2-iodohexadecanal induces autophagy during goiter involution.

BACKGROUND: Iodine plays an important role in thyroid physiology and biochemistry. The thyroid is capable of producing different iodolipids such as 2-iodohexadecanal (2-IHDA). Data from different laboratories have shown that 2-IHDA inhibits several thyroid parameters and it has been postulated as intermediary on the action of iodide function. OBJECTIVE: To explore different mechanisms involved during the involution of the hyperplastic thyroid gland of Wistar rats towards normality induced by 2-IHDA. METHODS: Goiter was induced by the administration of MMI for 10 days, then the treatment was discontinued and Wistar rats were injected with 2-IHDA or KI. RESULTS: During involution, 2-IHDA treatment reduced PCNA expression compared to spontaneous involution. KI treatment caused an increase of Caspase-3 activity and TUNEL-positive cells. In contrast, 2-IHDA failed to alter this value but induced an increase of LC3B expression. KI but not 2-IHDA led to an increase in peroxides levels, catalase and glutathione peroxidase activity. CONCLUSIONS: We demonstrated that 2-IHDA, in contrast to iodide, did not lead to an increase in oxidative stress or apoptosis induction, indicating that the involution triggered by 2-IHDA in Wistar rats, is primarily due to the inhibition of cell proliferation and the induction of autophagy.

Radiosensitivity enhancement of human thyroid carcinoma cells by the inhibitors of histone deacetylase sodium butyrate and valproic acid.

Radiotherapy is one of the leading treatments for clinical cancer therapy. External beam radiotherapy has been proposed as an adjuvant treatment for patients bearing differentiated thyroid cancer refractory to conventional therapy. Our purpose was to study the combined effect of HDAC inhibitors (HDACi) and ionizing irradiation in thyroid cancer cell lines (Nthy-ori 3-1, WRO, TPC-1 and 8505c). HDACi radiosensitized thyroid cancer cells as evidenced by the reduction of survival fraction, whereas they had no effect in the normal cells. HDACi enhanced radiation-induced cell death in WRO cells. Gamma-H2AX foci number increased and persisted long after ionizing exposure in the HDACi-treated cells (WRO and TPC-1). Moreover, the expression of the repair-related gene Ku80 was differentially modulated only in the cancer cells, by the compounds at the protein and/or mRNA levels. We present in vitro evidence that HDACi can enhance the radiosensitivity of human thyroid cancer cells.

NFE2-Related Transcription Factor 2 Coordinates Antioxidant Defense with Thyroglobulin Production and Iodination in the Thyroid Gland.

BACKGROUND: The thyroid gland has a special relationship with oxidative stress. While generation of oxidative substances is part of normal iodide metabolism during thyroid hormone synthesis, the gland must also defend itself against excessive oxidation in order to maintain normal function. Antioxidant and detoxification enzymes aid thyroid cells to maintain homeostasis by ameliorating oxidative insults, including during exposure to excess iodide, but the factors that coordinate their expression with the cellular redox status are not known. The antioxidant response system comprising the ubiquitously expressed NFE2-related transcription factor 2 (Nrf2) and its redox-sensitive cytoplasmic inhibitor Kelch-like ECH-associated protein 1 (Keap1) defends tissues against oxidative stress, thereby protecting against pathologies that relate to DNA, protein, and/or lipid oxidative damage. Thus, it was hypothesized that Nrf2 should also have important roles in maintaining thyroid homeostasis. METHODS: Ubiquitous and thyroid-specific male C57BL6J Nrf2 knockout (Nrf2-KO) mice were studied. Plasma and thyroids were harvested for evaluation of thyroid function tests by radioimmunoassays and of gene and protein expression by real-time polymerase chain reaction and immunoblotting, respectively. Nrf2-KO and Keap1-KO clones of the PCCL3 rat thyroid follicular cell line were generated using CRISPR/Cas9 technology and were used for gene and protein expression studies. Software-predicted Nrf2 binding sites on the thyroglobulin enhancer were validated by site-directed in vitro mutagenesis and chromatin immunoprecipitation. RESULTS: The study shows that Nrf2 mediates antioxidant transcriptional responses in thyroid cells and protects the thyroid from oxidation induced by iodide overload. Surprisingly, it was also found that Nrf2 has a dramatic impact on both the basal abundance and the thyrotropin-inducible intrathyroidal abundance of thyroglobulin (Tg), the precursor protein of thyroid hormones. This effect is mediated by cell-autonomous regulation of Tg gene expression by Nrf2 via its direct binding to two evolutionarily conserved antioxidant response elements in an upstream enhancer. Yet, despite upregulating Tg levels, Nrf2 limits Tg iodination both under basal conditions and in response to excess iodide. CONCLUSIONS: Nrf2 exerts pleiotropic roles in the thyroid gland to couple cell stress defense mechanisms to iodide metabolism and the thyroid hormone synthesis machinery, both under basal conditions and in response to excess iodide.

