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. 2018 Feb 15;131(4):jcs208728. doi: 10.1242/jcs.208728

Fig. 5.

Fig. 5.

Complex laminin and FGF2-dependent salivary proacinar organoids require mesenchymal cells. (A) Schematic demonstrating isolation and culture of enriched or purified E16 epithelial clusters. GF, growth factor. (B) E16 enriched epithelial clusters were seeded in laminin-111 on porous polycarbonate filters floating on simple medium containing either FGF2 or EGF. (C) Brightfield images show enriched epithelial clusters or purified epithelial clusters after mesenchyme depletion cultured in Matrigel with FGF2 for 7 days. (D,E) E16 enriched epithelial clusters or E16 purified epithelial clusters (-Mes) were seeded in Matrigel and grown with FGF2. Laminin-111, FGF2 and mesenchyme when together lead to AQP5 expression being retained in organoids, as shown by staining for the markers EpCAM (epithelium, green), AQP5 (proacinar/acinar cells, red) and vimentin (mesenchyme, cyan) with DAPI (nuclei, blue). Quantitative analysis demonstrates (F) EpCAM is variably preserved in all conditions, while (G) AQP5 is retained with FGF2 and basement membrane only when mesenchyme is also present. *P<0.05, **P<0.01, ***P<0.001 (one-way ANOVA with Tukey post-hoc test between each condition). n=3 experiments. Scale bars: 50 μm (B,D); 200 μm (C).