Nakano Hisako, Yonekawa Hiromichi, Shinohara Kunio
Department of Laboratory Animal Science, Tokyo Metropolitan Institute of Medical Science, Tokyo, Japan.
Int J Radiat Oncol Biol Phys. 2007 Jul 1;68(3):883-91. doi: 10.1016/j.ijrobp.2007.03.007.
To determine the threshold level for the initiation of apoptosis by studying the quantitative aspect of p53 response to DNA damage in individual cells, to better understand the process in X-ray-induced p53-dependent apoptosis.
Time-sequential changes in p53 protein level were obtained for X-irradiated MOLT-4 cells using flow cytometry and analyzed.
The changes in the cellular frequency distribution pattern of p53 content could be divided into two parts at a certain p53 level. The p53 vs. side-scatter in flow cytometry showed the sequential changes of p53 increase followed by an increase in cell death. On the basis of these results we determined a threshold level of p53 for the initiation of apoptosis. The level was estimated to be (1.08 +/- 0.05) x 10(5) molecules per cell, which was approximately threefold higher than the mean content of control cells. The minimum times for p53 level to reach this threshold level were independent of X-ray dose and 1.4-1.6 h. The times for the signal transduction from the p53 accumulation to disruption of the mitochondrial membrane potential, caspase-3 activation, and cell death were 1.6, 2.1, and 2.8 h, respectively.
The threshold level of p53 for the initiation of apoptosis and the time sequence in the course of apoptotic events were determined in X-irradiated MOLT-4 cells.
通过研究单个细胞中p53对DNA损伤反应的定量方面，确定细胞凋亡启动的阈值水平，以更好地理解X射线诱导的p53依赖性细胞凋亡过程。
使用流式细胞术获得X射线照射的MOLT-4细胞中p53蛋白水平的时间序列变化并进行分析。
在一定的p53水平下，p53含量的细胞频率分布模式变化可分为两部分。流式细胞术中p53与侧向散射光显示p53先增加随后细胞死亡增加的序列变化。基于这些结果，我们确定了细胞凋亡启动的p53阈值水平。该水平估计为每个细胞(1.08±0.05)×10⁵个分子，约为对照细胞平均含量的三倍。p53水平达到该阈值水平的最短时间与X射线剂量无关，为1.4 - 1.6小时。从p53积累到线粒体膜电位破坏、caspase-3激活和细胞死亡的信号转导时间分别为1.6、2.1和2.8小时。
在X射线照射的MOLT-4细胞中确定了细胞凋亡启动的p53阈值水平以及凋亡事件过程中的时间序列。
