Fig. 2.

Soluble factors from primary salivary mesenchyme cannot substitute for mesenchyme cells to support AQP5 proacinar epithelial cells in culture. (A) Schematic of initial conditions for epithelial clusters cultured without direct contact with the mesenchyme when separated by a filter or when cultured with mesenchymal cell conditioned medium for 7 days. (B) E16 epithelial clusters grown in Matrigel on a polycarbonate filter were separated from primary mesenchyme cells, which were seeded on a coverslip below. ICC and confocal imaging revealed loss of AQP5 expression with partial preservation of EpCAM⁺ epithelium after 7 days. (C) E16 epithelial clusters were grown in Matrigel or laminin-111 in the presence of concentrated conditioned medium (cm) collected from E16 primary salivary mesenchyme. The conditioned medium (D) partially preserved the EpCAM⁺ epithelial phenotype in the presence of Matrigel (mat) or laminin (lam) but (E) poorly rescued AQP5 expression, as determined by staining for the markers EpCAM (epithelium, green) and AQP5 (proacinar/acinar cells, red) with DAPI (nuclei, blue). **P<0.01, ***P<0.001 (one-way ANOVA with a Tukey post-hoc test was applied to determine the statistically relevant differences between each condition). n=3 experiments. Epi, epithelial clusters; Mes, mesenchyme. Scale bars: 50 μm.