Search

Thyroid hormones (TH) are crucial for brain development; their deficiency during neurodevelopment impairs neural cell differentiation and causes irreversible neurological alterations. Understanding TH action, and in particular the mechanisms regulating TH availability in the prenatal human brain is essential to design therapeutic strategies for neurological diseases due to impaired TH signaling during neurodevelopment. We aimed at the identification of cells involved in the regulation of TH availability in the human brain at fetal stages. To this end, we studied the distribution of the TH transporters monocarboxylate transporter 8 (MCT8) and organic anion-transporting polypeptide 1C1 (OATP1C1), as well as the TH-metabolizing enzymes types 2 and 3 deiodinases (DIO2 and DIO3). Paraffin-embedded human brain sections obtained from necropsies of thirteen fetuses from 14 to 38 gestational weeks were analyzed by immunohistochemistry and in situ hybridization. We found these proteins localized along radial glial cells, in brain barriers, in Cajal-Retzius cells, in migrating fibers of the brainstem and in some neurons and glial cells with particular and complex spatiotemporal patterns. Our findings point to an important role of radial glia in controlling TH delivery and metabolism and suggest two additional novel pathways for TH availability in the prenatal human brain: the outer, and the inner cerebrospinal fluid–brain barriers. Based on our data we propose a model of TH availability for neural cells in the human prenatal brain in which several cell types have the ability to autonomously control the required TH content., The IdiPAZ Biobank is supported by Instituto de Salud Carlos III, Spanish Health Ministry (Retic RD09/0076/00073) and Farmaindustria, through the Cooperation Program in Clinical and Translational Research of the Community of Madrid. We thank Drs. Soledad Bárez-López, José Miguel Cosgaya, Estrella Rausell and Ana Montero-Pedrazuela for the careful reading of the manuscript and their helpful suggestions, and Javier Pérez for his help on the artwork. This work was supported by Grants from the Spanish Plan Nacional de I+D+i (Grant numbers SAF2014-54919-R and SAF2017-86342-R to A.GF and J.B), and the Center for Biomedical Research on Rare Diseases (Ciberer), Instituto de Salud Carlos III, Madrid, Spain and the Sherman Foundation (OTR02211 to A.GF). D.LE is a recipient of a fellowship from “Fellowship Training Program for Advanced Human Capital, BECAS CHILE” from the National Commission for Scientific and Technological Research (CONICYT), Gobierno de Chile. The cost of this publication has been paid in part by FEDER funds.

The type 2 iodothyronine-deiodinase (D2) enzyme converts T(4) to T(3), and mice deficient in this enzyme [D2 knockout (D2KO) mice] have decreased T(3) derived from T(4) in skeletal muscle despite normal circulating T(3) levels. Because slow skeletal muscle is particularly susceptible to changes in T(3) levels, we expected D2 inactivation to result in more pronounced slow-muscle characteristics in the soleus muscle, mirroring hypothyroidism. However, ex vivo studies of D2KO soleus revealed higher rates of twitch contraction and relaxation and reduced resistance to fatigue. Immunostaining of D2KO soleus showed that these properties were associated with changes in muscle fiber type composition, including a marked increase in the number of fast, glycolytic type IIB fibers. D2KO soleus muscle fibers had a larger cross-sectional area, and this correlated with increased myonuclear accretion in myotubes formed from D2KO skeletal muscle precursor cells differentiated in vitro. Consistent with our functional findings, D2KO soleus gene expression was markedly different from that in hypothyroid wild-type (WT) mice. Comparison of gene expression between euthyroid WT and D2KO mice indicated that PGC-1α, a T(3)-dependent regulator of slow muscle fiber type, was decreased by ∼50% in D2KO soleus. Disruption of Dio2 in the C2C12 myoblast cell line led to a significant decrease in PGC-1α expression and a faster muscle phenotype upon differentiation. These results indicate that D2 loss leads to significant changes in soleus contractile function and fiber type composition that are inconsistent with local hypothyroidism and suggest that reduced levels of PCG-1α may contribute to the observed phenotypical changes.

Proliferation of pancreatic acinar cells is a critical process in the pathophysiology of pancreatic diseases, because limited or defective proliferation is associated with organ dysfunction and patient morbidity. In this context, elucidating the signalling pathways that trigger and sustain acinar proliferation is pivotal to develop therapeutic interventions promoting the regenerative process of the organ. In this study we used genetic and pharmacological approaches to manipulate both local and systemic levels of thyroid hormones to elucidate their role in acinar proliferation following caerulein-mediated acute pancreatitis in mice. In addition, molecular mechanisms mediating the effects of thyroid hormones were identified by genetic and pharmacological inactivation of selected signalling pathways.In this study we demonstrated that levels of the thyroid hormone 3,3',5-triiodo-l-thyronine (T3) transiently increased in the pancreas during acute pancreatitis. Moreover, by using genetic and pharmacological approaches to manipulate both local and systemic levels of thyroid hormones, we showed that T3 was required to promote proliferation of pancreatic acinar cells, without affecting the extent of tissue damage or inflammatory infiltration.Finally, upon genetic and pharmacological inactivation of selected signalling pathways, we demonstrated that T3 exerted its mitogenic effect on acinar cells via a tightly controlled action on different molecular effectors, including histone deacetylase, AKT, and TGFβ signalling.In conclusion, our data suggest that local availability of T3 in the pancreas is required to promote acinar cell proliferation and provide the rationale to exploit thyroid hormone signalling to enhance pancreatic regeneration. Copyright © 2019 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Background: One of the most widely used risk stratification systems for estimating individual patients' risk of persistent or recurrent differentiated thyroid cancer (DTC) is the American Thyroid Association (ATA) guidelines. The 2015 ATA version, which has increased the number of patients considered at low or intermediate risk, has been validated in several retrospective, single-center studies. The aims of this study were to evaluate the real-world performance of the 2015 ATA risk stratification system in predicting the response to treatment 12 months after the initial treatment and to determine the extent to which this performance is affected by the treatment center in which it is used. Methods: A prospective cohort of DTC patients collected by the Italian Thyroid Cancer Observatory web-based database was analyzed. We reviewed all records present in the database and selected consecutive cases that satisfied inclusion criteria: (i) histological diagnosis of DTC, with the exclusion of noninvasive follicular thyroid neoplasm with papillary-like nuclear features; (ii) complete data of the initial treatment and pathological features; and (iii) results of 1-year follow-up visit (6-18 months after the initial treatment), including all data needed to classify the estimated response to treatment. Results: The final cohort was composed of 2071 patients from 40 centers. The ATA risk of persistent/recurrent disease was classified as low in 1109 patients (53.6%), intermediate in 796 (38.4%), and high in 166 (8.0%). Structural incomplete responses were documented in only 86 (4.2%) patients: 1.5% in the low-risk, 5.7% in the intermediate-risk, and 14.5% in the high-risk group. The baseline ATA risk class proved to be a significant predictor of structural persistent disease, both for intermediate-risk (odds ratio [OR] 4.67; 95% confidence interval [CI] 2.59-8.43) and high-risk groups (OR 16.48; CI 7.87-34.5). Individual center did not significantly influence the prediction of the 1-year disease status. Conclusions: The ATA risk stratification system is a reliable predictor of short-term outcomes in patients with DTC in real-world clinical settings characterized by center heterogeneity in terms of size, location, level of care, local management strategies, and resource availability.