Effects of 2-iodohexadecanal in the physiology of thyroid cells.

Iodide has direct effects on thyroid function. Several iodinated lipids are biosynthesized by the thyroid and they were postulated as intermediaries in the action of iodide. Among them, 2-iodohexadecanal (2-IHDA) has been identified and proposed to play a role in thyroid autoregulation. The aim of this study was to compare the effect of iodide and 2-IHDA on thyroid cell physiology. For this purpose, FRTL-5 thyroid cells were incubated with the two compounds during 24 or 48 h and several thyroid parameters were evaluated such as: iodide uptake, intracellular calcium and H2O2 levels. To further explore the molecular mechanism involved in 2-IHDA action, transcript and protein levels of genes involved in thyroid hormone biosynthesis, as well as the transcriptional expression of these genes were evaluated in the presence of iodide and 2-IHDA. The results obtained indicate that 2-IHDA reproduces the action of excess iodide on the "Wolff-Chaikoff" effect as well as on thyroid specific genes transcription supporting its role in thyroid autoregulation.

2-Iodohexadecanal inhibits thyroid cell growth in part through the induction of let-7f microRNA.

UNLABELLED: It is well known that pituitary TSH exerts the major task in the regulation of thyroid function. However, this gland is capable of certain degree of autonomy, independently of TSH control. Iodine plays an important role in thyroid physiology and biochemistry. The thyroid is capable of producing different iodolipids such as 2-iodohexadecanal (2-IHDA). It was shown that this iodolipid mimic some of the inhibitory effects of excess iodide on several thyroid parameters. OBJECTIVES: To identify the miRNAs regulated by 2-IHDA in rat thyroid cells and likely characterize their role in thyroid cell proliferation and function. RESULTS: FRTL-5 cells were grown in the presence of TSH and treated with 2-IHDA. Among the miRNAs up-regulated by 2-IHDA we focused on miR-let-7f and miR-138. When we transfected the miRNAs, miR-let-7f but not miR-138 overexpression inhibited proliferation of FRTL 5 cells, while miR-let-7f inhibition restored cell growth in 2-IHDA treated cultures. Analysis of cell cycle by flow cytometric DNA analysis revealed that miR-let-7f inhibition reduced the percentage of 2-IHDA treated cells in G1 phase and an increased of the percentage of cells in S phase was observed upon anti-let-7f transfection. The expresion of Cyclin D1 and Cyclin D3 were reduced after the transfection of miR-let-7f and miR-138, respectively. In in vivo studies we observed that miR-let-7f and miR-138 were up regulated by 2-IHDA during goiter involution. CONCLUSION: These results suggest that the inhibitory effects of 2-IHDA on FRTL-5 thyroid cell proliferation are mediated in part through the induction of let-7f microRNA.

Inhibitory effects of 2-iodohexadecanal on FRTL-5 thyroid cells proliferation.

