Sulfation of thyroid hormone in chicken

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Thyroid hormone sulfation in chicken and axolotl

Next to deiodination, glucuronidation and sulfation of the phenolic hydroxyl group are important pathways of iodothyronine metabolism. Although in general conjugation serves to facilitate the urinary and biliary excretion of lipophilic substances, sulfation of thyroid hormones has another important function. Sulfation protects the tissues against an excess of active thyroid hormone and it can also be used as a reservoir from which active hormone is released by sulfatases when and where it is required. Interaction between sulfation and deiodination has already been described in several mammalian tissues. After an initial characterization of the sulfotransferases we determined the changes in sulfation during chicken embryonic development and induced metamorphosis of the axolotl, a neotenous amphibian. Sulfation assays demonstrate the presence of sulfotransferases in liver, kidney and brain cytosol in one-day-old chicken. Similar to the situation in mammals, the enzyme(s) show a substrate preference for 3,3'-diiodothyronine (3,3'-T-2) over 3,5,3'-triiodothyronine (T-3) and 3,3',5'-triiodothyronine (rT(3)) over thyroxine (T-4). Measurements of sulfation using 3,3'-T2 as substrate in tissues from 16-day-old embryonic chickens (E16) to one-day-old posthatch chicks (C1)demonstrate that sulfation activity varies independently in liver, kidney and brain, suggesting that the activity is regulated in a tissue specific manner. We also detected sulfation activity in liver, kidney and bl ain of the axolotl. The highest activity was found in the liver, hence we used this tissue to study the profile of activity during induced metamorphosis. We found that the activity in the liver remains stable during the full length of metamorphosis.status: publishe

Involvement of thyroid hormones in chicken embryonic brain development

Thyroid hormones (THs) play an important role in vertebrate brain development by stimulating and coordinating cell proliferation, migration and differentiation. Several TH-responsive genes involved in these processes have been identified, but the information is mainly derived from studies of late brain development, while relatively little is known about the more early stages, prior to the onset of embryonic TH secretion. We have chosen the chick embryo to investigate the role of THs in both late and early brain development. T4 and T3 are found in chicken brain from the earliest stages tested (day 4). Indirect clues for the involvement of T3 in brain development are found in the ontogenetic expression profiles of proteins regulating its bioavailability and action, including TH transporters, deiodinases and TH-receptors. All of them are expressed in whole embryos tested on day 2 of incubation and in developing brain tested from day 4 onwards. Their distribution patterns vary over time and according to the brain area and cell type studied. Hypothyroidism induced during the second half of incubation disturbs cell migration in the cerebellum, providing more direct evidence for the requirement for THs during the later stages of brain development. Direct morphological proof for the requirement for THs during the first half of incubation is still missing, but microarray analysis of telencephalon shows a clearly divergent gene expression profile in hypothyroid embryos. In vivo knockdown of TH transporters and deiodinases in chick embryos cultured ex ovo provides an excellent tool to study the role of THs in early brain development in more detail.status: publishe

cDNA cloning, functional expression, and characterization of chicken sulfotransferases belonging to the SULT1B and SULT1C families

A search of the chicken expressed sequence tag (EST) database identified 2 cDNA clones that appeared to represent members of the SULT1B and SULT1C enzyme families. These cDNAs were fully sequenced and found to contain full-length inserts. Phylogenetic analysis of the derived amino acid sequences clearly placed them as the first members of the chicken SULT1B and SULT1C families, respectively, to be identified, and we propose they be named SULT1B1 and SULT1C1. (CHICK)SULT1B1 shares approximately 60% amino acid sequence identity with mammalian SULT1B enzymes, whereas the closest neighbor to (CHICK)SULT1C1 was the ortholog (RAT)SULT1C1, with 68% identity. We cloned these cDNAs into the bacterial expression vectors from the pET series. Transformed Escherichia coli cells strongly expressed the recombinant proteins. Purification of the recombinant enzymes from E. coli was accomplished by a three-step procedure involving ammonium sulfate precipitation, anion exchange chromatography, and affinity chromatography. The purified enzymes displayed subunit molecular weights of approximately 35,000Da on SDS-PAGE, as predicted, and were both able to sulfate a wide range of compounds, including xenobiotics and endogenous substrates such as iodothyronines. Detailed kinetic analysis showed SULT1C1 was more prolific in that it was able to sulfate dopamine, tyramine, and apomorphine, which SULT1B1 was not. 2-Bromophenol was the best substrate for both enzymes. We also raised antibodies against these proteins, which were able to detect the SULTs by ELISA, and which were able to strongly inhibit the recombinant enzymes. This is the first detailed characterization of sulfotransferases from the chicken, and it demonstrates that the avian and mammalian SULT1 enzymes are closely related in both structure and function.status: publishe

