Presence Of T3 Research Articles

Nuclear receptor corepressor 1 levels differentially impact the intracellular dynamics of mutant thyroid hormone receptors associated with resistance to thyroid hormone syndrome

Thyroid hormone receptor α1 (TRα1) undergoes nucleocytoplasmic shuttling and mediates gene expression in response to thyroid hormone (T3). In Resistance to Thyroid Hormone Syndrome α (RTHα), certain TRα1 mutants have higher affinity for nuclear corepressor 1 (NCoR1) and may form stable complexes that are not released in the presence of T3. Here, we examined whether NCoR1 modulates intranuclear mobility and nuclear retention of TRα1 or RTHα-associated mutants in transfected human cells, as a way of analyzing critical structural components of TRα1 and to further explore the correlation between mutations in TRα1 and aberrant intracellular trafficking. We found no significant difference in intranuclear mobility, as measured by fluorescence recovery after photobleaching, between TRα1 and select RTHα mutants, irrespective of NCoR1 expression. Nuclear-to-cytoplasmic fluorescence ratios of RTHα mutants, however, varied from TRα1 when NCoR1 was overexpressed, with a significant increase in nuclear retention for A263V and a significant decrease for A263S and R384H. In NCoR1-knockout cells, nuclear retention of A263S, A263V, P389R, A382P, C392X, and F397fs406X was significantly decreased compared to control (wild-type) cells. Luciferase reporter gene transcription mediated by TRα1 was significantly repressed by both NCoR1 overexpression and NCoR1 knockout. Most RTHα mutants showed minimal induction regardless of NCoR1 levels, but T3-mediated transcriptional activity was decreased for R384C and F397fs406X when NCoR1 was overexpressed, and also decreased for N359Y in NCoR1-knockout cells. Our results suggest a complex interaction between NCoR1 and RTHα mutants characterized by aberrant intracellular localization patterns and transcriptional activity that potentially arise from variable repressor complex stability, and may provide insight into RTHα pathogenesis on a molecular and cellular level.

Effects of Melatonin and 3,5,3'-Triiodothyronine on the Development of Rat Granulosa Cells.

Melatonin, as an endocrine neurotransmitter, can promote the development of the ovary. Meanwhile, it also has protective effect on the ovary as an antioxidant. Thyroid hormone (TH) is essential for normal human reproductive function. Many studies have shown that 3,5,3'-triiodothyronine (T3) regulates the development of ovarian granulosa cells. However, little is known about the specific mechanisms by which melatonin combines with T3 to regulate granulosa cell development. The aim of present study was to investigate the effects and the possible mechanisms of melatonin and T3 on ovarian granulosa cell development. In the present study, cell development and apoptosis were detected by CCK8, EdU and TUNEL, respectively. The levels of related proteins were analyzed by Western blotting. The results showed that oxidative stress (OS) and reactive oxygen species (ROS) were induced by H2O2 in granulosa cells, and cell apoptosis was also increased accompanied with the decreased cellular proliferation and viability. Melatonin protects granulosa cells from H2O2-induced apoptosis and OS by downregulating ROS levels, especially in the presence of T3. Co-treatment of cell with melatonin and T3 also promotes the expression of GRP78 and AMH, while inhibiting CHOP, Caspase-3, and P16. It was demonstrated that melatonin alone or in combination with T3 had positive effect on the development of granulosa cells. In addition, the AMPK/SIRT1 signaling pathway is involved in the process of melatonin/T3 promoting granulosa cell development.

Impact of the thyroid hormone T3 and its nuclear receptor TRα1 on colon cancer stem cell phenotypes and response to chemotherapies

