金丝桃苷对宫颈癌Hela细胞凋亡及抗氧化能力的影响
|
摘要
目的探索金丝桃苷对宫颈癌Hela细胞凋亡、抗氧化能力的影响。方法采用不同浓度的金丝桃苷(0、25、50、100、200μmol/L)处理体外培养Hela细胞12 h、24 h、48 h,噻唑蓝(MTT)法检测Hela细胞各浓度不同时间点细胞增殖情况以筛选出适宜的金丝桃苷浓度与作用时间点;giemsa染色观察比较观察组(金丝桃苷:100μmol/L)与对照组(空白对照)Hela细胞24 h形态变化;流式细胞仪检测Hela细胞凋亡情况;免疫组化检测caspase-3与caspase-8蛋白表达;ELISA法检测Hela细胞MDA、谷胱甘肽(GSH)的水平,SOD、过氧化氢酶(CAT)活力;qPCR检测Hela细胞VEGF、Bcl-2、Bax、p53表达水平。结果随着金丝桃苷作用时间的延长,Hela细胞存活率逐渐降低并且存在剂量依赖效应;观察组Hela细胞数目减少、光泽度降低、细胞体积变小且形状发生改变;观察组Hela细胞凋亡率明显高于对照组;观察组Hela细胞caspase-3与caspase-8蛋白表达量高于对照组;观察组Hela细胞MDA含量高于对照组,而GSH水平显著低于对照组;观察组Hela细胞SOD、CAT活力显著低于对照组;观察组Hela细胞VEGF、Bcl-2表达量降低,而Bax、p53表达水平高于对照组。结论金丝桃苷可以通过调控凋亡相关基因和蛋白的表达、促进细胞凋亡、降低抗氧化能力,从而抑制宫颈癌Hela细胞生长。
Objective To explore the effects of Hyperoside on apoptosis and antioxidant capacity of cervical cancer Hela cells. Methods Hela cells were cultured in vitro for 12 h,24 h and 48 h by treatment of different concentrations of Hyperoside(0,25,50,100,200 μmol/L).Methyl thiazolyl tetrazolium(MTT) assay was used to detect the proliferation of Hela cells at different concentrations and time points to select appropriate hyperoside concentration and time point.Giemsa staining was used to observe and compare the 24 h morphological changes of Hela cells between observation group(hyperside:100 μmol/L) and control group(blank control).The apoptosis of Hela cells was detected by flow cytometry.The protein expression levels of caspase-3 and caspase-8 were detected by immunohistochemistry.The levels of MDA and glutathione(GSH) and activities of SOD and catalase(CAT) of Hela cells were detected by ELISA.The expression levels of VEGF,Bcl-2,Bax and p53 in Hela cells were detected by qPCR. Results With the prolongation of action of hyperoside,the survival rate of Hela cells WAS gradually decreased and showed dose-dependent effects.The number of Hela cells was decreased,the glossiness was decreased and the cell volume became smaller and the shape changed in observation group.The apoptosis rate of Hela cells in observation group was significantly higher than that in control group.The protein expression levels of caspase-3 and caspase-8 of Hela cells in observation group were higher than those in control group.The MDA content of Hela cells in observation group was higher than that in control group while the GSH level was significantly lower than that in control group.Theactivities of SOD and CAT of Hela cells in observation group were significantly lower than those in control group.The expression levels of VEGF and Bcl-2 in Hela cells were decreased while the expression levels of Bax and p53 were higher than those in control group.Conclusion Hyperoside can inhibit the growth of cervical cancer Hela cells by regulating the expressions of apoptosis-related genes and proteins,promoting apoptosis and reducing antioxidant capacity.
