Conclusions
The addition of 1 ng/ml TGF-β1 maintained the developmental potential of embryonic SMG epithelia separated from mesenchyme for 24 h. This suggests that 1 ng/ml TGF-β1 may partially compensate for the role of mesenchyme during the separation phase, although its compensation is limited in extent.
Methods
The ex vivo model was established by separating E13 mouse SMG epithelia and mesenchyme, culturing them independently for 24 h, recombining them, and observing branching morphogenesis. Microarray analysis was performed to evaluate the transcriptome of epithelia treated with and without 1 ng/ml TGF-β1. Differential gene expression, pathway enrichment, and protein-protein interaction networks were analyzed. Quantitative real-time polymerase chain reaction, Western blot, and immunofluorescence were employed to validate the mRNA and protein levels.
Objective
The objective of this study was to investigate the impact of transforming growth factor β1 (TGF-β1) on epithelial development using an ex vivo model of submandibular gland (SMG) epithelial-mesenchymal separation. Materials and
Results
Recombined SMGs using separated epithelia and mesenchyme that were cultured for 24 h showed a significant inhibition of epithelial development compared to SMGs recombined immediately after separation. The level of TGF-β1 decreased in the SMG epithelia after epithelia-mesenchyme separation. Epithelia that were separated from mesenchyme for 24 h and pretreated with 1 ng/ml TGF-β1 continued to develop after recombination with mesenchyme, while epithelia without 1 ng/ml TGF-β1 treatment did not. Microarray analysis suggested pathway enrichment related to epithelial development and an upregulation of Sox2 in the 1 ng/ml TGF-β1-treated epithelia. Further experiments validated the phosphorylation of SMAD2 and SMAD3, upregulation of SOX2 and genes associated with epithelial development, including Prol1, Dcpp1, Bhlha15, Smgc, and Bpifa2. Additionally, 1 ng/ml TGF-β1 inhibited epithelial apoptosis by improving the BCL2/BAX ratio and reducing cleaved caspase 3. Conclusions: The addition of 1 ng/ml TGF-β1 maintained the developmental potential of embryonic SMG epithelia separated from mesenchyme for 24 h. This suggests that 1 ng/ml TGF-β1 may partially compensate for the role of mesenchyme during the separation phase, although its compensation is limited in extent.