Thyroid and Hydrogen Peroxide / 한양의대학술지

The thyroid gland produces high concentrations of hydrogen peroxide (H2O2) which is used for the biosynthesis of the thyroid hormones, triiodothyronine and thyroxine. The most important step in thyroid hormone synthesis is oxidation and organification of iodide, which itself is a very potent oxidant. To oxidize iodide, thyrocytes generate high intracellular concentrations of H2O2 using the NADPH-dependent dual oxidase enzymes (DUOX) and remove used H2O2 by various well-characterized antioxidant systems, such as those involving glutathione peroxidases and peroxiredoxins. Although thyrocytes have very intricate and efficient systems to regulate H2O2 generation and removal, leakage of H2O2 may be inevitable because of the high intracellular concentrations of H2O2 in thyrocytes. Leakage of the highly reactive oxidant H2O2 is likely to cause the damage to various intracellular proteins and DNA that has been demonstrated to be a causative factor in the induction of various diseases, such as thyroid cancer, chronic thyroiditis and hyperthyroidism. Therefore, ongoing investigation of the systems regulating H2O2 production and elimination in thyrocytesis essential for understanding the regulation and process of thyroid hormone synthesis and also for elucidating pathogenetic mechanisms underlying various autoimmune thyroid diseases and the initiation and promotion of thyroid cancers.

Effect of High Concentration of Estradiol on Thyroid Specific Genes Expression and Cell Growth / 대한내분비학회지

BACKGROUND: Since various thyroid diseases have dominant prevalence in women, it has been suggested that female sex hormone have important role on thyroid cell physiology. Interestingly, many thyroid disorders are newly diagnosed or changed their course around the period of high estrogen status, such as pregnancy. In this study, we questioned whether high concentration of estrogen could modulate thyroid cell function. METHODS: We treated normal rat thyroid FRTL-5 cell line with different time and concentration of estradiol. Using cell count, FACscan, and Northern blot analysis, we compared the changes of cell growth, cell cycle progression and thyroid specific genes expression. To evaluate the influence of thyroid stimulating hormone (TSH), all experiment was designed as two different sets, with (6H) or without TSH (5H). RESULTS: The concentration of 10-1000 nM estradiol had definite stimulatory function on thyroid cell growth in 5H condition as concentration dependent manner. FACscan revealed the increased cell growths were related to G1/S progression. The Pax-8, TTF-1 and NIS gene expressions were dramatically increased in 10-1000 nM of estradiol, too. With TSH (6H), however, we could not find any cell growth stimulating effects with 10-1000 nM of estradiol. CONCLUSION: High concentration of estradiol is one of important control factor for thyroid growth and thyroid specific genes expression, especially in 5H condition. It indicate that exposure to high concentration of female sex hormone, such as pregnancy, can be a direct stimulating factor to various thyroid function and related to autoimmune or nodular thyroid diseases around the period of pregnancy.

Effect of Estrogen on H2O2 Induced Apoptosis of FRTL-5 Cells / 대한내분비학회지

BACKGROUND: Understanding the pathways and controlling mechanisms of thyrocyte apoptosis is important for the elucidation of the pathogenesis of goiter or thyroid cancer. A system for evaluating apoptosis, in FRTL-5 cells, triggered by hydrogen peroxide (H2O2), a highly likely apoptogenic signal in physiologic condition, was be set up to see the effects of TSH and estrogen on H2O2-induced apoptosis. METHOD: DNA laddering was used in the optimization process or the conditions of the set-up of system for the evaluation of apoptosis in the FRTL-5 cells. To quantify the apoptosis under the optimized conditions, histone-bound DNA fragments in the cytoplasm were measured by ELISA. RESULTS: 1) The optimized conditions for induction of apoptosis in the FRTL-5 cells by H2O2 were; observation of DNA laddering 18~24 hrs after the addition of 0.3 mM H2O2 to cells maintained in TSH-free, low serum containing media (5H1 or 5H0 media) for 48 hrs. 2) Exposure of the FRTL-5 cells to TSH (1 mU/L) for more than 48 hrs (6H0 media). before the addition of H2O2 significantly decreased the degree of apoptosis, compared to cells maintained under TSH-free conditions (0.98+/-0.21 vs. 2.27 0.11 arbitrary unit, p<0.05), whereas exposure for 24 hrs. did not. 3) Exposure of the FRTL-5 cells to high dose 17- estradiol (1-100 M) significantly decreased the degree of H2O2-induced apoptosis in a dose dependent manner. The addition of serum (1%) blunted the effects of estrogen on H2O2-induced apoptosis, and TSH totally abrogated the estrogen effect.Physiologic doses of estrogen (10~100 nM) showed no suppressive effects on H2O2-induced apoptosis in FRTL-5 cells. CONCLUSION: A system for evaluating apoptosis in FRTL-5 cells triggered by hydrogen peroxide (H2O2), a highly likely apoptogenic signal in physiologic condition, was set up, and found for the first time that high dose estrogen suppressed the H2O2-induced apoptosis in FRTL-5 cells

