MCT8 Deficiency in Male Mice Mitigates the Phenotypic Abnormalities Associated With the Absence of a Functional Type 3 Deiodinase

Abstract

Mice deficient in the type 3 deiodinase (D3KO mice) manifest impaired clearance of thyroid hormone (TH), leading to elevated levels of TH action during development. This alteration causes reduced neonatal viability, growth retardation, and central hypothyroidism. Here we examined how these phenotypes are affected by a deficiency in the monocarboxylate transporter 8 (MCT8), which is a major contributor to the transport of the active thyroid hormone, T3, into the cell. MCT8 deficiency eliminated the neonatal lethality of type 3 deiodinase (D3)-deficient mice and significantly ameliorated their growth retardation. Double-mutant newborn mice exhibited similar peripheral thyrotoxicosis and increased brain expression of T3-dependent genes as mice with D3 deficiency only. Later in neonatal life and adulthood, double-mutant mice manifested central and peripheral TH status similar to mice with single MCT8 deficiency, with low serum T4, elevated serum TSH and T3, and decreased T3-dependent gene expression in the hypothalamus. In double-mutant adult mice, both thyroid gland size and the hypothyroidism-induced rise in TSH were greater than those in mice with single D3 deficiency but less than those in mice with MCT8 deficiency alone. Our results demonstrate that the marked phenotypic abnormalities observed in the D3-deficient mouse, including perinatal mortality, growth retardation, and central hypothyroidism in adult animals, require expression of MCT8, confirming the interdependent relationship between the TH transport into cells and the deiodination processes.

Figure 1.

MCT8 deficiency rescues the neonatal lethality of D3KO mice. Genotype frequency distribution at age P2 (A) and ages P5 and P15 (B) of male offspring resulting from fathers heterozygous for the D3 mutation and mothers heterozygous for both D3 and MCT8 mutations. Bars represent the f_observed values and the dashed lines indicate the calculated f_expected values. The P values indicated were determined by the χ² test. Genotypes are named WT, KO, or Het (heterozygous). Additional details can be found in Supplemental Table 1.

Figure 2.

Growth of mice of all four genotypes. The BW of male WT, MCT8KO, D3KO, and D3/MCT8 double-KO mice is represented at P2 (A), P5 (B), and as a overall growth curve (C). Each data point represents the mean ± SEM of determinations in 5–22 mice. *, P < .05, **, P < .01, ***, P < .001, as determined by a one-way-ANOVA followed by Tukey's test (A and B) or a two-way-ANOVA with a random mouse effect.

Figure 3.

Thyroid status parameters of mice of all four genotypes at age P2. Serum T₄ (A), serum TSH (B), hepatic Dio1 mRNA (C), hypothalamic Hr mRNA (D), and cerebrocortical Hr mRNA (E) in P2 WT, MCT8KO, D3/MCT8 double-KO, and D3KO male mice. Data represent the mean ± SEM of determinations in three to eight mice. *, P < .05, **, P < .01, ***, P < .001, as determined by a one-way-ANOVA followed by Tukey's test.

Figure 4.

Thyroid status parameters of mice of all four genotypes at age P5. Serum T₄ (A), serum TSH (B), hepatic Dio1 mRNA (C), hypothalamic Hr mRNA (D), and cerebrocortical Hr mRNA (E) in P5 WT, MCT8KO, D3/MCT8 double-KO, and D3KO male mice. Data represent the mean ± SEM of determinations in five to eight mice. *, P < .05, **, P < .01, ***, P < .001, as determined by a one-way-ANOVA followed by Tukey's test.

Figure 5.

Thyroid status parameters of mice of all four genotypes at age P21. Serum T₃ (A), serum T₄ (B), serum TSH (C), hepatic Dio1 mRNA (D), hypothalamic Hr mRNA (E), and cerebrocortical Hr mRNA (F) in P21 WT, MCT8KO, D3/MCT8 double-KO, and D3KO male mice. Data represent the mean ± SEM of determinations in six to eight mice. *, P < .05, **, P < .01, ***, P < .001, as determined by a one-way-ANOVA followed by Tukey's test.

Figure 6.

Thyroid status parameters of mice of all four genotypes at age P90. Serum T₃ (A), serum T₄ (B), serum TSH (C), hepatic Dio1 mRNA (D), hypothalamic Hr mRNA (E), and cerebrocortical Hr mRNA (F) in P21 WT, MCT8KO, D3/MCT8 double-KO, and D3KO male mice. Data represent the mean ± SEM of determinations in six to eight mice. *, P < .05, **, P < .01, ***, P < .001, as determined by a one-way-ANOVA followed by Tukey's test.

Figure 7.

Effects of an induced change in TH status on the pituitary and hypothalamus of P120 male mice of all four genotypes. A, Effect of induced hypothyroidism on serum TSH. B, Effect of hypothyroidism and T₃ treatment on pituitary expression of Tshb mRNA. C and D, Effect of T₃ treatment on Hr mRNA expression in the pituitary and hypothalamus, respectively. Data represent the mean ± SEM of determinations in four to seven mice. *, P < .05, **, P < .01, ***, P < .001, as determined by a one-way-ANOVA followed by Tukey's test (A and B) or by the Student's t test (C and D).

Figure 8.

Thyroid gland size in short-term induced hypothyroidism. A, Representative transversal midsections of paraffin-embedded thyroid glands of mice with induced hypothyroidism. Scale bar, 100 μm. B, Representative photographs of the thyroid gland of P120 mice with induced hypothyroidism. Scale bar, 5 mm. Pictures were taken laterally and only one thyroid lobe is shown, as indicated by the arrow. C, Quantification of midsection areas (see Materials and Methods) of thyroid glands after short-term induced hypothyroidism. *, P < .05, **, P < .01, ***, P < .001, as determined by a one-way-ANOVA followed by Tukey's test.

Figure 9.

Ontogeny of thyroid status parameters. Summary of data representing serum T4 (A), serum T3 (B), serum TSH (C), hepatic Dio1 mRNA (D), and hypothalamic Hr mRNA (E) across developmental stages. Data for each genotype is represented as a percentage of the values obtained for WT mice (dotted line).