Endocrine Abstracts

Introduction: Since 2012, different heterozygous mutations in the THRA gene have been described in patients with resistance to thyroid hormone alpha (RTHα). The associating symptoms are reminiscent of untreated congenital hypothyroidism (growth retardation, psycho-neuromotor disorders, delayed bone development and bradycardia) but with raised T3/T4 ratio and normal TSH levels. All genetic abnormalities act in a dominant negative (DN) manner against functional receptors due to reduced T3-binding or defective interaction with corepressors or coactivators of the ligand-binding domain. Therefore, RTHα patients present variable sensitivity to TH treatment. We previously described that zebrafish embryos expressing a DN form of thraa recapitulate the key features of RTHα, and that zebrafish and human receptors are functionally interchangeable.

Methods: In this work, we present a simplified model obtained by direct mRNA microinjection into zebrafish eggs of several human THRA variants (D211G, A263V, A382PfsX7, E403X and F397fs406X). Using a series of molecular and analytical approaches (WISH, IHC, ELISA) we studied the embryonic development of cardiovascular, skeletal and nervous systems, which are directly involved in the T3-dependent TRα action. Additionally, thyroid and pituitary function, as well as the T3 and T4 contents were analysed.

Results: All THRA-injected embryos show defects in several tissues of variable entity: altered cardiovascular development, incomplete formation of cartilages and bones, reduced number motorneurons that is reflected by defective zebrafish locomotion. Furthermore, changes in deiodinases expression (high dio2, low dio3a and dio3b levels) in presence of normal thyroid and pituitary markers, could explain the high T3/T4 ratio observed in THRA-injecting embryos, as in RTHα cases. Interestingly, the treatment with high T3 dose efficiently rescue most of these defects only in D211G- and A263V-injected embryos, conserving part of the T3-binding activity and transactivation properties.

Conclusion: Zebrafish represent a new ‘biotool’ to delineate the functional impact of new THRA variants in vivo.