Effects of Long-Term Exposure to Iodine Excess on the Apoptosis of Thyrocytes in Wistar Rats

Abstract

Objective: To explore the effects of long-term exposure to more than adequate and excessive iodine on the apoptosis of thyrocytes in Wistar rats.

Methods: Wistar rats, divided into four groups: Group 0 (control, treated with 4 μg/day of iodine), Group 1 (1.5x control dose, i. e. 6 μg/day of iodine), Group 2 (3x control dose, i. e. 12 μg/day of iodine), and Group 3 (6x control dose, i. e. 24 μg/day of iodine) had oral iodine intake for 1, 2, 4, and 8 months before being sacrificed for study. After the initial treatment course of 8 months, some rats in each group continued to have an iodine intake at 4 μg/day for extra 3 months. Urinary iodine concentration was measured by arsenic/cerium catalyzing spectrophotography. In addition, the early apoptosis was assessed by flow cytometry with Annexin V-FITC staining, while the late apoptosis was evaluated by TUNEL microscopy. Furthermore, the rate of cell death and cell cycle kinetics were determined by flow cytometry with propidium iodide (PI) staining. Moreover, the level of intracellular reactive oxygen species (ROS) quantified by the fluorescence intensity was measured via flow cytometry with a DCFH-DA probe. Immunohistochemistry technique was also used for the detection of the expression of Bcl-2 and Bax.

Results: The apoptotic rate and ROS production in thyrocytes were significantly elevated in Groups 2 and 3 after 4 and 8 months of treatment compared to the control group (P<0.05). Groups 2 and 3 were also found to have greatly higher percentage of cells in S phase after 4 and 8 months of exposure (5%–6% vs. 3%, P<0.05), with a significant reduction in cells in G0G1-phase (64%–67% vs. 80%, P<0.05). These differences disappeared after normal iodine intake for extra 3 months. The expressions of Bcl-2 and Bax among different groups remained unchanged. Positive correlations were observed between the apoptotic rate, ROS level and urinary iodine concentration after 4 and 8 months of excessive iodine exposure (r is in the range of 0.736˜0.855, all of Ps<0.01).

Conclusion: The experimental data indicated that more apoptosis of thyrocytes after exposure to excessive iodine was attributed to the increased free radical production, other than regulated Bcl-2 and Bax expression. The effects of iodine excess are reversible.