Objective To explore the effects of Hyperoside on apoptosis and antioxidant capacity of cervical cancer Hela cells. Methods Hela cells were cultured in vitro for 12 h,24 h and 48 h by treatment of different concentrations of Hyperoside(0,25,50,100,200 μmol/L).Methyl thiazolyl tetrazolium(MTT) assay was used to detect the proliferation of Hela cells at different concentrations and time points to select appropriate hyperoside concentration and time point.Giemsa staining was used to observe and compare the 24 h morphological changes of Hela cells between observation group(hyperside:100 μmol/L) and control group(blank control).The apoptosis of Hela cells was detected by flow cytometry.The protein expression levels of caspase-3 and caspase-8 were detected by immunohistochemistry.The levels of MDA and glutathione(GSH) and activities of SOD and catalase(CAT) of Hela cells were detected by ELISA.The expression levels of VEGF,Bcl-2,Bax and p53 in Hela cells were detected by qPCR. Results With the prolongation of action of hyperoside,the survival rate of Hela cells WAS gradually decreased and showed dose-dependent effects.The number of Hela cells was decreased,the glossiness was decreased and the cell volume became smaller and the shape changed in observation group.The apoptosis rate of Hela cells in observation group was significantly higher than that in control group.The protein expression levels of caspase-3 and caspase-8 of Hela cells in observation group were higher than those in control group.The MDA content of Hela cells in observation group was higher than that in control group while the GSH level was significantly lower than that in control group.Theactivities of SOD and CAT of Hela cells in observation group were significantly lower than those in control group.The expression levels of VEGF and Bcl-2 in Hela cells were decreased while the expression levels of Bax and p53 were higher than those in control group.Conclusion Hyperoside can inhibit the growth of cervical cancer Hela cells by regulating the expressions of apoptosis-related genes and proteins,promoting apoptosis and reducing antioxidant capacity.
引文
[1] 杨诗婷,王晓倩,廖广辉.金丝桃苷的药理作用机制研究进展[J].中国现代应用药学,2018,35(6):947-951
[2] 高健美,牛爽,李柯,等.金丝桃苷对D-半乳糖衰老模型小鼠的抗衰老作用[J].中药药理与临床,2017,33(1):57-59
[3] 李春阳,付琳,黄午阳,等.蓝莓叶槲皮素和金丝桃苷的HPLC-MS鉴定及在内皮细胞中的舒血管和抗炎作用研究[J].现代食品科技,2017,33(1):20-25
[4] 丁兰,陈志武,郭岩.金丝桃苷对大鼠脑基底动脉的舒张作用及其作用机制[J].安徽医科大学学报,2016,51(11):1625-1629
[5] 周琦,吴小华,刘继红,等.宫颈癌诊断与治疗指南(第四版)[J].中国实用妇科与产科杂志,2018,34(6):613-622
[6] su V,Jerónimo J.Saving the World′s Women from Cervical Cancer.[J].New England Journal of Medicine,2016,374(26):2509-2511