Effect of Estrogen on H2O2 Induced Apoptosis of FRTL-5 Cells / 대한내분비학회지

BACKGROUND: Understanding the pathways and controlling mechanisms of thyrocyte apoptosis is important for the elucidation of the pathogenesis of goiter or thyroid cancer. A system for evaluating apoptosis, in FRTL-5 cells, triggered by hydrogen peroxide (H2O2), a highly likely apoptogenic signal in physiologic condition, was be set up to see the effects of TSH and estrogen on H2O2-induced apoptosis. METHOD: DNA laddering was used in the optimization process or the conditions of the set-up of system for the evaluation of apoptosis in the FRTL-5 cells. To quantify the apoptosis under the optimized conditions, histone-bound DNA fragments in the cytoplasm were measured by ELISA. RESULTS: 1) The optimized conditions for induction of apoptosis in the FRTL-5 cells by H2O2 were; observation of DNA laddering 18~24 hrs after the addition of 0.3 mM H2O2 to cells maintained in TSH-free, low serum containing media (5H1 or 5H0 media) for 48 hrs. 2) Exposure of the FRTL-5 cells to TSH (1 mU/L) for more than 48 hrs (6H0 media). before the addition of H2O2 significantly decreased the degree of apoptosis, compared to cells maintained under TSH-free conditions (0.98+/-0.21 vs. 2.27 0.11 arbitrary unit, p<0.05), whereas exposure for 24 hrs. did not. 3) Exposure of the FRTL-5 cells to high dose 17- estradiol (1-100 M) significantly decreased the degree of H2O2-induced apoptosis in a dose dependent manner. The addition of serum (1%) blunted the effects of estrogen on H2O2-induced apoptosis, and TSH totally abrogated the estrogen effect.Physiologic doses of estrogen (10~100 nM) showed no suppressive effects on H2O2-induced apoptosis in FRTL-5 cells. CONCLUSION: A system for evaluating apoptosis in FRTL-5 cells triggered by hydrogen peroxide (H2O2), a highly likely apoptogenic signal in physiologic condition, was set up, and found for the first time that high dose estrogen suppressed the H2O2-induced apoptosis in FRTL-5 cells

Effect of Class 2 Transactivator on Expression of Thyroid Transcription Factor-1 in FRTL-5 Cells / 대한내분비학회지

BACKGROUND: Gamma-interferon (gamma-IFN) is known to suppress thyroperoxidase (TPO), thyroglobulin (Tg), thyrotropin receptor (TSHR) mRNA expression via unclarified mechanism. Thyroid transcription factor-1 (TTF-1) is a nuclear protein involved in the maximal expression and tissue specific expression of thyroid-specific antigens (TPO, Tg, TSHR, NIS) in thyrocytes. Although It's plausible that gamam-IFN induced suppression of thyroid-specific antigen expression may be mediated by decrease of TTF-1 expression, such an effect has not been documented yet. In this study we investigated the effect of gamma-IFN on the expression of TTF-1 in the rat thyroid cell, FRTL-5, and determined whether such an effect is mediated by sclass 2 transactivator (CIITA). METHEODS: The mRNA expression of TTF-1 was quantitated by northern blot analysis after treatment of gamma-IFN, and after expression of CIITA in FRTL-5 cells. Four different promoter constructs were made by cloning into the pRSV-luciferase vector, each contained 5'flanking sequence of different lengths (-5.18 kb, -4.11 kb, -1.94 kb, -1.15 kb) of rat TTF-1 gene. Effects of gamma-IFN and CIITA on promoter activities were analyzed by luciferase assay in FRTL-5 cells into which each promoter construct had been transfected by DEAE-dextran method. RESULTS: Steady state TTF-1 mRNA level at 48 h after gamma-IFN treatment (100 U/mL) was significantly decreased from that of the pre-treatment level (1.65+/-0.16 vs. unit, p<0.05). In all 4 promoter constructs gamma-IFN significantly suppressed promoter activities compared to the vector only transfected cells. CIITA expression in FRTL-5 cells significantly decreased the steady state TTF-1 mRNA level when compared to that in mock-transfected cells (1.69+/-0.31 vs. 1.17+/-0.44 arbitrary unit, p<0.05). CIITA expression in FRTL-5 cells caused suppression of promoter activities in -5.18 kb and -4.11 kb constructs, but had no effects on those activities in -1.94 kb; and -1.15 kb constructs. CONCLUSION: gamma-IFN, directly and indirectly via CIITA expression, suppressed the transcription of TTF-1 gene in the FRTL-5 cells. It may be one of the mechanisms involved in the gamma-IFN-induced suppression of thyroid-specific protein expressions in thyrocytes 1.25+/-0.27 arbitrary

The signal pathways involved in the effects of thyroid stimulating antibody on the secretory function of thyrocytes / 中国实用内科杂志

Objective To investigate the signal pathways involved in the effects of thyroid stimulating antibody(TSAb)on the secretory function of cultured thyrocytes.Methods (1)With the method of enzyme-linked immunosorbent assay (ELISA),the PKA and PKC activities were observed.(2)Inhibitors or activators of protein kinases (PKA and PKC) were used to activate or block the signal pathways and the secretion of T3 measured by radioimmunoassay.Results (1)TSAb activated both PKA and PKC in thyrocytes in a dose-,time-dependent manner(.P.

Regulating Effect of Nitrogen Monoxide on Gene Expression of Sodium/Iodide Symporter in Human Thyrocyte / 中国药房

OBJECTIVE:To investigate the influence of nitrogen monoxide(NO) donor,sodium nitroprusside(SNP),on mRNA expression of sodium/iodide symporter(NIS) in thyrocyte membrane cultured in vitro in order to interpret molecular mechanism that NO inhibits the uptake of iodide.METHODS:The effect of SNP at different concentrations(0,10-6,10-5,10-4,10-3 mmol?L-1) on mRNA expression of NIS in thyrocyte by using cell culture method and RT-PCR with optical density ratio of NIS/?-actin as index.RESULTS:Optical density ratio of NIS/?-actin in each group decreased in order after treated with SNP(0,10-6,10-5,10-4,10-3 mmol?L-1).NO inhibited mRNA expression of NIS in a dose-dependent manner(P