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目的 研究放线菌酮 (CHX)诱导小鼠肝细胞凋亡的作用机制。方法 利用琼脂糖凝胶电泳、流式细胞术、电镜、活细胞荧光染色、HE染色等方法观察CHX诱导肝细胞凋亡的情况。结果 ( 1)2 0mg kg以上剂量的CHX作用 1h即可出现典型的肝细胞凋亡 ,溴化乙啶和吖啶橙荧光染色为桔黄色 ,HE染色可见核固缩、浓染以及凋亡小体形成 ,凋亡细胞周围无炎性细胞浸润。电镜观察可见肝细胞体积变小 ,染色质固缩。 ( 2 )电泳结果显示 :不同剂量的CHX处理的小鼠其肝细胞NDA呈典型的“Ladder”电泳条带。 ( 3 )流式细胞术分析 :在CHX 2 0mg kg ,作用 1h以后即可见到肝细胞凋亡峰 (AP峰 )。随CHX剂量增加和作用时间的延长 ,凋亡率也逐渐增高 ,凋亡细胞的NDA含量减少。 ( 4)C myc基因表达量随CHX剂量增加和作用时间的延长而增加 ,但bcl 2基因表达呈下降趋势。结论 CHX可以快速诱导小鼠肝细胞凋亡 ,在凋亡过程中C myc基因表达量增加 ,bcl 2基因表达量降低 ,细胞凋亡率与CHX剂量和作用时间有一定的线性关系
Aim To study the mechanism of hepatocellular apoptosis induced by cycloheximide in mice. Methods The apoptosis of hepatocytes induced by CHX was observed by agarose gel electrophoresis, flow cytometry, electron microscopy, fluorescent staining of living cells and HE staining. Results (1) Typical hepatocytes apoptosis occurred at a dose of 20 mg / kg CHX for 1 h. Fluorescent staining with ethidium bromide and acridine orange was orange, nuclear condensation was observed with HE staining, and the number of apoptotic cells was small Body formation, no inflammatory cell infiltration around the apoptotic cells. Electron microscopy shows that hepatocytes become smaller and chromatin shrinks. (2) The result of electrophoresis showed that the NDA of liver cells of CHX-treated mice showed typical “Ladder” electrophoresis bands. (3) Flow cytometry analysis: The hepatocyte apoptosis peak (AP peak) could be seen after CHX 20 mg kg for 1 h. With the increase of CHX dose and the prolongation of action time, the apoptotic rate also increased gradually and the NDA content of apoptotic cells decreased. (4) The expression of Cmyc gene increased with the dose of CHX and the prolongation of action time, but the expression of bcl 2 gene decreased. Conclusions CHX can induce hepatocellular apoptosis in mice rapidly. The expression of C-myc gene increases and the expression of bcl-2 decreases during apoptosis. The apoptosis rate has a certain linear relationship with the dose and duration of CHX