H2O2 INDUCES APOPTOSIS OF PIG THYROCYTES IN CULTURE

Apoptosis might be involved in the reduction of the thyroid cell popul ation in physiopathological situations such as goitre involution and a utoimmune deleterious processes. Up to now, little attention has been paid to the apoptotic phenomenon in the normal thyroid gland the speci alized metabolism of which is expected to generate reactive oxygen spe cies. Indeed, thyroid cells have the capacity to synthesize H2O2. In t his study, we have analyzed the capacity of H2O2 to trigger apoptosis of pig thyrocytes in culture to try to determine whether thyrocytes ex hibit a particular resistance to apoptosis induced by an oxidative str ess. We show that exposure of thyrocytes cultured as monolayers to exo genous H2O2 induced cell death with characteristics of apoptosis. The effect of H2O2 was concentration-dependent; apoptotic cells were alrea dy observed alter exposure to 50 mu M H2O2. At high concentrations (mi llimolar range), H2O2 exerted toxic effects leading to rapid cell disr uption. Within the first hour after the onset of exposure to 50-300 mu M H2O2, early signs of apoptosis, i.e. DNA fragmentation, appeared in a low (0.1-1%) but definite fraction of thyrocytes. The proportion of adherent cells exhibiting DNA fragmentation remained fairly constant after 6, 15 and 24 h. During the 24-h period, an increasing number of cells detached from the culture dish and up to 30-40% of cells in susp ension displayed apoptotic features. The fraction of cells that lost c ontact with the culture dish amounted to up to 25% 24 h alter addition of 300 mu M H2O2. In conclusion, as reported for other cell types, lo w H2O2 concentrations are capable of triggering apoptosis in thyrocyte s cultured as monolayers. Thyrocytes that undergo apoptosis secondaril y lose contact with neighbour cells and the substratum; cell detachmen t from the monolayer probably happens within 1-2 h after initiation of DNA fragmentation. Our data show that the apoptotic commitment can ta ke place many hours after initiation of the oxidative stress.