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徐文, 陈景元, 王枫. 锰诱导PC12细胞凋亡与p-38MAPKs的关系[J]. 中国公共卫生, 2004, 20(12): 1527-1529.
引用本文: 徐文, 陈景元, 王枫. 锰诱导PC12细胞凋亡与p-38MAPKs的关系[J]. 中国公共卫生, 2004, 20(12): 1527-1529.
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XU Wen, CHEN Jing-yuan, WANG Feng. Relationship between apoptosis and molecular mechanism of p-38 MAPKS activation in manganese-treated PC12 cells[J]. Chinese Journal of Public Health, 2004, 20(12): 1527-1529.
Citation: XU Wen, CHEN Jing-yuan, WANG Feng. Relationship between apoptosis and molecular mechanism of p-38 MAPKS activation in manganese-treated PC12 cells[J]. Chinese Journal of Public Health, 2004, 20(12): 1527-1529.
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锰诱导PC12细胞凋亡与p-38MAPKs的关系

Relationship between apoptosis and molecular mechanism of p-38 MAPKS activation in manganese-treated PC12 cells

    摘要
  • 摘要:
      目的   以鼠嗜铬神经瘤细胞(PC12)为模型, 筛选锰对神经细胞增殖抑制作用的时间及剂量, 观察细胞形态学、细胞周期和生化指标改变与丝裂原活化蛋白激酶(p38MAPKs)活化表达间的关系。
      方法   用200, 400, 600, 800μmol/LMnCl2的培养液, 分别作用对数生长期PC12细胞1, 2, 3, 4d后, 用噻唑蓝比色(MTT)筛选锰的细胞毒性剂量; 流式细胞仪检测细胞周期分布; 透射电镜观察细胞形态学变化; 琼脂糖凝胶电泳检测MnCl2对PC12细胞基因组DNA的影响.蛋白印迹(western-blot)法检测p-p38。
      结果   MTT实验结果显示, 200~800μmol/LMnCl2作用1, 2, 3, 4d对PC12有显著的抑制作用, 呈剂量和时间依赖趋势, 600μmol/LMnCl2作用4d对PC12的抑制率可达50%以上.流式细胞仪检测实验表明: 600μmol/LMnCl2作用4d将PC12细胞周期阻滞在S期, 诱导细胞凋亡, 与电镜结果一致, 同样条件下细胞DNA碎片化.Western-blot实验显示600μmol/LMnCl2作用1, 2, 3, 4dp-p38逐渐升高, 3d时较对照组增加66倍(n=3, P < 005), 200, 400, 600μmol/LMnCl2作用4d时, 磷酸化蛋白38(p-p38)也逐渐升高, 400μmol/LMnCl2作用4d时较对照组升高47倍(n=3, P < 005)。
      结论   锰通过MEK3/6磷酸化下游p38, 上调p-p38, 诱导细胞凋亡, 细胞增殖抑制。

     

    Abstract:
      Objective   To employ PC12 cell line as a model in vitro to test the concentration and time of maganese on proliferation arrest, and to observe the changes of cell morphology, cell cycle and biochemical marker and further to study the relationships between above aspects and activiations of p38MA PKs pathway.PC cells in logarithm period incubated in culture media with 200, 400, 600, 800 μmol/L manganese(MnCl2)for 1, 2, 3, 4 days respectively.
      Methods   Cell viability was examined by MTT.Cell cycle was monitored by FCM(flow cytometry).Morphological changes of PC12 cells was investigated by transmisssion electron microscope.Agarose gel electrophoresis was used to test the genomic DNA.Western-blot was used to test p-p38.
      Results   MTT revealed that 200, 400, 600, 800 μmol/L MnCl2 could suppress the proliferation of PC12 cells in dose and time-dependent trend.The cell inhibited ratio on the fourth day in 600 μmol/L MnCl2 approached 50% or more.FCM showed that the cell cycle of the PC12 cells could be inhibited in Speriod at the concentration of 600 μmol/L MnCl2 on the 4th day and apoptosis was observed through FCM and transmisssion electron microscope Biochemical hallmark of DNA fragments was observed.Western-blottests showed the p-p38 in 600 μmol/L MnCl2 culture medium was increasing gradually on the 1st, 2nd, 3rd and 4th day.the p-p38 on the 3rd day was 616 times higher than that of control group(n=3, P < 0.05).The p-p38 was enhanced by degrees in 200, 400, 600μmol/L MnCl2-treated PC12 cells in 4 days.The p-p38 of 400 μmol/L MnCl2 treated group on the 4th day was 4.7 times higher than that of control group(n=3, P < 0.05).
      Conclusion   Manganese upregulated p-p38 through MEK 3/6 downstream to induce proliferation arrest and apoptotic cell death.

     

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