Limits...
Dynamic tensile forces drive collective cell migration through three-dimensional extracellular matrices.

Gjorevski N, Piotrowski AS, Varner VD, Nelson CM - Sci Rep (2015)

Bottom Line: Moreover, cell movements are highly correlated and in phase with ECM deformations.Migrating cohorts use spatially localized, long-range forces and consequent matrix alignment to navigate through the ECM.These results suggest biophysical forces are critical for 3D collective migration.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemical &Biological Engineering, Princeton University, Princeton, NJ 08544, USA.

ABSTRACT
Collective cell migration drives tissue remodeling during development, wound repair, and metastatic invasion. The physical mechanisms by which cells move cohesively through dense three-dimensional (3D) extracellular matrix (ECM) remain incompletely understood. Here, we show directly that migration of multicellular cohorts through collagenous matrices occurs via a dynamic pulling mechanism, the nature of which had only been inferred previously in 3D. Tensile forces increase at the invasive front of cohorts, serving a physical, propelling role as well as a regulatory one by conditioning the cells and matrix for further extension. These forces elicit mechanosensitive signaling within the leading edge and align the ECM, creating microtracks conducive to further migration. Moreover, cell movements are highly correlated and in phase with ECM deformations. Migrating cohorts use spatially localized, long-range forces and consequent matrix alignment to navigate through the ECM. These results suggest biophysical forces are critical for 3D collective migration.

No MeSH data available.


Tensile forces drive collective migration by activating mechanically sensitive intracellular signaling and transcription factors.(a) Phase contrast images showing collective migration from representative tissues treated with DMSO (control) or blebbistatin (12.5 μM). (b) Quantification of cohort length from tissues treated with DMSO or blebbistatin. Mean ± s.e.m. of three replicates is shown, n = 60 tissues per group. **P < 0.01, Student’s t-test. Immunofluorescence staining for FAK pY397 and phospho-p130Cas in representative DMSO (c) and blebbistatin-treated (d) migrating cohorts. (e) Confocal image showing immunofluorescence staining for MRTF-A localization (green) and nuclei (red) in a representative tissue. (f) Quantification of the localization of MRTF-A (nucleus/cytoplasm) in tissues. Levels of nuclear and cytoplasmic MRTF-A were quantified by measuring signal intensity in the two compartments. Mean ± s.e.m. of three replicates is shown. Migrating cohort (invasion): n = 14; quiescent body: n = 11. **P < 0.01, Student’s t-test. (g) Confocal image showing immunofluorescence staining for MRTF-A localization (green) and nuclei (red) in representative tissues treated with DMSO or blebbistatin. (h) Quantification of the localization of MRTF-A (nucleus/cytoplasm) in tissues treated with DMSO or blebbistatin. Levels of nuclear and cytoplasmic MRTF-A were quantified by measuring signal intensity in the two compartments. Mean ± s.e.m. of three replicates is shown. MRTF-A DMSO: n = 29; MRTF-A blebbistatin: n = 21; DAPI DMSO: n = 10; DAPI blebbistatin: n = 12. **P < 0.01, Student’s t-test. (i) Phase contrast images showing collective migration from representative tissues treated with DMSO or CCG-1423 (10 μM). (j) Quantification of cohort length from tissues treated with DMSO or CCG-1423. Mean ± sem. of three replicates is shown. Invading cohorts treated with DMSO: n=80; invading cohorts treated with CCG-1423: n = 57. **P < 0.01, Student’s t-test. (k) Frequency maps showing collective invasion from 34 tissues (three independent replicates) transfected with scrambled shRNA (shScr) or shMRTF-A (two constructs). (l) Quantification of cohort length from tissues transfected with shScr or shMRTF-A. Mean ± s.e.m. of three replicates is shown, n = 50 tissues per group. ***P < 0.001, Student’s t-test. All images are representative of three independent replicates in which >50 tissues were monitored. Scale bars, 50 μm (a,i,k), 25 μm (c,d,e,g).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4499882&req=5