UNLABELLED: Although thyroid gland function is mainly under the control of pituitary TSH, other factors, such as iodine, play a role in this process. The thyroid is capable of producing different iodolipids such as 6-iodo-deltalactone and 2-iodohexadecanal (2-IHDA). It was shown that these iodolipids mimic some of the inhibitory effects of excess iodide on several thyroid parameters. OBJECTIVES: To study the effect of 2-IHDA on cell proliferation and apoptosis in FRTL-5 cells. RESULTS: FRTL-5 cells were grown in the presence of TSH and treated with increasing concentrations of KI and 2-IHDA (0.5, 5, 10 and 33 µM) for 24, 48 and 72 h. Whereas KI inhibited cell proliferation only at 33 µM after 72 h of treatment, 2-IHDA inhibited in a time and concentration dependent manner. Analysis of cell cycle by flow cytometric DNA analysis revealed an accumulation of cells in G1 phase induced by 2-IHDA. The expression of cyclin A, cyclin D1 and cyclin D3 were reduced after treatment with 2-IHDA whereas CDK4 and CDK6 proteins were not modified. 2-IHDA induced a dynamic change in cytoplasmic to nuclear accumulation of p21 and p27 causing these proteins to be accumulated mostly in the nucleus. We also observed evidence of a pro-apoptotic effect of 2-IHDA at highest concentrations. No significant effect of KI was observed. CONCLUSION: These results suggest that the inhibitory effects of 2-IHDA on FRTL-5 thyroid cell proliferation are mediated by cell cycle arrest in G1/S phase and cell death by apoptosis.

Assessment of biological effectiveness of boron neutron capture therapy in primary and metastatic melanoma cell lines.

PURPOSE: In order to optimize the effectiveness of Boron Neutron Capture Therapy (BNCT), Relative Biological Effectiveness (RBE) and Compound Biological Effectiveness (CBE) were determined in two human melanoma cell lines, M8 and Mel-J cells, using the amino acid p-boronophenylalanine (BPA) as boron carrier. MATERIALS AND METHODS: The effects of BNCT on the primary amelanotic cell line M8 and on the metastatic pigmented melanoma cell line Mel-J were studied using colony formation assay. The RBE values were determined using both a gamma ray source, and the neutron beam from the Nuclear Reactor of the National Atomic Energy Commission (RA-3). For the determination of the RBE, cells were irradiated with increasing doses of both sources, between 1 and 8 Gy; and for the determination of CBE factors, the cells were pre-incubated with BPA before irradiation. Afterwards, the cell surviving fraction (SF) was determined for each treatment. RESULTS: Marked differences were observed between both cell lines. Mel-J cells were more radioresistant than the M8 cell line. The clonogenic assays showed that for a SF of 1%, the RBE values were 1.3 for M8 cells and 1.5 for Mel-J cells. Similarly, the CBE values for a 1% SF were 2.1 for M8 and 3 for Mel-J cell lines. For the endpoint of 0.1% of SF the RBE values obtained were 1.2 for M8 and 1.4 for Mel-J cells. Finally, CBE values calculated for a 0.1% were 2 and 2.6 for M8 and Mel-J cell lines respectively. In order to estimate the uptake of the non-radioactive isotope Boron 10 ((10)B), a neutron induced autoradiographic technique was performed showing discrepancies in (10)B uptake between both cell lines. CONCLUSIONS: These obtained in vitro results are the first effectiveness factors determined for human melanoma at the RA-3 nuclear reactor and show that BNCT dosimetry planning for patients could be successfully performed using these new factors.

Protective effect of an antithyroid compound against γ-radiation-induced damage in human colon cancer cells.

We have previously reported the radioprotective effect of propylthiouracil (PTU) on thyroid cells. The aim of the present study was to analyze whether tumor cells and normal cells demonstrate the same response to PTU. Human colon carcinoma cells were irradiated with γ-irradiation with or without PTU. We evaluated the clonogenic survival, intracellular reactive oxygen species levels, catalase, superoxide dismutase and glutathione peroxidase activities, and apoptosis by nuclear cell morphology and caspase-3 activity assays. Cyclic AMP (cAMP) levels were measured by radioimmunoassay. PTU treatment increased surviving cell fraction at 2 Gy (SF2) from 56.9 ± 3.6 in controls to 75.0 ± 3.5 (p < 0.05) and diminished radiation-induced apoptosis. In addition, we observed that the level of antioxidant enzymes' activity was increased in cells treated with PTU. Moreover, pretreatment with PTU increased intracellular levels of cAMP. Forskolin (p < 0.01) and dibutyryl cAMP (p < 0.05) mimicked the effect of PTU on SF2. Co-treatment with H89, an inhibitor of protein kinase A, abolished the radioprotective effect of PTU. PTU reduces the toxicity of ionizing radiation by increasing cAMP levels and also possibly through a reduction in apoptosis levels and in radiation-induced oxidative stress damage. We therefore conclude that PTU protects both normal and cancer cells during exposure to radiation in conditions mimicking the radiotherapy.