Colorectal cancers (CRCs) are highly heterogeneous and show a hierarchical organization, with cancer stem cells (CSCs) responsible for tumor development, maintenance, and drug resistance. Our previous studies showed the importance of thyroid hormone-dependent signaling on intestinal tumor development and progression through action on stem cells. These results have a translational value, given that the thyroid hormone nuclear receptor TRα1 is upregulated in human CRCs, including in the molecular subtypes associated with CSC features. We used an established spheroid model generated from the human colon adenocarcinoma cell line Caco2 to study the effects of T3 and TRα1 on spheroid formation, growth, and response to conventional chemotherapies. Our results show that T3 treatment and/or increased TRα1 expression in spheroids impaired the response to FOLFIRI and conferred a survival advantage. This was achieved by stimulating drug detoxification pathways and increasing ALDH1A1-expressing cells, including CSCs, within spheroids. These results suggest that clinical evaluation of the thyroid axis and assessing TRα1 levels in CRCs could help to select optimal therapeutic regimens for patients with CRC.Proposed mechanism of action of T3/TRα1 in colon cancer spheroids. In the control condition, TRα1 participates in maintaining homeostatic cell conditions. The presence of T3 in the culture medium activates TRα1 action on target genes, including the drug efflux pumps ABCG2 and ABCB1. In the case of chemotherapy FOLFIRI, the increased expression of ABC transcripts and proteins induced by T3 treatment is responsible for the augmented efflux of 5-FU and Irinotecan from the cancer cells. Taken together, these mechanisms contribute to the decreased efficacy of the chemotherapy and allow cells to escape the treatment. Created with BioRender.com.

Resposta Vascular da Triiodotironina sobre Anéis de Aortas Isoladas: Contribuição de Mecanismos Redox

Abstract Background Vascular dysfunction constitutes the etiology of many diseases, such as myocardial infarction and hypertension, with the disruption of redox homeostasis playing a role in the imbalance of the vasomotor control mechanism. Our group previously has shown that thyroid hormones exert protective effects on the aortic tissue of infarcted rats by improving angiogenesis signaling. Objective Investigate the role of triiodothyronine (T3) on vascular response, exploring its effects on isolated aortas and whether there is an involvement of vascular redox mechanisms. Methods Isolated aortic rings (intact- and denuded-endothelium) precontracted with phenylephrine were incubated with T3 (10-8, 10-7, 10-6, 10-5, and 10-4 M), and tension was recorded using a force-displacement transducer coupled with an acquisition system. To assess the involvement of oxidative stress, aortic rings were preincubated with T3 and subsequently submitted to an in vitro reactive oxygen species (ROS) generation system. The level of significance adopted in the statistical analysis was 5%. Results T3 (10-4 M) promoted vasorelaxation of phenylephrine precontracted aortic rings in both intact- and denuded-endothelium conditions. Aortic rings preincubated in the presence of T3 (10-4 M) also showed decreased vasoconstriction elicited by phenylephrine (1 µM) in intact-endothelium preparations. Moreover, T3 (10-4 M) vasorelaxation effect persisted in aortic rings preincubated with NG-nitro-L-arginine methylester (L-NAME, 10 µM), a nonspecific NO synthase (NOS) inhibitor. Finally, T3 (10-4 M) exhibited, in vitro, an antioxidant role by reducing NADPH oxidase activity and increasing SOD activity in the aorta’s homogenates. Conclusion T3 exerts dependent- and independent-endothelium vasodilation effects, which may be related to its role in maintaining redox homeostasis.

Vascular Response of Triiodothyronine on Isolated Aortic Rings: Contribution of Redox Mechanisms.

Vascular dysfunction constitutes the etiology of many diseases, such as myocardial infarction and hypertension, with the disruption of redox homeostasis playing a role in the imbalance of the vasomotor control mechanism. Our group previously has shown that thyroid hormones exert protective effects on the aortic tissue of infarcted rats by improving angiogenesis signaling. Investigate the role of triiodothyronine (T3) on vascular response, exploring its effects on isolated aortas and whether there is an involvement of vascular redox mechanisms. Isolated aortic rings (intact- and denuded-endothelium) precontracted with phenylephrine were incubated with T3 (10-8, 10-7, 10-6, 10-5, and 10-4 M), and tension was recorded using a force-displacement transducer coupled with an acquisition system. To assess the involvement of oxidative stress, aortic rings were preincubated with T3 and subsequently submitted to an in vitro reactive oxygen species (ROS) generation system. The level of significance adopted in the statistical analysis was 5%. T3 (10-4 M) promoted vasorelaxation of phenylephrine precontracted aortic rings in both intact- and denuded-endothelium conditions. Aortic rings preincubated in the presence of T3 (10-4 M) also showed decreased vasoconstriction elicited by phenylephrine (1 µM) in intact-endothelium preparations. Moreover, T3 (10-4 M) vasorelaxation effect persisted in aortic rings preincubated with NG-nitro-L-arginine methylester (L-NAME, 10 µM), a nonspecific NO synthase (NOS) inhibitor. Finally, T3 (10-4 M) exhibited, in vitro, an antioxidant role by reducing NADPH oxidase activity and increasing SOD activity in the aorta's homogenates. T3 exerts dependent- and independent-endothelium vasodilation effects, which may be related to its role in maintaining redox homeostasis.

