Fig. 4.

Normal epithelium transiently protrudes and disseminates into collagen I but reestablishes a complete basement membrane. (A) Representative frames of a DIC time-lapse movie of a normal organoid grown in collagen I. (A′) Higher magnification of a transition from a protrusive to a smooth border with ECM. (A′′) Higher magnification of the reintegration of disseminated cells with the epithelial group. (B) Representative frames from a confocal time-lapse movie of a normal organoid grown in collagen I. (B′) Higher magnification of transient protrusions within single myoepithelial cells. Arrows indicate individual protrusions, retractions, and epithelial reorganization. (B′′) Higher magnification of a multicellular extension of myoepithelial cells (blue arrows) at the leading front. (C–G) Normal epithelia in collagen reform a multicomponent basement membrane. (C–D′ and F and F′) Localization of actin, DAPI, and laminin 111 in a merge of all channels (C, D, and F) and in a single channel of laminin 111 (C′, D′, and F′) in a normal organoid with a single-cell protrusion (C′), a multicellular extension (D′), and a normal organoid after reorganization (F′). All the Insets highlight negative correlation of cell protrusion (C, D, and F) and laminin 111 (C′, D′, and F′) at the leading front. (E and E′ and G and G′) Localization of actin, DAPI, and collagen IV in a merge of all channels (E and G) and in a single channel of collagen IV in a normal organoid (E′ and G′) with multicellular extensions (E′) and after reorganization (G′). (H and H′) Localization of DAPI and laminin 332 in a merge of two channels (H) and a single channel of laminin 332 (H′). (I–K′) Tumor epithelia display incomplete basement membrane coverage. Single channels show the localization of actin (I, J, and K), laminin 111(I′), collagen IV (J′), and laminin 332 (K′) in tumor organoids in collagen I. Red and green arrowheads indicate actin-based protrusions and signals of basement membrane components, respectively.