RESUMO
Necrosis from freezing manifested an orderly pattern of DNA fragmentations, including the apoptosis signature of 200 bp ladder, in three different cell populations, despite pancaspase suppression by zVAD-fmk. Immediately on thawing, all three populations had 100% dead cell indices and 2.2, 1.6, and 1.1 megabase fragmentations, which marked the point of death. Kilobase and 200 bp DNA ladder fragmentations manifested later together with overt necrotic morphologies. CpG oligodeoxynucleotides (ODNs) complementary to highly conserved GCn(x)GC motifs inhibited the megabase fragmentations and retarded their electrophoretic mobility (gel shift), indicating ODN-DNA binding, which is known to confer site-specific resistance to cleavage. Cleavage specificity was confirmed using EDTA-CpG ODN conjugates to direct free-radical-producing transitional element, vanadyl(4), to the binding sites to reproduce the megabase fragmentations in normal cells. Specific orderly fragmentations in necrosis suggested a necrosis-apoptosis convergence after death has been committed.
