天然产物gochnatiolide A对人前列腺癌DU145细胞增殖和凋亡的影响及机制研究
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摘要
目的研究兔儿风属植物中天然产物gochnatiolide A抗前列腺癌DU145细胞的作用及机制。方法采用CCK-8法和克隆形成实验检测gochnatiolideA对DU145细胞增殖的影响;DAPI染色法观察细胞凋亡形态;流式细胞仪检测gochnatiolide A对DU145细胞凋亡和周期的影响;Western blotting法检测gochnatiolide A对DU145细胞内活化的多聚腺苷二磷酸核糖聚合酶(cleaved-PARP)、活化的半胱氨酸蛋白酶9(cleaved Caspase-9)、p53、Bcl-2、核转录因子κB p65(NF-κB p65)和核转录因子κB抑制蛋白α、β(IKKα、IKKβ)表达的影响。结果与对照组比较,gochnatiolideA能够显著抑制DU145细胞增殖,其12、24、48 h的半数抑制浓度(IC_(50))值分别为4.15、2.80、1.74μmol/L。Gochnatiolide A可以浓度依赖性地促进细胞凋亡,阻滞细胞周期于G2期和S期;可以显著促进凋亡蛋白cleaved-PARP、cleaved Caspase-9和p53蛋白表达,抑制Bcl-2蛋白表达,抑制NF-κB通路中NF-κB p65、IKKα和IKKβ蛋白的表达。结论 Gochnatiolide A具有很好的抑制DU145细胞增殖及促进凋亡的作用,其机制可能与抑制NF-κB通路有关。
Objective To investigate the effects and mechanism of gochnatiolide A from Ainsliaea in anti-prostate cancer. Methods The activities of gochnatiolide A on the proliferation of DU145 cells were tested by CCK-8 and colony formation experiments.Apoptosis morphological changes of cells were observed by DAPI staining. The apoptosis and cell cycle were evaluated by flow cytometry. The expressions of cleaved Poly ADP ribose polymerase(cleaved-PARP), cleaved cystein-containing aspartate specific protease 9(cleaved Caspase-9), tumor suppressor protein(p53), b cell lymphoma-2(Bcl-2), transcription factors-κB p65(NF-κB p65)and IκB kinase α, β(IKKα, IKKβ) proteins in DU145 cells were investigated by Western blotting. Results The natural product gochnatiolide A significantly inhibited the proliferation of prostate cancer DU145 cells, with IC_(50) values of 4.15, 2.80 and 1.74 μmol/L at 12 h, 24 h, and 48 h, respectively. Meanwhile, gochnatiolide A promoted DU145 cells apoptosis and induced cell cycle arrest at G2 and S phases in a concentration dependent manner. In addition, gochnatiolide A also up-regulated apoptosis proteins cleaved-PARP,cleaved Caspase-9 and p53 proteins, and down-regulated Bcl-2 protein. Comparing the control group, the expression of NF-κB p65, IKKαand IKKβ proteins in the NF-κB pathway were decreased after treatment with gochnatiolide A. Conclusion The natural product gochnatiolide A remarkably inhibited the proliferation and promoted apoptosis in DU145 cells, and the possible mechanism was related to the inhibition of NF-κB pathway.
Objective To investigate the effects and mechanism of gochnatiolide A from Ainsliaea in anti-prostate cancer. Methods The activities of gochnatiolide A on the proliferation of DU145 cells were tested by CCK-8 and colony formation experiments.Apoptosis morphological changes of cells were observed by DAPI staining. The apoptosis and cell cycle were evaluated by flow cytometry. The expressions of cleaved Poly ADP ribose polymerase(cleaved-PARP), cleaved cystein-containing aspartate specific protease 9(cleaved Caspase-9), tumor suppressor protein(p53), b cell lymphoma-2(Bcl-2), transcription factors-κB p65(NF-κB p65)and IκB kinase α, β(IKKα, IKKβ) proteins in DU145 cells were investigated by Western blotting. Results The natural product gochnatiolide A significantly inhibited the proliferation of prostate cancer DU145 cells, with IC_(50) values of 4.15, 2.80 and 1.74 μmol/L at 12 h, 24 h, and 48 h, respectively. Meanwhile, gochnatiolide A promoted DU145 cells apoptosis and induced cell cycle arrest at G2 and S phases in a concentration dependent manner. In addition, gochnatiolide A also up-regulated apoptosis proteins cleaved-PARP,cleaved Caspase-9 and p53 proteins, and down-regulated Bcl-2 protein. Comparing the control group, the expression of NF-κB p65, IKKαand IKKβ proteins in the NF-κB pathway were decreased after treatment with gochnatiolide A. Conclusion The natural product gochnatiolide A remarkably inhibited the proliferation and promoted apoptosis in DU145 cells, and the possible mechanism was related to the inhibition of NF-κB pathway.