In this paper, we propose a new joint approach for analyzing affiliation (two-mode) networks by using factorial methods and blockmodeling. In our recent work, we pursued the joint use of a given factorial method, i.e., MCA/MFA, and a clustering method, namely blockmodeling, but through distinct steps. Here we look for a strategy that permits us to apply the two methods simultaneously. To this aim, we propose a method that allows us to group individuals and variables simultaneously and directly for binary matrices, namely cluster correspondence analysis (cluster CA). This method can be adopted when dealing with affiliation matrices having a binary structure. Hence, we look at the way network positions (clusters) can be incorporated in cluster CA to verify if cluster CA can properly represent specific network structures. We illustrate our proposal through an empirical application on an affiliation network of stage co-productions.

Melanoma is the most lethal form of skin cancer and its incidence is growing worldwide. In the last ten years, the therapeutic scenario of this disease has been revolutionized by the introduction of targeted therapies and immune-checkpoint inhibitors. However, in patients with many lesions and bulky tumors, in which surgery is no longer feasible, there is a need for new treatment options. Here we report, for the first time to our knowledge, a clinical case where a melanoma patient harboring the SMO p.Gln216Arg mutation has been treated with imiquimod, showing a complete and durable response. To better explain this outstanding response to the treatment, we transfected a melanoma cell line (MeWo) with the SMO p.Gln216Arg mutation in order to evaluate its role in response to the imiquimod treatment. Moreover, to better demonstrate that the antitumor activity of imiquimod was due to its role in suppressing the oncogenic SMO signaling pathway, independently of its immune modulating function, an in vivo experiment has been performed. This clinical case opens up a new scenario for the treatment of melanoma patients identifying a new potentially druggable target.

Recent research has shown that job crafting, which describes individuals' attempts to craft a job to make it correspond more to personal inclinations, skills, and abilities, can generate significant work and nonwork benefits for individuals. Using the theoretical lens of activation theory, we examined whether professionals are prompted to cognitively craft their jobs in response to the increasing perception of precarisation of their profession, measured in terms of job insecurity and perceived external prestige. We adopted a mixed methods approach among professional accountants operating in Southern Italy and the results indicated the presence of two curvilinear relationships. More specifically, we found that accountants were more likely to engage in cognitive crafting when experiencing moderate levels of job insecurity (rather than high or low) and in the presence of both low and high levels of perceived external prestige (rather than a moderate level). Implications for theory and practice are discussed.

Thyroid hormones (THs) mediate pleiotropic cellular processes involved in metabolism, cellular proliferation, and differentiation. The intracellular hormonal environment can be tailored by the type 1 and 2 deiodinase enzymes D2 and D3, which catalyze TH activation and inactivation respectively. In many cellular systems, THs exert well-documented stimulatory or inhibitory effects on cell proliferation; however, the molecular mechanisms by which they control rates of cell cycle progression have not yet been entirely clarified. We previously showed that D3 depletion or TH treatment influences the proliferation and survival of basal cell carcinoma (BCC) cells. Surprisingly, we also found that BCC cells express not only sustained levels of D3 but also robust levels of D2. The aim of the present study was to dissect the contribution of D2 to TH metabolism in the BCC context, and to identify the molecular changes associated with cell proliferation and survival induced by TH and mediated by D2 and D3.We used the CRISPR/Cas9 technology to genetically deplete D2 and D3 in BCC cells and studied the consequences of depletion on cell cycle progression and on cell death. Cell cycle progression was analyzed by fluorescence activated cell sorting analysis of synchronized cells, and the apoptosis rate by annexin V incorporation.Mechanistic investigations revealed that D2 inactivation accelerates cell cycle progression thereby enhancing the proportion of S-phase cells and cyclin D1 expression. Conversely, D3 mutagenesis drastically suppressed cell proliferation and enhanced apoptosis of BCC cells. Furthermore, the basal apoptotic rate was oppositely regulated in D2- and D3-depleted cells.Our results indicate that BCC cells constitute an example in which the TH signal is finely tuned by the concerted expression of opposite-acting deiodinases. The dual regulation of D2 and D3 expression plays a critical role in cell cycle progression and cell death by influencing cyclin D1-mediated entry into the G1-S phase. These findings reinforce the concept that TH is a potential therapeutic target in human BCC.

The deiodinase family of enzymes mediates the activation and inactivation of thyroid hormone. The role of these enzymes in the regulation of the systemic concentrations of thyroid hormone is well established and underpins the treatment of common thyroid diseases. Interest in this field has increased in the past 10 years as the deiodinases became implicated in tissue development and homeostasis, as well as in the pathogenesis of a wide range of human diseases. Three deiodinases have been identified, namely, types 1, 2 and 3 iodothyronine deiodinases, which differ in their catalytic properties and tissue distribution. Notably, the expression of these enzymes changes during the lifetime of an individual in relation to the different needs of each organ and to ageing. The systemic homeostatic role of deiodinases clearly emerges during changes in serum concentrations of thyroid hormone, as seen in patients with thyroid dysfunction. By contrast, the role of deiodinases at the tissue level allows thyroid hormone signalling to be finely tuned within a given cell in a precise time–space window without perturbing serum concentrations of thyroid hormone. This Review maps the overall functional role of the deiodinases and explores challenges and novel opportunities arising from the expanding knowledge of these ‘master’ components of the thyroid homeostatic system. This Review discusses the two main functions of the human deiodinases: the homeostatic control of plasma concentrations of thyroid hormone and the control of intracellular T3 concentrations.

Recent thyroid cancer guidelines found it reasonable to use local therapies during treatment with tyrosine kinase inhibitors (TKIs) in selected patients with oligoprogressive disease, namely, in the presence of a single progressing lesion in an otherwise TKI-responsive metastatic cancer. However, there is a lack of experience in the management of oligoprogressive thyroid cancers. This report illustrates the case of one patient with oligoprogressive thyroid cancer during therapy with lenvatinib. We found that the application of local ablative therapy in oligoprogressive disease prolonged the progression-free survival and thus extended the time to therapy interruption. However, the optimal care for TKI-treated oligoprogressive cancers remains unclear and needs to be investigated in prospective trials.

Thyroid hormone (TH) is a key metabolic regulator that acts by coordinating short- and long-term energy needs. Accordingly, significant metabolic changes are observed depending on thyroid status. Although it is established that hyperthyroidism augments basal energy consumption, thus resulting in an enhanced metabolic state, the net effects on cellular respiration and generation of reactive oxygen species (ROS) remain unclear. To elucidate the effects of augmented TH signal in muscle cells, we generated a doxycycline-inducible cell line in which the expression of the TH-activating enzyme, type 2 deiodinase (D2), is reversibly turned on by the "Tet-ON" system. Interestingly, increased intracellular TH caused a net shift from oxidative phosphorylation to glycolysis and a consequent increase in the extracellular acidification rate. As a result, mitochondrial ROS production, and both the basal and doxorubicin-induced production of cellular ROS were reduced. Importantly, the expression of a set of antioxidant genes was up-regulated, and, among them, the mitochondrial scavenger Sod2 was specifically induced at transcriptional level by D2-mediated TH activation. Finally, we observed that attenuation of oxidative stress and increased levels of SOD2 are key elements of the differentiating cascade triggered by TH and D2, thereby establishing that D2 is essential in coordinating metabolic reprogramming of myocytes during myogenic differentiation. In conclusion, our findings indicate that TH plays a key role in oxidative stress dynamics by regulating ROS generation. Our novel finding that TH and its intracellular metabolism act as mitochondrial detoxifying agents sheds new light on metabolic processes relevant to muscle physiology.