[7] 杜伟平,李颖,李芳芹.人乳头瘤病毒高危型感染与宫颈癌的研究进展[J].国际检验医学杂志,2015,36(7):967-969
[8] 张英,侯炜,林洪生.中医药治疗恶性肿瘤临床研究成果与思考[J].中医杂志,2014,55(6):523-525
[9] 高健美,李德芬,牛爽,等.金丝桃苷对过氧化氢诱导的A549细胞氧化损伤的保护作用[J].中国实验方剂学杂志,2017,23(11):128-133
[10] 李莹,周燏,孙蕾清,等.金丝桃苷增强人γδT细胞增殖及杀伤功能研究[J].中国免疫学杂志,2016,32(4):524-527
[11] 王丽敏,杨玉,张明远,等.金丝桃苷对耐长春新碱结肠癌HCT8/VCR细胞耐药逆转作用[J].中国实验方剂学杂志,2016,22(22):92-96
[12] Zhang N,Ying M D,Wu Y P,et al.Hyperoside,a Flavonoid Compound,Inhibits Proliferation and Stimulates Osteogenic Differentiation of Human Osteosarcoma Cells[J].Plos One,2014,9(7):98973
[13] 孔亚,王佳佳,卢宗亮,等.桔梗皂苷D诱导乳腺癌MDA-MB-231细胞凋亡[J].中国肿瘤生物治疗杂志,2016,23(3):350-354
[14] Gregoraszczuk E L,Agnieszka R M,Janusz R,et al.Effect of Chemotherapeutic Drugs on Caspase-3 Activity,as a Key Biomarker for Apoptosis in Ovarian Tumor Cell Cultured as Monolayer.A Pilot Study[J].Iranian Journal of Pharmaceutical Research Ijpr,2015,14(4):1153-1161
[15] Li C P,Li J H,He S Y,et al.Roles of Fas/Fasl,Bcl-2/Bax,and Caspase-8 in rat nonalcoholic fatty liver disease pathogenesis[J].Genetics & Molecular Research Gmr,2014,13(2):3991-3999
[16] Qin S,Zhang B,Xiao G,et al.Fibronectin protects lung cancer cells against docetaxel-induced apoptosis by promoting Src and caspase-8 phosphorylation.[J].Tumor Biology,2016,37(10):1-12
[17] 任毅,李平华,陈潇.金丝桃苷通过上调Caspase-3、NF-κB表达抑制眼睑麟状癌细胞增殖和转移的影响[J].中药药理与临床,2017,33(1):30-33
[18] 袁宏中,曹玉婷,李琳娜,等.SM-1通过激活前胱天蛋白酶-3诱导人胃腺癌BGC-823细胞凋亡发挥抗肿瘤作用[J].军事医学,2016,40(4):326-330
[19] Zhu A K,Zhou H,Xia J Z,et al.Ziyuglycoside II-induced apoptosis in human gastric carcinoma BGC-823 cells by regulating Bax/Bcl-2 expression and activating caspase-3 pathway[J].Brazilian Journal of Medical & Biological Research,2013,46(8):670-675
[20] Wu J H,Yao Y L,Gu T,et al.MiR-421 regulates apoptosis of BGC-823 gastric cancer cells by targeting caspase-3[J].Asian Pacific Journal of Cancer Prevention Apjcp,2014,15(13):5463
[21] Guon T E,Chung H S.Hyperoside and rutin of Nelumbo nucifera induce mitochondrial apoptosis through a caspase-dependent mechanism in HT-29 human colon cancer cells[J].Oncology Letters,2016,11(4):2463
[22] 朱玉侠,赵明星,姜登鸽,等.抑癌基因p53、凋亡抑制基因Bcl-2、促凋亡基因Bax在胃癌及癌前病变中的表达[J].胃肠病学和肝病学杂志,2016,25(9):1040-1043
[23] Li L,Wu W,Huang W,et al.NF-κB RNAi decreases the Bax/Bcl-2 ratio and inhibits TNF-α-induced apoptosis in human alveolar epithelial cells.[J].Inflammation Research,2013,62(4):387
[24] Liang S,Sun K,Yue W,et al.Role of Cyt-C/caspases-9,3,Bax/Bcl-2 and the FAS death receptor pathway in apoptosis induced by zinc oxide nanoparticles in human aortic endothelial cells and the protective effect by alpha-lipoic acid[J].Chem Biol Interact,2016,258:40-51