The thyroid hormone enhances mouse embryonic fibroblasts reprogramming to pluripotent stem cells: role of the nuclear receptor corepressor 1.

Pluripotent stem cells can be generated from somatic cells by the Yamanaka factors Oct4, Sox2, Klf4 and c-Myc. Mouse embryonic fibroblasts (MEFs) were transduced with the Yamanaka factors and generation of induced pluripotent stem cells (iPSCs) was assessed by formation of alkaline phosphatase positive colonies, pluripotency gene expression and embryod bodies formation. The thyroid hormone triiodothyronine (T3) enhances MEFs reprogramming. T3-induced iPSCs resemble embryonic stem cells in terms of the expression profile and DNA methylation pattern of pluripotency genes, and of their potential for embryod body formation and differentiation into the three major germ layers. T3 induces reprogramming even though it increases expression of the cyclin kinase inhibitors p21 and p27, which are known to oppose acquisition of pluripotency. The actions of T3 on reprogramming are mainly mediated by the thyroid hormone receptor beta and T3 can enhance iPSC generation in the absence of c-Myc. The hormone cannot replace Oct4 on reprogramming, but in the presence of T3 is possible to obtain iPSCs, although with low efficiency, without exogenous Klf4. Furthermore, depletion of the corepressor NCoR (or Nuclear Receptor Corepressor 1) reduces MEFs reprogramming in the absence of the hormone and strongly decreases iPSC generation by T3 and also by 9cis-retinoic acid, a well-known inducer of reprogramming. NCoR depletion also markedly antagonizes induction of pluripotency gene expression by both ligands. Inclusion of T3 on reprogramming strategies has a potential use in enhancing the generation of functional iPSCs for studies of cell plasticity, disease and regenerative medicine.

Thyroid hormone upregulates LAMP2 expression and lysosome activity

Thyroid hormone (T3)-induced autophagy and its biological significance have been extensively investigated in recent years. However, limited studies to date have focused on the important role of lysosomes in autophagy. In this study, we explored the effects of T3 on lysosomal protein expression and trafficking in detail. Our findings showed that T3 activates rapid lysosomal turnover and expression of numerous lysosomal genes, including TFEB, LAMP2, ARSB, GBA, PSAP, ATP6V0B, ATP6V0D1, ATP6V1E1, CTSB, CTSH, CTSL, and CTSS, in a thyroid hormone receptor-dependent manner. In a murine model, LAMP2 protein was specifically induced in mice with hyperthyroidism. T3-promoted microtubule assembly was significantly disrupted by vinblastine, resulting in accumulation of the lipid droplet marker PLIN2. In the presence of the lysosomal autophagy inhibitors bafilomycin A1, chloroquine and ammonium chloride, we observed substantial accumulation of LAMP2 but not LAMP1 protein. T3 further enhanced the protein levels of ectopically expressed LAMP1 and LAMP2. Upon knockdown of LAMP2, cavities of lysosomes and lipid droplets accumulated in the presence of T3, although the changes in LAMP1 and PLIN2 expression were less pronounced. More specifically, the protective effect of T3 against ER stress-induced death was abolished by knockdown of LAMP2. Our collective results indicate that T3 not only promotes lysosomal gene expression but also LAMP protein stability and microtubule assembly, leading to enhancement of lysosomal activity in digesting any additional autophagosomal burden.

Steroid-receptor coactivator complexes in thyroid hormone-regulation of Xenopus metamorphosis.

Anuran metamorphosis is perhaps the most drastic developmental change regulated by thyroid hormone (T3) in vertebrate. It mimics the postembryonic development in mammals when many organs/tissues mature into adult forms and plasma T3 level peaks. T3 functions by regulating target gene transcription through T3 receptors (TRs), which can recruit corepressor or coactivator complexes to target genes in the absence or presence of T3, respectively. By using molecular and genetic approaches, we and others have investigated the role of corepressor or coactivator complexes in TR function during the development of two highly related anuran species, the pseudo-tetraploid Xenopus laevis and diploid Xenopus tropicalis. Here we will review some of these studies that demonstrate a critical role of coactivator complexes, particularly those containing steroid receptor coactivator (SRC) 3, in regulating metamorphic rate and ensuring the completion of metamorphosis.

