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目的:观察尾叶香茶菜二萜类化合物B(DB)对人宫颈癌细胞株Hela细胞的生长抑制作用并探讨其机制。方法:处于对数生长期的Hela细胞分为正常对照组,0.5、1.0、2.0、4.0、8.0、16.0、32.0mg.L-1DB实验组,MTT法观察不同剂量DB作用24和48h后Hela细胞增殖抑制率,相差显微镜观察DB作用后Hela细胞形态的改变,RT-PCR和Western blotting分别观察DB作用后Hela细胞P53、P21、CDK2mRNA和蛋白表达水平。结果:MTT检测,24h时2.0、4.0、8.0DB组Hela细胞增殖抑制率分别为18.18%、28.89%和32.84%,48h时2.0、4.0、8.0DB组Hela细胞增殖抑制率分别为26.64%、47.03%和51.90%,呈时间-剂量依赖性(P<0.05或P<0.01);形态学观察,DB作用后Hela细胞变圆、体积缩小,有部分细胞漂浮;2.0、4.0和8.0mg.L-1DB分别作用Hela细胞24和48h后,P53和P21mRNA和蛋白表达水平明显高于正常对照组(P<0.01),CDK2mRNA和蛋白表达水平明显低于正常对照组(P<0.01)。结论:DB可抑制Hela细胞生长,其机制可能与上调细胞周期相关基因p53、p21的表达,下调CDK2表达,从而影响细胞从G1期进入S期有关。
OBJECTIVE: To observe the inhibitory effect of diterpenoid B (DB) on the growth of human cervical cancer Hela cells and to explore its mechanism. Methods: Hela cells in logarithmic growth phase were divided into normal control group, 0.5,1.0,2.0,4.0,8.0,16.0,32.0 mg.L-1DB experimental group, MTT method was used to observe the effect of different doses of DB on Hela cells Proliferation inhibition rate and phase contrast microscope were used to observe the morphological changes of Hela cells after treated with DB. The mRNA and protein expressions of P53, P21 and CDK2 in Hela cells were detected by RT-PCR and Western blotting respectively. Results: The MTT assay showed that the inhibitory rates of proliferation of Hela cells in 2.0, 4.0 and 8 DB groups were 18.18%, 28.89% and 32.84% at 24 h, respectively. The inhibition rates of Hela cells in 2.0, 4.0 and 8 DB groups were 26.64% and 47.03 (P <0.05 or P <0.01). The morphology of Hela cells became round after treatment with DB, the volume of the Hela cells decreased and some cells floated. The cells of 2.0, 4.0 and 8.0 mg.L- 1D respectively, the mRNA and protein expressions of P53 and P21 in Hela cells were significantly higher than those in normal control group (P <0.01). The expression of CDK2 mRNA and protein were significantly lower than those in normal control group (P <0.01). Conclusion: DB can inhibit the growth of Hela cells. The mechanism may be related to up-regulating the expression of p53 and p21 and down-regulating the expression of CDK2, thus affecting the cell cycle from G1 phase to S phase.