摘要
目的:探讨芍药苷-6-氧-苯磺酸酯(CP-25)通过Toll样受体4(TLR-4)/核因子κB(NF-κB)信号通路对卵巢癌细胞增殖、凋亡的影响。方法:CCK-8实验检测人卵巢癌细胞A2780、SK-OV-3、HO8910及正常卵巢细胞HOSEpiC的活性。A2780细胞分为对照组(0.01%二甲基亚砜)、CP-25低(0.5 mg/ml CP-25)、中(1 mg/ml CP-25)、高(2 mg/ml CP-25)浓度组和CP-25高浓度+LPS组(2 mg/ml CP-25和0.1μg/ml TLR-4激活剂LPS)。CCK-8和克隆形成实验评估细胞增殖能力;流式细胞术和TUNEL染色分析细胞凋亡情况;Western blot分析增殖蛋白Ki67、凋亡相关蛋白(Cleaved caspase-3、Caspase-3、Bax、Bcl-2)、TLR-4、p-NF-κB p65、NF-κB p65蛋白表达水平;免疫荧光染色观察TLR-4、NF-κB p65蛋白表达。结果:与HOSEpiC细胞相比,A2780、SK-OV-3、HO8910细胞活性随CP-25浓度增加而逐渐减弱(P<0.05),且CP-25在低、中、高浓度下对A2780细胞的生长有50%左右的抑制作用,故采用A2780细胞进行实验。与对照组相比,CP-25低、中、高浓度组A2780细胞克隆形成率显著下降,细胞凋亡率和凋亡指数明显上升,Ki67、Bcl-2、TLR-4、p-NF-κB p65表达水平和细胞核NF-κB p65水平显著降低,Cleaved caspase-3/Caspase-3、Bax表达显著升高(P<0.05),且呈现浓度依赖性。与CP-25高浓度组相比,CP-25高浓度+LPS组细胞活性、克隆形成率明显升高,细胞凋亡率和凋亡指数明显降低,Ki67、Bcl-2、TLR-4、p-NF-κB p65表达水平和细胞核NF-κB p65水平明显升高,Cleaved caspase-3/Caspase-3和Bax表达明显降低(P<0.05)。结论:CP-25通过抑制TLR-4/NF-κB信号通路抑制卵巢癌细胞增殖并促进其凋亡。
Objective:To investigate impacts of paeoniflorin-6′-O-benzene sulfonate(CP-25)on proliferation and apoptosis of ovarian cancer cells through Toll-like receptor 4(TLR-4)/nuclear factor-κB(NF-κB)signaling pathway.Methods:CCK-8 assay was performed to measure the cell activity of human ovarian cancer cells A2780,SK-OV-3,HO8910 and normal ovarian cells HOSEpiC.A2780 cells were separated into control group(0.01%DMSO),CP-25 low(0.5 mg/ml CP-25),medium(1 mg/ml CP-25)and high(2 mg/ml CP-25)concentration groups and CP-25 high concentration+LPS group(2 mg/ml CP-25 and 0.1μg/ml TLR-4 activator LPS).Cell proliferation was evaluated by CCK-8 and clone formation assays;cell apoptosis was analyzed by flow cytometry and TUNEL staining;proliferating protein Ki67,apoptosis-related proteins(Cleaved caspase-3,Caspase-3,Bax,Bcl-2),TLR-4,p-NF-κB p65,NF-κB p65 protein expression levels were analyzed by Western blot;protein expressions of TLR-4 and NF-κB p65 were observed by immunofluorescence staining.Results:Compared with HOSEpiC cells,viabilities of A2780,SK-OV-3 and HO8910 cells were reduced gradually with the increased of CP-25 concentration(P<0.05),and CP-25 had about 50%inhibitory effect on the growth of A2780 cells at low,medium and high concentrations,so A2780 cells were used for the experiments.Compared with control group,the clone formation rate of A2780 cells in low,medium and high concentration groups of CP-25 reduced obviously,while apoptosis rate and apoptosis index rose obviously,expression levels of Ki67,Bcl-2,TLR-4,p-NF-κB p65 and nuclear NF-κB p65 were reduced obviously,while expression of Cleaved caspase-3/Caspase-3 and Bax were increased obviously in a concentration-dependent manner(P<0.05).Compared with CP-25 high concentration group,cell viability and clone formation rate were significantly increased in CP-25 high concentration+LPS group,while apoptosis rate and apoptosis index were decreased significantly,expression levels of Ki67,Bcl-2,TLR-4,p-NF-κB p65 and nuclear NF-κB p65 were increased significant
作者
袁小波
彭小珊
周丽丽
唐静
YUAN Xiaobo;PENG Xiaoshan;ZHOU Lili;TANG Jing(Medical Transformation Center of Xiangtan First People's Hospital,Xiangtan 411100,China;School of Health Medicine,Chongqing Youth Vocational and Technical College,Chongqing 400712,China;Department of Nutrition,Xiangtan First People's Hospital,Xiangtan 411100,China;School of Nursing,Hunan Institute of Communications Engineering,Hengyang 421009,China)
出处
《中国免疫学杂志》
CAS
CSCD
北大核心
2024年第8期1677-1683,共7页
Chinese Journal of Immunology
