Maternal Thyroid Hormones are transcriptionally active during embryo-fetal development

Thyroid hormones (THs) are essential for the metabolic homeostasis of organs and tissues in mammals as well as lower organisms. Most of their biological actions are mediated by thyroid hormone receptors (TRs) binding to TH-responsive elements (TREs) in the regulatory regions of target genes. T4 and T3 have been detected in the rat embryo and fetal brain before the onset of fetal thyroid function (FTF), whereas nuclear T3 receptor has been found in human fetal brain at early and mid gestation: maternal THs can interact with embryonic TRs. This has been supported by the finding of maternal T4, essential for the development of the brain and other foetal organs, in coelomic and amniotic fluid. The direct evidence of their interaction with embryonic TRs during the early and late stages of embryogenesis was still lacking. To address this issue, a transgenic mouse model expressing ubiquitously the LacZ reporter gene (encoding beta-galactosidase [beta-gal]) under a TRE has been generated to trace maternal TH transcription activity during early and late embryo-fetal development. Euthyroid transient transgenic embryos, in vivo, did not show any betagal-staining at embryonic day (E) 9.5, whereas it was observed as early as E11.5- 12.5 in different primordia (CNS, sense organs, intestine) while the FTF is still inactive. At E11.5-12.5, the nuclear receptor TR alpha1 IHC shows an intense labelling in the central nervous system primordia and colocalizes with betagal-staining: the detection of betagal expression strengthens the idea that maternal THs might play a critical role in cell migration and formation of neuronal layer, as well as in neuronal and glial cell differentiation and synaptogenesis. The telencephalic vesicles are labelled by TRalpha1 IHC but not by betagal, while TRbeta1 IHC displays a similar intense labeling of cell bodies in the CNS with exception of the spinal cord. The TRs IHC staining observed is intense and colocalizes with betagal-staining not only in small intestine, follicular of the vibrissae and in the cartilagine primordial. In the cerebral cortex, TRalpha1 is detected in the cortical plate and in the subventricular zone, but an intense immunoreactivity is also observed in the hippocampus, in the striatum, in the developing ear and spinal cord. These findings support the idea that maternal THs cross the placental barrier and are transcriptionally active during embryo-fetal development. At late stage of development, when the FTF (E17.5) is active, beta gal expression is detected in different primordia compared to its expression in the early stages (small intestine, follicular nerves of the vibrissae, bones, skin, muscle, blood vessels). Furthermore, to confirm that betagal-staining was due to maternal THs, we induced a hypothyroidism status in TRE2x transgenic female mice by treatment with 0.1% MMI and 1% KClO4 solution in addition to a low iodine diet (LID) and the minimal detectable concentrations TSH, total T3 and T4 assays confirmed it. As expected, betagal-staining was undetectable at early stages (E12.5) in embryos from hypothyroid female mice, while TRs’ expression was unchanged. Importantly, treatment with T3 in the hypothyroid mice rescued betagal-expression in the diencephalum promordium of E12.5 transgenic embryos, producing a similar pattern to the observed in the embryos from euthyroid mothers: T3 is able to switch on the TRE2x transactivation and betagal-expression at the early stage of development when FTF is still not active. This study shows a tissue-specific expression of thyroid hormones receptors and the synergic action between maternal TH and embryonic TRs, regulating the transcription of target genes involved in the development of CNS, intestine and other embryo-fetal primordia before FTF is active; further, we showed that betagal expression is completely absent under hypothyroidism despite the presence of TRs and is rescued with T3 treatment. Our transgenic mouse may be a reliable tool to investigate molecular mechanisms underlying maternal TH action during early and late embryo-fetal development representing a model to analyse maternal thyroid failure like maternal hypothyroidism, transient gestational hypothyroxinemia or hyperthyroidism (high levels of T3 and T4).