α-鹅膏蕈碱诱导小鼠肝细胞凋亡的实验研究  

Hepatocytes apoptosis in mice induced by alpha-amanitin

作　　者：徐赛群[1] 朱小寒[1] 刘斌[1] 程宗勇[1] 

机构地区：[1]中南大学湘雅医学院湘雅二医院消化内科,长沙410011

出　　处：《中华急诊医学杂志》2005年第2期115-118,共4页Chinese Journal of Emergency Medicine

基　　金：湖南省发展计划委员会重点资助课题 ( 2 0 0 2 772 )

摘　　要：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 。Objective To investigate whether alpha-amanitin could induce hepatocytes apoptosis,and whether the apoptosis is dose-dependent or time-dependent.Methods Thirty Kun-ming mice were randomly divided into A,B,C,D,E,F group,5 mice in every group.Alpha-amanitin was injected into peritoneal cavity with 0.8,1.2 and 2.0 μg/g weight respectively in groups B to D,and group A injected with saline.The experimental final time was 24 h.Alpha-amanitin was injected the same way in group E and F with 2.0 μg/g weight,but the experimental final time were 16 h and 36 h respectively.Hepatocyte apoptosis was detected by light microscopy with HE staining,transmission electron microscopy(TEM),TdT-mediated duTP nick end labeling(TUNEL)method,and DNA“ladder”demonstrated by agarose gel electrophoresis.Results Experimental mice liver slices showed typical morphological change of apoptosis under light microscopy and transmission electron microscopy(TEM).DNA extracts from mice hepatocytes demonstrated typical DNA“ladder”under agarose gel electrophoresis.The nuclei of apoptotic hepatocytes were stained with brown under the light microscope by TUNEL method.Apoptosis index was 1.08%,4.88%,7.36%or 9.20% in group A to D(P<0.05),respectively,which reflected dose-dependent effect.However,it was 1.08%,5.68%,9.20%and 14.44% in group A,D,E or F(P<0.05),respectively,which reflected time-dependent effect.Conclusion Alpha-amanitin could induce hepatocyte apoptosis in mice,and there are dose-dependent effect and time-dependent within limited dose,and hepatocyte apoptosis rate increases with increasement of AMA dose and reacting time.

关 键 词：肝细胞凋亡 鼠肝 小鼠 剂量依赖性 经腹 诱导 腹腔注射 体重 果实 琼脂糖凝胶电泳 

分 类 号：R285[医药卫生—中药学]