Fig. 6.

SMG initiation is enhanced by addition of retinol and blocked by pharmacological inhibitor of RARs. (A-C) Mandibular explants were cultured with either vehicle control (A), 10 µM retinol (B) or 5 µM BMS493 (C), followed by whole-mount E-cadherin immunostaining and confocal microscopy. Bilateral patches of dense E-cadherin staining at base of tongue in control and retinol samples, but not in BMS493 samples, indicates that SMG initiation is blocked by RAR inhibitors. (D-I) Volume projections (D-F) and surface rendering (G-I) through the region of SMG gland initiation (rotated 90° from image plane for frontal view) are shown. (J) SMG epithelium volume quantitation indicates that retinol treatment increases initial bud size, whereas treatment with RAR inhibitor blocks initiation. Bar chart shows the average volume and standard error for each group (n=8 glands from four mandibles for each condition). (K) Contingency table for the presence or absence of initiated glands (TRUE versus FALSE) for each prospective site in the experiment (n=8 possible sites in four mandibles for each condition). (L-N) E-cadherin-stained frozen sections of cultured mandible explants reveal that inhibition of RAR blocks epithelial thickening (N). (O) qPCR of Fgf10 RNA in mandibles cultured on control or BMS493-treated medium reveals that treatment with RAR inhibitor causes a mild reduction in Fgf10 expression (n=3 mandibles). Yellow arrowheads indicate epithelial invagination into underlying mesenchyme, yellow asterisks indicate absence of epithelial invagination. *P<0.05. Scale bars: 100 µm.