Metastatic cancer patients generally respond well to treatment with tyrosine kinase inhibitors (TKIs). However, TKI resistance occurs in almost all cases and often leads to a change in treatment. Recent guidelines, including thyroid cancer, raised the possibility of locally treating TKI-resistant oligoprogressive disease, i.e., one or a few progressing lesions in an otherwise treatment-responsive metastatic cancer, thereby obviating the need to change the ongoing TKI. To determine the benefits of this intervention, we reviewed studies on the use of LAT for TKI-treated oligoprogressive cancers. We found that in non-small cell lung cancer at least, LAT prolongs disease control and the duration of exposure to a TKI irrespective of the LAT used. Moreover, we reviewed the local ablative therapies (LATs) that are feasible for the local control of oligoprogressive thyroid cancer. Lastly, we report two illustrative cases of patients with oligoprogressive thyroid cancer treated with two different LATs while on therapy with TKIs. Both LATs extended the duration of disease control and the time of exposure to the ongoing TKI, thereby indicating that LAT is a favorable option for TKI-treated oligoprogressive thyroid cancer. Prospective randomized studies are needed to verify the benefit of LATs in terms of progression-free and overall survival in this increasingly frequent clinical setting.

The thyroid hormone–inactivating (TH-inactivating) enzyme type 3 iodothyronine deiodinase (D3) is an oncofetal protein that is rarely expressed in adult life but has been shown to be reactivated in the context of proliferation and neoplasms. D3 terminates TH action within the tumor microenvironment, thereby enhancing cancer cell proliferation. However, the pathological role of D3 and the contribution of TH metabolism in cancer have yet to be fully explored. Here, we describe a reciprocal regulation between TH action and the cancer-associated microRNA-21 (miR21) in basal cell carcinoma (BCC) skin tumors. We found that, besides being negatively regulated by TH at the transcriptional level, miR21 attenuates the TH signal by increasing D3 levels. The ability of miR21 to positively regulate D3 was mediated by the tumor suppressor gene GRHL3, a hitherto unrecognized D3 transcriptional inhibitor. Finally, in a BCC mouse model, keratinocyte-specific D3 depletion markedly reduced tumor growth. Together, our results establish TH action as a critical hub of multiple oncogenic pathways and provide functional and mechanistic evidence of the involvement of TH metabolism in BCC tumorigenesis. Moreover, our results identify a miR21/GRHL3/D3 axis that reduces TH in the tumor microenvironment and has potential to be targeted as a therapeutic approach to BCC.

Thyroid hormone is a pleiotropic factor that controls many cellular processes in multiple cell types such as cancer stem cells (CSC). Thyroid hormone concentrations in the blood are stable, but the action of the deiodinases (D2–D3) provides cell-specific regulation of thyroid hormone activity. Deregulation of deiodinase function and thyroid hormone status has been implicated in tumorigenesis. Therefore, we investigated the role of thyroid hormone metabolism and signaling in colorectal CSCs (CR-CSC), where deiodinases control cell division and chemosensitivity. We found that increased intracellular thyroid hormone concentration through D3 depletion induced cell differentiation and sharply mitigated tumor formation. Upregulated BMP4 expression and concomitantly attenuated Wnt signaling accompanied these effects. Furthermore, we demonstrate that BMP4 is a direct thyroid hormone target and is involved in a positive autoregulatory feedback loop that modulates thyroid hormone signaling. Collectively, our findings highlight a cell-autonomous metabolic mechanism by which CR-CSCs exploit thyroid hormone signaling to facilitate their self-renewal potential and suggest that drug-induced cell differentiation may represent a promising therapy for preventing CSC expansion and tumor progression. Cancer Res; 76(5); 1237–44. ©2015 AACR.

Thyroid cancer metastasizes in 4% of cases. Approximately two-thirds of these patients are refractory to radioactive iodine-131 (RAI) therapy and have a poor 10-year survival prognosis. Treatment with tyrosine kinase inhibitors (TKIs) may be administered in selected RAI-refractory patients. However, these agents are often associated with adverse events, including vomiting. We report the case of a patient affected by RAI-refractory thyroid cancer with lung and intracranial metastases undergoing treatment with the antiangiogenic TKI lenvatinib, and with teriparatide replacement therapy for postsurgical hypoparathyroidism. Due to lenvatinib-related vomiting, which did not respond to therapy, conventional oral calcium supplementation failed to maintain normal serum calcium levels and the patient had repeated episodes of hypocalcemia. Subcutaneous teriparatide injections restored serum calcium levels, and thus lenvatinib therapy could be continued. This experience indicates that hormone replacement with teriparatide is a feasible option for cancer patients affected by hypoparathyroidism not treatable with oral calcium supplementation.

The type 2 iodothyronine deiodinase (D2) is essential for feedback regulation of TSH by T4. We genetically inactivated in vivo D2 in thyrotrophs using a mouse model of Cga-driven cre recombinase. Pituitary D2 activity was reduced 90% in the Cga-cre D2 knockout (KO) mice compared with control Dio2fl/fl mice. There was no growth or reproductive phenotype. Basal TSH levels were increased 1.5- to 1.8-fold, but serum T4 and T3 were not different from the controls in adult mice. In hypothyroid adult mice, suppression of TSH by T4, but not T3, was impaired. Despite mild basal TSH elevation, the TSH increase in response to hypothyroidism was 4-fold reduced in the Cga-cre D2KO compared with control mice despite an identical level of pituitary TSH α- and β-subunit mRNAs. In neonatal Cga-cre D2KO mice, TSH was also 2-fold higher than in the controls, but serum T4 was elevated. Despite a constant TSH, serum T4 increased 2–3-fold between postnatal day (P) 5 and P15 in both genotypes. The pituitary, but not cerebrocortical, D2 activity was markedly elevated in P5 mice decreasing towards adult levels by P17. In conclusion, a congenital severe reduction of thyrotroph D2 causes a major impairment of the TSH response to hypothyroidism. This would be deleterious to the compensatory adaptation of the thyroid gland to iodine deficiency.

The goal of evidence-based practice guidelines is to optimize the management of emerging diseases, such as differentiated thyroid cancer (DTC). The aim of this study was to assess therapeutic approaches for DTC in Italy and to see how closely these practices conformed to those recommended in the 2009 American Thyroid Association (ATA) guidelines.The Italian Thyroid Cancer Observatory was established to collect data prospectively on thyroid cancers consecutively diagnosed in participating centers (uniformly distributed across the nation). Data on the initial treatment of all pathologically confirmed DTC cases present in the database from January 1, 2013 (database creation) to January 31, 2016, were analyzed.A total of 1748 patients (77.2% females; median age 48.1 years [range 10-85 years]) were enrolled in the study. Most (n = 1640; 93.8%) were papillary carcinomas (including 84 poorly differentiated/aggressive variants); 6.2% (n = 108) were follicular and Hürthle cell carcinomas. The median tumor diameter was 11 mm (range 1-93 mm). Tumors were multifocal in 613 (35%) and presented extrathyroidal extension in 492 (28%) cases. Initial treatments included total thyroidectomy (involving one or two procedures; n = 726; 98.8%) and lobectomy (n = 22; 1.2%). A quarter of the patients who underwent total thyroidectomy had unifocal, intrathyroidal tumors ≤1 cm (n = 408; 23.6%). Neck dissection was performed in 40.4% of the patients (29.5% had central compartment dissection). Radioiodine remnant ablation (RRA) was performed in 1057 (61.2%) of the 1726 patients who underwent total thyroidectomy: 460 (41.2%) of the 983 classified by 2009 ATA guideline criteria as low-risk, 570 (87.1%) of the 655 as intermediate-risk, and 82 (93.1%) of the 88 as high-risk patients (p 0.001). RRA was performed in 44% of the cases involving multifocal DTCs measuring ≤1 cm.The treatment approaches for DTCs used in Italy display areas of inconsistency with those recommended by the 2009 ATA guidelines. Italian practices were characterized by underuse of thyroid lobectomy in intrathyroidal, unifocal DTCs ≤1 cm. The use of RRA was generally consistent with risk-stratified recommendations. However, its frequent use in small DTCs (≤1 cm) that are multifocal persists, despite the lack of evidence of benefit. These data provide a baseline for future assessments of the impact of international guidelines on DTC management in Italy. These findings also illustrate that the dissemination and implementation of guideline recommendations, and the change in practice patterns, require ongoing education and time.