P9.50: Novel MLTR Classification of Portal Vein Thrombosis For Purposes of Liver Transplantation

Portal vein thrombosis (PVT) is a common complication of decompensated liver cirrhosis and occurs in 0.6-26% of patients on waiting lists. In a population of patients with liver cirrhosis, PVT develops within a year in 4.6-12.8%, in 10.2-20% within 5 years, and within 8-10 years in 38.7%. The Liver Transplant Center of the Sklifosovsky Institute in Moscow is the leading transplant center in Russia, which performs liver transplants for portal vein thrombosis - such patients occur on the waiting list of the center up to 30%. The practice of the center led to the need to create its own classification, which would allow making decisions on the waiting list and intraoperatively. For the purposes of liver transplantation, especially if transplantation is planned from a living related donor, it is possible to propose a classification of portal vein thrombosis that would be applicable to address tactical issues in liver transplantation. A variant of such a classification, which may be called MLTR, is given in Table 1. Table 1. - MLTR classification of portal vein thrombosis Letter Designation M Thrombosis of the mesenteric veins 0 - no thrombosis 1 - non-occlusive thrombosis of one of the veins 2 - occlusive thrombosis of one of the veins L Thrombosis of the splenic veins 0 - no thrombosis 1 - non-occlusive thrombosis 2 - occlusive thrombosis T Thrombosis of the portal vein 0 - no thrombosis 1 - non-occlusive thrombosis 2 - occlusive thrombosis R Thrombosis of the branches of the portal vein 0 - no thrombosis 1 - non-occlusive thrombosis 2 - occlusive thrombosis In the presence of calcification, the letters ca are added, in the presence of cavernous transformation ct. Visually, the classification is presented in fig. 1.The developed classification was introduced into the work of the liver transplantation center of the N.V. Sklifosovsky. The clinical material consisted of 650 patients of the liver transplantation center of the N.V. Sklifosovsky operated from 2000 to 2020. Patients with PVT M0L0T1R0 accounted for 25% of the total number of patients, with M1L0T2R0 - half, with M1L1T2R0-1 - 25%. Based on the developed classification, the tactics of surgical treatment of patients was determined as follows. The developed classification was used to determine intraoperative tactics. In the presence of M1-2L0-1, the retropancreatic portal vein and its confluence were mobilized, for which the head of the pancreas was mobilized. This technique made it possible in some cases to perform eversion thrombinthymectomy without performing plastic or bypass interventions. In the presence of T3, thrombinthymectomy was not performed; it was planned to perform shunting or prosthetics of the portal vein. The presence of R1-2 prior to transplantation was of little hemodynamic significance. Thus, we can conclude that the developed classification was used by us to make a decision during liver transplantation. The authors would like to thank Maria Kozhevnikova for her help in illustrating our work.

Stem Cells from Healthy and Tendinopathic Human Tendons: Morphology, Collagen and Cytokines Expression and Their Response to T3 Thyroid Hormone.

The aim of this study was to investigate the effect of triiodothyronine (T3) on tendon specific markers and cytokines expression of stem cells extracted from human tendons. Indeed, thyroid hormones have been reported to be protective factors, maintaining tendons’ homeostasis, whereas tendinopathy is believed to be related to a failed healing response. Healthy and tendinopathic human tendons were harvested to isolate tendon stem/progenitor cells (TSPCs). TSPCs obtained from pathological samples showed gene expression and morphological modifications at baseline in comparison with cells harvested from healthy tissues. When cells were maintained in a medium supplemented with T3 (10−6 M), only pathological populations showed a significant upregulation of tenogenic markers (DCN, TNC, COL1A1, COL3A1). Immunostaining revealed that healthy cells constantly released type I collagen, typical of tendon matrix, whereas pathological ones overexpressed and secreted type III collagen, typical of scarred and impaired tissue. Pathological cells also overexpressed pro- and anti-inflammatory cytokines, suggesting an impaired balance in the presence of T3, without STAT3 activation. Moreover, DKK-1 was significantly high in the culture medium of pathological cell cultures and was reversed by T3. This study opens perspectives on the complex biochemical alteration of cells from pathological tendons, which may lead to the chronic disease context with an impaired extracellular matrix.

