Distribution of apoptosis-like cells in sea urchin early embryogenesis

Extract

It has been reported that the number of cells per embryo increases from the cleavage stage to the pluteus stage. Also, it has been reported that the number of cells per embryo from the early gastrula stage to the mid-gastrula stage increases very slightly (Mizoguchi, 1999). A detailed analysis of cell proliferation during this period would thus seem to be necessary.

On the other hand, Roccheri et al. (1997) reported spontaneous apoptosis at the early pluteus stage, especially in the regions of arm and intestine. However, it is unknown whether apoptosis occurs before the pluteus stage.

Using the Tumor, Neuro and/or Cardio TACS in situ apoptosis detection kit (terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labelling: TUNEL method, Trevigen, USA), the Comet assay kit (Trevigen, USA) and DAPI (4,6-diamidino-2-phenylinodole dihydrochloride) staining, we investigated the distribution of apoptosis-like cells in sea urchins during early embryogenesis to clarify the relationship between cell death and morphogenesis.

The embryos of the sea urchin Hemicentrotus pulcherrimus were used in the present study. Three methods of detection of apoptosis signals in the embryos were performed: (1) defects in chromosomes and punctured nuclear envelopes of cells were detected by DAPI staining; (2) apoptosis-like cells were stained a brown colour by the TUNEL method; (3) some clear tails which detected the fragmentation of DNA were found by the Comet assay. These signals indicate apoptosis-like cells.