Resumo

Cerebellar development is greatly affected by thyroid hormones (THs) and their levels influence its ontogenesis and cellular maturation. The morphogen sonic hedgehog (Shh) was demonstrated to act in a similar way, being an important factor for cerebellum development, regulating mainly neuronal proliferation, however the role in astroglial cells remains unknown. Here, we investigate the role of Shh in astroglial cells (ACs) and the interaction of this signaling pathway with THs. Our in vivo analysis demonstrated that ACs present Shh signaling pathway components, including its receptors, Ptch and Smo, and its downstream effectors, Gli1, Gli2 and Gli3. Ptch was mainly localized in immature ACs (nestin+ cells) suggesting a potential role of Shh signaling in ACs development. Further, nuclear translocation of Gli1, a hallmark of Shh pathway activation, was mainly identified in migratory ACs, suggesting function for this pathway in astroglial migration. Gli2 was also localized in ACs at P0, but its colocalization at P7 is negligible. In hypothyroid cerebella, we observed diminished Gli2 translocation in all cells, whether ACs or neurons. In vitro, we observed that astrocytes do present Shh signaling pathway components. Shh-treated ACs increase proliferation (Ki67 and 3H thymidine assays; n=6; p<0,0001), cellular viability (n=7; p=0,0176) and cell area (n=3; p=0,0003). Blocking Shh function promotes cell death (n=3). Interaction between Shh and THs was analyzed by exposing ACs cultures to T3 or T4. THs greatly increased ACs proliferation, especially T4 (n=6; p<0,0001). Further, immunocolocalization and Western blotting assays revealed that T4 increases Gli1 nuclear translocation (n=6; p<0,0001), in contrast to T3 that reduces it. Taken together, our results suggest a novel role for Shh signaling pathway in cerebellar development, markedly modulated by THs, and corroborate for a role of THs as pivotal orchestrators of cerebellar ACs development.