[25] Sharifi S,Barar J,Hejazi M S,et al.Doxorubicin Changes Bax /Bcl-xLRatio,Caspase-8 and 9 in Breast Cancer Cells[J].Adv Pharm Bull,2015,5(3):351-359
[26] Peng H,Zhang Q,Li J,et al.Apatinib inhibits VEGF signaling and promotes apoptosis in intrahepatic cholangiocarcinoma:[J].Oncotarget,2016,7(13):17220
[27] Li Z L,Liu J C,Hu J,et al.Protective effects of hyperoside against human umbilical vein endothelial cell damage induced by hydrogen peroxide[J].Journal of Ethnopharmacology,2012,139(2):388
[28] Sumaira S,Khan M R,Khan R A.Hepatoprotective effects of methanol extract ofCarissa opacaleaves on CCl4-induced damage in rat[J].BMC Complementary and Alternative Medicine,2011,11(1):1-9
[29] Zhou Y H,Yu J P,Liu Y F,et al.Effects of Ginkgo biloba Extract on Inflammatory Mediators (SOD,MDA,TNF-α,NF-κBp65,IL-6) in TNBS-InducedColitis in Rats[J].Mediators of Inflammation,2017,2006(05):1-9
[30] Balabanli B,Balaban T.Investigation into the Effects of Boron on Liver Tissue Protein Carbonyl,MDA,and Glutathione Levels in Endotoxemia[J].Biological Trace Element Research,2015,167(2):1-5
[2] 高健美,牛爽,李柯,等.金丝桃苷对D-半乳糖衰老模型小鼠的抗衰老作用[J].中药药理与临床,2017,33(1):57-59
[3] 李春阳,付琳,黄午阳,等.蓝莓叶槲皮素和金丝桃苷的HPLC-MS鉴定及在内皮细胞中的舒血管和抗炎作用研究[J].现代食品科技,2017,33(1):20-25
[4] 丁兰,陈志武,郭岩.金丝桃苷对大鼠脑基底动脉的舒张作用及其作用机制[J].安徽医科大学学报,2016,51(11):1625-1629
[5] 周琦,吴小华,刘继红,等.宫颈癌诊断与治疗指南(第四版)[J].中国实用妇科与产科杂志,2018,34(6):613-622
[6] su V,Jerónimo J.Saving the World′s Women from Cervical Cancer.[J].New England Journal of Medicine,2016,374(26):2509-2511
[7] 杜伟平,李颖,李芳芹.人乳头瘤病毒高危型感染与宫颈癌的研究进展[J].国际检验医学杂志,2015,36(7):967-969
[8] 张英,侯炜,林洪生.中医药治疗恶性肿瘤临床研究成果与思考[J].中医杂志,2014,55(6):523-525
[9] 高健美,李德芬,牛爽,等.金丝桃苷对过氧化氢诱导的A549细胞氧化损伤的保护作用[J].中国实验方剂学杂志,2017,23(11):128-133
[10] 李莹,周燏,孙蕾清,等.金丝桃苷增强人γδT细胞增殖及杀伤功能研究[J].中国免疫学杂志,2016,32(4):524-527
[11] 王丽敏,杨玉,张明远,等.金丝桃苷对耐长春新碱结肠癌HCT8/VCR细胞耐药逆转作用[J].中国实验方剂学杂志,2016,22(22):92-96
[12] Zhang N,Ying M D,Wu Y P,et al.Hyperoside,a Flavonoid Compound,Inhibits Proliferation and Stimulates Osteogenic Differentiation of Human Osteosarcoma Cells[J].Plos One,2014,9(7):98973
[13] 孔亚,王佳佳,卢宗亮,等.桔梗皂苷D诱导乳腺癌MDA-MB-231细胞凋亡[J].中国肿瘤生物治疗杂志,2016,23(3):350-354
[14] Gregoraszczuk E L,Agnieszka R M,Janusz R,et al.Effect of Chemotherapeutic Drugs on Caspase-3 Activity,as a Key Biomarker for Apoptosis in Ovarian Tumor Cell Cultured as Monolayer.A Pilot Study[J].Iranian Journal of Pharmaceutical Research Ijpr,2015,14(4):1153-1161
[15] Li C P,Li J H,He S Y,et al.Roles of Fas/Fasl,Bcl-2/Bax,and Caspase-8 in rat nonalcoholic fatty liver disease pathogenesis[J].Genetics & Molecular Research Gmr,2014,13(2):3991-3999
[16] Qin S,Zhang B,Xiao G,et al.Fibronectin protects lung cancer cells against docetaxel-induced apoptosis by promoting Src and caspase-8 phosphorylation.[J].Tumor Biology,2016,37(10):1-12