Triiodotironina não influencia na diferenciação condrogênica de células tronco do tecido adipose de ratas jovens

ABSTRACT: The objective of this study was to investigate the in vitro action of triiodothyronine (T3) on the chondrogenic differentiation of adipose tissue-derived stem cells (ASCs) of female rats, with different time periods and doses. ASCs were extracted from female Wistar rats and were cultured in chondrogenic medium with and without the presence of T3. Five groups were established: 1) ASCs without T3; and 2,3,4,5) ASCs with 0.01, 1, 100 and 1,000 nM T3, respectively). After 7, 14 and 21 days, cell morphology, chondrogenic matrix formation, and expression of Sox9, aggrecan, collagen II, and collagen X were evaluated. The Student-Newman-Keuls test was used. ASCs showed CD54, CD73, and CD90 before chondrogenic differentiation. The hormone treatment did not alter chondrogenic matrix formation, Sox9 expression at 14 or 21 days, or expression of collagen II or collagen X at any time. However, the 0.01, 1, and 1000 nM T3 doses decreased Sox9 expression at 7 days. In conclusion, chondrogenic differentiation of ASCs of female rats is not influenced by T3.

T3-Dependent Transcriptional Programming by TRβ in Thyroid Cells Requires SWI/SNF Chromatin Remodelers

Abstract Transcriptional regulation in response to thyroid hormone (T3) is a dynamic and cell-type specific process that maintains cellular homeostasis and identity. A bimodal switch model, where T3 binding alters the co-regulator profile of a constitutively DNA-bound thyroid hormone receptor (TR) to affect downstream gene expression, is widely used to describe the interaction between TRβ and chromatin. To test this model on a genome-wide scale, we used an integrated genomics approach to profile and characterize the cistrome of TRβ by CUT&RUN, map changes in chromatin accessibility by ATAC-seq, and capture the transcriptomic changes in response to T3 by RNA-seq in the normal thyroid cell line, Nthy-ORI. Our CUT&RUN data demonstrated that T3 binding causes significant shifts in TRβ genomic occupancy; these shifts are associated with differential chromatin accessibility. Most of the T3-induced differentially expressed genes have a TRβ binding site associated with the proximal protomer region within one kilobase of the transcriptional start site, suggesting that these are direct TRβ regulatory target genes. Remarkably, the majority of TRβ binding sites were found in transgenic regions and distal regulatory elements. In order to identify the co-regulatory proteins that are required for execution of a T3-dependent transcriptional program in our thyroid cells, we used a TRβ-miniTurboID fusion construct to perform a proximity ligation assay followed by mass spectrometry. We identified 1,138 nuclear proteins that interact with TRβ. Of these proteins, 75 interact preferentially in the presence of T3 and 68 in the absence of T3. All of the core SWI/SNF complex subunits from both of the major subtypes (BAF and PBAF) were identified as TRβ. Interestingly, we found that the PBAF-specific subunit, PBRM1, was significantly enriched in the presence of T3. BAF complex-specific subunits, such as ARID1A, were also present in our dataset. To test whether TRβ could differentially recruit BAF and PBAF complexes to its binding sites, we performed CUT&RUN targeting BRG1, PBRM, and ARID1A to determine the degree of co-occupancy with TRβ. Based on our comprehensive genomic and proteomic analyses, we propose a new model for selective recruitment of BAF and PBAF SWI/SNF complexes to TRβ binding sites for differential functions in regulating chromatin accessibility.

Role of AMPK in the protective effects exerted by triiodothyronine in ischemic-reperfused myocardium.

Recent studies have provided evidence that triiodothyronine (T3) might play an effective role in the recovery of ischemic myocardium, through the preservation of mitochondrial function and the improvement of energy substrate metabolism. To this respect, it has been suggested that T3 could activate AMP-activated protein kinase (AMPK), the cellular 'fuel-gauge' enzyme, although its role has yet to be elucidated. The aim of the present study was to investigate the effects produced by acute treatment with T3 (60 nM) and the pharmacological inhibition of AMPK by compound C on isolated rat left atria subjected to 75 min simulated ischemia-75 min reperfusion. Results showed that T3 increased AMPK activation during simulated ischemia-reperfusion, while compound C prevented it. At the end of simulated reperfusion, acute T3 treatment increased contractile function recovery and cellular viability conservation. Mitochondrial ultrastructure was better preserved in the presence of T3 as well as mitochondrial ATP production rate and tissue ATP content. Calcium retention capacity, a parameter widely used as an indicator of the resistance of mitochondrial permeability transition pore (MPTP) to opening, and GSK-3β phosphorylation, a master switch enzyme that limits MPTP opening, were increased by T3 administration. All these beneficial effects exerted by T3 acute treatment were prevented when compound C was co-administrated. The present study provided original evidence that T3 enhances intrinsic activation of AMPK during myocardial ischemia-reperfusion, being this enzyme involved, at least in part, in the protective effects exerted by T3, contributing to mitochondrial structure and function preservation, post-ischemic contractile recovery and conservation of cellular viability.