引文
[1]孙颖浩,高旭.前列腺癌诊断和治疗的百年演变史[J].上海医学,2017(7):391-395.
[2]Cjd W,Glaser A,Hu J C,et al.Survival and complications following surgery and radiation for localized prostate cancer:An international collaborative review[J].Eur Urol,2017,doi:10.1016/j.eururo.2017.05.055
[3]Newman D J,Cragg G M.Newman D J,et al.Natural products as sources of new drugs over the 30 years from1981 to 2010.[J].J Nat Prod,2012,75(3):311-335.
[4]Blunt J W,Copp B R,Munro M H,et al.Marine natural products[J].Nat Prod Rep,2008,25(1):35-94.
[5]Charan R D,Garson M J,Brereton I M,et al.Haliclonacyclamines A and B,cytotoxic alkaloids from the tropical marine sponge Haliclona sp[J].Tetrahedron,1996,52(27):9111-9120.
[6]Strapasson R L B,Cervi A C,Carvalho J E,et al.Bioactivity-guided isolation of cytotoxic sesquiterpene lactones of Gochnatia polymorpha ssp.floccosa[J].Phytother Res,2012,26(7):1053-1056.
[7]Xiong H P,Wu Z J,Chen D S,et al.A dimeric sesquiterpene,gochnatiolide A[J].Acta Crystallogr E,2009,doi:10.1107/S1600536809042743.
[8]Cheng Z,Yuan X,Qu Y,et al.Bruceine D inhibits hepatocellular carcinoma growth by targetingβ-catenin/Jagged1 pathways[J].Cancer Lett,2017,doi:10.1016/j.canlet.2017.06.014.
[9]Khan G N,Kim E J,Shin T S,et al.Heterogeneous cell types in single-cell-derived clones of MCF7 and MDA-MB-231 cells[J].Anticancer Res,2017,37(5):2343-2354.
[10]余展鹏,宋方茗,蔡琰,等.黄芩素对鼻咽癌CNE2细胞增殖和凋亡的影响[J].中草药,2018,49(4):879-884.
[11]葛宇清,杨波,程汝滨,等.隐丹参酮对K562细胞凋亡的影响及其机制[J].中草药,2013,44(22):3188-3194.
[12]高美玲,孙彦君,付露,等.小叶莲中graphislactone A的分离制备及其抗肿瘤作用机制研究[J].中草药,2018,49(1):167-172.
[13]Lv C,Zeng H W,Wang J X,et al.The antitumor natural product tanshinone IIA inhibits protein kinase C and acts synergistically with 17-AAG[J].Cell Death Dis,2018,doi:10.1038/s41419-017-0247-5.
[14]Xie Q,Wu G Z,Yang N,et al.Delavatine A,an unusual isoquinoline alkaloid exerts anti-inflammation on LPS-induced proinflammatory cytokines production by suppressing NF-κB activation in BV-2 microglia[J].Biochem Bioph Res Commun,2018,doi:10.1016/j.bbrc.2018.05.144.
[15]李小江,李洋洋,牟睿宇,等.中医药治疗前列腺癌骨转移的研究进展[J].中草药,2018,49(4):965-969.
[16]Lin Y E,Jia Y,Ke J I,et al.Traditional Chinese medicine in the prevention and treatment of cancer and cancer metastasis[J].Oncol Lett,2015,10(3):1240-1250.
[17]张元.鬼臼毒素衍生物CIP-36抗肿瘤多药耐药作用机制研究[D].石家庄:河北医科大学,2010.
[18]Dong Y,Cao A,Shi J,et al.Tangeretin,a citrus polymethoxyflavonoid,induces apoptosis of human gastric cancer AGS cells through extrinsic and intrinsic signaling pathways[J].Oncol Rep,2014,31(4):1788-1794.