iodine - thyroid - apoptosis

References

• 1 Bilodeau JF, Faure R, Piedboeuf B. et al . Hyperoxia induces S-phase cell-cycle arrest and p21(Cip1/Waf1)-independent Cdk2 inhibition in human carcinoma T47D-H3 cells.  Exp Cell Res. 2000;  256 347-357
• 2 Bjorkman U, Ekholm R, Denef J-F. Cytochemical localization of hydrogen peroxide in isolated thyroid follicles.  J Ultrastruct Res. 1981;  74 105-115
• 3 Braverman LE, Ingbar SH. Changes in thyroidal function during adaptation to large doses ofiodide.  J Clin Invest. 1963;  42 1216-1231
• 4 Burikhanov RB, Matsuzaki S. Excess iodine induces apoptosis in the thyroid of goitrogen pretreated rats in vivo.  Thyroid. 2000;  10 123-129
• 5 Ekholm R, Bjorkman U. Glutathione Peroxidase Degrades Intracellular Hydrogen Peroxide and Thereby Inhibits Intracellular Protein Iodination in Thyroid Epithelium.  Endocrinol. 1997;  138 2871-2878
• 6 Eng PH, Cardona GR, Fang SL. et al . Escape from the acute Wolff-Chaikoff effect is associated with a decrease in thyroid sodium/iodide symporter messenger ribonucleic acid and protein.  Endocrinol. 1999;  140 3404-3410
• 7 Gao TS, Hu FN, Teng WP. Effects of mild and moderate excessive iodine supplementation on thyroid function and morphology in non-iodine deficiency rat model.  Chin J Intern Med. 2003;  42 705-708
• 8 Golstein J, Dumont JE. Cytotoxic effects of iodide on thyroid cells:difference between rat thyroid FRTL-5 cells and primary dog thyrocyte responsiveness.  J Endocrinol Invest. 1996;  19 119-126
• 9 Johannes B, Jan AP. Molecular events associated with reactive oxygen species and cell cycle progression in mammalian cells.  Gene. 2004;  337 1-13
• 10 Laurberg P, Bulow Pedersen I, Knudsen N. et al . Environmental iodine intake affects the type of nonmalignant thyroid disease.  Thyroid. 2001;  11 457-469
• 11 Lennon SV, Martin SJ, Cotter TG. Dose-dependent induction of apoptosis in human tumour cell lines by widely diverging stimuli.  Cell Prolif. 1991;  24 203-207
• 12 Lin LX, Ye ZK, Nie XL. et al . Effect of iodine on the apoptosis and expressions of bax, bcl-2 mRNA in rats.  Clin J Endomiol. 2005;  24 245-247
• 13 Man N, Guan HX, Shan ZY. et al . Long-term effects of high iodine intake: inhibition of thyroid iodine uptake and organification in Wistar rats.  Natl Med J China. 2006;  83 3420-3424
• 14 McGowan AJ, Fernandes RS, Samali A. et al . Anti-oxidants and apoptosis.  Biochem Soc Trans. 1996;  24 229-235
• 15 Mooij P, Wit HJ, Bloot AM. et al . Iodine deficiency induces thyroid autoimmune reactivity in Wistar rats.  Endocrinol. 1993;  133 1197-1204
• 16 Riou C, Remy C, Rabilloud R. et al . H2O2 induces apoptosis of pig thyrocytes in culture.  J Endocrinol. 1998;  156 315-322
• 17 Singh PP, Kumar GP, Laloraya M. Regulation of Superoxide Anion Radical-Superoxide Dismutase System in the Avian Thyroid by TSH with Reference to Thyroid Hormonogenesis.  Biochem Biophys Res Commun. 1997;  239 212-216
• 18 Smerdely P, Pitsiavas V, Boyages SC. Evidence that the inhibitory effects of iodide on thyroid cell proliferation are due to arrest of the cell cycle at G0G1 and G2M phases.  Endocrinol. 1993;  133 2881-2888
• 19 Strasser A, O’Connor L, Dixit VM. Apoptosis signaling.  Annu Rev Biochem. 2000;  69 217-245
• 20 Sugawara M, Sugawara Y, Wen K. et al . Generation of oxygen free radicals in thyroid cells and inhibition of thyroid peroxidase.  Exp Biol Med(Maywood). 2002;  227 141-146
• 21 Tamura M, Kimura H, Koji T. et al . Role of apoptosis of thyrocytes in a rat model of goiter: A possible involvement of fas system.  Endocrinol. 1998;  139 3646-3653
• 22 Teng W, Shan Z, Teng X. et al . Effect of iodine intake on thyroid diseases in China.  N Engl J Med. 2006;  354 2783-2793
• 23 Uyttersprot N, Pelgrins N, Carrasco N. et al . Moderate doses of iodide in vivo inhibit cell proliferation and the expression of thyroperoxidase and Na+/I- symporter mRNAs in dog thyroid.  Mol Cell Endocrinol. 1997;  131 195-203
• 24 Vitale M, Di Matola T, D’Ascoli F. et al . Iodide excess induces apoptosis in thyroid cells through a p53-independent mechanism involving oxidative stress.  Endocrinol. 2000;  141 598-605
• 25 Wolff J, Chaikoff IL. Plasma inorganic iodide as a homeostatic regulator of thyroid function.  J Biol Chem. 1948;  154 555-564
• 26 Zhang N, Tong Y, Shan Z. et al . Effect of chronic mild and moderate iodine excess on the anti-oxidant capability of thyroid in Wistar rats with or without iodine dificiency.  Natl Med J China. 2006;  86 1274-1278
• 27 Zheng H, Wang Y, Wang M. Study on anti-oxidant capability of mice with different iodine intake.  Chin J Ctrl Endem Dis. 2002;  17 219-221

Correspondence