6 Iodo-δ-lactone: a derivative of arachidonic acid with antitumor effects in HT-29 colon cancer cells.

BACKGROUND: IL-δ (5-hydroxy-6 iodo-8,11,14-eicosatrienoic delta lactone) an iodinated arachidonic acid (AA) derivative, is one of the iodolipids biosynthesized by the thyroid. Although IL-δ regulates several thyroid parameters such as cell proliferation and goiter growth it was found that this iodolipid inhibits the growth of other non thyroid cell lines. OBJECTIVES: To study the effect of IL-δ on cell proliferation and apoptosis in the colon cancer cell line HT-29. RESULTS: Treatment with IL-δ reduced cell viability in a concentration-dependent manner: 1µM 20%, 5µM 25%, 10µM 31%, 50µM 47% and caused a significant decrease of PCNA expression (25%). IL-δ had pro-apoptotic effects, evidenced by morphological features of programmed cell death such as pyknosis, karyorrhexis, cell shrinkage and cell blebbing observed by fluorescence microscopy, and an increase in caspase-3 activity and in Bax/Bcl-2 ratio (2.5 after 3h of treatment). Furthermore, IL-δ increased ROS production (30%) and lipid peroxidation levels (19%), suggesting that apoptosis could be a result of increased oxidative stress. A maximum increase in c-fos and c-jun protein expression in response to IL-δ was observed 1h after initiation of the treatment. IL-δ also induced a tumour growth delay of 70% compared to the control group in NIH nude mice implanted with HT-29 cells. CONCLUSION: Our study shows that IL-δ inhibits cell growth and induces apoptosis in the colon cancer cell line, HT-29 and opens the possibility that IL-δ could be a potential useful chemotherapy agent.

Protection against radiation-induced damage of 6-propyl-2-thiouracil (PTU) in thyroid cells.

Many epidemiologic studies have shown that the exposure to high external radiation doses increases thyroid neoplastic frequency, especially when given during childhood or adolescence. The use of radioprotective drugs may decrease the damage caused by radiation therapy and therefore could be useful to prevent the development of thyroid tumors. The aim of this study was to investigate the possible application of 6-propyl-2-thiouracil (PTU) as a radioprotector in the thyroid gland. Rat thyroid epithelial cells (FRTL-5) were exposed to different doses of γ irradiation with or without the addition of PTU, methimazole (MMI), reduced glutathione (GSH) and perchlorate (KClO4). Radiation response was analyzed by clonogenic survival assay. Cyclic AMP (cAMP) levels were measured by radioimmunoassay (RIA). Apoptosis was quantified by nuclear cell morphology and caspase 3 activity assays. Intracellular reactive oxygen species (ROS) levels were measured using the fluorescent dye 2',7'-dichlorofluorescein-diacetate. Catalase, superoxide dismutase and glutathione peroxidase activities were also determined. Pretreatment with PTU, MMI and GSH prior to irradiation significantly increased the surviving cell fraction (SF) at 2 Gy (P < 0.05), while no effect was observed with KClO4. An increase in extracellular levels of cAMP was found only in PTU treated cells in a dose and time-dependent manner. Cells incubated with agents that stimulate cAMP (forskolin and dibutyril cAMP) mimicked the effect of PTU on SF. Moreover, pretreatment with the inhibitor of protein kinase A, H-89, abolished the radioprotective effect of PTU. PTU treatment diminished radiation-induced apoptosis and protected cells against radiation-induced ROS elevation and suppression of the antioxidant enzyme's activity. PTU was found to radioprotect normal thyroid cells through cAMP elevation and reduction in both apoptosis and radiation-induced oxidative stress damage.

