05).Conclusion:1.Phenylformin inhibits the proliferation,migration and invasion of glioma cells and promotes their apoptosis.2.Phenylmetformin reduces SKP2 gene and protein levels in glioma cells.3.Phenylmetformin may inhibit the proliferation,migration and invasion of glioma cells by down-regulating the expression of glioma cells' SKP2 and activating AMPK/mTOR signaling pathway,which may be the potential mechanism of the effect of phenylmetformin on glioma cells.