基于高内涵技术的补骨脂肝毒性成分筛选研究  

Hepatotoxic components in Psoraleae Fructus based on high-content screening technology

作　　者：尚慧莹 魏雪 程宏愽 马增春 涂博丹 肖成荣 刘献 高月[1,2] SHANG Huiying;WEI Xue;CHENG Hongbo;MA Zengchun;TU Bodan;XIAO Chengrong;LIU Xian;GAO Yue(Tianjin University of Traditional Chinese Medicine,Tianjin 301617,China;Institute of Radiation Medicine,Academy of Military Medical Sciences,Academy of Military Sciences,Beijing 100850,China;Guangdong Pharmaceutical University,Guangzhou Guangdong 510006,China)

机构地区：[1]天津中医药大学,天津301617 [2]军事科学院军事医学研究院辐射医学研究所,北京100850 [3]广东药科大学,广东广州510006

出　　处：《中国药物警戒》2024年第1期74-82,共9页Chinese Journal of Pharmacovigilance

基　　金：国家自然科学基金资助项目(82192911)。

摘　　要：目的采用高内涵技术筛选补骨脂潜在肝毒性成分及其可能的作用机制。方法采用CCK-8法测定补骨脂中异补骨脂二氢黄酮、补骨脂二氢黄酮甲醚、补骨脂二氢黄酮、异补骨脂查尔酮、4’-O-甲基补骨脂查尔酮、补骨脂宁、补骨脂素、异补骨脂素、补骨脂定、补骨脂酚10种成分的IC_(50)值。在相同浓度药物处理HepG 2细胞24 h后,进行高内涵检测,通过阳性细胞的平均荧光强度评价各组细胞活性氧、还原型谷胱甘肽、线粒体膜电位及线粒体超氧化物水平4个指标的变化情况,初步筛选补骨脂中主要肝毒性成分。结果CCK-8细胞活力实验结果显示,异补骨脂二氢黄酮、补骨脂二氢黄酮甲醚、补骨脂二氢黄酮、异补骨脂查尔酮、4’-O-甲基补骨脂查尔酮、补骨脂宁、异补骨脂素、补骨脂定、补骨脂酚对Hep G 2细胞的IC_(50)值分别为52.69、42.72、83.63、42.29、57.43、110.80、1420.00、23.58和25.34μmol·L^(-1)。高内涵结果显示,在20μmol·L^(-1)浓度下,补骨脂二氢黄酮甲醚、补骨脂二氢黄酮、异补骨脂查尔酮、补骨脂定和补骨脂酚可显著提高细胞内活性氧水平;异补骨脂查尔酮、4’-O-甲基补骨脂查尔酮、补骨脂宁、异补骨脂素、补骨脂定和补骨脂酚均可导致细胞中还原型谷胱甘肽水平保护性升高;异补骨脂查尔酮、补骨脂定和补骨脂酚均显著降低线粒体膜电位,造成线粒体超氧化物累积。结论异补骨脂查尔酮、补骨脂定和补骨脂酚是补骨脂中造成线粒体功能损伤的主要成分,具有潜在的肝脏毒性,其具体作用机制有待进一步研究。Objective To screen the potential hepatotoxic components in Psoraleae Fructus and explore the possible mechanisms using high-content screening technology.Methods The IC_(50)values of ten components in Psoraleae Fructus,including isobavachin,bavachinin,bavachin,isobavachalcone,4’-O-methyl-bavachalcone,corylin,psoralen,ispsoralen,psoralidin,and bakuchiol were determined by CCK-8 cell viability assays.After treating HepG 2 cells with the same concentration of individual drug for 24 h,high-content screening was performed.The potential hepatotoxic components were revealed by evaluating the average fluorescence intensity of positive cells of four indicators,including reactive oxygen species,reduced glutathione,mitochondrial membrane potential and mitochondrial superoxide.Results CCK-8 cell viability assays showed that the IC_(50)values of isobavachin,bavachinin,bavachin,isobavachalcone,4’-O-methylbavachalcone,corylin,ispsoralen,psoralidin,and bakuchiol were 52.69,42.72,83.63,42.29,57.43,110.80,1420.00,23.58 and 25.34μmol·L^(-1),respectively.The high-content screening analysis showed that bavachinin,bavachin,isobavachalcone,psoralidin and bakuchiol could significantly increase the level of reactive oxygen species at the same concentration of 20μmol·L^(-1);Isobavachalcone,4’-O-methyl-bavachalcone,corylin,ispsoralen,psoralidin,bakuchiol can lead to a protective increase in the level of reduced glutathione;Additionally,isobavachalcone,psoralidin and bakuchiol could prominently decrease the level of mitochondrial membrane potential and promoted the accumulation of mitochondrial superoxide.Conclusion Isobavachalcone,psoralidin and bakuchiol were the main mitotoxic components in Psoraleae Fructus,which promoted Psoraleae Fructus-induced liver injuries.The specific mechanisms of the three components remains to be investigated in the future.

关 键 词：补骨脂 异补骨脂查尔酮 补骨脂定 补骨脂酚 肝毒性 高内涵筛选技术 线粒体膜电位 线粒体超氧化物 

分 类 号：R978[医药卫生—药品] R994.1[医药卫生—药学]