Context:A substantial proportion of athyreotic levothyroxine (LT4)-treated patients experience hypothyroid-like symptoms. During LT4 replacement, levels of the active hormone triiodothyronine (T3) strictly depend on type 2-deiodinase (D2)-mediated activation of LT4. The Thr92Ala polymorphism and the 258 G/A in the DIO2 gene have been associated with various clinical conditions.Objectives:To investigate the effects of DIO2 polymorphisms in thyroid hormone homeostasis.Design:We compared the presurgical hormonal status of thyroidectomized LT4-treated patients who had a similar thyroid-stimulating hormone (TSH) level with their postsurgery status and analyzed their DIO2 genotype in a subgroup of 102/140 (72.8%) of patients. We measured the enzymatic properties of Thr92Ala in living cells and in relevant generated mouse models.Subjects and methods:A total of 140 thyroidectomized subjects were included. Serum free T3 (FT3), free thyroxine, and TSH levels were directly measured. Immunohistochemistry and immunoblotting were performed for D2 protein.Results:The DIO2 genotyping revealed an association between low FT3 values and Thr92Ala. Specifically, the mean postsurgery FT3 levels were significantly lower in patients carrying the mutated allele(s) than in wild-type patients, in whom FT3 postsurgical levels were similar to presurgery levels. The −258 G/A variation was not associated with hormonal alteration. We found that endogenous wild-type D2 and Thr92Ala share the same subcellular localization but differ in protein stability. Importantly, Thr92Ala reduced D2-mediated thyroxine to T3 conversion.Conclusions:Thyroidectomized patients carrying Thr92Ala are at increased risk of reduced intracellular and serum T3 concentrations that are not adequately compensated for by LT4, thus providing evidence in favor of customized treatment of hypothyroidism in athyreotic patients.

Thyroid hormone (TH) regulates such crucial biological functions as normal growth, development and metabolism of nearly all vertebrate tissues. In skeletal muscle, TH plays a critical role in regulating the function of satellite cells, the bona fide skeletal muscle stem cells. Deiodinases (D2 and D3) have been found to modulate the expression of various TH target genes in satellite cells. Regulation of the expression and activity of the deiodinases constitutes a cell-autonomous, pre-receptor mechanism that controls crucial steps during the various phases of myogenesis. Here, we review the roles of deiodinases in skeletal muscle stem cells, particularly in muscle homeostasis and upon regeneration. We focus on the role of T3 in stem cell functions and in commitment towards lineage progression. We also discuss how deiodinases might be therapeutically exploited to improve satellite-cell-mediated muscle repair in skeletal muscle disorders or injury.

Thyroid hormone (TH) metabolism is mediated by deiodinases, a family of thioredoxin fold-containing enzymes that remove iodide from thyroxine and its derivatives. The coordinated action of deiodinases allows target cells to modulate rapidly their own TH availability in response to different cues. Type 3 deiodinase (D3), the physiological inactivator of TH, is an oncofetal protein whose re-activation in adult tissues has been correlated with hyperproliferative states and with human solid tumors. This suggests a link between deiodinase-mediated TH metabolism and carcinogenesis.D3 is overexpressed in basal cell carcinomas (BCCs) and sustains the proliferation of BCC cells. It exerts a similar function in colon cancer, which suggests that attenuating the TH signal is part of a widespread neoplastic program. Here, recent advances in D3 research, particularly as regards the role of D3 regulation and function in solid tumors are reviewed.Given the vast array of TH's physiological and cellular functions, unraveling TH metabolism in cancer biology is a promising challenge for the development of new therapies for human cancer.

Background Thyroid hormone influences gene expression in virtually all vertebrates. Its action is initiated by the activation of T4 to T3, an outer ring deiodination reaction that is catalyzed by the type 1 or the type 2 iodothyronine selenodeiodinases (D1 or D2). Inactivation of T4 and T3 occurs via inner ring deiodination catalyzed by the type 3 iodothyronine selenodeiodinases (D3). The T4 concentration is generally quite stable in human plasma, with T3 levels also remaining constant. Deiodinase actions are tightly regulated in both pre- and post-natal life when they are required to make local adjustments of intracellular T3 concentrations in a precise spatio- and temporal manner. Although all the signals governing the dynamic expression of deiodinases in specific cell types are not known, many important regulatory factors have been deciphered. Scope of review This review provides striking examples from the recent literature illustrating how the expression of D2 and D3 is finely tuned during maturation of different organs, and how their action play a critical role in different settings to control intracellular T3 availability. Major conclusions Emerging evidence indicates that in various cell contexts, D2 and D3 are expressed in a dynamic balance, in which the expression of one enzyme is coordinately regulated with that of the other to tightly control intracellular T3 levels commensurate with cell requirements at that time. General significance Deiodinases control TH action in a precise spatio-temporal fashion thereby providing a novel mechanism for the local paracrine and autocrine regulation of TH action. This remarkable tissue-specific regulation of intracellular thyroid status remains hidden due to the maintenance of constant circulating TH concentrations by the hypothalamic–pituitary–thyroid axis. This article is part of a Special Issue entitled Thyroid hormone signalling., Highlights ► Deiodinases contribute to the control of TH availability in different cell types. ► Deiodinase actions are part of the TH-mediated control of differentiation. ► Deiodinases are critical in cochlear and retinal maturation in pre/post-natal life. ► In muscle cells, deiodinases are required for myogenesis and muscle regeneration.

Contains fulltext : 118008.pdf (Publisher’s version ) (Open Access) Ankyloblepharon, ectodermal defects, cleft lip/palate (AEC) syndrome is a rare autosomal dominant disorder caused by mutations in the p63 gene, essential for embryonic development of stratified epithelia. The most severe cutaneous manifestation of this disorder is the long-lasting skin fragility associated with severe skin erosions after birth. Using a knock-in mouse model for AEC syndrome, we found that skin fragility was associated with microscopic blistering between the basal and suprabasal compartments of the epidermis and reduced desmosomal contacts. Expression of desmosomal cadherins and desmoplakin was strongly reduced in AEC mutant keratinocytes and in newborn epidermis. A similar impairment in desmosome gene expression was observed in human keratinocytes isolated from AEC patients, in p63-depleted keratinocytes and in p63 null embryonic skin, indicating that p63 mutations causative of AEC syndrome have a dominant-negative effect on the wild-type p63 protein. Among the desmosomal components, desmocollin 3, desmoplakin and desmoglein 1 were the most significantly reduced by mutant p63 both at the RNA and protein levels. Chromatin immunoprecipitation experiments and transactivation assays revealed that p63 controls these genes at the transcriptional level. Consistent with reduced desmosome function, AEC mutant and p63-deficient keratinocytes had an impaired ability to withstand mechanical stress, which was alleviated by epidermal growth factor receptor inhibitors known to stabilize desmosomes. Our study reveals that p63 is a crucial regulator of a subset of desmosomal genes and that this function is impaired in AEC syndrome. Reduced mechanical strength resulting from p63 mutations can be alleviated pharmacologically by increasing desmosome adhesion with possible therapeutic implications.

