Mori C, Nakamura N, Kimura S, Irie H, Takigawa T, Shiota K
Department of Anatomy, Faculty of Medicine, Kyoto University, Japan.
Anat Rec. 1995 May;242(1):103-10. doi: 10.1002/ar.1092420114.
Programmed cell death is an essential event during mammalian morphogenesis which eliminates unnecessary cells to accomplish histogenesis and organogenesis. Cell death in interdigital spaces of the developing limb is a classical example of morphogenetic cell death. We investigated whether classical programmed cell death in the interdigital tissue of the developing limb in mice is apoptosis with fragmentation of nuclear DNA and also examined sequentially the occurrence of programmed cell death and cell proliferation in the developing limb of mouse fetuses to analyze their interrelation.
We examined the occurrence of apoptotic cell death in the developing limbs of mouse fetuses by using Nile blue sulphate staining, agarose gel electrophoresis for detecting DNA laddering, and a cytochemical labeling of DNA fragmentation. We also labeled proliferating cells using BrdU/anti-BrdU immunohistochemistry and examined the interrelation between apoptotic programmed cell death and cell proliferation.
DNA ladders, a biochemical evidence of apoptosis, were detected in DNA extracts from the interdigital tissue of day 13 mouse fetuses by agarose gel electrophoresis. Programmed cell death and DNA fragmentation were detected by Nile blue staining and cytochemical labeling of DNA fragmentation, respectively, in the interdigital mesoderm and in the regions of presumptive joints of the digit. BrdU/anti-BrdU immunohistochemistry for identifying proliferating S-phase cells revealed that interdigital mesenchymal cells cease DNA synthesis before programmed cell death and DNA fragmentation begin.
We confirmed that both cytological apoptotic alterations and fragmentation of nuclear DNA occur in the interdigital tissue and presumptive joint areas of fetal mouse limbs, and they appear to play a significant role in the separation of digits as well as the formation of joint cavities.
程序性细胞死亡是哺乳动物形态发生过程中的一个重要事件，它通过消除多余细胞来完成组织发生和器官发生。发育中肢体指间间隙的细胞死亡是形态发生性细胞死亡的一个经典例子。我们研究了小鼠发育中肢体指间组织中的经典程序性细胞死亡是否是伴有核DNA片段化的凋亡，并依次检查了小鼠胎儿发育中肢体程序性细胞死亡和细胞增殖的发生情况，以分析它们之间的相互关系。
我们通过使用硫酸尼罗蓝染色、用于检测DNA梯状条带的琼脂糖凝胶电泳以及DNA片段化的细胞化学标记，来检查小鼠胎儿发育中肢体凋亡性细胞死亡的发生情况。我们还使用BrdU/抗BrdU免疫组织化学标记增殖细胞，并检查凋亡性程序性细胞死亡与细胞增殖之间的相互关系。
通过琼脂糖凝胶电泳在第13天小鼠胎儿指间组织的DNA提取物中检测到DNA梯状条带，这是凋亡的生化证据。分别通过尼罗蓝染色和DNA片段化的细胞化学标记，在指间中胚层和指的假定关节区域检测到程序性细胞死亡和DNA片段化。用于识别增殖S期细胞的BrdU/抗BrdU免疫组织化学显示，指间间充质细胞在程序性细胞死亡和DNA片段化开始之前停止DNA合成。
我们证实，在胎儿小鼠肢体的指间组织和假定关节区域发生了细胞学上的凋亡改变和核DNA片段化，它们似乎在指的分离以及关节腔的形成中发挥了重要作用。
