A STUDY OF THE BIOLOGICAL OXIDATION IN APOPTOSIS

Abstract

Intranuclear oxidative stress was imposed on human Chang liver cells by the induction of nuclear internalization of vanadyl(4) via an acidification prepulse, which was shown by nuclear microscopy. This allowed reactive oxygen species (ROS) to be liberated intranuclearly with realkalinization. DNA damage was shown within 5-10 minutes by detecting megabase level DNA fragmentations in pulsed field gel electrophoresis (PFGE). There was corroborative self-inflicted destruction of euchromatin domains with self-expressed light G-band resolutions in mitotic chromosomes. The megabase (Mb) cleavage produced 2.2 Mb, 1.6 Mb, 1.1 Mb and 0.68 Mb bands which reassembled the pattern from CpG specific cleavages by rare cutter restriction endonuclease Notl. The megabase cleavage sites at or in close proximity with CpG islands were suggested. The CpG islands are known to be at 5' promoter regions of all housekeeping genes. Oxidative stress was imposed on human Chang liver cells by downregulating the cellular main detoxification defence. Sulfation and glutathione antioxidative detoxification defence are known interdependent processes. 2,6-dichloro-4-nitrophenol (DCNP), inhibition of sulfation, led to the induction of a burst of ROS which was shown by flow cytometry. Glutathione was depleted, suggesting impairment of both sulfation and oxidative defence. Interestingly, this indirect oxidative induced apoptosis with the similar two characteristics: ( 1 ). Early megabase level DNA fragmentation with CpG specific cleavage patterns. (2). Proliferation-specific apoptosis. Oxidative stress produced early megabase level DNA fragmentations with a CpG-specific pattern. Apoptosis occurred following early DNA fragmentations. This apoptosis seemed to be associated with proliferative processes as shown by conjoint chromosomal and apoptotic condensations in the same cell and apoptosis targeting proliferative cells.