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北京大学学报（医学版） ›› 2025, Vol. 57 ›› Issue (5): 926-933. doi: 10.19723/j.issn.1671-167X.2025.05.017

• 论著 • 上一篇    下一篇

左金丸中吴茱萸生物碱和小檗碱联合用药对HepG2细胞毒性的影响

高亚东1,2, 朱安1,3, 李璐迪1, 李盈姿1, 王旗1,4,5,*()   

  1. 1. 北京大学公共卫生学院毒理学系，北京 100191
    2. 福建省疾病预防控制中心福建省人兽共患病研究重点实验室，福州 350012
    3. 福建医科大学基础医学院消化道恶性肿瘤教育部重点实验室，福州 350108
    4. 国家中医药管理局中药配伍减毒重点研究室，北京 100191
    5. 食品安全毒理学研究与评价北京市重点实验室，北京 100191
  • 收稿日期:2024-02-07 出版日期:2025-10-18 发布日期:2025-05-07
  • 通讯作者: 王旗
  • 基金资助:
    国家重点研发计划专项(2018YFC1704500); 国家重点研发计划专项(2018YFC1704506)

Effect of the combination of alkaloids from Euodiae Fructus and berberine in Zuojin Pill on cytotoxicity in HepG2 cells

Yadong GAO1,2, An ZHU1,3, Ludi LI1, Yingzi LI1, Qi WANG1,4,5,*()   

  1. 1. Department of Toxicology, Peking University School of Public Health, Beijing 100191, China
    2. Fujian Provincial Key Laboratory of Zoonosis Research, Fujian Provincial Center for Disease Control and Prevention, Fuzhou 350012, China
    3. Key Laboratory of Ministry of Education for Gastrointestinal Cancer, School of Basic Medical Sciences, Fujian Medical University, Fuzhou 350108, China
    4. Key Laboratory of State Administration of Traditional Chinese Medicine for Compatibility Toxicology, Beijing 100191, China
    5. Beijing Key Laboratory of Toxicological Research and Risk Assessment for Food Safety, Beijing 100191, China
  • Received:2024-02-07 Online:2025-10-18 Published:2025-05-07
  • Contact: Qi WANG
  • Supported by:
    the National Key Research and Development Program of China(2018YFC1704500); the National Key Research and Development Program of China(2018YFC1704506)

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摘要： 目的: 研究左金丸中吴茱萸生物碱和小檗碱联合用药对肝细胞毒性的影响，探讨联合用药的减毒机制。方法: 参照左金丸中各化学成分吸收入血的峰浓度(maximum concentration，Cmax)确定成分配伍比例，采用HepG2细胞模型研究吴茱萸中肝毒性成分吴茱萸碱(evodiamine，EVO)、去氢吴茱萸碱(dehydroevodiamine，DHED)与小檗碱联合用药48 h的细胞毒性。实验分为溶剂对照组、EVO组、DHED组、小檗碱组，以及EVO或DHED与小檗碱的联合用药组。以细胞计数试剂盒法(cell counting kit-8，CCK-8)测定细胞存活率，并采用联合指数判断药物的毒性作用。检测细胞上清液中谷丙转氨酶(alanine transaminase，ALT)、谷草转氨酶(aspartate aminotransferase，AST)、乳酸脱氢酶(lactate dehydrogenase，LDH)、碱性磷酸酶(alkaline phosphatase，ALP)活性及总胆红素(total bilirubin，TBIL)含量。采用Western blot检测胆汁酸转运体胆盐输出泵(bile salt export pump，BSEP)和多耐药相关蛋白2(multidrug resistance-associated protein 2，MRP2)的表达水平。检测HepG2细胞丙二醛(malondialdehyde，MDA)含量和超氧化物歧化酶(superoxide dismutase，SOD)活性。结果: 与EVO或DHED组相比，EVO 1 μmol/L与小檗碱10 μmol/L联用、DHED 50 μmol/L与小檗碱35 μmol/L联用，均可显著提高HepG2细胞的存活率(P＜0.01)，其联合指数值分别为77.89和4.49，远大于1。联用小檗碱可降低细胞上清液中ALT、AST、LDH和ALP活性，并减少TBIL含量(P＜0.05、P＜0.01)。与EVO组相比，EVO联用小檗碱可上调BSEP和MRP2蛋白表达；与DHED组相比，DHED联用小檗碱可显著下调BSEP和MRP2蛋白表达(P＜0.01)。EVO或DHED与小檗碱联用均可显著减少MDA含量(P＜0.05、P＜0.01)。结论: 一定配比的小檗碱与EVO或DHED联用，对其HepG2细胞毒性具有拮抗作用，其减毒作用的机制与调节胆汁酸转运体表达、减轻脂质过氧化损伤有关。

