The TSH receptor (TSHR) is the critical target for antibody production in Graves' disease (GD). Insulin-like growth factor 1 receptor (IGF1R) has been proposed as a second autoantigen in complications of GD such as orbitopathy. We attempted to induce orbital tissue remodeling in mice undergoing immunizations with plasmids encoding TSHR and IGF1R delivered by in vivo skeletal muscle electroporation, a procedure known to give a sustained, long-term antibody response. Female BALB/c mice were challenged with TSHR A-subunit or IGF1Rα subunit plasmid by injection and electroporation. Mice challenged with TSHR A-subunit plasmid resulted in high frequency (75%) of hyperthyroidism and thyroid-stimulating antibodies. But strikingly, immunization with TSHR A-subunit plasmid also elicited antibody to IGF1Rα subunit. Mice challenged in the same manner with IGF1Rα subunit plasmid produced strong antibody responses to IGF1R, but did not undergo any changes in phenotype. Simultaneous challenge by double antigen immunization with the two plasmids in distant anatomical sites reduced the incidence of hyperthyroidism, potentially as a consequence of antigenic competition. Thyroid glands from the TSHR A-subunit plasmid-challenged group were enlarged with patchy microscopic infiltrates. Histological analysis of the orbital tissues demonstrated moderate connective tissue fibrosis and deposition of Masson's trichrome staining material. Our findings imply that immunization with TSHR A-subunit plasmid leads to generation of IGF1R antibodies, which together with thyroid-stimulating antibodies may precipitate remodeling of orbital tissue, raising our understanding of its close association with GD.
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- Abstract: Thyroid* x
- Abstract: Digestion x
- Abstract: Thyroxine x
- Abstract: Thyroglobulin x
- Abstract: Thyroiditis x
- Abstract: Thyrotoxicosis x
- Abstract: Hypothyroidism x
- Abstract: Hyperthyroidism x
- Abstract: TSHR x
- Abstract: Metabolism x
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Shuang-Xia Zhao, Shanli Tsui, Anthony Cheung, Raymond S Douglas, Terry J Smith and J Paul Banga
Lei Zhang, Carol Paddon, Mark D Lewis, Fiona Grennan-Jones and Marian Ludgate
Since TSH receptor (TSHR) expression increases during adipogenesis and signals via cAMP/phospho-cAMP-response element binding protein (CREB), reported to be necessary and sufficient for adipogenesis, we hypothesised that TSHR activation would induce preadipocyte differentiation. Retroviral vectors introduced constitutively active TSHR (TSHR*) into 3T3L1 preadipocytes; despite increased cAMP (RIA) and phospho-CREB (western blot) there was no spontaneous adipogenesis (assessed morphologically, using oil red O and QPCR measurement of adipogenesis markers). We speculated that Gβγ signalling may be inhibitory but failed to induce adipogenesis using activated Gsα (gsp*). Inhibition of phosphodiesterases did not promote adipogenesis in TSHR* or gsp* populations. Furthermore, differentiation induced by adipogenic medium with pioglitazone was reduced in TSHR* and abolished in gsp* expressing 3T3L1 cells. TSHR* and gsp* did not inactivate PPARγ (PPARG as listed in the HUGO database) by phosphorylation but expression of PPARγ1 was reduced and PPARγ2 undetectable in gsp*. FOXO1 phosphorylation (required to inactivate this repressor of adipogenesis) was lowest in gsp* despite the activation of AKT by phosphorylation. PROF is a mediator that facilitates FOXO1 phosphorylation by phospho-Akt. Its transcript levels remained constantly low in the gsp* population. In most measurements, the TSHR* cells were between the gsp* and control 3T3L1 preadipocytes. The enhanced down-regulation of PREF1 (adipogenesis inhibitor) permits retention of some adipogenic potential in the TSHR* population. We conclude that Gsα signalling impedes FOXO1 phosphorylation and thus inhibits PPARγ transcription and the alternative promoter usage required to generate PPARγ2, the fat-specific transcription factor necessary for adipogenesis.
