硫酸铍致小鼠肝毒性的线粒体损伤机制

发布时间：2018-10-10 17:14

【摘要】：目的：通过整体动物染毒硫酸铍（BeSO4·4H2O），观察硫酸铍对小鼠肝脏的毒性作用，，探讨硫酸铍致小鼠肝毒性的线粒体损伤的可能机制。方法： 1．选取6周龄健康成年昆明（KM）雄性小鼠30只，随机分为3组，每组10只，设阴性对照组和两个染毒组，对照组按0.1ml/10g（体重）腹腔注射灭菌生理盐水，低、高剂量染毒组分别腹腔注射1mg/kg和2mg/kg硫酸铍溶液。隔天染毒，持续2周。观察染毒期间小鼠的一般情况变化。 2．断颈放血法处死小鼠，收集血液，自动生化分析仪检测小鼠血清谷丙转氨酶（ALT）和谷草转氨酶（AST）含量；分离完整肝脏，计算肝脏脏器系数；肝组织采用苏木素-伊红（HE）染色，光镜下观察肝脏病理组织学变化。 3．小鼠肝线粒体悬液采用差速离心法制备。分别用荧光染料罗丹明123（Rh123）、二氯二氢荧光素-乙酰乙酸酯（DCFH-DA）负载肝线粒体，通过荧光分光光度法测定线粒体膜电位（Δψm）变化和线粒体活性氧（ROS）含量；分别用分光光度法测定线粒体吸光度（A520）值反映线粒体通透性转变孔道（PTP）开放程度、线粒体细胞色素C（Cytc）含量、线粒体丙二醛（MDA）含量和线粒体谷胱甘肽过氧化物酶（GSH-Px）活性；采用紫外分光光度法测定线粒体超氧化物歧化酶（SOD）活性。结果： 1．对照组小鼠和染毒组小鼠一般体征无明显差异；与对照组比较，染毒组体重差异无统计学意义（P0.05）。 2．与对照组比较，染毒组小鼠肝脏脏器系数增高、血清ALT和AST水平升高（P0.05）；病理学检查发现，对照组肝细胞结构正常；低剂量染毒组小鼠肝细胞水肿,有灶性坏死等病变；高剂量染毒组小鼠肝细胞胞浆成空泡状，有广泛变性、坏死等病理改变。 3．与对照组比较，染毒组线粒体Δψm下降（P0.05）；线粒体SOD活性和GSH-Px活性降低（P0.05）；染毒组线粒体PTP开放明显（P0.05）；染毒组线粒体Cytc含量、线粒体ROS含量和线粒体MDA含量增高（P0.05）。结论： 1．硫酸铍可致小鼠肝功能异常、肝组织病理形态学异常，具有明显肝毒性。 2．硫酸铍可致小鼠肝线粒体Δψm下降、PTP开放、Cytc释放增加、SOD和GSH-Px活性下降，ROS和MDA含量生成增加。 3.肝线粒体功能障碍和氧化损伤可能是硫酸铍致小鼠肝毒性的主要原因，线粒体可能是硫酸铍致肝毒性的作用靶点。
[Abstract]:Objective: to observe the toxicity of beryllium sulfate (BeSO4 4H2O) to mice liver and explore the possible mechanism of mitochondrial damage induced by beryllium sulfate. Methods: 1. Thirty healthy male Kunming (KM) mice aged 6 weeks were randomly divided into 3 groups, 10 in each group, with negative control group and two exposure groups. The control group was injected intraperitoneally with sterilized saline according to 0.1ml/10g (body weight). In high dose group, beryllium sulfate solution (1mg/kg) and 2mg/kg (beryllium sulfate) were injected intraperitoneally respectively. Poisoned the next day for 2 weeks. To observe the general changes of mice during exposure. 2. The mice were killed by blood release method, blood was collected, serum alanine aminotransferase (ALT) and alanine aminotransferase (AST) levels were detected by automatic biochemical analyzer, and the intact liver was isolated. Liver tissue was stained with hematoxylin-eosin (HE) and histopathological changes were observed under light microscope. 3. Mouse liver mitochondria suspension was prepared by differential centrifugation. Hepatic mitochondria were loaded with fluorescent dye Rhodamine 123 (Rh123) and dichlorodihydrofluorescein acetate (DCFH-DA). The changes of mitochondrial membrane potential (螖蠄 m) and the content of reactive oxygen species (ROS) in mitochondria were measured by fluorescence spectrophotometry. The mitochondrial absorbance (A520) was measured by spectrophotometry to reflect the opening degree of mitochondrial permeability transition pore (PTP), mitochondrial cytochrome C (Cytc) content, mitochondrial malondialdehyde (MDA) content and mitochondrial glutathione peroxidase (GSH-Px) activity. The activity of mitochondrial superoxide dismutase (SOD) was determined by ultraviolet spectrophotometry. Results: 1. There was no significant difference in general physical signs between the control group and the exposed group, and there was no significant difference in body weight between the control group and the control group (P0.05). 2. Compared with the control group, there was no significant difference in body weight between the control group and the control group. Liver organ coefficient increased, serum ALT and AST levels increased in the exposed group (P0.05); pathological examination showed that the structure of hepatocytes in the control group was normal; liver cell edema, focal necrosis and other pathological changes were found in the low dose exposure group. Compared with the control group, the mitochondria 螖蠄 m decreased (P0.05), the activity of mitochondrial SOD and GSH-Px decreased (P0.05). The content of mitochondrial Cytc, mitochondrial ROS and mitochondrial MDA were significantly increased in exposed group (P0.05). Conclusion: 1. Beryllium sulfate can induce abnormal liver function and pathological morphology of liver tissue in mice. 2. Beryllium sulfate could induce the decrease of 螖蠄 m, the opening of PTP, the increase of Cytc release, the decrease of SOD and GSH-Px activity, and the increase of ROS and MDA production in mouse liver mitochondria. The dysfunction and oxidative damage of liver mitochondria may be the main causes of hepatotoxicity induced by beryllium sulfate in mice, and mitochondria may be the target of hepatotoxicity induced by beryllium sulfate.
【学位授予单位】：南华大学
【学位级别】：硕士
【学位授予年份】：2012
【分类号】：R114

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