Endocrine Abstracts

Anaemia in RTHα patients correlates with documented abnormal erythropoiesis and reduced haematocrit in TRα-null or mutant mice, but the underlying mechanisms and characterization of the defect remain elusive. In a previous work, we described that the injection of several hTRα variants in zebrafish eggs caused a marked reduction of circulating erythrocytes at later stages of the erythropoiesis. To characterise the involvement of TRα during erythrocyte development, we analysed by qRT-PCR and in situ hybridization (ISH) the expression of different haematopoietic markers. We microinjected the zebrafish eggs at 1–2 cells stage with 80pg/embryos of the purified mRNAs of several hTRα variants, and from the 6-hpf, the injected embryos were treated with the control vehicle, or with 20 μM-T3, added in the harvested water. By qRT-PCR, the expression of genes involved in the specification of lateral mesoderm (cdx4, bmp2b, smad5, chd), induction and proliferation of haematopoietic stem cells (HSC) (tal1, lmo2, gata1, gata2) and proliferation of erythrocytes (zin, grx5, cia, cha) appeared significantly reduced in all hTRα-injected embryos. Consistent with previous results, the treatment with high T3 doses can rescue the DN-activity of the missense variants D211G and A263V, whereas the truncated receptors A382PfsX7, E403X and F397fs406X are unaffected by TH treatment. Furthermore, we analysed the specification of HSC in embryos at 8-somite stage, by ISH. The vast majority of the hTRα-injected embryos showed a reduced or undetectable expression of early erythropoiesis markers, such as c-myb, pu.1, tal1 and gata1 in anterior or posterior lateral mesoderm. At 36-hpf, we also observed a dramatic reduction of gata1 in the intermediated cell mass of hTRα-injected embryos. Therefore, our results highlight a relevant role for TRα during the early phases of the primitive haematopoiesis, when the HSC start to proliferate and differentiate.

Research founding: IAI-Ricerca Corrente RF-2011 05C102.