p63 and p73 are highly homologous members of the p53 family that originated by gene duplication at the invertebrate-to-vertebrate transition. We characterize here a previously unreported gene, Transformation-related protein 63 regulated (Tprg), located upstream of the p63 gene in the vertebrate genome, with striking similarity to Transformation related protein 63 regulated like (Tprgl), an uncharacterized gene located upstream of p73, suggesting that p63/Tprg and p73/Tprgl are embedded in a paralogue region originated from a single duplication event. Tprg is predominantly expressed in the epithelial compartment of the skin, more abundantly in differentiated cells. Consistent with its relative higher expression in differentiated keratinocytes, finely tuned p63 expression levels are required for optimal Tprg expression in primary keratinocytes. p63 is essential for Tprg expression as shown in p63-knockdown keratinocytes; however, high levels of p63 result in Tprg downregulation. p63 directly binds in vivo to a canonical p63-binding site in an evolutionary conserved genomic region located in Tprg intron 4. This genomic region is sufficient to function as a p63-inducible enhancer in promoter studies. Thus, we demonstrate that the Tprg gene is predominantly expressed in skin, is physically associated with the p63 gene during evolution, and directly regulated by p63 through a long-distance enhancer located within the Tprg locus.

et al., Guideline advice of many societies on the management of subclinical hypothyroidism in pregnancy suggests treatment when TSH serum levels exceed 2.5 mU/l. Justification of this procedure is based on limited experience, mainly from studies carried out in patients with positive thyroid-specific antibodies and higher TSH levels that classically define the condition in the non-pregnant state. Taking into account a lack of clear understanding of the regulation of thyroid hormone transport through the utero-placental unit and in the absence of foetal markers to monitor the adequacy of thyroxine treatment, this review attempts to discuss currently available data and suggests a more cautious approach.

By producing T3 from T4, type 2 iodothyronine deiodinase (D2) catalyzes the first step in the cascade underlying the effect exerted by thyroid hormone. Type 2 iodothyronine deiodinase mRNA is expressed at high levels in human heart but is barely detectable in the corresponding rodent tissue. Although the heart is a major target of thyroid hormone, the role of cardiac D2 and the factors that regulate its expression are unknown.Here we report that the human Dio2 promoter is very sensitive to the cardiac transcription factors Nkx-2.5 and GATA-4. Nkx-2.5 transactivates a 6.5-kb human (h)Dio2-chloramphenicol acetyltransferase construct, with maximal induction reached with a 633-bp proximal promoter region. Interestingly, despite 73% identity with the corresponding human region, the rat Dio2 promoter is much less responsive to Nkx-2.5 induction. Using EMSA, we found that two sites in the human promoter (C and D) specifically bind Nkx-2.5. In coexpression studies, GATA-4 alone was a poor inducer of the hDio2 promoter; however in synergy with Nkx-2.5, it activated D2 reporter gene expression in the human, but not the rat promoter. Functional analysis showed that both C and D sites are required for the complete Nkx-2.5 response and for the Nkx-2.5/GATA-4 synergistic effect. In neonatal rat primary myocardiocytes, most of the hDio2-chloramphenicol acetyltransferase activity was suppressed by mutation of the Nkx-2.5 binding sites. Finally, a mutant Nkx-2.5 protein (N188K), which causes, in heterozygosity, congenital heart diseases, did not transactivate the Dio2 promoter and interfered with its activity in cardiomyocytes, possibly by titrating endogenous Nkx-2.5 protein away from the promoter.In conclusion, this study shows that Nkx-2.5 and GATA-4 play prime roles in Dio2 gene regulation in the human heart and suggests that it is their synergistic action in humans that causes the differential expression of the cardiac Dio2 gene between humans and rats.

The type 3 iodothyronine selenodeiodinase (D3) is an integral membrane protein that inactivates thyroid hormones. By using immunofluorescence cytochemistry confocal microscopy of live or fixed cells transiently expressing FLAG-tagged human D3 or monkey hepatocarcinoma cells expressing endogenous D3, we identified D3 in the plasma membrane. It co-localizes with Na,K-ATPase alpha, with the early endosomal marker EEA-1 and clathrin, but not with two endoplasmic reticulum resident proteins. Most of the D3 molecule is extracellular and can be biotinylated with a cell-impermeant probe. There is constant internalization of D3 that is blocked by sucrose or methyl-beta-cyclodextrin-containing medium. Exposing cells to a weak base such as primaquine increases the pool of internalized D3, suggesting that D3 is recycled between plasma membrane and early endosomes. Such recycling could account for the much longer half-life of D3 (12 h) than the thyroxine activating members of the selenodeiodinase family, type 1 (D1; 8 h) or type 2 (D2; 2 h) deiodinase. The extracellular location of D3 gives ready access to circulating thyroid hormones, explaining its capacity for rapid inactivation of circulating thyroxine and triiodothyronine in patients with hemangiomas and its blockade of the access of maternal thyroid hormones to the human fetus.

Negli ultimi anni le riviste cliniche e di management hanno moltiplicato il numero di studi sul tema delle reti sociali e del capitale sociale dei professionisti nel settore sanitario. La complessita dei task, l’elevata incertezza e la crescente necessita di rispondere in tempi adeguati alle richieste dei pazienti hanno reso di vitale importanza la gestione delle interdipendenze tra organizzazioni, gruppi e singoli professionisti in questo settore. La vastita di contributi richiede un’analisi sistematica della letteratura. Questo articolo presenta una review dei principali lavori scientifici che riguardano il tema delle reti sociali e del capitale sociale tra professionisti in sanita. L’analisi riguarda 76 articoli scientifici selezionati con l’ausilio della piattaforma ISI Web of Knowledge. L’articolo riassume gli antecedenti e le conseguenze delle reti sociali tra i professionisti. La conoscenza di questi antecedenti e di queste conseguenze puo essere utile a policy maker e manager aziendali per progettare le aziende sanitarie e facilitare l’introduzione di qualsiasi innovazione organizzativa.

Summary Precise control of the thyroid hormone (T3)-dependent transcriptional program is required by multiple cell systems, including muscle stem cells. Deciphering how this is achieved and how the T3 signal is controlled in stem cell niches is essentially unknown. We report that in response to proliferative stimuli such as acute skeletal muscle injury, type 3 deiodinase (D3), the thyroid hormone-inactivating enzyme, is induced in satellite cells where it reduces intracellular thyroid signaling. Satellite cell-specific genetic ablation of dio3 severely impairs skeletal muscle regeneration. This impairment is due to massive satellite cell apoptosis caused by exposure of activated satellite cells to the circulating TH. The execution of this proapoptotic program requires an intact FoxO3/MyoD axis, both genes positively regulated by intracellular TH. Thus, D3 is dynamically exploited in vivo to chronically attenuate TH signaling under basal conditions while also being available to acutely increase gene programs required for satellite cell lineage progression., Graphical Abstract, Highlights • D3 is induced in proliferating satellite cells thereby reducing thyroid signaling • D3 depletion causes massive cell apoptosis in vitro and in vivo • Apoptosis requires FoxO3, a TH target gene • Satellite cells customize TH signature and adapt it to their functional needs, Deiodinases inactivate thyroid hormone (TH) signaling, allowing a precise control of TH action at the cellular level. Dentice et al. find that type 3 deiodinase acts as a survival factor during skeletal muscle repair, and attenuation of TH signaling is required to prevent muscle stem cell apoptosis and promote lineage progression in vivo.

