Hepatocytes apoptosis in mice induced by alpha-amanitin

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(P0.05),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(P0.05),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.