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Propulsion and navigation within the advancing monolayer sheet.

Kim JH, Serra-Picamal X, Tambe DT, Zhou EH, Park CY, Sadati M, Park JA, Krishnan R, Gweon B, Millet E, Butler JP, Trepat X, Fredberg JJ - Nat Mater (2013)

Bottom Line: Here we show that such a relationship between motion and stress is far from direct.Using monolayer stress microscopy, we probed migration velocities, cellular tractions and intercellular stresses in an epithelial cell sheet advancing towards an island on which cells cannot adhere.We found that cells located near the island exert tractions that pull systematically towards this island regardless of whether the cells approach the island, migrate tangentially along its edge, or paradoxically, recede from it.

View Article: PubMed Central - PubMed

Affiliation: School of Public Health, Harvard University, Boston, Massachusetts 02115, USA.

ABSTRACT
As a wound heals, or a body plan forms, or a tumour invades, observed cellular motions within the advancing cell swarm are thought to stem from yet to be observed physical stresses that act in some direct and causal mechanical fashion. Here we show that such a relationship between motion and stress is far from direct. Using monolayer stress microscopy, we probed migration velocities, cellular tractions and intercellular stresses in an epithelial cell sheet advancing towards an island on which cells cannot adhere. We found that cells located near the island exert tractions that pull systematically towards this island regardless of whether the cells approach the island, migrate tangentially along its edge, or paradoxically, recede from it. This unanticipated cell-patterning motif, which we call kenotaxis, represents the robust and systematic mechanical drive of the cellular collective to fill unfilled space.

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Kenotactic tractions are evident in human mammary epithelial cells MCF10A vector, but are attenuated in MCF10A 14–3–3ζ, which disrupts adherens junctionsA,E: Phase contrast images of nontransformed human mammary epithelial cell line, MCF10A, vector control (A) and cells overexpressing 14–3–3ζ which have decreased expression of cell-cell junctional markers (E)23. B,F: Traction vectors, <T⃗>, averaged over an ensemble of 4 monolayers corresponding to cell types in panels (A,E) (see Methods). C,G: Color maps of x-component of tractions,<Tx>. D,H: Color maps of tractions normal to the frustrated edge, <Tn>. In case of nontransformed MCF10A vector cells, tractions near the frustrated edge are largest and oriented toward the edge (B,C,D). In case of MCF10A 14–3–3ζ cells, however, both the magnitude and alignment of tractions near the edge are attenuated (F,G,H). I: Normal component of tractions at the frustrated edge normalized by root-mean-square (RMS) traction across the entire maps, , for three cell types, MDCK (black), MCF10A vector (blue) and MCF10A 14–3–3ζ cells (red) (see Methods). *:  of 14–3–3ζ transfected MCF10a cells is smaller than that of vector-transfected MCF10A cells or that of MDCK cells (mean +/− standard error of the mean; p< 0.05 by Kruskal-Wallis test). J: The alignment angle, φ, between traction vectors at the frustrated edge and normal vectors to the edge for three cell types in panel (I). MDCK and MCF10A vector cells are seen to exert tractions highly oriented toward the frustrated edge, which are largest at that edge (I, J). In contrast, MCF10A 14–3–3ζ cells exert tractions in smaller extent toward the edge, the alignment angle of which are widely distributed, as if they are not frustrated by the edge (I, J). Scale bar in panels (A,E): 100μm. Each bar in (I) include observations from 6 monolayers of MDCK cells and 4 monolayers per each MCF10A cell type. Distributions in (J) have more than 7,000 observations.
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Figure 4: Kenotactic tractions are evident in human mammary epithelial cells MCF10A vector, but are attenuated in MCF10A 14–3–3ζ, which disrupts adherens junctionsA,E: Phase contrast images of nontransformed human mammary epithelial cell line, MCF10A, vector control (A) and cells overexpressing 14–3–3ζ which have decreased expression of cell-cell junctional markers (E)23. B,F: Traction vectors, <T⃗>, averaged over an ensemble of 4 monolayers corresponding to cell types in panels (A,E) (see Methods). C,G: Color maps of x-component of tractions,<Tx>. D,H: Color maps of tractions normal to the frustrated edge, <Tn>. In case of nontransformed MCF10A vector cells, tractions near the frustrated edge are largest and oriented toward the edge (B,C,D). In case of MCF10A 14–3–3ζ cells, however, both the magnitude and alignment of tractions near the edge are attenuated (F,G,H). I: Normal component of tractions at the frustrated edge normalized by root-mean-square (RMS) traction across the entire maps, , for three cell types, MDCK (black), MCF10A vector (blue) and MCF10A 14–3–3ζ cells (red) (see Methods). *: of 14–3–3ζ transfected MCF10a cells is smaller than that of vector-transfected MCF10A cells or that of MDCK cells (mean +/− standard error of the mean; p< 0.05 by Kruskal-Wallis test). J: The alignment angle, φ, between traction vectors at the frustrated edge and normal vectors to the edge for three cell types in panel (I). MDCK and MCF10A vector cells are seen to exert tractions highly oriented toward the frustrated edge, which are largest at that edge (I, J). In contrast, MCF10A 14–3–3ζ cells exert tractions in smaller extent toward the edge, the alignment angle of which are widely distributed, as if they are not frustrated by the edge (I, J). Scale bar in panels (A,E): 100μm. Each bar in (I) include observations from 6 monolayers of MDCK cells and 4 monolayers per each MCF10A cell type. Distributions in (J) have more than 7,000 observations.