Type 3 deiodinase (D3), the physiologic inactivator of thyroid hormones, is induced during tissue injury and regeneration. This has led to the hypotheses that D3 impacts injury tolerance by reducing local T3 signaling and contributes to the fall in serum triiodothyronine (T3) observed in up to 75% of sick patients (termed the low T3 syndrome). Here we show that a novel mutant mouse with hepatocyte-specific D3 deficiency has normal local responses to toxin-induced hepatonecrosis, including normal degrees of tissue necrosis and intact regeneration, but accelerated systemic recovery from illness-induced hypothyroxinemia and hypotriiodothyroninemia, demonstrating that peripheral D3 expression is a key modulator of the low T3 syndrome.

The proliferation and differentiation of muscle precursor cells require myogenic regulatory factors and chromatin modifiers whose concerted action dynamically regulates access to DNA and allows reprogramming of cells towards terminal differentiation. Type 2 deiodinase (D2), the thyroid hormone (TH)-activating enzyme, is sharply upregulated during myoblast differentiation, whereas type 3 deiodinase (D3), the TH-inactivating enzyme, is downregulated. The molecular determinants controlling synchronized D2 and D3 expression in muscle differentiation are completely unknown. Here, we report that the histone H3 demethylating enzyme (LSD-1) is essential for transcriptional induction of D2 and repression of D3. LSD-1 relieves the repressive marks (H3-K9me2-3) on the Dio2 promoter and the activation marks (H3-K4me2-3) on the Dio3 promoter. LSD-1 silencing impairs the D2 surge in skeletal muscle differentiation while inducing D3 expression thereby leading to a global decrease in intracellular TH production. Furthermore, endogenous LSD-1 interacts with FoxO3a, and abrogation of FoxO3-DNA binding compromises the ability of LSD-1 to induce D2. Our data reveal a novel epigenetic control of reciprocal deiodinases expression and provide a molecular mechanism by which LSD-1, through the opposite regulation of D2 and D3 expression, acts as a molecular switch that dynamically finely tunes the cellular needs of active TH during myogenesis.

Background: NKX2-1 mutations have been described in several patients with primary congenital hypothyroidism, respiratory distress, and benign hereditary chorea, which are classical manifestations of the brain-thyroid-lung syndrome (BTLS). Methods: The NKX2-1 gene was sequenced in the members of a Brazilian family with clinical features of BTLS, and a novel monoallelic mutation was identified in the affected patients. We introduced the mutation in an expression vector for the functional characterization by transfection experiments using both thyroidal and lung-specific promoters. Results: The mutation is a deletion of a cytosine at position 834 (ref. sequence NM-003317) (c.493delC) that causes a frameshift with formation of an abnormal protein from amino acid 165 and a premature stop at position 196. The last amino acid of the nuclear localization signal, the whole homeodomain, and the carboxy-terminus of NKX2-1 are all missing in the mutant protein, which has a premature stop codon at position 196 (p.Arg165Glyfs*32). The p.Arg165Glyfs*32 mutant does not bind DNA, and it is unable to transactivate the thyroglobulin (Tg) and the surfactant protein-C (SP-C) promoters. Interestingly, a dose-dependent dominant negative effect of the p.Arg165Glyfs*32 was demonstrated only on the Tg promoter, but not on the SP-C promoter. This effect was also noticed when the mutation was tested in presence of PAX8 or cofactors that synergize with NKX2-1 (P300 and TAZ). The functional effect was also compared with the data present in the literature and demonstrated that, so far, it is very difficult to establish a specific correlation among NKX2-1 mutations, their functional consequence, and the clinical phenotype of affected patients, thus suggesting that the detailed mechanisms of transcriptional regulation still remain unclear. Conclusions: We describe a novel NKX2-1 mutation and demonstrate that haploinsufficiency may not be the only explanation for BTLS. Our results indicate that NKX2-1 activity is also finely regulated in a tissue-specific manner, and additional studies are required to better understand the complexities of genotype-phenotype correlations in the NKX2-1 deficiency syndrome. © Copyright 2013, Mary Ann Liebert, Inc. 2013.

The major product secreted by the thyroid is thyroxine (T4), whereas most of the biologically active triiodothyronine (T3) derives from the peripheral conversion of T4 into T3. The deiodinase enzymes are involved in activation and inactivation of thyroid hormones. Type 1 and type 2 deiodinase (D1 and D2) convert T4 into T3 whereas D3 degrades T4 and T3 into inactive metabolites and is thus the major physiological thyroid hormone inactivator. The hypothalamic-pituitary-thyroid axis maintains circulating thyroid hormone levels constant, while the deiodinases tissue-specifically regulate intracellular thyroid status by controlling thyroid hormone action in a precise spatio-temporal fashion. Here we review the data related to the recent identification of a paraneoplastic syndrome called “consumptive hypothyroidism”, which exemplifies how deiodinases alter substantially the concentration of thyroid hormone in blood. This syndrome results from the aberrant uncontrolled expression of D3 that can induce a severe form of hypothyroidism by inactivating T4 and T3 in defined tumor tissue. This rare thyroid hormone insufficiency generally affects patients in the first years of life, and has distinct features in terms of diagnosis, treatment and prognosis with respect to other forms of hypothyroidism.

Background & Aims Activation of the β-catenin/T-cell factor (TCF) complex occurs in most colon tumors, and its actions correlate with the neoplastic phenotype of intestinal epithelial cells. Type 3 deiodinase (D3), the selenoenzyme that inactivates thyroid hormone (3,5,3′ triiodothyronine [T3]), is frequently expressed by tumor cells, but little is known about its role in the regulation of T3 signaling in cancer cells. Methods We measured D3 expression in 6 colon cancer cell lines and human tumors and correlated it with the activity of the β-catenin/TCF complex. We also determined the effects of D3 loss on local thyroid hormone signaling and colon tumorigenesis. Results We show that D3 is a direct transcriptional target of the β-catenin/TCF complex; its expression was higher in human intestinal adenomas and carcinomas than in healthy intestinal tissue. Experimental attenuation of β-catenin reduced D3 levels and induced type 2 deiodinase (the D3 antagonist that converts 3,5,3′,5′ tetraiodothyronine into active T3) thereby increasing T3-dependent transcription. In the absence of D3, excess T3 reduced cell proliferation and promoted differentiation in cultured cells and in xenograft mouse models. This occurred via induction of E-cadherin, which sequestered β-catenin at the plasma membrane and promoted cell differentiation. Conclusions Deiodinases are at the interface between the β-catenin and the thyroid hormone pathways. Their synchronized regulation of intracellular T3 concentration is a hitherto unrecognized route by which the multiple effects of β-catenin are generated and may be targeted to reduce the oncogenic effects of β-catenin in intestinal cells.

Thyroid hormone (TH) is a pleiotropic agent that has widespread biological functions, i.e., it controls cellular growth, tissue development and homeostasis and neoplastic transformation. Suitable TH levels are critical for the development of various types of tissues and are essential for the regulation of metabolic processes throughout life. The serum concentrations of TH affect its biological activity. Moreover, at tissue level, TH action is regulated by the expression and activity of deiodinases, i.e., the enzymes that mediate the metabolic pathways by activating and/or inactivating TH. The type I and II deiodinases (D1 and D2) initiate TH action by converting thyroxine (T4) into the active TH form (T3), whereas type III deiodinase (D3) mediates the local attenuation of TH by converting T4 and T3 into the inactive metabolites rT3 and T2, respectively. The deiodinase system is a potent mechanism of pre-receptoral control of TH action; it is often altered in such pathological conditions as cancer. D3 is widely expressed in embryonic tissues and in placenta, where it blocks excessive maternal-to-fetal transfer of TH. In contrast, during late neonatal and adult life, D3 is expressed mainly in the central nervous system and skin. Interestingly, D3 expression is re-activated in various types of human cancers. Here we review recent evidence that D3 expression plays a crucial role in human carcinogenesis, and speculate as to its complex role in the regulation of cell proliferation in several neoplastic contexts. It is conceivable that the local modulation of TH action via deiodinases is a powerful molecular tool to manipulate the intracellular TH status, thus influencing the growth and maintenance of selected hormone-dependent cancers.