[19]Wang Y Y,Yang Y X,Zhao R,et al.Bardoxolone methyl induces apoptosis and autophagy and inhibits epithelialto-mesenchymal transition and stemness in esophageal squamous cancer cells[J].Drug De Dev Ther,2015,doi:10.2147/DDDT.S73493.
[20]唐惠玲.NF-κB信号传导与疾病研究进展[J].黑龙江医药科学,2012,35(5):34-36.
[2]Cjd W,Glaser A,Hu J C,et al.Survival and complications following surgery and radiation for localized prostate cancer:An international collaborative review[J].Eur Urol,2017,doi:10.1016/j.eururo.2017.05.055
[3]Newman D J,Cragg G M.Newman D J,et al.Natural products as sources of new drugs over the 30 years from1981 to 2010.[J].J Nat Prod,2012,75(3):311-335.
[4]Blunt J W,Copp B R,Munro M H,et al.Marine natural products[J].Nat Prod Rep,2008,25(1):35-94.
[5]Charan R D,Garson M J,Brereton I M,et al.Haliclonacyclamines A and B,cytotoxic alkaloids from the tropical marine sponge Haliclona sp[J].Tetrahedron,1996,52(27):9111-9120.
[6]Strapasson R L B,Cervi A C,Carvalho J E,et al.Bioactivity-guided isolation of cytotoxic sesquiterpene lactones of Gochnatia polymorpha ssp.floccosa[J].Phytother Res,2012,26(7):1053-1056.
[7]Xiong H P,Wu Z J,Chen D S,et al.A dimeric sesquiterpene,gochnatiolide A[J].Acta Crystallogr E,2009,doi:10.1107/S1600536809042743.
[8]Cheng Z,Yuan X,Qu Y,et al.Bruceine D inhibits hepatocellular carcinoma growth by targetingβ-catenin/Jagged1 pathways[J].Cancer Lett,2017,doi:10.1016/j.canlet.2017.06.014.
[9]Khan G N,Kim E J,Shin T S,et al.Heterogeneous cell types in single-cell-derived clones of MCF7 and MDA-MB-231 cells[J].Anticancer Res,2017,37(5):2343-2354.
[10]余展鹏,宋方茗,蔡琰,等.黄芩素对鼻咽癌CNE2细胞增殖和凋亡的影响[J].中草药,2018,49(4):879-884.
[11]葛宇清,杨波,程汝滨,等.隐丹参酮对K562细胞凋亡的影响及其机制[J].中草药,2013,44(22):3188-3194.
[12]高美玲,孙彦君,付露,等.小叶莲中graphislactone A的分离制备及其抗肿瘤作用机制研究[J].中草药,2018,49(1):167-172.
[13]Lv C,Zeng H W,Wang J X,et al.The antitumor natural product tanshinone IIA inhibits protein kinase C and acts synergistically with 17-AAG[J].Cell Death Dis,2018,doi:10.1038/s41419-017-0247-5.
[14]Xie Q,Wu G Z,Yang N,et al.Delavatine A,an unusual isoquinoline alkaloid exerts anti-inflammation on LPS-induced proinflammatory cytokines production by suppressing NF-κB activation in BV-2 microglia[J].Biochem Bioph Res Commun,2018,doi:10.1016/j.bbrc.2018.05.144.
[15]李小江,李洋洋,牟睿宇,等.中医药治疗前列腺癌骨转移的研究进展[J].中草药,2018,49(4):965-969.
[16]Lin Y E,Jia Y,Ke J I,et al.Traditional Chinese medicine in the prevention and treatment of cancer and cancer metastasis[J].Oncol Lett,2015,10(3):1240-1250.
[17]张元.鬼臼毒素衍生物CIP-36抗肿瘤多药耐药作用机制研究[D].石家庄:河北医科大学,2010.
[18]Dong Y,Cao A,Shi J,et al.Tangeretin,a citrus polymethoxyflavonoid,induces apoptosis of human gastric cancer AGS cells through extrinsic and intrinsic signaling pathways[J].Oncol Rep,2014,31(4):1788-1794.
[19]Wang Y Y,Yang Y X,Zhao R,et al.Bardoxolone methyl induces apoptosis and autophagy and inhibits epithelialto-mesenchymal transition and stemness in esophageal squamous cancer cells[J].Drug De Dev Ther,2015,doi:10.2147/DDDT.S73493.
[20]唐惠玲.NF-κB信号传导与疾病研究进展[J].黑龙江医药科学,2012,35(5):34-36.