组蛋白去乙酰化酶抑制剂Apicidine对宫颈癌HeLa细胞的抑制作用及机制研究

目的:探讨组蛋白去乙酰化酶抑制剂Apicidine对宫颈癌细胞的杀伤作用和机制。方法:用不同浓度的Apicidine作用于体外培养的宫颈癌细胞株HeLa和正常呼吸道上皮细胞REC;用MTT法检测细胞生长存活率;用流式细胞术(FCM)定量检测细胞凋亡及细胞周期的变化;用RT-PCR法和Western blot法分析Apicidine作用后宫颈癌细胞中p21WAF1和p53表达的变化。结果:Apicidine纳摩尔级浓度即能有效地抑制宫颈癌细胞增殖,对正常细胞无明显抑制效应。FCM结果表明,Apicidine能显著诱导宫颈癌细胞发生凋亡,对正常细胞无明显促凋亡作用;加入2μmol/L Apicidine作用24h和48h后宫颈癌细胞HeLa凋亡率分别为(10.11±0.63)%和(23.28±0.34)%,差异有显著性(P<0.05),正常呼吸道上皮细胞REC凋亡率分别为(5.81±0.92)%和(6.8±0.55)%,差异无显著性(P>0.05);FCM结果亦表明,Apicidine主要引起G0/G1期细胞周期阻滞,1μmol/L Apicidine处理24h后HeLa细胞G0/G1期细胞显著增多,由(46.8±3.2)%增至(69.5±6.1)%,差异有统计学意义(P<0.05);RT-PCR和Western blot结果显示,Apici-dine能显著诱导p21WAF1RNA和蛋白水平表达,对p53表达无明显影响。上述结果都呈现明显的量-效与时-效关系。结论:Apicidine在体外能有效地抑制人宫颈癌细胞生长,对人正常细胞无明显影响,其抗肿瘤生长机制之一可能是通过上调p21WAF1蛋白水平和引起G0/G1期细胞周期阻滞实现的。

Inhibitory effect of Apicidine on ovarian carcinoma cells and its mechanism.

Objective:To investigate antitumor activity of histone deacetylase inhibitor Apicidine against human cervical cancer cells and its mechanism.Methods:Human cervical cancer Hela cells and normal respiratory endothelial cells REC were treated with different concentration of Apicidine for different periods of time.After treatment,viable cell rate was measured by MTT assay.Cell apoptosis and changes in cell cycle were examined by means of flow cytometry(FCM).p21WAF1 and p53 expression was assessed by RT-PCR and Western blot.Results:Apicidine inhibited the proliferation of cervical cancer cells significantly at nanomolar concentrations,while there was no evident inhibitory effect of Apicidine on normal cells.FCM assay showed that Apicidine could induce apoptosis in Hela cells,but not in normal cells.After treatment with 2 μmol/L Apicidine for 24 h and 48 h,the apoptotic rate of Hela cells was(10.11±0.63) % and(23.28±0.34) %,(P<0.05),whereas the apoptotic rate of REC was(5.81±0.92)% and(6.8±0.55)%,(P<0.05).FCM assay also showed that Apicidine mainly caused cell cycle arrest at G0/G1 phase.After treatment with 1μmol/L Apicidine for 24 h,the number of G0/G1 phase Hela cells markedly increased from(46.8±3.2) % to(69.5±6.1) %,(P<0.05).In addition,Apicidine could greatly induce mRNA and protein expression of p21WAF1 but no changes in p53 expression as shown by RT-PCR and Western blot.Above results are in a time-and dose-dependent fashions.Conclusion:Apicidine is able to effectively inhibit the growth of cervical cancer cells in vitro,while there is no evident inhibitory effect of Apicidine on normal cells.One of the antitumor mechanisms of Apicidine is most likely through induction of p21WAF1 expression and subsequent arrest of cell cycling at G1 phase.