Multiple mechanisms regulate H3 acetylation of enhancers in response to thyroid hormone

Hormone-dependent activation of enhancers includes histone hyperacetylation and mediator recruitment. Histone hyperacetylation is mostly explained by a bimodal switch model, where histone deacetylases (HDACs) disassociate from chromatin, and histone acetyl transferases (HATs) are recruited. This model builds on decades of research on steroid receptor regulation of transcription. Yet, the general concept of the bimodal switch model has not been rigorously tested genome wide. We have used a genomics approach to study enhancer hyperacetylation by the thyroid hormone receptor (TR), described to operate as a bimodal switch. H3 acetylation, HAT and HDAC ChIP-seq analyses of livers from hypo- and hyperthyroid wildtype, TR deficient and NCOR1 disrupted mice reveal three types of thyroid hormone (T3)-regulated enhancers. One subset of enhancers is bound by HDAC3-NCOR in the absence of hormone and constitutively occupy TR and HATs irrespective of T3 levels, suggesting a poised enhancer state in absence of hormone. In presence of T3, HDAC3-NCOR1 dissociates from these enhancers leading to histone hyperacetylation, suggesting a histone acetylation rheostat function of HDACs at poised enhancers. Another subset of enhancers, not occupied by HDACs, is hyperacetylated in a T3-dependent manner, where TR is recruited to chromatin together with HATs. Lastly, a subset of enhancers, is not occupied directly by TR yet require TR for histone hyperacetylation. This indirect enhancer activation involves co-association with TR bound enhancers within super-enhancers or topological associated domains. Collectively, this demonstrates various mechanisms controlling hormone-dependent transcription and adds significant details to the otherwise simple bimodal switch model.

Multiple mechanisms regulate H3 acetylation of enhancers in response to thyroid hormone

Hormone-dependent activation of enhancers includes histone hyperacetylation and mediator recruitment. Histone hyperacetylation is mostly explained by a bimodal switch model, where histone deacetylases (HDACs) disassociate from chromatin, and histone acetyl transferases (HATs) are recruited. This model builds on decades of research on steroid receptor regulation of transcription. Yet, the general concept of the bimodal switch model has not been rigorously tested genome wide. We have used a genomics approach to study enhancer hyperacetylation by the thyroid hormone receptor (TR), described to operate as a bimodal switch. H3 acetylation, HAT and HDAC ChIP-seq analyses of livers from hypo- and hyperthyroid wildtype, TR deficient and NCOR1 disrupted mice reveal three types of thyroid hormone (T3)-regulated enhancers. One subset of enhancers is bound by HDAC3-NCOR1 in the absence of hormone and constitutively occupy TR and HATs irrespective of T3 levels, suggesting a poised enhancer state in absence of hormone. In presence of T3, HDAC3-NCOR1 dissociates from these enhancers leading to histone hyperacetylation, suggesting a histone acetylation rheostat function of HDACs at poised enhancers. Another subset of enhancers, not occupied by HDACs, is hyperacetylated in a T3-dependent manner, where TR is recruited to chromatin together with HATs. Lastly, a subset of enhancers, is not occupied directly by TR yet requires TR for histone hyperacetylation. This indirect enhancer activation involves co-association with TR bound enhancers within super-enhancers or topological associated domains. Collectively, this demonstrates various mechanisms controlling hormone-dependent transcription and adds significant details to the otherwise simple bimodal switch model.

