Figure - PMC

Skip to main content

An official website of the United States government

Here's how you know

Here's how you know

Official websites use .gov
A .gov website belongs to an official government organization in the United States.

Secure .gov websites use HTTPS
A lock ( ) or https:// means you've safely connected to the .gov website. Share sensitive information only on official, secure websites.

View full-text article in PMC

. 2016 Feb 1;27(3):451–465. doi: 10.1091/mbc.E15-09-0615

Search in PMC
Search in PubMed
View in NLM Catalog
Add to search

© 2016 Bleaken et al. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).

“ASCB®,” “The American Society for Cell Biology®,” and “Molecular Biology of the Cell®” are registered trademarks of The American Society for Cell Biology.

PMC Copyright notice

FIGURE 7: — Functional properties of invading cells in response to injury. (A–G) The 3D Matrigel invasion assay of ex vivo–wounded explants at (A–D) day 6 in culture and (E–G) day 2 in culture. (A) A 3D reconstruction of a confocal Z-stack of cords of cells in the injury invasion assay labeled for vimentin (green) and F-actin (red), emphasizing the extension of vimentin-rich cells into the Matrigel matrix in leader-cell positions. Bar, 30 μm. (B, C) Cords of invading cells in the Matrigel invasion assay were immunolabeled for cortactin (red) and vimentin (green) and viewed by confocal microscopy. This projection image of a confocal Z-stack shows that the processes extended by the vimentin-rich mesenchymal cells into the Matrigel also expressed cortactin (red). Bar, 5 μm. (D) Labeling for MMP-9 (green) in cells at the leading edges of the cords extended into Matrigel and was distributed in discrete vesicular structures in the cells (F-actin; red) and also in a punctate pattern just beyond the cells in the Matrigel matrix, where it can act to degrade the matrix for invasion. A representative single optical plane captured by confocal microscopy. Bar, 10 μm. (E–G) Vimentin-rich cells at the invading front (green) also express αSMA (red), often with a punctate distribution (arrowhead). Labeling for F-actin (purple) was mostly absent from the areas with a strong distribution of αSMA puncta, suggesting that this actin population was nonfilamentous. A representative single optical plane captured by confocal microscopy. Bar, 10 μm. (H) The individual roles of MMP-2/9 and vimentin in invasion after wounding were evaluated using an inhibitor approach in the Matrigel-Transwell chemoinvasion assay. This study included the MMP-2 inhibitor MMP-2i, the MMP-9 inhibitor MMP-9i, the combined MMP-2/9 inhibitor MMP-2/9i, and the vimentin inhibitor WFA. Invaded cells from at least 10 capsules were counted and quantified using MetaMorph or NIS Elements image analysis software. Relative numbers of invaded cells were plotted on graphs ±SEM. MMP-2 and MMP-9 function were required for invasion of cells in the Matrigel-Transwell assay. MMP-2 inhibited invasion by 99%, MMP-9 by 96%, and MMP2/9 by 84% in the Matrigel-Transwell assays. Inhibiting vimentin function with WFA suppressed invasion in the Matrigel-Transwell assay by 98%.