Biosynthesis of 3-iodothyronamine (T1AM) is dependent on the sodium-iodide symporter and thyroperoxidase but does not involve extrathyroidal metabolism of T4

Abstract

3-Iodothyronamine (T(1)AM) is an endogenous thyroid hormone derivative with unknown biosynthetic origins. Structural similarities have led to the hypothesis that T(1)AM is an extrathyroidal metabolite of T(4). This study uses an isotope-labeled T(4) [heavy-T(4) (H-T(4))] that can be distinguished from endogenous T(4) by mass spectrometry, which allows metabolites to be identified based on the presence of this unique isotope signature. Endogenous T(1)AM levels depend upon thyroid status and decrease upon induction of hypothyroidism. However, in hypothyroid mice replaced with H-T(4), the isotope-labeled H-T(3) metabolite is detected, but no isotope-labeled T(1)AM is detected. These data suggest that T(1)AM is not an extrathyroidal metabolite of T(4), yet is produced by a process that requires the same biosynthetic factors necessary for T(4) synthesis.

Fig. 1.

Structures of T₄ and T₁AM.

Fig. 2.

Establishment of a hypothyroid and TH replacement mouse model. Total serum T₄ levels were measured by RIA after treatment with 0.1% MMI and 0.2% KClO₄ and replacement with 20 ng/g T₄ for 2–5 wk (A) or replacement with 0–500 ng/g T₄ for 3 wk (B); n = 3–5 per group; *, P < 0.05; **, P < 0.01. ND, Not detectable.

Fig. 3.

H-T₄ replacement in a hypothyroid mouse model. A, Total serum T₄ levels as measured by RIA after 2 wk of treatment with 0.1% MMI and 0.2% KClO₄ and replacement with T₄ or H-T₄; n = 4–5 per group. B, Body weights of euthyroid, hypothyroid, T₄, and H-T₄-replaced mice. Hypothyroid, T₄ and H-T₄-replaced mice were treated with 0.1% MMI and 0.2% KClO₄ starting on d 1 and continuing through d 36. At d 15, replaced mice began receiving daily injections of either T₄ or H-T₄; n = 4–8 per group. ND, Not detectable.

Fig. 4.

Representative LC-MS/MS chromatogram for T₁AM, T₃, and T₄ from euthyroid and 2-wk hypothyroid mouse livers. The m/z transitions represented are: T₁AM, 356 to 339; d₄-T₁AM, 360 to 343; T₃, 652 to 606; ¹³C₆-T₃, 658 to 612; T₄, 778 to 732; and ¹³C₆-T₄, 784 to 738. Solid black line, Endogenous analyte; dashed line, internal standard. Representative traces of three samples analyzed for each group. cps, Counts per second.

Fig. 5.

T₁AM levels decrease in mouse liver as a function of time of 0.1% MMI and 0.2% KClO₄ treatment, as measured by LC-MS/MS; n = 3–5 per time point.

Fig. 6.

Representative LC-MS/MS chromatogram for T₁AM, T₃, and T₄ from T₄ and H-T₄-replaced mouse livers. The m/z transitions represented are: T₁AM, 356 to 339; d₄-T₁AM, 360 to 343; H-T₁AM, 365 to 347; T₃, 652 to 606; ¹³C₆-T₃, 658 to 612; H-T₃, 662 to 615; T₄, 778 to 732; ¹³C₆-T₄, 784 to 738; and H-T₄, 788 to 741. Solid black line, Unlabeled analyte (endogenous analyte for T₁AM and H-T₄-replaced group, and metabolite of exogenous T₄ in T₄-replaced group); dashed line, internal standard; gray line, heavy metabolite from H-T₄. cps, Counts per second.

Fig. 7.

Liver T₄, T₃, T₁AM and H-T₄, H-T₃, and H-T₁AM levels in euthyroid, hypothyroid, and hypothyroid mice with T₄ or H-T₄ replacement, as measured by LC-MS/MS. Inducing hypothyroidism for 2 wk decreases endogenous T₄, T₃, and T₁AM. Replacement with T₄ results in normal T₄ and increased T₃ with respect to euthyroid but no increase in T₁AM with respect to hypothyroid. Replacement with H-T₄ results in measurable H-T₄ and H-T₃, no increase in endogenous T₄, T₃ or T₁AM, and no detectable H-T₁AM. H-T₄ and H-T₃ were only detected in mice replaced with H-T₄; n = 3 for euthyroid, hypothyroid, and H-T₄ replaced; n = 2 for T₄ replaced.