NEDD8 共价修饰抑制因子MLN4924 对人卵巢癌细胞株SKOV3 增殖和凋亡的影响

目的:探讨神经前体细胞发育相关受控表达分子8(NEDD8)共价修饰抑制因子MLN4924 对人卵巢癌细胞增殖和凋亡的影响,并分析其可能机制。方法:使用不同浓度MLN4924(0、0.125、0.25、0.5 μmol/ L) 处理人卵巢癌细胞株SKOV3 4 h,免疫印迹法检测PAR3、HER2、Neddylated-cullins、P鄄I资B琢的表达水平;同时取细胞培养上清用ELISA 检测IL-6 的分泌。不同浓度MLN4924 处理SKOV3 细胞72 h,CCK8 法检测细胞增殖抑制率,流式细胞仪检测细胞周期和细胞凋亡率。结果:MLN4924 作用4 h 时,能够使Neddylated-cullins 下降、P-κBα积聚,IL-6 分泌减少,而PAR3、HER2 表达水平变化不显著。MLN4924 在0.125、0.25 和0.5 μmol/ L 的浓度下处理SKOV3 细胞72 h 时,对细胞增殖有明显的抑制作用,并且随着剂量的加大,抑制效果也显著提高;同时加药组S 期细胞百分率相较于对照组明显升高,且形成大量四倍体,使之没有完整的细胞周期;细胞凋亡率也随MLN4924 剂量增大而升高。结论:NEDD8 修饰特异性抑制剂MLN4924 能够明显抑制人卵巢癌细胞株SKOV3 的增殖,诱导细胞周期阻滞和凋亡。上述效应可能与NF-κB 通路活性受到抑制和IL-6 表达下降有关。

Effects of NEDD8 covalent modification inhibitor MLN4924 on proliferation and apoptosis of human ovarian cancer cell line SKOV3

Objective:To explore the effects of neural precursor cell-expressed developmentally down regulated 8(NEDD8)covalent modification on ovarian cancer cell proliferation and apoptosis,and the possible underlying mechanisms.Methods:Use different concentrations of MLN4924 (0,0.125,0.25,0. 5 mol/ L) and human ovarian cancer cell line SKOV3,the expression levels of PAR3,HER2,Neddylated-cullins, P-κBαwere detected by Western blot and the secretion of IL-6 was measured by ELISA after 4 h treatment.The cell proliferation was determined by CCK-8 staining and the cell cycle and apoptosis were analyzed by flow cytometry after 72 h treatment.Results:MLN4924 inhibits the proliferation of SKOV3 cells in a dose-dependent manner,which is associated with reduced IL-6 secretion.Western blot revealed that MLN4924 dose-dependently inhibits the neddylation of cullins (reduced neddylated-cullins) and induced the accumulation of P-κBα,whereas the expression levels of PARs and HER2 remained largely unchanged.At the same time MLN4924 dose-dependently induced cell cycle prove at S phase and the formation of a large number of tetraploids.Furthermore,apoptosis increased with the dose of MLN4924.Conclusion:NEDD8 covalent modification specific inhibitor MLN4924 can significantly inhibit the proliferation of SKOV3 cells and induce cell cycle arrest and apoptosis.The above inhibitory effects may partially result from impaired P-κBαdegradation and IL-6 secretion.