基于斑马鱼模型的重楼不同制备样本肝毒性比较和评价

doi:10.19803/j.1672-8629.20250828

摘要/Abstract

摘要： 目的探究重楼不同制备样本在斑马鱼系统中肝毒性方面的差异,确认重楼皂苷Ⅰ（Polyphyllin Ⅰ, PPⅠ）在重楼整体肝毒性中的作用,为临床安全用药提供实验参考。方法采用甲醇提取重楼药材,经正丁醇萃取获得总皂苷粗品,进一步通过柱层析分离得到剔除重楼皂苷Ⅰ的提取物（Paris Extract without Polyphyllin Ⅰ, PE-PPⅠ）。利用斑马鱼模型评估不同样本毒性效应,包括10%致死浓度（LC₁₀）测定、肝脏组织病理切片观察、尼罗红染色、吖啶橙染色以及肝脏荧光形态学分析;同时检测血清中甘油三酯、总胆固醇、高密度脂蛋白和低密度脂蛋白水平;并通过实时荧光定量PCR检测apoa2、cyp7a1、cyp27a1、hmgcs1、fabp4、cd36基因mRNA表达水平。结果在特定质量浓度下,重楼甲醇提取物（Methanol Extract, ME）、正丁醇萃取物（n-Butanol Extract, NBE）、PE-PPⅠ均可诱导斑马鱼肝脏损伤,但毒性强度与病理特征存在显著差异：NBE高、低剂量组表现出较强毒性,表现为广泛的肝脏组织损伤,伴随明显肝细胞凋亡及肝脏面积代偿性增大,并伴有显著脂质代谢紊乱;ME组主要呈现局灶性肝细胞损伤,脂质累积程度中等;PE-PPⅠ组仅引起轻微肝脏病变。RT-qPCR结果表明,脂蛋白代谢相关核心基因表达显著上调,胆汁酸合成限速酶基因呈现抑制特征,胆固醇合成调控基因表达上调,脂肪转运关键因子呈现显著上调趋势。结论重楼皂苷Ⅰ是重楼肝毒性的关键活性成分之一,可能通过干扰脂质代谢通路,导致肝脏脂质蓄积、肝细胞凋亡及组织结构损伤。

关键词: 重楼皂苷Ⅰ, 肝毒性, 斑马鱼, 甲醇提取物, 安全性评价

Abstract: Objective To investigate the differences in hepatotoxicity between diverse prepared samples of Paris polyphylla in a zebrafish system and confirm the role of PolyphyllinⅠ(PPⅠ) in the overall hepatotoxicity of the herb so as to provide data for elucidating the toxic material basis of PPⅠ and its safe applications. Methods Methanol was used to extract Paris polyphylla medicinal materials while crude total saponins were obtained via n-Butanol extraction. More separation by column chromatography yielded the Paris extract without PolyphyllinⅠ(PE-PPⅠ). On this basis, a zebrafish model was used to evaluate the toxicity of different samples, involving the determination of lethal concentration 10% (LC₁₀), observation of histopathological sections of livers, Nile Red staining, acridine orange staining, and fluorescence morphological analysis of livers. Meanwhile, the serum levels of triglycerides (TG), total cholesterol (TC), high-density lipoprotein (HDL), and low-density lipoprotein (LDL) were detected. Real-time quantitative PCR (RT-qPCR) was used to measure the mRNA expression levels of such genes as apoa2, cyp7a1, cyp27a1, hmgcs1, fabp4 and cd36. Results At specific mass concentrations, the methanol extract (ME), n-Butanol extract (NBE), and PE-PPⅠ of Paris polyphylla all induced liver injury in zebrafish, but there were significant differences in toxicity and pathology. In the high- and low-dose NBE groups, toxicity was relatively strong, characterized by extensive liver tissue damage that was accompanied by obvious hepatocyte apoptosis, compensatory enlargement of the liver size, and significant lipid metabolism disorders. In the ME group, focal hepatocyte damage with moderate lipid accumulation was observed. In the PE-PPⅠ group, only mild liver lesions occurred. RT-qPCR results indicated that the expression levels of core genes related to lipoprotein metabolism were significantly upregulated, the rate-limiting enzyme genes for bile acid synthesis were inhibited, the regulatory genes for cholesterol synthesis were upregulated, and that the key factors for fat transport trended upward. Conclusion PolyphyllinⅠ is one of the key active components responsible for the hepatotoxicity of Paris polyphylla, which may interfere with lipid metabolism pathways, leading to liver lipid accumulation, hepatocyte apoptosis, and structural damage to liver tissues.

Key words: PolyphyllinⅠ, Hepatotoxicity, Zebrafish, Methanol Extract, Safety Evaluation

中图分类号:

于蔚洁, 刘雨欣, 林瑞超, 李向日, 赵崇军. 基于斑马鱼模型的重楼不同制备样本肝毒性比较和评价[J]. 中国药物警戒, 2026, 23(4): 390-397.

YU Weijie, LIU Yuxin, LIN Ruichao, LI Xiangri, ZHAO Chongjun. Comparison and Evaluation of Hepatotoxicity of Different Processed Samples of Paris polyphylla Based on a Zebrafish Model[J]. Chinese Journal of Pharmacovigilance, 2026, 23(4): 390-397.

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https://zgywjj.magtechjournal.com/CN/Y2026/V23/I4/390

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