Figure 9: Cell contractility, and epithelial and substrate topography participate in tuft formation in the absence of EpCAM.
From: Contractile forces at tricellular contacts modulate epithelial organization and monolayer integrity
(a) Statistical analysis of the number of tuft-like structures detected along the 3D microfabricated villus structures in control (Caco2 shNT), EpCAM-depleted cells (Caco2 shEpCAM) and 2h- blebbistatin-treated EpCAM-depleted cells (Caco2 shEpCAM#1+Blebbistatin 50 μM). Three independent replicates have been performed. One-way analysis of variance with unpaired t-test, *P<0.0001. n(Caco2 shNT)=196 villi, n(Caco2 shEpCAM)=210 villi, n(Caco2 shEpCAM+Blebistatin)=210 villi. Caco2 shNT(mean number of tufts per villus)=5, Caco2 shEpCAM(mean number of tufts per villus)=12, Caco2 shEpCAM+Blebbistatin 50 μM(mean number of tufts per villus)=6. (b–d) Confocal microscopy analysis of myosin-IIa (b), myosin-IIb (c) and P-MLC2 (d) distribution in control (Caco2 shNT) and EpCAM-silenced (Caco2 shEpCAM) cells that were grown on villous PDMS inserts or in 3D Matrigel cultures for 21 days. Transversal xy views are presented. Scale bars, 50 μm. (e) Schemes recapitulating the cellular and epithelial phenotypes observed in control (Caco2 shNT) or EpCAM-depleted (Caco2 shEpCAM) conditions when cells are grown in 2D, 3D synthetic villi or 3D Matrigel cultures. Contractile apparatus (blue) and predictated tensile forces (red arrows) are presented.