f3: Tensile forces drive collective migration by activating mechanically sensitive intracellular signaling and transcription factors.(a) Phase contrast images showing collective migration from representative tissues treated with DMSO (control) or blebbistatin (12.5 μM). (b) Quantification of cohort length from tissues treated with DMSO or blebbistatin. Mean ± s.e.m. of three replicates is shown, n = 60 tissues per group. **P < 0.01, Student’s t-test. Immunofluorescence staining for FAK pY397 and phospho-p130Cas in representative DMSO (c) and blebbistatin-treated (d) migrating cohorts. (e) Confocal image showing immunofluorescence staining for MRTF-A localization (green) and nuclei (red) in a representative tissue. (f) Quantification of the localization of MRTF-A (nucleus/cytoplasm) in tissues. Levels of nuclear and cytoplasmic MRTF-A were quantified by measuring signal intensity in the two compartments. Mean ± s.e.m. of three replicates is shown. Migrating cohort (invasion): n = 14; quiescent body: n = 11. **P < 0.01, Student’s t-test. (g) Confocal image showing immunofluorescence staining for MRTF-A localization (green) and nuclei (red) in representative tissues treated with DMSO or blebbistatin. (h) Quantification of the localization of MRTF-A (nucleus/cytoplasm) in tissues treated with DMSO or blebbistatin. Levels of nuclear and cytoplasmic MRTF-A were quantified by measuring signal intensity in the two compartments. Mean ± s.e.m. of three replicates is shown. MRTF-A DMSO: n = 29; MRTF-A blebbistatin: n = 21; DAPI DMSO: n = 10; DAPI blebbistatin: n = 12. **P < 0.01, Student’s t-test. (i) Phase contrast images showing collective migration from representative tissues treated with DMSO or CCG-1423 (10 μM). (j) Quantification of cohort length from tissues treated with DMSO or CCG-1423. Mean ± sem. of three replicates is shown. Invading cohorts treated with DMSO: n=80; invading cohorts treated with CCG-1423: n = 57. **P < 0.01, Student’s t-test. (k) Frequency maps showing collective invasion from 34 tissues (three independent replicates) transfected with scrambled shRNA (shScr) or shMRTF-A (two constructs). (l) Quantification of cohort length from tissues transfected with shScr or shMRTF-A. Mean ± s.e.m. of three replicates is shown, n = 50 tissues per group. ***P < 0.001, Student’s t-test. All images are representative of three independent replicates in which >50 tissues were monitored. Scale bars, 50 μm (a,i,k), 25 μm (c,d,e,g).

Mentions: To determine whether active cellular contractility was required for collective migration, we blocked cytoskeletal tension by treating the invading tissues with blebbistatin (Fig. 3a) or Y27632 (Fig. S1a), which inhibit myosin ATPase and Rho kinase, respectively. Disrupting cell-generated forces significantly impaired the extent of collective migration (Fig. 3b; Fig. S1b). Conversely, enhancing cellular contractility by treating with LPA, an activator of Rho, increased the extent of migration (Fig. S1a).


Dynamic tensile forces drive collective cell migration through three-dimensional extracellular matrices.

Gjorevski N, Piotrowski AS, Varner VD, Nelson CM - Sci Rep (2015)

Tensile forces drive collective migration by activating mechanically sensitive intracellular signaling and transcription factors.(a) Phase contrast images showing collective migration from representative tissues treated with DMSO (control) or blebbistatin (12.5 μM). (b) Quantification of cohort length from tissues treated with DMSO or blebbistatin. Mean ± s.e.m. of three replicates is shown, n = 60 tissues per group. **P < 0.01, Student’s t-test. Immunofluorescence staining for FAK pY397 and phospho-p130Cas in representative DMSO (c) and blebbistatin-treated (d) migrating cohorts. (e) Confocal image showing immunofluorescence staining for MRTF-A localization (green) and nuclei (red) in a representative tissue. (f) Quantification of the localization of MRTF-A (nucleus/cytoplasm) in tissues. Levels of nuclear and cytoplasmic MRTF-A were quantified by measuring signal intensity in the two compartments. Mean ± s.e.m. of three replicates is shown. Migrating cohort (invasion): n = 14; quiescent body: n = 11. **P < 0.01, Student’s t-test. (g) Confocal image showing immunofluorescence staining for MRTF-A localization (green) and nuclei (red) in representative tissues treated with DMSO or blebbistatin. (h) Quantification of the localization of MRTF-A (nucleus/cytoplasm) in tissues treated with DMSO or blebbistatin. Levels of nuclear and cytoplasmic MRTF-A were quantified by measuring signal intensity in the two compartments. Mean ± s.e.m. of three replicates is shown. MRTF-A DMSO: n = 29; MRTF-A blebbistatin: n = 21; DAPI DMSO: n = 10; DAPI blebbistatin: n = 12. **P < 0.01, Student’s t-test. (i) Phase contrast images showing collective migration from representative tissues treated with DMSO or CCG-1423 (10 μM). (j) Quantification of cohort length from tissues treated with DMSO or CCG-1423. Mean ± sem. of three replicates is shown. Invading cohorts treated with DMSO: n=80; invading cohorts treated with CCG-1423: n = 57. **P < 0.01, Student’s t-test. (k) Frequency maps showing collective invasion from 34 tissues (three independent replicates) transfected with scrambled shRNA (shScr) or shMRTF-A (two constructs). (l) Quantification of cohort length from tissues transfected with shScr or shMRTF-A. Mean ± s.e.m. of three replicates is shown, n = 50 tissues per group. ***P < 0.001, Student’s t-test. All images are representative of three independent replicates in which >50 tissues were monitored. Scale bars, 50 μm (a,i,k), 25 μm (c,d,e,g).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4499882&req=5