Thyroid hormone deiodination in birds

Because the avian thyroid gland secretes almost exclusively thyroxine (T4), the availability of receptor-active 3,3',5-triiodothyronine (T3) has to be regulated in the extrathyroidal tissues, essentially by deiodination. Like mammals and most other vertebrates, birds possess three types of iodothyronine deiodinases (D1, D2, and D3) that closely resemble their mammalian counterparts, as shown by biochemical characterization studies in several avian species and by cDNA cloning of the three enzymes in chicken. The tissue distribution of these deiodinases has been studied in detail in chicken at the level of activity and mRNA expression. More recently specific antibodies were used to study cellular localization at the protein level. The abundance and distribution of the different deiodinases shows substantial variation during embryonic development and postnatal life. Deiodination in birds is subject to regulation by hormones from several endocrine axes, including thyroid hormones, growth hormone and glucocorticoids. In addition, deiodination is also influenced by external parameters, such as nutrition, temperature, light and also a number of environmental pollutants. The balance between the outer and inner ring deiodination resulting from the impact of all these factors ultimately controls T3 availability.status: publishe

Placental growth factor 2-A potential therapeutic strategy for chronic myocardial ischemia

OBJECTIVES: We investigated whether sustained infusion of recombinant human placental growth factor-2 (rhPlGF-2) improves myocardial perfusion and left ventricular (LV) function in a porcine model of ischemic cardiomyopathy (ICM). METHODS: We induced myocardial ischemia using a flow-limiting stent in the LAD. Four weeks later, we randomized pigs with confirmed myocardial dysfunction to blinded rhPlGF-2 administration (PlGF2, 15 μg/kg/day, 14 days) or PBS (CON). At 8 weeks, we measured hemodynamics, contractile function and regional perfusion at rest and during stress using MRI and microspheres. We evaluated neovascularization post mortem. RESULTS: RhPlGF-2 administration increased PlGF serum levels more than 63-fold (83 3 ± 361 versus 11 ± 5 pg/ml CON, P<0.05) without adverse effects. After 4weeks, rhPlGF-2 significantly enhanced perfusion in the ischemic region at rest (0.83 ± 0.32 versus 0.58 ± 0.21 ml/min/g CON, P<0.05) and during hyperemia (1.50 ± 0.50 versus 1.02 ± 0.46 ml/min/g CON, P<0.05). Consequently, regional contractile function in rhPlGF-2-treated pigs improved at rest (37 ± 15% versus 23 ± 9% CON, P<0.05) and during high dose dobutamine stress (53 ± 31% versus 27 ± 16% CON, P<0.05). Enhanced perfusion translated into a greater improvement in LV ejection fraction and in preload-recruitable stroke work in rhPlGF-2-treated animals than in CON (52 ± 11 versus 41 ± 9%, and 76 ± 24 versus 41 ± 21 mmHg, respectively, P<0.05 for both), which was associated with significantly greater vascular density in the ischemic region. CONCLUSIONS: In chronic ICM, systemic rhPlGF-2 administration significantly enhances regional myocardial perfusion, contractile function at rest and during stress, and induces a prominent recovery of global cardiac function. PlGF-2 protein infusion is safe and may represent a promising therapy in chronic ICM.publisher: Elsevier articletitle: Placental growth factor 2 — A potential therapeutic strategy for chronic myocardial ischemia journaltitle: International Journal of Cardiology articlelink: http://dx.doi.org/10.1016/j.ijcard.2015.10.177 content_type: article copyright: Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.status: publishe

Enhanced Antitumor Efficacy of PhAc-ALGP-Dox, an Enzyme-Activated Doxorubicin Prodrug, in a Panel of THOP1-Expressing Patient-Derived Xenografts of Soft Tissue Sarcoma

Despite poor response rates and dose-limiting cardiotoxicity, doxorubicin (doxo) remains the standard-of-care for patients with advanced soft tissue sarcoma. We evaluated the efficacy of two tetrapeptidic doxo prodrugs (PhAc-ALGP-Dox or CBR-049 and CBR-050) that are locally activated by enzymes expressed in the tumor environment, in ten sarcoma patient-derived xenografts. Xenograft models were selected based on expression of the main activating enzyme, i.e., thimet oligopeptidase (THOP1). Mice were either randomized to vehicle, doxo, CBR-049 and CBR-050 or control, doxo, aldoxorubicin (aldoxo) and CBR-049. Treatment efficacy was assessed by tumor volume measurement and histological assessment of ex-mouse tumors. CBR-049 showed significant tumor growth delay compared to control in all xenografts investigated and was superior compared to doxo in all but one. At the same time, CBR-049 showed comparable efficacy to aldoxo but the latter was found to have a complex safety profile in mice. CBR-050 demonstrated tumor growth delay compared to control in one xenograft but was not superior to doxo. For both experimental prodrugs, strong immunostaining for THOP1 was found to predict better antitumor efficacy. The prodrugs were well tolerated without any adverse events, even though molar doses were 17-fold higher than those administered and tolerated for doxo

Placental growth factor increases regional myocardial blood flow and function in a new porcine model of chronic myocardial ischemia

Liu X., Caluwe E., Reyns G., Verhamme P., Pokreisz P., Vandenwijngaert S., Dubois C., Zalewski J., Ghysels S., Maes F., Gillijns H., Pellens M., Vanden Driessche N., Patel A., Van de Werf F., Verbeken E., Bogaert J., Janssens S., ''Placental growth factor increases regional myocardial blood flow and function in a new porcine model of chronic myocardial ischemia'', Circulation, vol. 120, no. 18, suppl., pp. S837, November 2009 (American Heart Association scientific sessions 2009, November 14-18, 2009, Orlando, Florida, USA).status: publishe