[17] 任毅,李平华,陈潇.金丝桃苷通过上调Caspase-3、NF-κB表达抑制眼睑麟状癌细胞增殖和转移的影响[J].中药药理与临床,2017,33(1):30-33
[18] 袁宏中,曹玉婷,李琳娜,等.SM-1通过激活前胱天蛋白酶-3诱导人胃腺癌BGC-823细胞凋亡发挥抗肿瘤作用[J].军事医学,2016,40(4):326-330
[19] Zhu A K,Zhou H,Xia J Z,et al.Ziyuglycoside II-induced apoptosis in human gastric carcinoma BGC-823 cells by regulating Bax/Bcl-2 expression and activating caspase-3 pathway[J].Brazilian Journal of Medical & Biological Research,2013,46(8):670-675
[20] Wu J H,Yao Y L,Gu T,et al.MiR-421 regulates apoptosis of BGC-823 gastric cancer cells by targeting caspase-3[J].Asian Pacific Journal of Cancer Prevention Apjcp,2014,15(13):5463
[21] Guon T E,Chung H S.Hyperoside and rutin of Nelumbo nucifera induce mitochondrial apoptosis through a caspase-dependent mechanism in HT-29 human colon cancer cells[J].Oncology Letters,2016,11(4):2463
[22] 朱玉侠,赵明星,姜登鸽,等.抑癌基因p53、凋亡抑制基因Bcl-2、促凋亡基因Bax在胃癌及癌前病变中的表达[J].胃肠病学和肝病学杂志,2016,25(9):1040-1043
[23] Li L,Wu W,Huang W,et al.NF-κB RNAi decreases the Bax/Bcl-2 ratio and inhibits TNF-α-induced apoptosis in human alveolar epithelial cells.[J].Inflammation Research,2013,62(4):387
[24] Liang S,Sun K,Yue W,et al.Role of Cyt-C/caspases-9,3,Bax/Bcl-2 and the FAS death receptor pathway in apoptosis induced by zinc oxide nanoparticles in human aortic endothelial cells and the protective effect by alpha-lipoic acid[J].Chem Biol Interact,2016,258:40-51
[25] Sharifi S,Barar J,Hejazi M S,et al.Doxorubicin Changes Bax /Bcl-xLRatio,Caspase-8 and 9 in Breast Cancer Cells[J].Adv Pharm Bull,2015,5(3):351-359
[26] Peng H,Zhang Q,Li J,et al.Apatinib inhibits VEGF signaling and promotes apoptosis in intrahepatic cholangiocarcinoma:[J].Oncotarget,2016,7(13):17220
[27] Li Z L,Liu J C,Hu J,et al.Protective effects of hyperoside against human umbilical vein endothelial cell damage induced by hydrogen peroxide[J].Journal of Ethnopharmacology,2012,139(2):388
[28] Sumaira S,Khan M R,Khan R A.Hepatoprotective effects of methanol extract ofCarissa opacaleaves on CCl4-induced damage in rat[J].BMC Complementary and Alternative Medicine,2011,11(1):1-9
[29] Zhou Y H,Yu J P,Liu Y F,et al.Effects of Ginkgo biloba Extract on Inflammatory Mediators (SOD,MDA,TNF-α,NF-κBp65,IL-6) in TNBS-InducedColitis in Rats[J].Mediators of Inflammation,2017,2006(05):1-9
[30] Balabanli B,Balaban T.Investigation into the Effects of Boron on Liver Tissue Protein Carbonyl,MDA,and Glutathione Levels in Endotoxemia[J].Biological Trace Element Research,2015,167(2):1-5
© 2004-2018 中国地质图书馆版权所有 京ICP备05064691号 京公网安备11010802017129号 地址：北京市海淀区学院路29号 邮编：100083 电话：办公室：(+86 10)66554848；文献借阅、咨询服务、科技查新：66554700 |