Joachim M Weitzel, Torsten Viergutz, Dirk Albrecht, Rupert Bruckmaier, Marion Schmicke, Armin Tuchscherer, Franziska Koch and Björn Kuhla
During the transition between late gestation and early lactation, dairy cows experience severe metabolic stress due to the high energy and nutrient requirements of the fetus and the mammary gland. Additional thermal stress that occurs with rising temperatures during the ongoing climate change has further adverse implications on energy intake, metabolism and welfare. The thyroid hormone (TH)-mediated cellular signaling has a pivotal role in regulation of body temperature, energy intake and metabolic adaptation to heat. To distinguish between energy intake and heat stress-related effects, Holstein cows were first kept at thermoneutrality at 15°C followed by exposure to heat stress (HS) at 28°C or pair-feeding (PF) at 15°C for 6 days, in late pregnancy and again in early lactation. Herein, we focused on hepatic metabolic changes associated with alterations in the hypothalamic–pituitary–thyroid axis in HS and PF animals. T3 and T4 levels dropped with HS or PF; however, in HS animals, this decline was more pronounced. Thyroid-stimulating hormone (TSH) levels remain unaffected, while plasma cholesterol concentrations were lower in HS than PF animals. Hepatic marker genes for TH action (THRA, DIO1 and PPARGC1) decreased after HS and were lower compared to PF cows but only post-partum. Proteomics data revealed reduced hepatic amino acid catabolism ante-partum and a shift toward activated beta-oxidation and gluconeogenesis but declined oxidative stress defense post-partum. Thus, liver metabolism of HS and PF cows adapts differently to diminished energy intake both ante-partum and post-partum, and a different TH sensitivity is involved in the regulation of catabolic processes.
Dawn E W Livingstone, Sarah L Grassick, Gillian L Currie, Brian R Walker and Ruth Andrew
In obese humans, metabolism of glucocorticoids by 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1) and A-ring reduction (by 5α- and 5β-reductases) is dysregulated in a tissue specific manner. These changes have been recapitulated in leptin resistant obese Zucker rats but were not observed in high-fat fed Wistar rats. Recent data from mouse models suggest that such discrepancies may reflect differences in leptin signalling. We therefore compared glucocorticoid metabolism in murine models of leptin deficiency and resistance. Male ob/ob and db/db mice and their respective littermate controls (n=10–12/group) were studied at the age of 12 weeks. Enzyme activities and mRNA expression were quantified in snap-frozen tissues. The patterns of altered pathways of steroid metabolism in obesity were similar in ob/ob and db/db mice. In liver, 5β-reductase activity and mRNA were increased and 11β-HSD1 decreased in obese mice, whereas 5α-reductase 1 (5αR1) mRNA was not altered. In visceral adipose depots, 5β-reductase was not expressed, 11β-HSD1 activity was increased and 5αR1 mRNA was not altered in obesity. By contrast, in subcutaneous adipose tissue 11β-HSD1 and 5αR1 mRNA were decreased. Systematic differences were not found between ob/ob and db/db murine models of obesity, suggesting that variations in leptin signalling through the short splice variant of the Ob receptor do not contribute to dysregulation of glucocorticoid metabolism.
Corinne Caillaud, Mie Mechta, Heidi Ainge, Andreas N Madsen, Patricia Ruell, Emilie Mas, Catherine Bisbal, Jacques Mercier, Stephen Twigg, Trevor A Mori, David Simar and Romain Barrès
Erythropoietin (EPO) ameliorates glucose metabolism through mechanisms not fully understood. In this study, we investigated the effect of EPO on glucose metabolism and insulin signaling in skeletal muscle. A 2-week EPO treatment of rats fed with a high-fat diet (HFD) improved fasting glucose levels and glucose tolerance, without altering total body weight or retroperitoneal fat mass. Concomitantly, EPO partially rescued insulin-stimulated AKT activation, reduced markers of oxidative stress, and restored heat-shock protein 72 expression in soleus muscles from HFD-fed rats. Incubation of skeletal muscle cell cultures with EPO failed to induce AKT phosphorylation and had no effect on glucose uptake or glycogen synthesis. We found that the EPO receptor gene was expressed in myotubes, but was undetectable in soleus. Together, our results indicate that EPO treatment improves glucose tolerance but does not directly activate the phosphorylation of AKT in muscle cells. We propose that the reduced systemic inflammation or oxidative stress that we observed after treatment with EPO could contribute to the improvement of whole-body glucose metabolism.