Disruptive effects of two organotin pesticides on the thyroid signaling pathway in Xenopus laevis during metamorphosis

Organotin compounds are the ubiquitous environmental pollutants due to their wide industrial and agricultural applications and unexpected releasing into the environment, which show characteristic of endocrine disruptors to interfere with the synthesis, receptor binding or action of endogenous-hormones. Organotin pesticides (OTPs) are used in agriculture and may impact endocrine functions on organisms. Thyroid hormones (THs) play fundamental roles in regulating the basal metabolism and energy balance, while thyroid function can be impaired by environmental contaminants. Therefore, it is crucial to clarify the effects and mechanisms of OTPs on hypothalamus-pituitary-thyroid (HPT) axis. In this study, Xenopus laevis tadpoles at stage 51 were exposed to fentin hydroxide and fenbutatin oxide (0.04, 0.20 and 1.00 μg·L−1) for 21 days. It was found that both compounds caused inhibitory effects on metamorphic development of tadpoles (e.g., significant decrease in hindlimb length and retarding development). Triiodothyronine (T3) significantly decreased in tadpoles exposed to 0.20 μg/L and 1.00 μg/L of the two OTPs for 14 days or 21 days. The expressions of TH responsive genes trβ, bteb and dio2 were down-regulated, while tshβ and slc5a5 were up-regulated. Surface plasmon resonance (SPR) binding assays showed that fentin hydroxide had a moderate affinity to recombinant human thyroid hormone receptor β but fenbutatin oxide did not have. Result of the SPR assay was highly consistent with the luciferase reporter gene assays that fentin hydroxide suppressed the relative luciferase activity in the presence of T3 while fenbutatin oxide did not, demonstrating fentin hydroxide but not fenbutatin oxide displayed an antagonistic activity against T3-TR complex mediated transcriptional activation. Overall, the findings elucidated the mechanisms induced by OTPs along HPT axis. These results highlighted the adverse influences of organotin pesticides on thyroid hormone- dependent development in vertebrates and the need for more comprehensive investigations of their potential ecological risks.

Coordinated transcriptional regulation by thyroid hormone and glucocorticoid interaction in adult mouse hippocampus-derived neuronal cells.

The hippocampus is a well-known target of thyroid hormone (TH; e.g., 3,5,3’-triiodothyronine—T3) and glucocorticoid (GC; e.g., corticosterone—CORT) action. Despite evidence that TH and GC play critical roles in neural development and function, few studies have identified genes and patterns of gene regulation influenced by the interaction of these hormones at a genome-wide scale. In this study we investigated gene regulation by T3, CORT, and T3 + CORT in the mouse hippocampus-derived cell line HT-22. We treated cells with T3, CORT, or T3 + CORT for 4 hr before cell harvest and RNA isolation for microarray analysis. We identified 9 genes regulated by T3, 432 genes by CORT, and 412 genes by T3 + CORT. Among the 432 CORT-regulated genes, there were 203 genes that exhibited an altered CORT response in the presence of T3, suggesting that T3 plays a significant role in modulating CORT-regulated genes. We also found 80 genes synergistically induced, and 73 genes synergistically repressed by T3 + CORT treatment. We performed in silico analysis using publicly available mouse neuronal chromatin immunoprecipitation-sequencing datasets and identified a considerable number of synergistically regulated genes with TH receptor and GC receptor peaks mapping within 1 kb of chromatin marks indicative of hormone-responsive enhancer regions. Functional annotation clustering of synergistically regulated genes reveal the relevance of proteasomal-dependent degradation, neuroprotective effect of growth hormones, and neuroinflammatory responses as key pathways to how TH and GC may coordinately influence learning and memory. Taken together, our transcriptome data represents a promising exploratory dataset for further study of common molecular mechanisms behind synergistic TH and GC gene regulation, and identify specific genes and their role in processes mediated by cross-talk between the thyroid and stress axes in a mammalian hippocampal model system.

Thyroid Hormone Protects Primary Cortical Neurons Exposed to Hypoxia by Reducing DNA Methylation and Apoptosis.

Traumatic brain injury (TBI) is associated with disruption of cerebral blood flow leading to localized brain hypoxia. Thyroid hormone (TH) treatment, administered shortly after injury, has been shown to promote neural protection in rodent TBI models. The mechanism of TH protection, however, is not established. We used mouse primary cortical neurons to investigate the effectiveness and possible pathways of T3-promoted cell survival after exposure to hypoxic injury. Cultured primary cortical neurons were exposed to hypoxia (0.2% oxygen) for 7 hours with or without T3 (5 nM). T3 treatment enhanced DNA 5-hydroxymethylcytosine levels and attenuated the hypoxia-induced increase in DNA 5-methylcytosine (5-mc). In the presence of T3, mRNA expression of Tet family genes was increased and DNA methyltransferase (Dnmt) 3a and Dnmt3b were downregulated, compared with conditions in the absence of T3. These T3-induced changes decreased hypoxia-induced DNA de novo methylation, which reduced hypoxia-induced neuronal damage and apoptosis. We used RNA sequencing to characterize T3-regulated genes in cortical neurons under hypoxic conditions and identified 22 genes that were upregulated and 15 genes that were downregulated. Krüppel-like factor 9 (KLF9), a multifunctional transcription factor that plays a key role in central nervous system development, was highly upregulated by T3 treatment in hypoxic conditions. Knockdown of the KLF9 gene resulted in early apoptosis and abolished the beneficial role of T3 in neuronal survival. KLF9 mediates, in part, the neuronal protective role of T3. T3 treatment reduces hypoxic damage, although pathways that reduce DNA methylation and apoptosis remain to be elucidated.

