BACKGROUND: 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.
METHODS: 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.
RESULTS: 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.
CONCLUSIONS: 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.