Gisela Helfer and Qing-Feng Wu
Metabolic syndrome is a global public health problem and predisposes individuals to obesity, diabetes and cardiovascular disease. Although the underlying mechanisms remain to be elucidated, accumulating evidence has uncovered a critical role of adipokines. Chemerin, encoded by the gene Rarres2, is a newly discovered adipokine involved in inflammation, adipogenesis, angiogenesis and energy metabolism. In humans, local and circulating levels of chemerin are positively correlated with BMI and obesity-related biomarkers. In this review, we discuss both peripheral and central roles of chemerin in regulating body metabolism. In general, chemerin is upregulated in obese and diabetic animals. Previous studies by gain or loss of function show an association of chemerin with adipogenesis, glucose homeostasis, food intake and body weight. In the brain, the hypothalamus integrates peripheral afferent signals including adipokines to regulate appetite and energy homeostasis. Chemerin increases food intake in seasonal animals by acting on hypothalamic stem cells, the tanycytes. In peripheral tissues, chemerin increases cell expansion, inflammation and angiogenesis in adipose tissue, collectively resulting in adiposity. While chemerin signalling enhances insulin secretion from pancreatic islets, contradictory results have been reported on how chemerin links to obesity and insulin resistance. Given the association of chemerin with obesity comorbidities in humans, advances in translational research targeting chemerin are expected to mitigate metabolic disorders. Together, the exciting findings gathered in the last decade clearly indicate a crucial multifaceted role for chemerin in the regulation of energy balance, making it a promising candidate for urgently needed pharmacological treatment strategies for obesity.
T V Novoselova, R Larder, D Rimmington, C Lelliott, E H Wynn, R J Gorrigan, P H Tate, L Guasti, The Sanger Mouse Genetics Project, S O’Rahilly, A J L Clark, D W Logan, A P Coll and L F Chan
Melanocortin receptor accessory protein 2 (MRAP2) is a transmembrane accessory protein predominantly expressed in the brain. Both global and brain-specific deletion of Mrap2 in mice results in severe obesity. Loss-of-function MRAP2 mutations have also been associated with obesity in humans. Although MRAP2 has been shown to interact with MC4R, a G protein-coupled receptor with an established role in energy homeostasis, appetite regulation and lipid metabolism, the mechanisms through which loss of MRAP2 causes obesity remains uncertain. In this study, we used two independently derived lines of Mrap2 deficient mice (Mrap2 tm1a/tm1a) to further study the role of Mrap2 in the regulation of energy balance and peripheral lipid metabolism. Mrap2 tm1a/tm1a mice have a significant increase in body weight, with increased fat and lean mass, but without detectable changes in food intake or energy expenditure. Transcriptomic analysis showed significantly decreased expression of Sim1, Trh, Oxt and Crh within the hypothalamic paraventricular nucleus of Mrap2 tm1a/tm1a mice. Circulating levels of both high-density lipoprotein and low-density lipoprotein were significantly increased in Mrap2 deficient mice. Taken together, these data corroborate the role of MRAP2 in metabolic regulation and indicate that, at least in part, this may be due to defective central melanocortin signalling.