Estimación de la Incidencia de Cáncer de Tiroides en Capital Federal y el Gran Buenos Aires (período 2003-2011) / Incidences Rates of Thyroid Cancer in Buenos Aires (2003-2011)

La incidencia del cáncer de tiroides ha aumentado significativamente en las últimas décadas en el resto del mundo. En Argentina no existe un registro nacional de cáncer por lo que la incidencia del mismo no puede establecerse. Por lo tanto, nuestro objetivo fue estimar la incidencia de cáncer de tiroides en la población de la Ciudad Autánoma de Buenos aires y Gran Buenos Aires así como la relación por género y la histología en el período de 2003 hasta 2011. Asumiendo que la población de afiliados a la Obra Social de la Policía Federal Argentina es representativa de los habitantes de Buenos Aires y el conurbano calculamos que la incidencia es de 6,51 casos/100.000 habitantes/año, con un incremento en 25 años mayor al doble, con predominio del carcinoma papilar frente al folicular.

Thyroid cancer incidence has significantly risen worldwide in the last decades. In Argentina, there is no national cancer registry; therefore its incidence can not be established. The objective of this study was to estimate the incidence of thyroid cancer in the population of Buenos Aires City and suburbs, and the relationship between gender and histology over the period 2003-2011. Assuming that the population affiliated to the Social Security of the Argentine Federal Police is representative of the inhabitants of Buenos Aires City and suburbs, we estimate an incidence of 6.51 cases/100,000 population/year, with an increasing incidence of almost double from 1981-1986 to 2003-2011. An increase in papillary thyroid cancer was mainly responsible for this rising trend. Incidence rates were higher for females (11.76/100,000 women) compared to those for males (2.65/100,000 men). Among men and women of all ages, the highest rate of incidence was for tumor size < 1 cm.

Estimación de la Incidencia de Cáncer de Tiroides en Capital Federal y el Gran Buenos Aires (período 2003-2011) / Incidences Rates of Thyroid Cancer in Buenos Aires (2003-2011)

La incidencia del cáncer de tiroides ha aumentado significativamente en las últimas décadas en el resto del mundo. En Argentina no existe un registro nacional de cáncer por lo que la incidencia del mismo no puede establecerse. Por lo tanto, nuestro objetivo fue estimar la incidencia de cáncer de tiroides en la población de la Ciudad Autánoma de Buenos aires y Gran Buenos Aires así como la relación por género y la histología en el período de 2003 hasta 2011. Asumiendo que la población de afiliados a la Obra Social de la Policía Federal Argentina es representativa de los habitantes de Buenos Aires y el conurbano calculamos que la incidencia es de 6,51 casos/100.000 habitantes/año, con un incremento en 25 años mayor al doble, con predominio del carcinoma papilar frente al folicular.(AU)

Thyroid cancer incidence has significantly risen worldwide in the last decades. In Argentina, there is no national cancer registry; therefore its incidence can not be established. The objective of this study was to estimate the incidence of thyroid cancer in the population of Buenos Aires City and suburbs, and the relationship between gender and histology over the period 2003-2011. Assuming that the population affiliated to the Social Security of the Argentine Federal Police is representative of the inhabitants of Buenos Aires City and suburbs, we estimate an incidence of 6.51 cases/100,000 population/year, with an increasing incidence of almost double from 1981-1986 to 2003-2011. An increase in papillary thyroid cancer was mainly responsible for this rising trend. Incidence rates were higher for females (11.76/100,000 women) compared to those for males (2.65/100,000 men). Among men and women of all ages, the highest rate of incidence was for tumor size < 1 cm.(AU)

Biochemical changes during goiter induction by methylmercaptoimidazol and inhibition by delta-iodolactone in rat.