f3: Tensile forces drive collective migration by activating mechanically sensitive intracellular signaling and transcription factors.(a) Phase contrast images showing collective migration from representative tissues treated with DMSO (control) or blebbistatin (12.5 μM). (b) Quantification of cohort length from tissues treated with DMSO or blebbistatin. Mean ± s.e.m. of three replicates is shown, n = 60 tissues per group. **P < 0.01, Student’s t-test. Immunofluorescence staining for FAK pY397 and phospho-p130Cas in representative DMSO (c) and blebbistatin-treated (d) migrating cohorts. (e) Confocal image showing immunofluorescence staining for MRTF-A localization (green) and nuclei (red) in a representative tissue. (f) Quantification of the localization of MRTF-A (nucleus/cytoplasm) in tissues. Levels of nuclear and cytoplasmic MRTF-A were quantified by measuring signal intensity in the two compartments. Mean ± s.e.m. of three replicates is shown. Migrating cohort (invasion): n = 14; quiescent body: n = 11. **P < 0.01, Student’s t-test. (g) Confocal image showing immunofluorescence staining for MRTF-A localization (green) and nuclei (red) in representative tissues treated with DMSO or blebbistatin. (h) Quantification of the localization of MRTF-A (nucleus/cytoplasm) in tissues treated with DMSO or blebbistatin. Levels of nuclear and cytoplasmic MRTF-A were quantified by measuring signal intensity in the two compartments. Mean ± s.e.m. of three replicates is shown. MRTF-A DMSO: n = 29; MRTF-A blebbistatin: n = 21; DAPI DMSO: n = 10; DAPI blebbistatin: n = 12. **P < 0.01, Student’s t-test. (i) Phase contrast images showing collective migration from representative tissues treated with DMSO or CCG-1423 (10 μM). (j) Quantification of cohort length from tissues treated with DMSO or CCG-1423. Mean ± sem. of three replicates is shown. Invading cohorts treated with DMSO: n=80; invading cohorts treated with CCG-1423: n = 57. **P < 0.01, Student’s t-test. (k) Frequency maps showing collective invasion from 34 tissues (three independent replicates) transfected with scrambled shRNA (shScr) or shMRTF-A (two constructs). (l) Quantification of cohort length from tissues transfected with shScr or shMRTF-A. Mean ± s.e.m. of three replicates is shown, n = 50 tissues per group. ***P < 0.001, Student’s t-test. All images are representative of three independent replicates in which >50 tissues were monitored. Scale bars, 50 μm (a,i,k), 25 μm (c,d,e,g).
Mentions: To determine whether active cellular contractility was required for collective migration, we blocked cytoskeletal tension by treating the invading tissues with blebbistatin (Fig. 3a) or Y27632 (Fig. S1a), which inhibit myosin ATPase and Rho kinase, respectively. Disrupting cell-generated forces significantly impaired the extent of collective migration (Fig. 3b; Fig. S1b). Conversely, enhancing cellular contractility by treating with LPA, an activator of Rho, increased the extent of migration (Fig. S1a).

Bottom Line: Moreover, cell movements are highly correlated and in phase with ECM deformations.Migrating cohorts use spatially localized, long-range forces and consequent matrix alignment to navigate through the ECM.These results suggest biophysical forces are critical for 3D collective migration.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemical &Biological Engineering, Princeton University, Princeton, NJ 08544, USA.

ABSTRACT
Collective cell migration drives tissue remodeling during development, wound repair, and metastatic invasion. The physical mechanisms by which cells move cohesively through dense three-dimensional (3D) extracellular matrix (ECM) remain incompletely understood. Here, we show directly that migration of multicellular cohorts through collagenous matrices occurs via a dynamic pulling mechanism, the nature of which had only been inferred previously in 3D. Tensile forces increase at the invasive front of cohorts, serving a physical, propelling role as well as a regulatory one by conditioning the cells and matrix for further extension. These forces elicit mechanosensitive signaling within the leading edge and align the ECM, creating microtracks conducive to further migration. Moreover, cell movements are highly correlated and in phase with ECM deformations. Migrating cohorts use spatially localized, long-range forces and consequent matrix alignment to navigate through the ECM. These results suggest biophysical forces are critical for 3D collective migration.

No MeSH data available.