OR10-6 Thyroid Hormone Protects Primary Cortical Neurons Exposed to Hypoxia by Reducing DNA Methylation and Apoptosis

Traumatic Brain Injury (TBI) is associated with disruption of cerebral blood flow leading to localized brain hypoxia. Thyroid hormone (TH) treatment, when administered shortly after injury, has been shown to promote neural survival in rodent TBI models. The mechanism of TH protection, however, is not established. We used mouse primary cortical neurons, to investigate the effectiveness and mechanism of T3 promoted cell survival after exposure to hypoxic injury. Cultured primary cortical neurons were exposed to hypoxia (0.2% oxygen) for 7 hours, in either the presence or absence of T3 (5 nM). T3 treatment enhanced DNA 5-hydroxymethylcytosin (5-hmc) levels compared to the conditions without T3, and attenuated the hypoxia-induced increase in DNA 5-methylcytosin (5-mc). In the presence of T3, mRNA expression of Tet family genes was increased and DNA methyltransferase, (Dnmt) 3a and Dnmt3b, were downregulated, compared to conditions in the absence of T3. These T3-induced changes may reduce hypoxia-induced DNA de novo methylation. We utilized RNA-seq to characterize T3-regulated genes in cortical neurons under hypoxic conditions, and identified 23 genes that were upregulated and 15 genes that were downregulated. KLF9, krupple-like factor 9, a multifunctional transcription factor that plays a key role in CNS development, was highly upregulated by T3 treatment in hypoxic conditions. We show that knockdown of the KLF9 gene induced early apoptosis and abolished the beneficial role of T3 in neuronal survival. KLF9 mediates, in part, the neuronal protective role of T3. T3 treatment reduces hypoxic damage and acts through pathways that reduce DNA methylation and apoptosis.

Synthesis, crystal structure and physicochemical properties of organic-inorganic compounds KCS-3 and KCS-4

Abstract Organic-inorganic hybrid aluminosilicate compounds KCS-3 and KCS-4 were successfully synthesized by low-temperature hydrothermal reaction of mother gels composed of organically-bridged silane, fumed alumina, alkali metal hydroxide and water. Two bridged organosilanes i.e. 4,4′-bis(triethoxysilyl)-1,1′-biphenyl (BTEbP) and 1,4-bis(triethoxysilyl)benzene (BTEB) were used as a Si source for the synthesis of KCS-3 and KCS-4, respectively. The presence of T3 (≡O3Si–R, R: organic group) local structure, AlO4 tetrahedral unit and the organic groups originating from the employed organosilane were confirmed by 29Si, 27Al and 13C MAS NMR measurements. Their crystal structures were solved by the direct space method using the parallel tempering algorithm from powder X-ray diffraction data. The crystal structure of KCS-3 belongs to tetragonal symmetry (lattice constants: a = 10.22 A and c = 71.37 A, space group: I41/amd). Four aluminosilicate layers composed of 4-ring and 8-ring were stacked along c-axis, and adjacent layers were connected alternately by a biphenyl pillar (–C6H4–C6H4–) and a Q2 Si pillar (–OSi(OH)2O–). KCS-3 showed amphiphilic adsorption properties, reflecting the presence of hydrophilic- and lipophilic-inner spaces. On the other hand, the crystal structure of KCS-4 belongs to monoclinic symmetry (lattice constants: a = 10.05 A, b = 9.349 A, c = 5.052 A and β = 97.01°, space group: P21/c). A large amount of Li ions forming four-coordination environment is included in a layer-like framework, and adjacent layers were bridged by a phenylene group, that is, KCS-4 can be regarded as a hybrid-type lithosilicate.