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卵巢癌是女性生殖系统常见的恶性肿瘤之一,其中上皮性卵巢癌(epithelial ovarian cancer,EOC)的死亡率最高,占所有妇科疾病的90%左右,严重危害女性的生命健康[1-2]。绝大多数病人查出EOC时为晚期,且经治疗后存活率较低。EOC治疗大多以化疗与外科手术为主,而顺铂是治疗癌症最重要的化疗药物,本研究以顺铂为例,研究EOC细胞对其耐药机制。Panobinostat(LBH589)作为小分子抑制剂,是高效的组蛋白去乙酰化酶的一种,可以阻断多种疾病(如癌症)通路,降低癌细胞的存活率,诱导癌细胞的凋亡。作为促进细胞增殖、生长、抑制细胞凋亡的信号转导通路, 磷脂酰肌醇-3-激酶/蛋白激酶B(phosphatidylinositol-3-kinase/protein kinase B,PI3K/AKT)是细胞生存的重要通路之一。近年研究[2-4]表明PI3K/AKT通路与EOC细胞的耐药性有一定的相关性。本实验以EOC细胞为研究对象,探讨低剂量LBH589通过PI3K/AKT通路的诱导对EOC细胞凋亡的影响及低剂量LBH589对EOC细胞的逆转耐药机制。
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在同一条件下,对照组的EOC细胞的凋亡指数(3.86±1.26)%,明显低于实验组的(8.45±1.02)%(t=49.04,P < 0.01)。LBH589作为一种广谱HDAC抑制剂,含有低剂量LBH589的实验组比无LBH589作用的对照组凋亡指数明显升高(P < 0.01)(见图 1)。
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结果显示,含有低剂量LBH589实验组的EOC细胞的S期数量明显低于对照组(P < 0.01)(见表 1)。
分组 G1 S G2 对照组 53.36±1.02 42.26±1.35 20.29±1.32 实验组 63.32±1.32 18.27±1.66 9.12±1.05 t 10.34 19.42 11.47 P < 0.01 < 0.01 < 0.01 表 1 2组EOC细胞周期变化(ni=3;x±s;%)
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结果显示,实验组12、24、36、48 h抑制率分别为(12.35±0.27)%、(21.24±0.33)%、(33.54±0.26)%、(41.23±0.52)%,对照组抑制率始终为0。
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低剂量LBH589作用的EOC细胞中PI3K/AKT、p-PI3K、p-AKT的表达比对照组表达明显减少(P < 0.01),即低剂量LBH589会降低PI3K,AKT在卵巢癌细胞中的表达并抑制其磷酸化,从而逆转EOC细胞对顺铂的耐药性(见图 2、表 2)。
分组 PI3K AKT p-PI3K p-AKT 对照组 2.33±0.08 3.12±0.64 3.54±0.78 2.48±0.22 实验组 0.93±0.24 1.55±0.36 1.06±0.23 0.96±0.31 t 9.59 3.703 5.282 6.926 P < 0.01 < 0.05 < 0.01 < 0.01 表 2 2组蛋白表达量(ni=3;x±s)
低剂量LBH589通过PI3K/AKT途径诱导对上皮性卵巢癌细胞凋亡作用机制研究
Study on the mechanism of apoptosis induced by low-dose LBH 589 through PI3K/AKT pathway in epithelial ovarian cancer cells
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摘要:
目的探讨低剂量LBH589通过PI3K/AKT途径诱导对上皮性卵巢癌细胞凋亡作用机制的影响。 方法以上皮性卵巢癌细胞OVCAR-3为研究对象,使用TUNEL和流式细胞术,分别检测添加(实验组)与不添加(对照组)低剂量LBH589上皮性卵巢癌细胞的细胞凋亡情况与周期变化;MTT法检测低剂量LBH589对上皮性卵巢癌细胞增殖的抑制情况;Western blotting检测2组中PI3K和AKT表达情况以及其磷酸化的情况。 结果实验组细胞凋亡指数明显高于对照组的(3.86±1.26)%(P < 0.01),上皮性卵巢癌细胞S期数量(18.27±1.66)%,明显低于对照组的(42.26±1.35)%(P < 0.01),实验组12、24、36、48 h抑制率分别为(12.35±0.27)%、(21.24±0.33)%、(33.54±0.26)%、(41.23±0.52)%,对照组抑制率始终为0;实验组的EOC细胞对顺铂的敏感性显著增加(P < 0.01),逆转系数是对照组的3.26倍,明显抑制PI3K和AKT在卵巢癌细胞中的表达并抑制其磷酸化(P < 0.01)。 结论低剂量LBH589能增加上皮性卵巢癌细胞OVCAR-3的凋亡,并可能通过PI3K/AKT通路逆转顺铂耐药。 -
关键词:
- 卵巢肿瘤 /
- LBH589 /
- PI3K/AKT通路 /
- 细胞凋亡 /
- 逆转耐药性
Abstract:ObjectiveTo investigate the effects of low-dose LBH589 on the apoptosis of epithelial ovarian cancer cells induced by PI3K/AKT pathway. MethodsThe apoptosis and cycle of epithelial ovarian cancer cells(OVCAR-3 cells) in the experimental group with low-dose LBH589 inducing and control group without low-dose LBH589 inducing were detected using TUNEL and flow cytometry, respectively.The inhibition of low-dose LBH589 on epithelial overian cancer cell proliferation was detected using MTT assay, and the expression levels of PI3K, AKT p-PI3K and p-AKT in two groups were detected using Western blot ting. ResultsThe cell apoptosis index in experimental group was significantly higher than that in control group(P < 0.01), the number of epithelial ovarian cancer cells in S-phase in experimental group was significantly lower than that in control group(P < 0.01).The inhibitory rates of epithelial ovarian cancer cells in experimental group at 12, 24, 36 and 48 h were(12.35±0.27)%, (21.24±0.33)%, (33.54±0.26)% and(41.23±0.52)%, respectively.The inhibition rate in control group was 0.In experimental group, the sensitivity of EOC cells to cisplatin significantly increased(P < 0.01), and the reversal coefficient was 3.26 times of control group.Compared with the control group, the expression levels of PI3K, AKT, p-PI3K and p-AKT in ovarian cancer cells were significantly inhibited in experimental group(P < 0.01). ConclusionsThe low-dose LBH589 can increase the apoptosis of the epithelial ovarian cancer cells OVCAR-3, and may reverse the cisplatin resistance through the PI3K/AKT pathway. -
Key words:
- ovarian neoplasms /
- LBH589 /
- PI3K/AKT pathway /
- apoptosis /
- reversal of drug resistance
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表 1 2组EOC细胞周期变化(ni=3;x±s;%)
分组 G1 S G2 对照组 53.36±1.02 42.26±1.35 20.29±1.32 实验组 63.32±1.32 18.27±1.66 9.12±1.05 t 10.34 19.42 11.47 P < 0.01 < 0.01 < 0.01 表 2 2组蛋白表达量(ni=3;x±s)
分组 PI3K AKT p-PI3K p-AKT 对照组 2.33±0.08 3.12±0.64 3.54±0.78 2.48±0.22 实验组 0.93±0.24 1.55±0.36 1.06±0.23 0.96±0.31 t 9.59 3.703 5.282 6.926 P < 0.01 < 0.05 < 0.01 < 0.01 -
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