不同培养基条件下黄芪和附子及其配伍对Caco-2细胞毒性的影响
|
摘要
目的观察培养条件对MTT法评价细胞毒性结果的影响,同时观察黄芪、附子及其配伍对细胞活性的影响。方法取对数生长期的Caco-2细胞接种于96孔板,待细胞生长稳定后,分别采用完全培养基和DMEM培养基培养,以MTT法考察黄芪、附子及其配伍在不同给药时间下对Caco-2细胞毒性的影响。结果在完全培养基条件下,黄芪、附子及其配伍对细胞毒性的影响呈现出剂量依赖性关系,而在DMEM培养基的条件下黄芪和附子对细胞毒性影响不规律;与单独使用附子相比,黄芪-附子配伍能显著提高细胞存活率。结论完全培养基条件下MTT法能较好的反映药物对细胞毒性的影响;黄芪能降低附子的毒性。
Objective To observe the effects of culture conditions on the cytotoxicity of MTT, and observe the effects of Astragalus membranaceus and aconite root on cell viability. Methods Caco-2 cells from the logarithmic growth period were inoculated on 96 orifice plates. After the cell growth was stable, the total culture medium and DMEM medium were cultured respectively. The effects of Astragalus membranaceus, aconite and their compatibility on the toxicity of Caco-2 cells were investigated by MTT method.Results Under the condition of complete medium, Astragalus membranaceus, aconite and their compatibility have a dose-dependent relationship on cytotoxicity, and the effects of Astragalus membranaceus and Aconitum on cytotoxicity are irregular under the condition of DMEM medium, and the compatibility of Astragalus membranaceus and Aconitum can significantly increase the cell survival rate compared with the single use of aconite. Conclusion The MTT method can better reflect the effect of drugs on cytotoxicity under complete medium conditions. Astragalus can reduce the toxicity of Radix Aconiti Lateralis Preparata.
Objective To observe the effects of culture conditions on the cytotoxicity of MTT, and observe the effects of Astragalus membranaceus and aconite root on cell viability. Methods Caco-2 cells from the logarithmic growth period were inoculated on 96 orifice plates. After the cell growth was stable, the total culture medium and DMEM medium were cultured respectively. The effects of Astragalus membranaceus, aconite and their compatibility on the toxicity of Caco-2 cells were investigated by MTT method.Results Under the condition of complete medium, Astragalus membranaceus, aconite and their compatibility have a dose-dependent relationship on cytotoxicity, and the effects of Astragalus membranaceus and Aconitum on cytotoxicity are irregular under the condition of DMEM medium, and the compatibility of Astragalus membranaceus and Aconitum can significantly increase the cell survival rate compared with the single use of aconite. Conclusion The MTT method can better reflect the effect of drugs on cytotoxicity under complete medium conditions. Astragalus can reduce the toxicity of Radix Aconiti Lateralis Preparata.
引文
[1] 陈荣昌,孙桂波,张强.附子毒性研究进展[J].中国中药杂志,2013,38(8):1126.
[2] 刘会艳.黄芪及其制剂的药理作用和临床应用[J].内蒙古中医药,2012,31(22):44.
[3] 罗晓珍,于琴. 中药黄芪化学成份、药理活性与临床应用[J].中国保健营养,2013,23(6):1071.
[4] 张广平,解素花,朱晓光,等.附子相杀、相畏配伍减毒实验研究[J].中国中药杂志,2012,37(15):2215.
[5] 宋·魏岘.魏氏家藏方[M].北京:中国古籍出版社,1987:254.
[6] 骆梅娟,周至安.附子的毒性及临床应用浅析[J].广州中医药大学学报,2009,26(5):512.
[7] 孔小强,蒋且英,罗云,等.基于外翻肠囊模型的黄芪-附子配伍对附子 6 种生物碱肠吸收的影响研究[J].中草药,2017,48(23):4928.
[8] Silva R, Carmo H,Dinis-Oliveira R,et al.In vitro study of P-glycoprotein induction as an antidotal pathway to prevent cytotoxicity in Caco-2 cells[J].Arch Toxicol,2011,85(4):315.
[9] 蔡文涛.MTT法和CCK-8法检测中药抗病毒活性成分细胞毒性的比较[J].湖北大学学报(自然科学版),2017,39(3):305.
[10] 万文婷,李宁.CCK-8法与MTT法检测人前列腺癌PC3细胞活性的比较研究[J].时珍国医国药,2010,21(12):3041.
[11] 陈云芳,王胜,李冰.MTT法用于药物对L2细胞毒性的实验研究[J].现代生物医学进展.2012,12(8):5451.
[12] Mosmann T.Rapid colorinetric assay for cellular growth and survial: Application to proliferation and cytotoxicity assays[J].Journal of Immunological Methods,1983,65(1-2) :55.
[13] 刘涛,郭辰.毒理学研究中的体外细胞毒性评价[J].生命科学,2014,26(3):319.