Historically, adipose tissue was considered to be a passive storage vessel discharging nutrients in times of famine and accumulating fat in times of surfeit. This view changed with the identification of leptin as an adipocyte hormone. Leptin functions as an afferent signal in a negative feedback loop that regulates food intake and metabolism to maintain homeostatic control of adipose tissue mass. Before this, the existence of a system maintaining homeostatic control of energy balance was unclear. The identification of leptin has thus uncovered a new endocrine system that also links changes in nutrition to adaptive responses in most if not all other physiologic systems. Further studies have revealed a set of clinical syndromes caused by leptin deficiency, including lipodystrophy and hypothalamic amenorrhea. This work has led to new therapeutic approaches for a number of human conditions and has also established a conceptual framework for studying the pathogenesis of obesity.
Dawn E W Livingstone, Emma M Di Rollo, Tracy C-S Mak, Karen Sooy, Brian R Walker and Ruth Andrew
5α-Reductases irreversibly catalyse A-ring reduction of pregnene steroids, including glucocorticoids and androgens. Genetic disruption of 5α-reductase 1 in male mice impairs glucocorticoid clearance and predisposes to glucose intolerance and hepatic steatosis upon metabolic challenge. However, it is unclear whether this is driven by changes in androgen and/or glucocorticoid action. Female mice with transgenic disruption of 5α-reductase 1 (5αR1-KO) were studied, representing a ‘low androgen’ state. Glucocorticoid clearance and stress responses were studied in mice aged 6 months. Metabolism was assessed in mice on normal chow (aged 6 and 12 m) and also in a separate cohort following 1-month high-fat diet (aged 3 m). Female 5αR1-KO mice had adrenal suppression (44% lower AUC corticosterone after stress), and upon corticosterone infusion, accumulated hepatic glucocorticoids (~27% increased corticosterone). Female 5αR1-KO mice aged 6 m fed normal chow demonstrated insulin resistance (~35% increased area under curve (AUC) for insulin upon glucose tolerance testing) and hepatic steatosis (~33% increased hepatic triglycerides) compared with controls. This progressed to obesity (~12% increased body weight) and sustained insulin resistance (~38% increased AUC insulin) by age 12 m. Hepatic transcript profiles supported impaired lipid β-oxidation and increased triglyceride storage. Female 5αR1-KO mice were also predisposed to develop high-fat diet-induced insulin resistance. Exaggerated predisposition to metabolic disorders in female mice, compared with that seen in male mice, after disruption of 5αR1 suggests phenotypic changes may be underpinned by altered metabolism of glucocorticoids rather than androgens.
Valentina Pampanini, Daniela Germani, Antonella Puglianiello, Jan-Bernd Stukenborg, Ahmed Reda, Iuliia Savchuk, Kristín Rós Kjartansdóttir, Stefano Cianfarani and Olle Söder
Prenatal events such as intrauterine growth restriction can affect gonadal development of the offspring and have an impact on reproductive health. To investigate the effects of intrauterine growth restriction induced by uterine artery ligation on the postnatal rat testis. Pregnant rats underwent uterine artery ligation at day 19 of gestation. Offspring were killed at 5, 20 and 40 days post-partum (dpp). At killing, one gonad was snap-frozen in liquid nitrogen and processed for RNA and steroid extraction. The other gonad was formalin-fixed for histology. Gene expression was analyzed by TaqMan Low-Density Array. Intratesticular testosterone, estradiol and serum gonadotrophins were measured. Thirty genes were dysregulated in intrauterine growth-restricted rats compared to controls, among which markers of Sertoli cell and Leydig cell function, cell metabolism and growth factors. Testis weights were significantly reduced at 5 and 20 dpp in intrauterine growth-restricted rats and caught-up by 40 dpp. Accordingly, Sertoli cell number was significantly lower in 5 dpp intrauterine growth-restricted rats. At 20 dpp, intratesticular testosterone was significantly increased in intrauterine growth-restricted rats, whereas serum gonadotrophins were unchanged. IUGR altered the gene expression in the rat testes up to peripubertal age and reduced testis size and Sertoli cell number in neonatal age. Multiple mechanisms encompassing genetic changes and steroid production by the testis may be involved in the catch-up growth phase that restored testis size by 40 dpp. Permanent consequences on organ function and gamete integrity cannot be excluded and deserve further investigations.