BACKGROUND: We have demonstrated that the administration of delta-iodolactone (i.e., 5-iodo-delta lactone) of arachidonic acid (IL-delta), a mediator in thyroid autoregulation, prevents goiter induction by methylmercaptoimidazol (MMI) in rats. Other studies have shown that transforming growth factor beta-1 (TGF-beta1) mimics some of the actions of excess iodide, but its participation in autoregulation is disputed. The present studies were performed to test the hypotheses that IL-delta decreases thyroid growth by inhibition of cell proliferation and/or by stimulation of apoptosis due to oxidative stress, that TGF-beta is stimulated by an excess of iodide and by IL-delta, and that c-Myc and c-Fos expression are upregulated during goiter induction and downregulated during goiter inhibition. METHODS: Rats were treated with MMI alone or together with iodide or IL-delta. Thyroid weight, cell number, cell proliferation, apoptosis, and oxidative stress were determined. Proliferating cell nuclear antigen (PCNA), TGF-beta1, TGF-beta3, c-Myc, and c-Fos were measured by Western blot. RESULTS: MMI caused a progressive increase in thyroid weight accompanied by an increase in cell number, asymmetry of the ploidy histograms, and PCNA, c-Fos, and c-Myc expression. In addition, an early increase of apoptosis was observed. Peroxides as well as glutathione peroxidase and catalase activities were also increased in goitrous animals. The inhibitory action of IL-delta on goiter formation was accompanied by the inhibition of cell proliferation evidenced by a significant decrease in cell number, PCNA expression, and asymmetry of the ploidy histograms. A transient stimulation of apoptosis after 7 days of treatment was also observed. MMI administration stimulated TGF-beta1 but not TGF-beta3 synthesis. IL-delta alone caused a slight increase of TGF-beta3 but not TGF-beta1, whereas potassium iodide (KI) stimulated both isoforms and MMI reversed KI effect on TGF-beta1 expression but not on TGF-beta3. CONCLUSIONS: The goiter inhibitory action of IL-delta is due to the inhibition of cell proliferation and the transient stimulation of apoptosis. This latter action does not involve oxidative stress. TGF-beta1 does not play a role in the autoregulatory pathway mediated by IL-delta. Iodide stimulates TGF-beta3 without the need of being organified. These results suggest that there may be more than one pathway involved in the autoregulatory mechanism.

6 Iodo-delta-lactone reproduces many but not all the effects of iodide.

BACKGROUND: Iodide has direct effects on thyroid function. Several iodinated lipids are biosynthesized by the thyroid and they were postulated as intermediaries in the action of iodide. Among them 6 iodo-delta-lactone (IL-delta) has been identified and proposed to play a role in thyroid autoregulation. The aim of this study was to compare the effect of iodide and IL-delta on several thyroid parameters. METHODS: Thyroid bovine follicles were incubated with the different compounds during three days. RESULTS: KI and IL-delta inhibited iodide uptake, total protein and Tg synthesis but only KI had an effect on NIS and Tg mRNAs levels. Both compounds inhibited Na+/K+ ATPase and deoxy-glucose uptake. As PAX 8, FOXE 1 and TITF1 are involved in the regulation of thyroid specific genes their mRNA levels were measured. While iodide inhibited the expression of the first two, the expression of TITF1 was stimulated by iodide and IL-delta had no effect on these parameters. CONCLUSION: These findings indicate that IL-delta reproduces some but not all the effects of excess iodide. These observations apply for higher micromolar concentrations of iodide while no such effects could be demonstrated at nanomolar iodide concentrations.

Role of transforming growth factor beta in the regulation of thyroid function and growth.

