Effects of antioxidants on induction of apoptosis in bursal cells of Fabricius during in vitro cultivation

After physically disrupting cell contacts, apoptosis of bursal cells of Fab ricius was induced during in vitro cultivation. The percentage of apoptotic cells increased with incubation time and approximately 70% cells represent ed apoptosis after 6 hr of incubation. The induction of apoptosis was signi ficantly inhibited by treatment of the cells with ascorbic acid (vitamin C) , but not with trolox, a vitamin E analog. An intense DNA ladder pattern wa s shown at 6 hr post-isolation, which is a biochemical hallmark of apoptosi s. Treatment of the cells with ascorbic acid inhibited the DNA fragmentatio n, but trolox did not. To monitor the intracellular production of reactive oxygen species (ROSs), the intensity of fluorescence emitted from DCFH-DA w as measured. The intensity of fluorescence from cells incubated for 0.5-2 h r was approximately 2-fold higher than that from cells at 0 hr. The relativ e intensity of fluorescence decreased immediately after the addition of asc orbic acid to the cells. The intensity from the cells treated with ascorbic acid was 20-30% of that from the control cells at each incubation time. Fo r trolox, the intensity was 50-70% of that from the control cells at each 1 to 2 hr incubation time. When ROSs-induced lipid peroxidation was assessed using cis-parinaric acid (PnA) as a monitor molecule, lipid peroxidation w as found to occur in the control cells after isolation of the bursal cells. Treatment of the cells with trolox reduced lipid peroxidation, but treatme nt with ascorbic acid enhanced peroxidation.