PTC596对脑胶质瘤细胞生物学行为的影响及机制研究

The influence and mechanism of PTC596 on the biological behavior of brain glioma

目的探讨BMI-1小分子抑制剂PTC596对脑胶质瘤细胞生物学行为的影响,并探索其机制。方法以0、0.25、0.5、1μmol/L的PTC596刺激脑胶质瘤U251细胞,CCK8法检测24及48h后的细胞存活率,克隆形成实验检测细胞克隆形成能力变化,划痕实验观察细胞迁移能力变化,流式细胞术检测细胞凋亡和细胞周期分布情况,荧光定量PCR法及westblot法检测U251细胞的BMI-1、MCL-1、XIAP、Bcl-2及CyclinD1表达量的变化。结果随着PTC596药物浓度的提高及刺激时间的延长,U251细胞存活率较对照组显著下降,差异均有统计学意义(all P<0.05)。各个PTC596药物浓度组,细胞克隆形成数目和迁移率均较对照组显著减少,差异均有统计学意义(all P<0.05);U251细胞凋亡率与对照组相比,差异均有统计学意义(all P<0.05);G0/G1期细胞比例高于对照组,差异均有统计学意义(all P<0.05)比例增加,S期及G2/M期细胞比例低于对照组,差异均有统计学意义(all P<0.05)。与对照组比较,0.5μmol/L PTC596作用的细胞中BMI-1、MCL-1、XIAP、Bcl-2及CyclinD1基因表达均下降,差异有统计学意义(均P<0.05),对应的蛋白表达量均减少。结论 PTC596抑制U251细胞增殖,减弱脑胶质瘤细胞迁移及克隆形成能力,诱导脑胶质瘤细胞凋亡并阻碍细胞生长周期进展,其机制与PTC596抑制BMI-1及其下游因子的表达及调控相关。

Objective To investigate the effect of BMI-1 small molecule inhibitor PTC596 on the biological behavior of glioma cells, and explore its mechanism. Methods UTC cells of glioma U251 were stimulated with PTC596 at 0, 0.25, 0.5, 1 μmol/L, CCK8 method was used to detect cell survival rate after 24 and 48 hours. The changes of cell clone formation ability were detected by clone formation test, and the changes of cell migration ability were observed by scratch test. Cell apoptosis and cell cycle distribution were detected by flow cytometry. The expression levels of BMI-1, Mcl-1, XIAP, Bcl-2 and CyclinD1 in U251 cells were detected by fluorescence quantitative PCR and Westblot. Results With the increase of PTC596 drug concentration and prolongation of stimulation time, the survival rate of U251 cells decreased significantly compared with the control group, and the differences were statistically significant(all P<0.05). In each PTC596 drug concentration group, the number of cell clone formation and migration rate were significantly reduced compared with the control group, the difference was statistically significant(all P<0.05);the apoptosis rate of U251 cells was statistically significant compared with the control group(All P<0.05);G0/G1 phase cell ratio is higher than the control group, the difference is statistically significant(all P<0.05) ratio increases, S phase and G2/M phase cell ratio is lower than the control group, the difference is both Statistical significance(all P<0.05). Compared with the control group, the expression of BMI-1, MCL-1, XIAP, Bcl-2 and CyclinD1 genes in cells treated with 0.5 μmol/L PTC596 all decreased, the difference was statistically significant(all P<0.05), and the corresponding protein expression The amount is reduced. Conclusion PTC596 inhibits the proliferation of U251 cells, weakens the ability of glioma cell migration and clone formation, induces glioma cell apoptosis and hinders the progress of the cell growth cycle. The mechanism and PTC596 inhibit the expression and regulation of BMI-1 and its downstream factors Related.