关键词: 吴茱萸碱, 去氢吴茱萸碱, 小檗碱, 中药毒性, 化学性与药物性肝损伤

Abstract: Objective: To investigate the hepatotoxicity of alkaloids from Euodiae Fructus combined with berberine (BBR) in Zuojin Pill, and to preliminarily explore the possible detoxification mechanism of the combination components. Methods: The combination ratio of components was determined by the maximum concentration (Cmax) of the chemical components in Zuojin Pill. HepG2 cell model was used to investigate the combined toxicity of the hepatotoxic components from Euodiae Fructus, such as evodiamine (EVO) or dehydroevodiamine (DHED), with BBR for 48 h. The experimental groups were set as follows: the vehicle control group, the EVO group, the DHED group, the BBR group, and the combination group of EVO or DHED with BBR. The cell counting kit-8 (CCK-8) method was used to determine the cell viability, and the combination index (CI) was used to determine the combined toxicity of the components. The alanine transaminase (ALT), aspartate aminotransferase (AST), lactate dehydroge-nase (LDH), and alkaline phosphatase (ALP) activities as well as total bilirubin (TBIL) content in the cell culture supernatant were detected. The protein expression levels of bile acid transporters, such as bile salt export pump (BSEP) and multidrug resistance-associated protein 2 (MRP2), were detected by Western blot. The intracellular malondialdehyde (MDA) content and superoxide dismutase (SOD) activity in HepG2 cells were detected. Results: Compared with EVO or DHED group, the combination of EVO 1 μmol/L with BBR 10 μmol/L or DHED 50 μmol/L with BBR 35 μmol/L significantly increased cell viability of HepG2 cells (P < 0.01), with CI values of 77.89 or 4.49, respectively, much greater than 1. Significant decreases in the activities of ALT, AST, LDH, ALP, and TBIL content in the cell culture supernatant were found in both combination groups (P < 0.05, P < 0.01). Compared with the EVO group, the combination of EVO with BBR upregulated the protein expression levels of BSEP and MRP2. Compared with the DHED group, the combination of DHED with BBR significantly downregulated the protein expression levels of BSEP and MRP2 (P < 0.01). Compared with EVO or DHED group, the combination of EVO or DHED with BBR significantly reduced the MDA content in HepG2 cells (P < 0.05, P < 0.01). Conclusion: A certain ratio of BBR combined with EVO or DHED had an antagonistic effect on HepG2 cytotoxicity, which might be related to regulating the expression of bile acid transpor-ters, and reducing lipid peroxidation damage.

Key words: Evodiamine, Dehydroevodiamine, Berberine, Toxicity of Chinese medicine, Chemical and drug induced liver injury

中图分类号: 

  • R114

图1

EVO、DHED和小檗碱处理48 h对HepG2细胞存活率的影响($\bar x \pm s$, n=3)"

图2

EVO、DHED和小檗碱处理48 h对HepG2细胞肝功能指标的影响($\bar x \pm s$, n=3)"

图3

EVO、DHED和小檗碱处理48 h对HepG2细胞BSEP和MRP2蛋白表达的影响($\bar x \pm s$, n=3)"

图4

EVO、DHED和小檗碱处理48 h对HepG2细胞MDA含量和SOD活性的影响($\bar x \pm s$, n=3)"

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[1] 高亚东,朱安,李璐迪,张涛,王硕,单丹萍,李盈姿,王旗. 吴茱萸碱对HepG2细胞毒性及其机制[J]. 北京大学学报（医学版）, 2021, 53(6): 1107-1114.
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ISSN 1671-167X
CN 11-4691/R
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