Transforming growth factor beta (TGF-beta) exists in nature as three isoforms. They exert their effects by binding to a type II receptor located at the cell membrane. The TGF-beta-type II receptor complex then recruits type I receptor, and this new complex stimulates the phosphorylation of Smads 2 and 3, which are subsequently transferred to the nucleus, where they regulate gene transcription. The thyroid gland expresses the TGF-beta1 gene mRNA and synthesizes the protein, which under physiologic conditions regulates thyroid growth and function. Different studies have demonstrated that TGF-beta1 inhibits cell proliferation and a number of functional parameters. These include cyclic adenosine monophosphate (AMP) formation, iodine uptake and organification, hormone secretion, and the expression of thyroglobulin, thyroid peroxidase, and Na(+)/I(-) symporter. The expression of the TGF-beta1 gene and protein may be stimulated by iodine under normal conditions. Since TGF-beta1 mimics some of the inhibitory actions of iodine, its participation in thyroid autoregulation has been proposed; however, this concept is still debated. In thyroid tumors, the inhibitory action of TGF-beta1 on cell proliferation is progressively lost as the tumor becomes more undifferentiated. The alterations in the signaling pathway of TGF-beta1 are not the same in tumors from different species. Even within the same species, such as the pig thyroid, the results may be different depending on whether monolayers or follicular suspensions are employed. The data suggest that it is not entirely possible to apply the results obtained in animal studies to normal or pathological human thyroid tissue. More studies are required to provide the information needed to develop treatments, based on targeting the signaling pathway of TGF-beta1, for undifferentiated thyroid cancer and other thyroid diseases.

Biodistribution of boron compounds in an animal model of human undifferentiated thyroid cancer for boron neutron capture therapy.

Undifferentiated thyroid carcinoma (UTC) is a rapidly growing, highly invasive malignant tumor that currently lacks any effective treatment. Boron neutron capture therapy (BNCT) has been investigated recently for some types of tumors including glioblastoma multiforme and malignant melanoma. In previous studies we have shown the selective uptake of p-boronophenylalanine (BPA) by undifferentiated thyroid cancer cells in vitro and in vivo, as well as the histologic cure of 50% of the nude mice transplanted with human UTC cells when treated with BPA and an appropriate neutron beam. The present studies were performed to further optimize this treatment through the investigation of a boronated porphyrin, both alone and in combination with BPA. In vitro studies with cells in culture showed that BOPP (tetrakis-carborane carboxylate ester of 2,4-bis-(alpha,beta-dihydroxyethyl)-deutero-porphyrin IX) is localized intracellularly, with a highest concentration in the 11500g (mitochondrial-enriched pellet) fraction. When BOPP was administered alone to NIH nude mice transplanted with UTC human cells, no significant tumor uptake or selectivity in our in vivo model was observed. In contrast, when BOPP was injected 5-7 days before BPA and the animals were sacrificed 60 min after administration of BPA, a significant increase in boron uptake by the tumor was found (38-45 ppm with both compounds vs 20 ppm with BPA alone). On day 5 the tissue boron selectivity ratios were tumor/blood approximately 3.8 and tumor/distal skin approximately 1.8. Other important ratios were tumor/thyroid approximately 6.6 and tumor/lung approximately 2.9. These results open the possibility of improving the efficacy of BNCT for the treatment of this so far "orphan" tumor.

Role of peroxidase inhibition by insulin in the bovine thyroid cell proliferation mechanism.

Monolayer primary cultures of thyroid cells produce, in the presence of insulin, a cytosolic inhibitor of thyroid peroxidase (TPO), lacto peroxidase (LPO), horseradish peroxidase (HRPO) and glutathione peroxidase (GPX). The inhibitor, localized in the cytosol, is thermostable and hydrophylic. Its molecular mass is less than 2 kDa. The inhibitory activity, resistant to proteolytic and nucleolytic enzymes, disappears with sodium metaperiodate treatment, as an oxidant of carbohydrates, supporting its oligosaccharide structure. The presence of inositol, mannose, glucose, the specific inhibition of cyclic AMP-dependent protein kinase and the disappearance of peroxidase inhibition by alkaline phosphatase and alpha-mannosidase in purified samples confirms its chemical structure as inositol phosphoglycan-like. Purification by anionic interchange shows that the peroxidase inhibitor elutes like the two subtypes of inositol phosphoglycans (IPG)P and A, characterized as signal transducers of insulin action. Insulin significantly increases the concentration of the peroxidase inhibitor in a thyroid cell culture at 48 h. The addition of both isolated substances to a primary thyroid culture produces, after 30 min, a significant increase in hydrogen peroxide (H2O2) concentration in the medium, concomitantly with the disappearance of the GPX activity in the same conditions. The presence of insulin or anyone of both products, during 48 h, induces cell proliferation of the thyroid cell culture. In conclusion, insulin stimulates thyroid cell division through the effect of a peroxidase inhibitor, as its second messenger. The inhibition of GPX by its action positively modulates the H2O2 level, which would produce, as was demonstrated by other authors, the signal for cell proliferation.

