异烟肼灌胃建立小鼠急性肝损伤模型的研究

 目的 探索应用异烟肼灌胃建立小鼠急性肝损伤模型，并初步阐明其机制。 方法 昆明种小鼠 50 只，随机分为异烟肼正常剂量造模组（常量组）、2 倍剂量造模组（2 倍量组）、5 倍剂量造模组（5 倍量组）、8 倍剂量造模组（8 倍量组）和空白对照组，每组各 10 只。各造模组分别以 90、180、450、720 mg/kg 剂量的异烟肼灌胃，18 h 后检测血清丙氨酸氨基转移酶（ALT）、天冬氨酸氨基转移酶（AST）水平并取肝组织进行光镜观察，探索导致明显急性肝损伤的相对合适剂量。另取昆明种小鼠 90 只，随机分为单纯造模组（40 只）、干预造模组（40 只）和空白对照组（10 只），各造模组以相对合适剂量异烟肼灌胃，其中干预造模组于造模前以甘利欣75 mg/kg 体重连续灌胃 5 d，18、36、54、72 h 后分别检测血清 ALT、超氧化物歧化酶（SOD）、丙二醛（MDA）、谷胱甘肽过氧物酶（GSH-Px）水平，并进行肝组织光镜和电镜观察，探索导致明显急性肝损伤的相对合适时间及可能机制。 结果 在探索相对合适剂量实验中，2 倍量组小鼠血清ALT、AST 分别为（101.6 ± 6.3）和（108.1 ± 14.9）U/L，明显高于空白对照组的（30.1 ± 3.6）、（35.3 ± 6.5）U/L 和常量组的（52.8 ± 5.3）、（53.9 ± 8.9）U/L，差异均有统计学意义（均P < 0.05），肝细胞出现广泛的脂肪变性和水肿，肝窦几乎消失；5 倍量组小鼠死亡 7 只，8 倍量组小鼠则全部死亡。在探索相对合适时间及发生机制实验中，应用2 倍剂量异烟肼灌胃后 18 h，单纯造模组小鼠血清 ALT、MDA 水平明显高于空白对照组，SOD、GSH-Px 水平明显低于空白对照组，肝细胞广泛损伤，细胞超微结构显著变化。 结论 应用 2 倍剂量（180 mg/kg）异烟肼灌胃可成功建立小鼠急性肝损伤模型。异烟肼所致急性肝损伤可能与自由基脂质过氧化反应密切相关。

Establishment of mouse model of acute liver injury by perfusing stomach with isoniazid

Objective To establish a mouse model of acute liver injury by perfusing stomach with isoniazid, and to explore its mechanism. Methods Fifty mice of Kunming-strain were randomly divided into normal, 2-fold, 5-fold, and 8-fold dosage model groups and control group with 10 mice in each. The mice in the model groups were perfused with 90, 180, 450, and 720 mg/kg isoniazid respectively into the stomach. Eighteen hours after the perfusion, the serum levels of ALT and AST were measured, and the liver tissues were obtained from the mice and observed under a light microscope to find a relatively suitable dosage that can induce a significant acute liver injury. Then, 90 mice of Kunming-strain were randomly divided into simple model (40 mice), intervention model (40 mice), and control groups (10 mice). The mice in the simple model group were perfused with the suitable dosage of isoniazid into the stomach, while the mice in the intervention model group were treated with Ganlixin (diammonium glycyrrhizinate) at a dosage of 75 mg/kg for 5 days successively before establishing the model. Afterwards, the serum levels of ALT, SOD, MDA, and GSH-Px were measured at 18, 36, 54, and 72 hours, and the liver tissues were obtained from the mice and observed under light and electron microscopes respectively to find a relatively suitable time for establishing a model of acute liver injury, and to explore its underlying mechanism. Results In the first experiment, the mean serum levels of ALT and AST in the 2-fold dosage group were 101.6 U/L ± 6.3 U/L and 108.1 U/L ± 14.9 U/L respectively, which were statistically significantly higher than those in the control (30.1 U/L ± 3.6 U/L and 35.3 U/L ± 6.5 U/L) and the normal dosage group (52.8 U/L ± 5.3 U/L and 53.9 U/L ± 8.9 U/L, P < 0.05). Light microscopy showed fatty degeneration and edema of the liver cells, and the hepatic sinus could be hardly detected. In the 5-fold dosage model group, 7 mice died; while in the 8-fold group, all the mice. In the second experiment, 18 hours after the perfusion with 2-fold dosage (180 mg/kg) of isoniazid into the stomach, the serum levels of ALT and MDA in the simple model group were significantly higher than those in the control, while the levels of SOD and GSH-Px were significantly lower. Histological examination showed a large amount of injured liver cells with significantly changed ultrastructure. Conclusions Mouse model of acute liver injury can be successively established by perfusing stomach with isoniazid of 2-fold dosage (180 mg/kg). Its mechanism may be closely related to lipid peroxidation of free radical.