Mentions: These findings seem not be restricted to our particular choice of experimental system. For example, when using rat pulmonary microvascular endothelial (RPME) cells22, which are spindle-shaped, the same kenotactic motif was evident (Supplementary Fig. S6). When using MCF10A mammary epithelial cells, and when overexpressing empty vector in those cells (Fig. 4A–D), the same motif was again evident, although overexpressing the oncogene 14–3–3ζ (Fig. 4E–H), which disrupts adherens junctions23, caused tractions near the frustrated edge to become not only smaller (Fig. 4I) but also less well aligned toward the frustrated edge (Fig. 4J). When we inhibited myosin II using blebbistatin (25μM), tractions far from the island decreased dramatically and the bare island, which comprises elastic gel, recoiled centripetally, thus indicating of release of monolayer tension; tractions near the frustrated edge were attenuated but remained well aligned toward the frustrated edge (Supplementary Fig. S7). Finally, using a crescent-like island shape, tractions vectors were seen to align toward the frustrated edge in a manner that was indifferent to the sign of edge curvature (Supplementary Fig. S8). Accordingly, kenotaxis is not to be confused with any mechanism of wound closure that is driven by hoop tension acting through the law of Laplace over some positive (convex) local radius of curvature, as in the purse-string mechanism.


Propulsion and navigation within the advancing monolayer sheet.

Kim JH, Serra-Picamal X, Tambe DT, Zhou EH, Park CY, Sadati M, Park JA, Krishnan R, Gweon B, Millet E, Butler JP, Trepat X, Fredberg JJ - Nat Mater (2013)

