Aortic effects of thyroid hormone in male mice

in Journal of Molecular Endocrinology
Correspondence should be addressed to J Mittag: jens.mittag@uni-luebeck.de
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It is well established that thyroid hormones are required for cardiovascular functions; however, the molecular mechanisms remain incompletely understood, especially the individual contributions of genomic and non-genomic signalling pathways. In this study, we dissected how thyroid hormones modulate aortic contractility. To test the immediate effects of thyroid hormones on vasocontractility, we used a wire myograph to record the contractile response of dissected mouse aortas to the adrenergic agonist phenylephrine in the presence of different doses of T3 (3,3′,5-triiodothyronine). Interestingly, we observed reduced vasoconstriction under low and high T3 concentrations, indicating an inversed U-shaped curve with maximal constrictive capacity at euthyroid conditions. We then tested for possible genomic actions of thyroid hormones on vasocontractility by treating mice for 4 days with 1 mg/L thyroxine in drinking water. The study revealed that in contrast to the non-genomic actions the aortas of these animals were hyperresponsive to the contractile stimulus, an effect not observed in endogenously hyperthyroid TRβ knockout mice. To identify targets of genomic thyroid hormone action, we analysed aortic gene expression by microarray, revealing several altered genes including the well-known thyroid hormone target gene hairless. Taken together, the findings demonstrate that thyroid hormones regulate aortic tone through genomic and non-genomic actions, although genomic actions seem to prevail in vivo. Moreover, we identified several novel thyroid hormone target genes that could provide a better understanding of the molecular changes occurring in the hyperthyroid aorta.

 

      Society for Endocrinology

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    Aortic response ex vivo recorded in a wire myograph to phenylephrine (PE) after 5 min prestimulation with solvent (0 nM T3), 1 nM T3 (A), 10 nM T3 (B) or 100 nM T3 (C). Please note that the 1 nM T3 curve serves as euthyroid reference control and is thus included for better comparison in (A, B and C). All values are mean ± s.e.m. of n = 8. ***P < 0.001 for top of curve. (D) Phosphorylation of ERK, AKT and AMPK after prestimulation of aortic rings for 10 min with DMSO, 100 nM T3 or 100 nM rT3 as assessed by Western blot. All values are mean ± s.e.m. of n = 9–10.

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    (A) Serum total T4 and T3 levels after 4 day treatment with 1 mg/L T4 in drinking water. Values are mean ± s.e.m. of n = 6. **P < 0.01; ***P < 0.001 with unpaired 2-tailed t test. (B) Aortic response ex vivo recorded in a wire myograph to phenylephrine (PE) in mice treated for 4 days with 1 mg/L T4 in drinking water as compared to controls. Values are mean ± s.e.m. of n = 10. *P < 0.05 for top of curve. (C) Aortic response ex vivo recorded in a wire myograph to phenylephrine (PE) in mice lacking TRβ as compared to controls. Values are mean ± s.e.m. of n = 5. *P < 0.05 for bottom of curve. (D) Phosphorylation of ERK, AKT and AMPK of aortic rings from wild type or TRβ knockout mice as assessed by Western blot. All values are mean ± s.e.m. of n = 4.

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    (A) Overview of the genomic effects of T4 on aortic gene expression in mice treated for 4 days with 1 mg/L T4 in drinking water as compared to controls as studied by micoarray analysis. (B) Weighted gene set enrichment analysis of the microarray results. The numbers on the bars indicate the number of genes found to be significantly changed in the respective pathway. (C) Unbiased hierarchical clustering of the microarray results with the top five candidate genes by P-value in (D), which were validated using qPCR in a different cohort (E). Values are mean ± s.e.m. of n = 4–5. *P < 0.05 with unpaired 2-tailed t test. (F) Correlation of gene expression as assessed by microarray with the individual total T3 and T4 levels of the respective animals of the top correlated genes with P < 0.0001. Tm100, transmembrane protein 100; Penk, preproenkephaline; Hr, hairless; Fbln1, fibulin 1; Elmo1, engulfment and cell motility 1; Pel3, HXXXD-type acyltransferase family protein; Maf, MAF bZIP transcription factor; Pola1, DNA Polymerase Alpha 1; Msantd2, Myb/SANT DNA Binding Domain Containing 2.

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