银杏内酯B对膀胱癌J82细胞增殖、凋亡的机制研究
Mechanism of ginkgolide B on proliferation and apoptosis of bladder cancer J82 cells
庞华 1智静涛 2边建华 1刘文杰1
作者信息
- 1. 102413 北京,北京核工业医院泌尿外科
- 2. 北京市房山区良乡医院泌尿外科
- 折叠
摘要
目的 探究银杏内酯B对人膀胱癌细胞增殖、凋亡的影响及核转录因子-κB(NF-κB)信号通路的调控机制.方法 体外培养人膀胱癌J82 细胞系,分为对照组、低/中/高浓度实验组和阳性药物组;对照组、银杏内酯B组、激活剂组和抑制剂组,干预 24 h.活细胞计数法测定细胞活力;5-乙炔基-2'脱氧尿嘧啶核苷法测定细胞增殖情况;Hoechst 33258 染色法测定细胞凋亡情况;RT-qPCR法测定半胱氨酸天冬氨酸蛋白酶-3(Caspase-3)和细胞周期素D1(Cyclin D1)的 mRNA表达水平;蛋白免疫印迹法测定Caspase-3、Cyclin D1 及 NF-κB通路相关蛋白表达水平.结果 与对照组相比,中/高浓度实验组和阳性药物组细胞活力、增殖率显著降低,细胞凋亡率显著升高(P<0.05);与阳性药物组相比,低/中浓度实验组细胞活力和增殖率显著升高,细胞凋亡率显著降低(P<0.05);实验选择高浓度实验组分别加入激活剂和抑制剂作为激活剂组和抑制剂组进行后续 NF-κB通路验证实验,发现与对照组相比,银杏内酯 B组 Caspase-3 mRNA 和蛋白表达水平显著升高,而Cyclin D1 mRNA和蛋白以及 p-IκBα和 p-NF-κB p65 蛋白表达水平显著降低(P<0.05);与银杏内酯B组相比,激活剂组细胞凋亡率及 Caspase-3 mRNA 和蛋白表达水平显著降低,细胞活力、增殖率、Cyclin D1 mRNA和蛋白以及p-IκBα和p-NF-κB p65 蛋白表达水平显著升高(P<0.05),抑制剂组的这些指标变化趋势与激活剂组相反(P<0.05).结论 银杏内酯 B可显著抑制人膀胱癌J82 细胞的增殖,诱导其凋亡,其作用机制与抑制 NF-κB通路的信号转导相关.
Abstract
Objective To explore the effect of ginkgolide B on the proliferation and apoptosis of human bladder cancer cells and the regulatory mechanism of nuclear factor-κB(NF-κB)signaling pathway.Methods Human bladder cancer J82 cell lines were cultured in vitro and divided into control group(no intervention),low/medium/high concentration experimental group(5,10 and 20 μmol/L ginkgolide B)and positive control group(100 nmol/L paclitaxel);control group(no inter-vention),ginkgolide B group(20 μmol/L ginkgolide B),activator group(20 μmol/L ginkgolide B+ 1 μmol/L NF-κB pathway activator Prostratin)and inhibitor group(20 μmol/L ginkgolide B+5 μmol/L NF-κB pathway inhibitor BAY 11-7082).They were intervened for 24 h.Cell viability,pro-liferation,apoptosis and mRNA expression levels of Caspase-3 and Cyclin D1 were measured by cell counting kit-8,5-ethynyl-2'deoxyuridine,Hoechst 33258 staining and real-time quantitative PCR,respectively.The expression levels of Caspase-3,Cyclin D1 and NF-κB pathway-related protein were detected by western blotting.Results Compared with the control group,the cell viability and proliferation rate of the medium/high concentration experimental group were significantly decreased,and the cell apoptosis rate was significantly increased(P<0.05).Compared with the positive con-trol group,the cell viability and proliferation rate of the low/medium concentration experimental group were significantly increased,and the apoptosis rate was significantly decreased(P<0.05).The high-concentration experimental group was selected to add activator and inhibitor as the activator group and inhibitor group for the follow-up NF-κB pathway verification experimen.t Compared with the control group,the ex-pression levels of Caspase-3 mRNA and protein in ginkgolide B group were significantly increased,while the expression levels of Cyclin D1 mRNA and protein,p-IκBα and p-NF-κB p65 protein were significantly decreased(P<0.05).Compared with ginkgolide B group,the apoptosis rate and Caspase-3 mRNA and protein expression levels in the activator group were signifi-cantly reduced,while the cell viability,proliferation rate,Cyclin D1 mRNA and protein,p-IκBα and p-NF-κB p65 protein ex-pression level were significantly increased(P<0.05).However,the change trends of these indicators in the inhibitor group were opposite to those in the activator group.Conclusions Ginkgolide B inhibits the proliferation and induces apoptosis of human bladder cancer J82 cells,and its mechanism is related to inhibiting the signal transduction of NF-κB pathway.
关键词
膀胱癌/银杏内酯B/核转录因子-κB信号通路/增殖/凋亡化Key words
Bladder cancer/Ginkgolide B/Nuclear transcription factor-κB signaling pathway/Proliferation/Apoptosis引用本文复制引用
出版年
2024
NETL
NSTL
万方数据