Consumptive hypothyroidism is a rare syndrome resulting from increased catabolism of T(4) and T(3) by increased type 3 iodothyronine deiodinase (D3) activity. Consumptive hypothyroidism has primarily been described as a paraneoplastic syndrome in infants as well as in two adults with D3-expressing tumors.The aim of the study was to report the third case of consumptive hypothyroidism in an adult and the first in an athyreotic patient.We present a 38-yr-old athyreotic female who was euthyroid on a stable therapeutic dose of thyroid hormone for many years and then developed marked hyperthyrotropinemia, coincident with the discovery of large D3-expressing hepatic vascular tumors. The patient also had low serum T(3) and elevated serum rT(3). Hyperthyrotropinemia transiently worsened after surgical resection of the vascular tumors and then persisted for 3 wk after the operation, despite further increases in levothyroxine therapy.The patient's vascular tumor and adjacent normal liver parenchyma were probed with a polyclonal antibody directed against D3.D3 immunostaining of the patient's vascular tumor was positive, with no significant immunoreactivity in the adjacent normal hepatic tissue.This is the third case report of consumptive hypothyroidism in an adult and the first in an athyreotic individual. This case demonstrates that hyperthyrotropinemia may persist after partial liver resection, possibly from the hepatic resection itself.

A novel oncogene has been recently found activated in human thyroid carcinomas. This oncogene was named RET/PTC and was shown to be a chimeric gene, product of the fusion of the tyrosinekinase domain or the RET proto-oncogene to the 5’-terminal region of another gene named H4 or D10S170. A paracentric inversion (10) (q 11.2-21) was demonstrated to be responsible for the H4/RET fusion. Thyroid tumors consist of a broad range of lesions ranging from the benign follicular adenomas to the differentiated papillary and follicular carcinomas and to the fatal anaplastic carcinomas. Papillary and follicular carcinomas have different clinical behaviours and are associated with different risk factors, thus they are probably related to different genetic events. In fact the activation of RET is restricted to the papillary subtype. Here we show the activation of the RET oncogene in 2 out of 10 new cases of papillary thyroid carcinomas.

Because of its large mass, relatively high metabolic activity and responsiveness to thyroid hormone, skeletal muscle contributes significantly to energy expenditure. Despite the presence of mRNA encoding the type 2 iodothyronine-deiodinase (D2), an enzyme that activates T4 to T3, very low or undetectable activity has been reported in muscle homogenates of adult humans and mice. With a modified D2 assay, using microsomal protein, overnight incubation and protein from D2 knockout mouse muscle as a tissue-specific blank, we examined slow- and fast-twitch mouse skeletal muscles for D2 activity and its response to physiological stimuli. D2 activity was detectable in all hind limb muscles of 8- to 12-wk old C57/BL6 mice. Interestingly, it was higher in the slow-twitch soleus than in fast-twitch muscles (0.40 ± 0.06 vs. 0.076 ± 0.01 fmol/min · mg microsomal protein, respectively, P < 0.001). These levels are greater than those previously reported. Hypothyroidism caused a 40% (P < 0.01) and 300% (P < 0.001) increase in D2 activity after 4 and 8 wk treatment with antithyroid drugs, respectively, with no changes in D2 mRNA. Neither D2 mRNA nor activity increased after an overnight 4 C exposure despite a 10-fold increase in D2 activity in brown adipose tissue in the same mice. The magnitude of the activity, the fiber specificity, and the robust posttranslational response to hypothyroidism argue for a more important role for D2-generated T3 in skeletal muscle physiology than previously assumed.

Activity of the sodium/iodide symporter (NIS) in lactating breast is essential for iodide (I-) accumulation in milk. Significant NIS upregulation was also reported in breast cancer, indicating a potential use of radioiodide treatment. All-trans-retinoic acid (tRA) is a potent ligand that enhances NIS expression in a subset of breast cancer cell lines and in experimental breast cancer models. Indirect tRA stimulation of NIS in breast cancer cells is very well documented; however, direct upregulation by tRA-activated nuclear receptors has not been identified yet. Aiming to uncover cis-acting elements directly regulating NIS expression, we screened evolutionary-conserved non-coding genomic sequences for responsiveness to tRA in MCF-7. Here, we report that a potent enhancer in the first intron of NIS mediates direct regulation by tRA-stimulated nuclear receptors. In vitro as well as in vivo DNA-protein interaction assays revealed direct association between retinoic acid receptor-α (RARα) and retinoid-X-receptor (RXR) with this enhancer. Moreover, using chromatin immunoprecipitation (ChIP) we uncovered early events of NIS transcription in response to tRA, which require the interaction of several novel intronic tRA responsive elements. These findings indicate a complex interplay between nuclear receptors, RNA Pol-II and multiple intronic RAREs in NIS gene, and they establish a novel mechanistic model for tRA-induced gene transcription. © The Author(s) 2010. Published by Oxford University Press.

…E quelli che s'inarnorono di pratica senza scientia sono come il nocchiere che entra navilio senza timone e bussola, e che mai ha certezza dove si vada… [He who loves practice without theory is like the sailor who boards ship without a rudder and compass and never knows where he may be cast.] (Leonardo Da Vinci) Thyroid endocrinology aims to advance our knowledge of the physiopathology of the thyroid gland. The thyroidologist, like all physicians concerned with internal medicine, lives in an environment of increasing specialization and multiplying subspecialties. This applies also to thyroid research, in which the divide between “basic” and “clinical” scientists is often wider than it should be. Consequently, the grand challenge that Frontiers in Thyroid Endocrinology faces is to provide a common platform for the opinions and rigorous investigations of both of them. Of course, the link will be the so-called “translational research,” in which both sides of the coin merge in the ultimate interest of the patient affected by thyroid disease. I will quote only a few examples of a non-exhaustive list of ideas and a few examples of “hot topics” that are jostling for attention. I am fully aware of the limits of this list, but my aim is to give a flavor of what and where the grand challenges are and what awaits us in the near future.

The FoxO3-dependent increase in type II deiodinase (D2), which converts the prohormone thyroxine (T4) to 3,5,3′-triiodothyronine (T3), is required for normal mouse skeletal muscle differentiation and regeneration. This implies a requirement for an increase in D2-generated intracellular T3 under these conditions, which has not been directly demonstrated despite the presence of D2 activity in skeletal muscle. We directly show that D2-mediated T4-to-T3 conversion increases during differentiation in C2C12 myoblast and primary cultures of mouse neonatal skeletal muscle precursor cells, and that blockade of D2 eliminates this. In adult mice given 125I-T4 and 131I-T3, the intracellular 125I-T3/131I-T3 ratio is significantly higher than in serum in both the D2-expressing cerebral cortex and the skeletal muscle of wild-type, but not D2KO, mice. In D1-expressing liver and kidney, the 125I-T3/131I-T3 ratio does not differ from that in serum. Hypothyroidism increases D2 activity, and in agreement with this, the difference in 125I-T3/131I-T3 ratio is increased further in hypothyroid wild-type mice but not altered in the D2KO. Notably, in wild-type but not in D2KO mice, the muscle production of 125I-T3 is doubled after skeletal muscle injury. Thus, D2-mediated T4-to-T3 conversion generates significant intracellular T3 in normal mouse skeletal muscle, with the increased T3 required for muscle regeneration being provided by increased D2 synthesis, not by T3 from the circulation.

