Selenium has a protective role in caspase-3-dependent apoptosis induced by H2O2 in primary cultured pig thyrocytes.
OBJECTIVE
Hydrogen peroxide (H2O2), necessary for thyroid hormonogenesis, is produced at the apical surface of the thyroid follicular epithelium. Excess H2O2 is potentially cytotoxic and may contribute to the development of hypothyroidism, e.g. in severe selenium deficiency. Yet it is unclear how H2O2 contributes to thyroid cell death.


METHODS
H2O2-induced apoptosis and necrosis were studied in primary cultured pig thyroid cells. Glutathione peroxidase (GPx) activity was altered by culture in low serum with or without selenite substitution. Apoptosis was evaluated by spectrofluorometric measurement of caspase-3-specific substrate cleavage, and by analysis of DNA fragmentation by agarose gel electrophoresis. Necrosis was detected by 51Cr release from prelabeled cells.


RESULTS
Exogenous H2O2 dose-dependently (100-400 micromol/l) activated caspase-3 within 3-12 h, and DNA degradation was observed after 24 h. The potency of H2O2 to induce apoptosis was low compared with that of staurosporine, a strong proapoptotic agent. H2O2-treated cells with reduced GPx activity showed increased caspase-3 activation. Incubation of serum-starved cells with selenite (10-100 nmol/l) normalized the GPx activity and reduced the activation of caspase-3 by H2O2. High H2O2 concentrations (400-800 micromol/l) were required to obtain necrosis. The H2O2-induced necrosis was exaggerated by both low GPx activity and catalase inhibition.


CONCLUSIONS
Cytotoxic effects of H2O2 on thyroid cells include caspase-3-dependent apoptosis that occurs at H2O2 concentrations insufficient to induce necrosis. Selenium deficiency aggravates the apoptotic response, probably due to impaired capacity of GPx to degrade H2O2.
Demelash
Abeba
A
Institute of Anatomy and Cell Biology, Sahlgrenska Academy at Gothenburg University, Gothenburg, Sweden. abeba.demelash@anatcell.gu.se
Karlsson
Jan-Olof
JO
Nilsson
Mikael
M
Björkman
Ulla
U
D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism*; D002478:Cells, Cultured; D002860:Chromium Radioisotopes / Q000378:metabolism; D053938:DNA Fragmentation / Q000187:drug effects; D004587:Electrophoresis, Agar Gel; D004789:Enzyme Activation / Q000187:drug effects; D005979:Glutathione Peroxidase / Q000378:metabolism; D006861:Hydrogen Peroxide / Q000494:pharmacology*; D012643:Selenium / Q000494:pharmacology*; D013050:Spectrometry, Fluorescence; D019311:Staurosporine / Q000494:pharmacology; D013552:Swine; D013961:Thyroid Gland / Q000166:cytology*
D002860:Chromium Radioisotopes; D006861:Hydrogen Peroxide; D005979:Glutathione Peroxidase; D053148:Caspase 3; D020169:Caspases; D012643:Selenium; D019311:Staurosporine