摘要
目的探究姜黄素联合5-氟尿嘧啶(5-FU)对胰腺癌细胞增殖抑制和促凋亡作用的机制。方法体外培养胰腺癌细胞,并分为:空白对照组、5-FU组(10μmol/L)、低剂量姜黄素联合5-FU组(姜黄素20μmol/L+5-FU10μmol/L)、高剂量姜黄素联合5-FU组(姜黄素60μmol/L+5-FU10μmol/L),采用MTT观察姜黄素联合5-FU对胰腺癌细胞生长的抑制作用,使用流式细胞仪检测细胞凋亡情况,使用Westen bolt和RT-PCR定性、半定量检测Bcl-2、Bax、P-mTOR、P-AMPK、caspase-3、caspase-8、caspase-9。结果MTT观察结果显示,相比空白对照组,5-FU组胰腺癌细胞存活百分比有一定程度的下降(P<0.05),低剂量姜黄素联合5-FU组、高剂量姜黄素联合5-FU组胰腺癌细胞存活百分比显著下降,差异有统计学意义(P<0.01),相比低剂量姜黄素联合5-FU组,高剂量姜黄素联合5-FU组胰腺癌细胞存活百分比下降(P<0.05);流式检验细胞凋亡率显示,低剂量姜黄素联合5-FU组、高剂量姜黄素联合5-FU组胰腺癌细胞增殖受到显著抑制,且随着姜黄素联合5-FU浓度的升高,抑制效果增强;相比空白对照组,5-FU组、低剂量姜黄素联合5-FU组、高剂量姜黄素联合5-FU组Bcl-2蛋白表达均显著降低,差异有统计学意义(P<0.01),Bax、PmTOR、P-AMPK、caspase-3、caspase-8、caspase-9蛋白表达显著升高,差异有统计学意义(P<0.01);相比5-FU组,低剂量姜黄素联合5-FU组、高剂量姜黄素联合5-FU组Bcl-2蛋白表达均显著降低,差异有统计学意义(P<0.01),Bax、P-mTOR、P-AMPK、caspase-3、caspase-8、caspase-9蛋白表达显著升高,差异有统计学意义(P<0.01)。结论姜黄素联合5-FU可以通过下调抑凋亡蛋白Bcl-2,上调促凋亡蛋白Bax、凋亡调控因子caspase-3、caspase-9和caspase-8蛋白表达,抑制AMPK/mTRO信号通路达到抑制胰腺癌细胞增殖、促进胰腺癌细胞凋亡的效果。
Objective To explore the role mechanism of curcumin combined with 5-fluorouracil(5-FU)on proliferation inhibition and apoptosis promotion of pancreatic cancer cells.Methods Pancreatic cancer cells were cultured in vitro and divided into blank control group,5-FU group(10μmol/L),low-dose curcumin combined with 5-FU group(20μmol/L curcumin+10μmol/L 5-FU)and high-dose curcumin combined with 5-FU group(60μmol/L curcumin+10μmol/L 5-FU).The inhibitory effects of curcumin combined with 5-FU on the growth of pancreatic cancer cells were observed by MTT.The cell apoptosis was detected by flow cytometry.Bcl-2,Bax,P-mTOR,P-AMPK,caspase-3,caspase-8 and caspase-9 were qualitatively and semi-quantitatively determined by Western bolt and RT-PCR.Results MTT observation showed that the survival percentage of pancreatic cancer cells in 5-FU group decreased to some extent compared with that in blank control group(P<0.05),and the survival percentage of pancreatic cancer cells significantly decreased in low-dose curcumin combined with 5-FU group and high-dose curcumin combined with 5-FU group(P<0.01).Compared with low-dose curcumin combined with 5-FU group,the survival percentage of pancreatic cancer cells decreased in high-dose curcumin combined with 5-FU group(P<0.05).Flow cytometry for apoptosis rate detection showed the proliferation of pancreatic cancer cells was significantly inhibited in low-dose curcumincombined with 5-FU group and high-dose curcumin combined with 5-FU group,and with the increase of concentration of curcumin combined with 5-FU,the inhibition effects increased.Compared with blank control group,the protein expression level of Bcl-2 significantly decreased in 5-FU group,low-dose curcumin combined with 5-FU group and high-dose curcumin combined with 5-FU group(P<0.01),and the protein expression levels of Bax,P-mTOR,P-AMPK,caspase-3,caspase-8 and caspase-9 significantly increased(P<0.01).Compared with 5-FU group,the protein expression level of Bcl-2 significantly decreased in low-dose curcumin combined with 5-FU group and high-dose curcumin combined with 5-FU group(P<0.01),and the protein expression levels of Bax,P-mTOR,P-AMPK,caspase-3,caspase-8 and caspase-9 significantly increased(P<0.01).Conclusion Curcumin combined with 5-FU can down-regulate anti-apoptotic protein Bcl-2,up-regulate the protein expression of pro-apoptotic protein Bax and apoptosis regulators caspase-3,caspase-9 and caspase-8,and inhibit AMPK/mTRO signaling pathwayso as to inhibit the proliferation and promote the apoptosis of pancreatic cancer cells.
作者
叶璇
张鹏
黄丹
Ye Xuan;Zhang Peng;Huang Dan(Tongji Medical College,Huazhong University of Science and Technology Hospital of Tongji,Wuhan 430050,China)
出处
《中国煤炭工业医学杂志》
2020年第3期230-235,共6页
Chinese Journal of Coal Industry Medicine
基金
湖北省卫生厅科研项目(编号:2015JX5B03)。