RA-induced prominence-specific response resulted in distinctive regulation of Wnt and osteogenesis [miRNA-seq]

Proper retinoic acid (RA) signaling is essential for normal craniofacial development. Both excessive RA and RA deficiency in early embryonic stage led to a variety of craniofacial malformations, e.g., cleft palate, which have been investigated extensively. Dysregulated Wnt and Shh signaling were shown to underlie the pathogenesis of RA-induced craniofacial defects. In our present study, we showed a spatiotemporal-specific effect of RA signaling in regulating early development of facial prominences. While inhibited the Wnt activities in E12.5/E13.5 mouse palatal shelves, early exposure of excessive RA induced Wnt signaling and Wnt-related gene expression in mouse embryonic Frontonasal/Maxillary processes. A conserved regulatory network of miR-484-Fzd5 was identified to play critical roles in RA regulated craniofacial development using RNA-seq. Overall design: In this work we investigated the changes in gene expression profile in the developing craniofacial prominences of RA induced CLP mouse model. Multiple dosages of all-trans RA (atRA) were applied to pregnant mice between early gestation (from E8.5 to E 10.5) to induce full penetrance of CL/P. The frontonasal prominence (FnP), a pair of maxillary processes (MxP) and a pair of mandibular processes (MdP) at E12.5 were collected and subjected to miRNA-sequencing. To minimize the chance of variance due to casualness in sampling, each of the fetus was collected from an independent conception with (n=4) or without RA exposure (n=4). The dysregulated craniofacial development was confirmed by morphological analysis before sampling.