中国中药杂志

2011, v.36(05) 628-632

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基于精密肝切片技术的野百合碱体外肝毒性初步研究
Evaluation of vitro hepatotoxicity of monocrotaline by precision-cut liver slice technique

回连强;高双荣;刘婷;曹春雨;郭静;郝然;易艳;李春英;赵雍;梁爱华;
HUI Lianqiang,GAO Shuangrong,LIU Ting,CAO Chunyu,GUO Jing,HAO Ran, YI Yan,LI Chunying,ZHAO Yong,LIANG Aihua(Institute of Chinese Materia Medica,China Academy of Chinese Medical Sciences,Beijing 100700,China)

摘要(Abstract):

目的:对精密肝切片技术进行实验条件的优化,并依此技术对野百合碱的体外肝毒性进行初步的研究探讨。方法:利用震荡切片机制备肝切片,通过对培养液、切片厚度、pH及不同培养温度等条件下,肝切片MTT还原能力的测定,建立肝切片培养技术,并将肝切片与野百合碱共培养,连续监测法测定GPT,GOT,LDH,GGT酶活浓度,BCA法测定蛋白含量,观察其对肝切片MTT还原能力及以上生化指标的影响。结果:精密肝切片厚度在200μm,pH 6.8,37℃,BPM培养液中肝切片MTT还原能力最强;野百合碱0.02,0.1,0.5 g.L-1与肝切片共培养24 h,LDH漏出率显著上升,蛋白含量显著下降;野百合碱3个剂量组与肝切片共培养6 h,对MTT还原能力及生化各项指标无明显影响。结论:大鼠精密肝切片在BPM培养基的优化培养条件下,MTT还原能力可维持24 h;野百合碱0.02,0.1,0.5 g.L-1与肝切片共培养24 h,显示出一定的肝脏毒性。
Objective:To modify the empirical method of precision-cut liver slice technique,and study the hepatotoxicity of monocrotaline by this technique.Method:Liver slices were prepared by the domestic shaking slicer.The technique of precision-cut liver slice was established by detecting MTT reduction used as the slice viability under different culture medium,thickness of slices,pH and culture temperature.After monocrotaline and liver slices co-culture for 6,24 h,the slice viability,enzyme activity of GPT,GOT,LDH,GGT and protein concentration were detected by MTT reduction,enzyme kinetics method and BCA protein assay method,respectively.Result:When the thickness of slices was 200 μm and pH of medium was 6.8,culture temperature was 37 ℃,BPM culture medium,the viability of slices could maintain on a steady level.LDH leakage was significantly increased and protein content was obviously decreased after monocrotaline co-culture for 24 h with final concentration 0.02,0.1 and 0.5 g·L-1.No statistically significant difference between control group and monocrotaline 3 dose groups was observed in the slice viability and the content of GPT,GOT,LDH,GGT and protein after monocrotaline co-culture for 6 h.Conclusion:The slice viability could retain 24 h in modified BPM medium surroundings;monocrotaline displayed liver toxicity in some degree after co-culture for 24 hours in 0.02,0.1 and 0.5 g·L-1 concentration.

关键词(KeyWords): 精密肝切片;野百合碱;肝脏毒性
precision-cut liver slice;monocrotaline;liver toxicity

Abstract:

Keywords:

基金项目(Foundation): 国家“重大新药创制”科技重大专项(2009ZX09301-005-08);; 国际科技合作项目(2006DFA31760);; 中国中医科学院自选课题(2006年度)

作者(Authors): 回连强;高双荣;刘婷;曹春雨;郭静;郝然;易艳;李春英;赵雍;梁爱华;
HUI Lianqiang,GAO Shuangrong,LIU Ting,CAO Chunyu,GUO Jing,HAO Ran, YI Yan,LI Chunying,ZHAO Yong,LIANG Aihua(Institute of Chinese Materia Medica,China Academy of Chinese Medical Sciences,Beijing 100700,China)

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