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目的 探讨α 鹅膏蕈碱 (α amanitinAMA)是否可诱导在体小鼠肝细胞凋亡 ,以及可能的剂量依赖性效应和时间依赖性效应。方法 30只健康雄性昆明小白鼠被分为 6组 ,每组 5只。A组经腹腔注射生理盐 ,B ,C ,D组分别经腹腔注射AMA 0 8,1 2 ,2 0 μg/g (体重 ) ,观察终止时点为 2 4h ,用于观察肝细胞凋亡剂量依赖性效应。E和F组经腹腔注射AMA 2 0 μg/g (体重 )。观察终止时点为 16h和 36h ,与A和D组一起用于对凋亡的时间依赖性效应进行分析。采用光学显微镜、透射电镜、琼脂糖凝胶电泳技术和TUNEL法检测小鼠肝细胞是否存在凋亡以及凋亡的效应。结果 实验组小鼠肝脏切片中均出现典型的凋亡形态学改变 ,琼脂糖凝胶电泳呈现DNA“梯带” ,TUNEL检测可见细胞核着色呈棕黄色的凋亡细胞。从剂量依赖性效应分析 ,A～D组的肝细胞凋亡指数分别为 1 0 8% ,4 88% ,7 36 %和 9 2 0 % ,差异具有显著性(P <0 0 5 )。从时间依赖性效应分析 ,A ,E ,D和F组的肝细胞凋亡指数分别为 1 0 8% ,5 6 8% ,9 2 0 %和14 4 4 % ,差异具有显著性 (P <0 0 5 )。结论 α 鹅膏蕈碱能诱导小鼠肝细胞凋亡 ,并且在一定剂量范围内具有剂量依赖性和时间依赖性效应 ,肝细胞凋亡率随剂量的增大和作用时间延长而增加
Objective To investigate whether α amanitin AMA can induce apoptosis of hepatocytes in mice, as well as possible dose-dependent and time-dependent effects. Methods 30 healthy male Kunming mice were divided into 6 groups, 5 in each group. In group A, normal saline was injected intraperitoneally. In groups B, C, and D, AMA 88,12, and 20 μg/g (body weight) were injected intraperitoneally, and the termination point was observed at 24 hours for observation of the dose of hepatocyte apoptosis. Dependency effect. The E and F groups were injected intraperitoneally with AMA 20 μg/g (body weight). Observations were terminated at 16 h and 36 h and used together with groups A and D to analyze the time-dependent effects of apoptosis. Optical microscopy, transmission electron microscopy, agarose gel electrophoresis, and TUNEL assay were used to detect the effects of apoptosis and apoptosis in mouse hepatocytes. Results The typical morphology of apoptosis was observed in the liver sections of the experimental group. Agarose gel electrophoresis showed DNA ladders. The TUNEL assay showed apoptotic cells with nuclear brown staining. From the dose-dependent effect analysis, the apoptotic indexes of hepatocytes in groups A-D were 10.8%, 488%, 736.6%, and 92.0%, respectively, with a significant difference (P < 0.05). From the time-dependent effect analysis, the apoptotic indexes of hepatocytes in groups A, E, D, and F were 108%, 56.8%, 92%, and 144.4%, respectively, and the difference was significant (P < 0 0 5 ). Conclusion α-amanitin can induce hepatocyte apoptosis in mice and has a dose-dependent and time-dependent effect within a certain dose range. The hepatocyte apoptosis rate increases with the increase of dose and the prolonged action time.