Typing of the locus DYS19 from DNA derived from fingernail clippings using PCR Concert rapid purification system.

DNA extracted from the fingernails of female victims of a violent or aggressive act may assist in the identification of the male. Sometimes with the current autosomal STR loci, however, the victim's profile may mask the perpetrator's DNA profile or the perpetrator's DNA may be substantially lower in quantity than that of the victim's DNA. Thus, under these conditions, no characterization is possible. In this paper, an alternative DNA extraction procedure was employed, and the application of an STR locus residing on the Y chromosome DYS19 was typed to allow for genetic characterization of the perpetrator in such cases.

Further studies on iodide uptake autoregulation in calf thyroid slices

La autorregulación tiroídea es mediada por un yodocompuesto orgánico de natureleza desconocida. Trabajos previos de diferentes laboratorios sugieren diversos yodocompuestos como posibles intermediarios, tales como hormonas tiroideas, vodolípidos y una yodoproteína. Recientes estudios propusieron que una yodoproteína estaría iunvolucrada en la autorregulación de la captación de yodo en tiroides de gato. El presente trabajo fue diseñado con el objeto de explorar esta hipótesis en tiroides bovina. La preincubación de cortes de tiroides bovina en presencia de KI 0.01mM produce una inhibición del 36-40% en la captación de I-125 (relación tiroides/medio, T/M). Este efecto no fue modificado por puromicina ni cicloheximida bajo condiciones en las cuales la síntesis proteica fue casi totalmente abolida. Por otra parte, estos dos compuestos no son capaces de alterar significativamente el perfil de incorporación de I-125 a los diferentes yodocompuestos tiroideos analizados por cromatografía en papel. Cuando se exploró la reversibilidad del efecto inhibitorio del yodo sobre la captación de I-125, se observó que después de 60 min de interrumpido el tratamiento con yoduro existió una recuperación parcial, en tanto que a los 90 min se retornó a los valores de captación basales. Estos resultados indican que una yodoproteína no sería el mediador de la autorregulación de yodo, en vaca. Por otra parte el yodocompuesto involucrado tendría una vida media relativamente corta

Further studies on iodide uptake autoregulation in calf thyroid slices

La autorregulación tiroídea es mediada por un yodocompuesto orgánico de natureleza desconocida. Trabajos previos de diferentes laboratorios sugieren diversos yodocompuestos como posibles intermediarios, tales como hormonas tiroideas, vodolípidos y una yodoproteína. Recientes estudios propusieron que una yodoproteína estaría iunvolucrada en la autorregulación de la captación de yodo en tiroides de gato. El presente trabajo fue diseñado con el objeto de explorar esta hipótesis en tiroides bovina. La preincubación de cortes de tiroides bovina en presencia de KI 0.01mM produce una inhibición del 36-40% en la captación de I-125 (relación tiroides/medio, T/M). Este efecto no fue modificado por puromicina ni cicloheximida bajo condiciones en las cuales la síntesis proteica fue casi totalmente abolida. Por otra parte, estos dos compuestos no son capaces de alterar significativamente el perfil de incorporación de I-125 a los diferentes yodocompuestos tiroideos analizados por cromatografía en papel. Cuando se exploró la reversibilidad del efecto inhibitorio del yodo sobre la captación de I-125, se observó que después de 60 min de interrumpido el tratamiento con yoduro existió una recuperación parcial, en tanto que a los 90 min se retornó a los valores de captación basales. Estos resultados indican que una yodoproteína no sería el mediador de la autorregulación de yodo, en vaca. Por otra parte el yodocompuesto involucrado tendría una vida media relativamente corta (AU)