红霉素和阿奇霉素诱导斑马鱼肝毒性及作用机制研究

Study on hepatotoxicity inducement and mechanisms of erythromycin and azithromycin in zebrafish

目的利用模式生物斑马鱼模型探索红霉素和阿奇霉素导致肝毒性及作用机制,并比较红霉素和阿奇霉素诱导肝毒性的差异。方法选用肝脏转基因斑马鱼Tg(fapb10:dsRed)作为实验动物,将发育正常的3dpf斑马鱼幼鱼暴露于不同浓度的红霉素和阿奇霉素溶液中72h后,统计各组斑马鱼死亡率,计算红霉素和阿奇霉素对斑马鱼的LD50;荧光显微镜下活体观察斑马鱼幼鱼给药后肝脏形态的变化;采用整体油红O染色观察肝脏脂肪含量变化;利用qRT-PCR检测肝脏病理标志基因和凋亡相关基因在转录水平的变化。结果红霉素和阿奇霉素对斑马鱼的LD50分别为3.82mmol/L和3.10mmol/L;活体观察显示红霉素和阿奇霉素均能导致幼鱼肝脏形态和荧光强度变化;整体油红O染色显示红霉素和阿奇霉素均能导致肝细胞脂肪堆积,并呈剂量依赖性;qRT-PCR结果显示红霉素和阿奇霉素均能影响脂肪肝、肝纤维化和细胞凋亡相关基因的表达。结论红霉素和阿奇霉素均能诱导斑马鱼肝毒性,作用机制可能与两者诱导肝细胞变性或凋亡有关,但两者影响的相关基因通路可能有所不同。

Objective To explore the hepatotoxicity and mechanisms of erythromycin and azithromycin in zebrafish,and compare and analyze the differences of erythromycin and azithromycin induced hepatotoxicity.Methods Zebrafish(Danio rerio)of transgenic Tg(fabp10:dsRed)were used,larvae at 3 dpf(day post fertilization)were exposed to the erythromycin and azithromycin solution for 72h at different concentrations.The mortality of zebrafish in each group and the LD50 of erythromycin and azithromycin on zebrafish were calculated.Viviperception of liver morphology was conducted using fluorescence imaging.Oil Red O(ORO)stain was employed to detect the hepatic steatosis in zebrafish larvae.Expression levels of some gene markers associated with liver pathology and apoptosis were investigated by qRT-PCR.Results The LD50 of erythromycin and azithromycin in zebrafish were 3.82mmol/L and 3.10mmol/L respectively.Fluorescence imaging and Oil Red O stain assays showed erythromycin and azithromycin induced liver degeneration,changed liver size and liver steatosis in larval zebrafish.The qRT-PCR analysis demonstrated erythromycin and azithromycin could affect the expression of genes related to fatty liver,liver fibrosis and apoptosis.Conclusion Erythromycin and azithromycin could induce liver toxicity in zebrafish,and the hepatotoxicity might be related to hepatocyte degeneration or apoptosis induced by erythromycin and azithromycin,while the related genes affected by them were different.