Characterization of TGF-beta inhibition of an in-vitro 3D mouse model of early post-implantation development

TGF-β signaling, in specific Nodal, is known to play a crucial role during early mouse embryonic development, regulating tissue patterning and axis formation. Using ETiX-embryoids I investigated the effects of TGF-β signaling inhibition, using small molecule inhibitor A83-01, on peri-implantation and early post-implantation stages. Inhibition showed a loss of pluripotency and extraembryonic marker expression, disruption in self-organization and basement membrane assembly, and impairment in lumen formation. These phenotypes were more severe than those observed in previous studies with Nodal mutants, suggesting that other members of the TGF-β family are involved during these stages. Results suggested that TGF-β signaling inhibition may be reinforcing cell-cell adhesion and disrupting polarization, leading to an impaired lumenogenesis. Further studies on the basement membrane and cell-cell adhesion would be required to investigate further into this hypothesis. Additionally, I observed that inhibition in ex utero cultured mouse embryos resulted in comparable phenotypes than those observed in the ETiX-embryoids, validating the use of this model to study early development and perform experimental manipulations. Future studies using knock-out lines and conditional KO approaches will be essential to further understand the precise role of TGF-β in lineage maintenance and self-organization.