The thyroid hormone plays a fundamental role in the development, growth, and metabolic homeostasis in all vertebrates by affecting the expression of different sets of genes. A group of thioredoxin fold-containing selenoproteins known as deiodinases control thyroid hormone action by activating or inactivating the precursor molecule thyroxine that is secreted by the thyroid gland. These pathways ensure regulation of the availability of the biologically active molecule T3, which occurs in a time-and tissue-specific fashion. In addition, because cells and plasma are in equilibrium and deiodination affects central thyroid hormone regulation, these local deiodinase-mediated events can also affect systemic thyroid hormone economy, such as in the case of non-thyroidal illness. Heightened interest in the field has been generated following the discovery that the deiodinases can be a component in both the Sonic hedgehog signaling pathway and the TGR-5 signaling cascade, a G-protein-coupled receptor for bile acids. These new mechanisms involved in deiodinase regulation indicate that local thyroid hormone activation and inactivation play a much broader role than previously thought.

The Sonic hedgehog (Shh) pathway plays a critical role in hair follicle physiology and is constitutively active in basal cell carcinomas (BCCs), the most common human malignancy. Type 3 iodothyronine deiodinase (D3), the thyroid hormone-inactivating enzyme, is frequently expressed in proliferating and neoplastic cells, but its role in this context is unknown. Here we show that Shh, through Gli2, directly induces D3 in proliferating keratinocytes and in mouse and human BCCs. We demonstrate that Gli-induced D3 reduces intracellular active thyroid hormone, thus resulting in increased cyclin D1 and keratinocyte proliferation. D3 knockdown caused a 5-fold reduction in the growth of BCC xenografts in nude mice. Shh-induced thyroid hormone degradation via D3 synergizes with the Shh-mediated reduction of the type 2 deiodinase, the thyroxine-activating enzyme, and both effects are reversed by cAMP. This previously unrecognized functional cross-talk between Shh/Gli2 and thyroid hormone in keratinocytes is a pathway by which Shh produces its proliferative effects and offers a potential therapeutic approach to BCC.

Mutations of the Ret receptor tyrosine kinase are responsible for inheritance of multiple endocrine neoplasia (MEN2A and MEN2B) and familial medullary thyroid carcinoma syndromes. Although several familial medullary thyroid carcinoma and most MEN2A mutations involve substitutions of extracellular cysteine residues, in most MEN2B cases there is a methionine-to-threonine substitution at position 918 (M918T) of the Ret kinase domain. The mechanism by which the MEN2B mutation converts Ret into a potent oncogene is poorly understood. Both MEN2A and MEN2B oncoproteins exert constitutive activation of the kinase. However, the highly aggressive MEN2B phenotype is not supported by higher levels of Ret-MEN2B kinase activity compared with Ret-MEN2A. It has been proposed that Ret-MEN2B is more than just an activated Ret kinase and that the M918T mutation, by targeting the kinase domain of Ret, might alter Ret substrate specificity, thus affecting Ret autophosphorylation sites and the ability of Ret to phosphorylate intracellular substrates. We show that the Ret-MEN2B mutation causes specific potentiated phosphorylation of tyrosine 1062 (Y1062) compared with Ret-MEN2A. Phosphorylated Y1062 is part of a Ret multiple effector docking site that mediates recruitment of the Shc adapter and of phosphatidylinositol-3 kinase (PI3K). Accordingly, we show that Ret-MEN2B is more active than Ret-MEN2A in associating with She and in causing constitutive activation of the Ras/mitogen-activated protein kinase and PI3K/Akt cascades. We conclude that the MEN2B mutation specifically potentiates the ability of Ret to autophosphorylate Y1062 and consequently to couple to the Ras/mitogen-activated protein kinase and the PI3K/Akt pathways. The more efficient triggering of these pathways may account for the difference between MEN2A and MEN2B syndromes.

1352 schoolchildren between 6-14 years old (699 males and 653 females) and 943 adults (176 males and 767 females) from eight villages of the province of Avellino were studied. All subjects were examined for thyroid size by at least two expert examiners. In most of them urine samples were collected for iodine determinations. 387 schoolchildren and 161 adults from Flumeri and Villanova were evaluated by thyroid echography. The prevalence of goiter was from 23.5 to 52.2% and the median urinary iodine excretion was from 42.3 to 66.2 micrograms/l in schoolchildren. In adults the prevalence of goiter was from 41.2 to 86.7% and the median urinary iodine excretion was from 37.1 to 53.7 micrograms/l. Our data showed a degree of iodine deficiency from low to moderate. The echography permitted to point out a greater prevalence of nodules than the thyroid palpation.

Non-shivering thermogenesis is required for survival of rodents during cold stress. Uncoupling protein-1 acts in brown adipose tissue (BAT) to transport protons, thus dissipating the proton gradient across the inner mitochondrial membrane. This permits respiration uncoupled from ATP synthesis. UCP-1 function is inhibited by the binding of purine nucleotides, with GTP/GDP being more potent than ATP/ADP. We used a cDNA subtraction analysis to identify cDNAs rapidly induced by cold exposure. One of these encodes rat guanosine monophosphate reductase (GMP-r). This was surprising in that previous data had suggested that this enzyme was absent in rodents. Rat GMP-r is 96% identical to human GMP-r, and its mRNA is increased 30-fold in BAT within 6 h of cold exposure. The gene is also expressed (but not cold-responsive) in muscle and kidney, but not in white fat. We speculate that the physiological function of the marked increase in BAT GMP-r during cold stress may be to deplete the brown adipocyte of guanine nucleotides, converting them to IMP, thus permitting enhanced UCP-1 function. This is a previously unrecognized regulatory aspect of thermogenesis, an essential physiological response of rodents to cold.

Salvatore D, Celetti A, Fabien N, Paulin C, Martelli ML, Battaglia C, Califano D, Monaco C, Viglietto G, Santoro M, Fusco A. Low frequency of p53 mutations in human thyroid tumors; p53 and Ras mutation in two out of fifty-six thyroid tumours. Eur J Endocrinol 1996;134:177–83. ISSN 0804–4643 Objective: p53 is a well-known nuclear phosphoprotein encoded by a suppressor gene known to be mutated in various kinds of human tumours. A relationship between p53 gene mutation and tumour progression seems to be a common feature of several neoplasias. Desing: In order to investigate the role of p53 mutations in human thyroid tumours, DNA samples derived from fifty-six neoplastic tissues, ranging from benign adenomas to undifferentiated carcinomas, were examined for the presence of p53 gene mutations. Methods: The analysis has been conducted using polymerase chain reaction (PCR) amplification of the exons 5–9 of the p53 gene followed by single strand conformation polymorphism (SSCP) and sequence analyses. Results: One anaplastic carcinoma and one papillary carcinoma showed p53 gene mutations in exons 5 and 8, respectively. A cell line established from the papillary carcinoma showed the same mutation present in the original tumour. Both p53 mutations were heterozygous. The p53 positive samples were analysed for other genetic alterations frequently detected in human thyroid carcinomas (mutations of the RET, TRK, and ras oncogenes): both p53-mutated samples proved to be mutated at level of codon 13 of the c-Ki-ras gene. Conclusions: Our data confirm that p53 gene alterations are rare in well-differentiated thyroid tumours, that they are an important requirement for the establishment in culture of human thyroid carcinoma cell lines, and that they can be associated with other genetic alterations, namely ras mutations, in the malignant progression of thyroid tumours. Alfredo Fusco, Dipartimento di Biologia e Patologia Cellulare e Molecolare, Facoltà di Medicina e Chirurgia, via S. Pansini 5, 80131, Napoli, Italy