A light and electron microscopic study on complete dissociation of rat ascites hepatoma cells under activation of neutral protease and calcium depletion

Summary

A neutral proteasea, which was extracted from rat ascites hepatoma cells and then purified by chromatography using TEAE-cellulose followed by hydroxyapatite (eluted in 0.12 M phosphate buffer), induced an electron microscopic alteration (disruption) of tight junctions resulting in a dissociation of the close contact in the apical portion of the cell islands, but simple apposition, intermediate junctions and desmosomes in the close contact in the inner portion of the cell islands remained intact. On the other hand, ethylenediamine tetraacetate (EDTA), as previously described, induced an electron microscopic alteration of simple apposition, intermediate junctions and desmosomes resulting in a dissociation of the close contact in the inner portion of the cell islands, but tight junctions remained unchanged, Thus, the protease could provoke a complete microscopic dissociation of the cells in the presence of EDTA. A similar complete dissociation of the cells, under calcium-depleted conditions, was also provoked by a factor (of peptide nature) capable of activating and releasing the protease in and from the cells, which was isolated from tumor tissue. In contrast, no or little dissociation of the cells was induced by a neutral proteaseb in the presence of EDTA. This protease was extracted from the cells and eluted in 0.1 M phosphate buffer on hydroxyapatite described above. It was therefore assumed that such tumor cell dissociation may be associated with the combined conditions of activation of a certain neutral protease and calcium depletion.