[14] 杨晓珊.CYP450 酶与 BCRP、MRP2 外排转运蛋白对单酯型和醇胺型乌头生物碱处置的作用及机制[D].南方医科大学硕士学位论文,2014
[15] Zhang GY,Ou RL,Li FY,et al.Regulation of drug-metabolizing enzymes and efflux transporters by Astragali radix decoction and its main bioactive compounds: Implication for clinical drug-drug interactions[J].J Ethnopharmacol,2016,108:104.
[16] Wang X,Wang H,Zhang A,et al. Metabolomics study on the toxicity of aconite root and its processed products using ultraperformance liquid-chromatography/electrospray-ionization synapt high-definition mass spectrometry coupled with pattern recognition approach andingenuity pathways analysis[J].J Proteome Res,2012,11(2):1284.
[17] Hidalgo IJ, Jibin L.Carrier-mediated transport and efflux mechanisms in Caco-2 cells[J].Adv Drug Deliv Rev,1996,22(1):53.
[18] Ehret MJ, Levin GM, Narasimhan M,et al.Venlafaxine induces P-glycoprotein in human Caco-2 cells[J].Hum Psychopharmacol,2007,22(1):49.
[19] Wang YF,Yang XF,Cheng B,et al.Protective effect of Astragalus polysaccharides on ATP binding cassette transporter A1in THP- 1 derived foam cells exposed to tumor necrosis factor- alpha[J].Phytother Res.2010,24( 3) :393.
[20] Tian QE,Li HD,Yan M,et al.Effects of Astragalus polysaccharides on P-glycoprotein efflux pump function and protein expression in H22 hepatoma cells in vitro[J].BMC Complement Altern Med,2012,(12):94.
[2] 刘会艳.黄芪及其制剂的药理作用和临床应用[J].内蒙古中医药,2012,31(22):44.
[3] 罗晓珍,于琴. 中药黄芪化学成份、药理活性与临床应用[J].中国保健营养,2013,23(6):1071.
[4] 张广平,解素花,朱晓光,等.附子相杀、相畏配伍减毒实验研究[J].中国中药杂志,2012,37(15):2215.
[5] 宋·魏岘.魏氏家藏方[M].北京:中国古籍出版社,1987:254.
[6] 骆梅娟,周至安.附子的毒性及临床应用浅析[J].广州中医药大学学报,2009,26(5):512.
[7] 孔小强,蒋且英,罗云,等.基于外翻肠囊模型的黄芪-附子配伍对附子 6 种生物碱肠吸收的影响研究[J].中草药,2017,48(23):4928.
[8] Silva R, Carmo H,Dinis-Oliveira R,et al.In vitro study of P-glycoprotein induction as an antidotal pathway to prevent cytotoxicity in Caco-2 cells[J].Arch Toxicol,2011,85(4):315.
[9] 蔡文涛.MTT法和CCK-8法检测中药抗病毒活性成分细胞毒性的比较[J].湖北大学学报(自然科学版),2017,39(3):305.
[10] 万文婷,李宁.CCK-8法与MTT法检测人前列腺癌PC3细胞活性的比较研究[J].时珍国医国药,2010,21(12):3041.
[11] 陈云芳,王胜,李冰.MTT法用于药物对L2细胞毒性的实验研究[J].现代生物医学进展.2012,12(8):5451.
[12] Mosmann T.Rapid colorinetric assay for cellular growth and survial: Application to proliferation and cytotoxicity assays[J].Journal of Immunological Methods,1983,65(1-2) :55.
[13] 刘涛,郭辰.毒理学研究中的体外细胞毒性评价[J].生命科学,2014,26(3):319.
[14] 杨晓珊.CYP450 酶与 BCRP、MRP2 外排转运蛋白对单酯型和醇胺型乌头生物碱处置的作用及机制[D].南方医科大学硕士学位论文,2014
[15] Zhang GY,Ou RL,Li FY,et al.Regulation of drug-metabolizing enzymes and efflux transporters by Astragali radix decoction and its main bioactive compounds: Implication for clinical drug-drug interactions[J].J Ethnopharmacol,2016,108:104.
[16] Wang X,Wang H,Zhang A,et al. Metabolomics study on the toxicity of aconite root and its processed products using ultraperformance liquid-chromatography/electrospray-ionization synapt high-definition mass spectrometry coupled with pattern recognition approach andingenuity pathways analysis[J].J Proteome Res,2012,11(2):1284.
[17] Hidalgo IJ, Jibin L.Carrier-mediated transport and efflux mechanisms in Caco-2 cells[J].Adv Drug Deliv Rev,1996,22(1):53.
[18] Ehret MJ, Levin GM, Narasimhan M,et al.Venlafaxine induces P-glycoprotein in human Caco-2 cells[J].Hum Psychopharmacol,2007,22(1):49.
[19] Wang YF,Yang XF,Cheng B,et al.Protective effect of Astragalus polysaccharides on ATP binding cassette transporter A1in THP- 1 derived foam cells exposed to tumor necrosis factor- alpha[J].Phytother Res.2010,24( 3) :393.
[20] Tian QE,Li HD,Yan M,et al.Effects of Astragalus polysaccharides on P-glycoprotein efflux pump function and protein expression in H22 hepatoma cells in vitro[J].BMC Complement Altern Med,2012,(12):94.