Kenotactic tractions are evident in human mammary epithelial cells MCF10A vector, but are attenuated in MCF10A 14–3–3ζ, which disrupts adherens junctionsA,E: Phase contrast images of nontransformed human mammary epithelial cell line, MCF10A, vector control (A) and cells overexpressing 14–3–3ζ which have decreased expression of cell-cell junctional markers (E)23. B,F: Traction vectors, <T⃗>, averaged over an ensemble of 4 monolayers corresponding to cell types in panels (A,E) (see Methods). C,G: Color maps of x-component of tractions,<Tx>. D,H: Color maps of tractions normal to the frustrated edge, <Tn>. In case of nontransformed MCF10A vector cells, tractions near the frustrated edge are largest and oriented toward the edge (B,C,D). In case of MCF10A 14–3–3ζ cells, however, both the magnitude and alignment of tractions near the edge are attenuated (F,G,H). I: Normal component of tractions at the frustrated edge normalized by root-mean-square (RMS) traction across the entire maps, , for three cell types, MDCK (black), MCF10A vector (blue) and MCF10A 14–3–3ζ cells (red) (see Methods). *:  of 14–3–3ζ transfected MCF10a cells is smaller than that of vector-transfected MCF10A cells or that of MDCK cells (mean +/− standard error of the mean; p< 0.05 by Kruskal-Wallis test). J: The alignment angle, φ, between traction vectors at the frustrated edge and normal vectors to the edge for three cell types in panel (I). MDCK and MCF10A vector cells are seen to exert tractions highly oriented toward the frustrated edge, which are largest at that edge (I, J). In contrast, MCF10A 14–3–3ζ cells exert tractions in smaller extent toward the edge, the alignment angle of which are widely distributed, as if they are not frustrated by the edge (I, J). Scale bar in panels (A,E): 100μm. Each bar in (I) include observations from 6 monolayers of MDCK cells and 4 monolayers per each MCF10A cell type. Distributions in (J) have more than 7,000 observations.
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Figure 4: Kenotactic tractions are evident in human mammary epithelial cells MCF10A vector, but are attenuated in MCF10A 14–3–3ζ, which disrupts adherens junctionsA,E: Phase contrast images of nontransformed human mammary epithelial cell line, MCF10A, vector control (A) and cells overexpressing 14–3–3ζ which have decreased expression of cell-cell junctional markers (E)23. B,F: Traction vectors, <T⃗>, averaged over an ensemble of 4 monolayers corresponding to cell types in panels (A,E) (see Methods). C,G: Color maps of x-component of tractions,<Tx>. D,H: Color maps of tractions normal to the frustrated edge, <Tn>. In case of nontransformed MCF10A vector cells, tractions near the frustrated edge are largest and oriented toward the edge (B,C,D). In case of MCF10A 14–3–3ζ cells, however, both the magnitude and alignment of tractions near the edge are attenuated (F,G,H). I: Normal component of tractions at the frustrated edge normalized by root-mean-square (RMS) traction across the entire maps, , for three cell types, MDCK (black), MCF10A vector (blue) and MCF10A 14–3–3ζ cells (red) (see Methods). *: of 14–3–3ζ transfected MCF10a cells is smaller than that of vector-transfected MCF10A cells or that of MDCK cells (mean +/− standard error of the mean; p< 0.05 by Kruskal-Wallis test). J: The alignment angle, φ, between traction vectors at the frustrated edge and normal vectors to the edge for three cell types in panel (I). MDCK and MCF10A vector cells are seen to exert tractions highly oriented toward the frustrated edge, which are largest at that edge (I, J). In contrast, MCF10A 14–3–3ζ cells exert tractions in smaller extent toward the edge, the alignment angle of which are widely distributed, as if they are not frustrated by the edge (I, J). Scale bar in panels (A,E): 100μm. Each bar in (I) include observations from 6 monolayers of MDCK cells and 4 monolayers per each MCF10A cell type. Distributions in (J) have more than 7,000 observations.
Mentions: These findings seem not be restricted to our particular choice of experimental system. For example, when using rat pulmonary microvascular endothelial (RPME) cells22, which are spindle-shaped, the same kenotactic motif was evident (Supplementary Fig. S6). When using MCF10A mammary epithelial cells, and when overexpressing empty vector in those cells (Fig. 4A–D), the same motif was again evident, although overexpressing the oncogene 14–3–3ζ (Fig. 4E–H), which disrupts adherens junctions23, caused tractions near the frustrated edge to become not only smaller (Fig. 4I) but also less well aligned toward the frustrated edge (Fig. 4J). When we inhibited myosin II using blebbistatin (25μM), tractions far from the island decreased dramatically and the bare island, which comprises elastic gel, recoiled centripetally, thus indicating of release of monolayer tension; tractions near the frustrated edge were attenuated but remained well aligned toward the frustrated edge (Supplementary Fig. S7). Finally, using a crescent-like island shape, tractions vectors were seen to align toward the frustrated edge in a manner that was indifferent to the sign of edge curvature (Supplementary Fig. S8). Accordingly, kenotaxis is not to be confused with any mechanism of wound closure that is driven by hoop tension acting through the law of Laplace over some positive (convex) local radius of curvature, as in the purse-string mechanism.

Bottom Line: Here we show that such a relationship between motion and stress is far from direct.Using monolayer stress microscopy, we probed migration velocities, cellular tractions and intercellular stresses in an epithelial cell sheet advancing towards an island on which cells cannot adhere.We found that cells located near the island exert tractions that pull systematically towards this island regardless of whether the cells approach the island, migrate tangentially along its edge, or paradoxically, recede from it.

View Article: PubMed Central - PubMed

Affiliation: School of Public Health, Harvard University, Boston, Massachusetts 02115, USA.

ABSTRACT
As a wound heals, or a body plan forms, or a tumour invades, observed cellular motions within the advancing cell swarm are thought to stem from yet to be observed physical stresses that act in some direct and causal mechanical fashion. Here we show that such a relationship between motion and stress is far from direct. Using monolayer stress microscopy, we probed migration velocities, cellular tractions and intercellular stresses in an epithelial cell sheet advancing towards an island on which cells cannot adhere. We found that cells located near the island exert tractions that pull systematically towards this island regardless of whether the cells approach the island, migrate tangentially along its edge, or paradoxically, recede from it. This unanticipated cell-patterning motif, which we call kenotaxis, represents the robust and systematic mechanical drive of the cellular collective to fill unfilled space.

Show MeSH
Related in: MedlinePlus