Tension, Free Space, and Cell Damage in a Microfluidic Wound Healing Assay
We use a novel, microfluidics-based technique to deconstruct the classical wound healing scratch assay, decoupling the contribution of free space and cell damage on the migratory dynamics of an epithelial sheet. This method utilizes multiple laminar flows to selectively cleave cells enzymatically, a...
| Main Authors: | , , |
|---|---|
| Other Authors: | |
| Format: | Article |
| Language: | en_US |
| Published: |
Public Library of Science
2012
|
| Online Access: | http://hdl.handle.net/1721.1/69108 https://orcid.org/0000-0002-7232-304X |
| _version_ | 1811078925138264064 |
|---|---|
| author | Murrell, Michael Kamm, Roger Dale Matsudaira, Paul T. |
| author2 | Massachusetts Institute of Technology. Department of Biology |
| author_facet | Massachusetts Institute of Technology. Department of Biology Murrell, Michael Kamm, Roger Dale Matsudaira, Paul T. |
| author_sort | Murrell, Michael |
| collection | MIT |
| description | We use a novel, microfluidics-based technique to deconstruct the classical wound healing scratch assay, decoupling the contribution of free space and cell damage on the migratory dynamics of an epithelial sheet. This method utilizes multiple laminar flows to selectively cleave cells enzymatically, and allows us to present a 'damage free' denudation. We therefore isolate the influence of free space on the onset of sheet migration. First, we observe denudation directly to measure the retraction in the cell sheet that occurs after cell-cell contact is broken, providing direct and quantitative evidence of strong tension within the sheet. We further probe the mechanical integrity of the sheet without denudation, instead using laminar flows to selectively inactivate actomyosin contractility. In both cases, retraction is observed over many cell diameters. We then extend this method and complement the enzymatic denudation with analogies to wounding, including gradients in signals associated with cell damage, such as reactive oxygen species, suspected to play a role in the induction of movement after wounding. These chemical factors are evaluated in combination with the enzymatic cleavage of cells, and are assessed for their influence on the collective migration of a non-abrasively denuded epithelial sheet. We conclude that free space alone is sufficient to induce movement, but this movement is predominantly limited to the leading edge, leaving cells further from the edge less able to move towards the wound. Surprisingly, when coupled with a gradient in ROS to simulate the chemical effects of abrasion however, motility was not restored, but further inhibited. |
| first_indexed | 2024-09-23T11:07:23Z |
| format | Article |
| id | mit-1721.1/69108 |
| institution | Massachusetts Institute of Technology |
| language | en_US |
| last_indexed | 2024-09-23T11:07:23Z |
| publishDate | 2012 |
| publisher | Public Library of Science |
| record_format | dspace |
| spelling | mit-1721.1/691082022-10-01T01:21:47Z Tension, Free Space, and Cell Damage in a Microfluidic Wound Healing Assay Murrell, Michael Kamm, Roger Dale Matsudaira, Paul T. Massachusetts Institute of Technology. Department of Biology Massachusetts Institute of Technology. Department of Mechanical Engineering Massachusetts Institute of Technology. Department of Mechanical Engineering Whitehead Institute for Biomedical Research Kamm, Roger Dale Murrell, Michael Kamm, Roger Dale Matsudaira, Paul T. We use a novel, microfluidics-based technique to deconstruct the classical wound healing scratch assay, decoupling the contribution of free space and cell damage on the migratory dynamics of an epithelial sheet. This method utilizes multiple laminar flows to selectively cleave cells enzymatically, and allows us to present a 'damage free' denudation. We therefore isolate the influence of free space on the onset of sheet migration. First, we observe denudation directly to measure the retraction in the cell sheet that occurs after cell-cell contact is broken, providing direct and quantitative evidence of strong tension within the sheet. We further probe the mechanical integrity of the sheet without denudation, instead using laminar flows to selectively inactivate actomyosin contractility. In both cases, retraction is observed over many cell diameters. We then extend this method and complement the enzymatic denudation with analogies to wounding, including gradients in signals associated with cell damage, such as reactive oxygen species, suspected to play a role in the induction of movement after wounding. These chemical factors are evaluated in combination with the enzymatic cleavage of cells, and are assessed for their influence on the collective migration of a non-abrasively denuded epithelial sheet. We conclude that free space alone is sufficient to induce movement, but this movement is predominantly limited to the leading edge, leaving cells further from the edge less able to move towards the wound. Surprisingly, when coupled with a gradient in ROS to simulate the chemical effects of abrasion however, motility was not restored, but further inhibited. Massachusetts Institute of Technology. Presidential Fellowship National Institutes of Health (U.S.). Biotechnology Training Fellowship Singapore-MIT Alliance for Research and Technology Massachusetts Institute of Biotechnology Training Grant Massachusetts Institute of Technology (Open-source Funding) 2012-02-15T17:20:45Z 2012-02-15T17:20:45Z 2011-09 2011-03 Article http://purl.org/eprint/type/JournalArticle 1932-6203 http://hdl.handle.net/1721.1/69108 Murrell, Michael, Roger Kamm, and Paul Matsudaira. “Tension, Free Space, and Cell Damage in a Microfluidic Wound Healing Assay.” Ed. Laurent Kreplak. PLoS ONE 6.9 (2011): e24283. Web. 15 Feb. 2012. https://orcid.org/0000-0002-7232-304X en_US http://dx.doi.org/10.1371/journal.pone.0024283 PLoS ONE Creative Commons Attribution http://creativecommons.org/licenses/by/2.5/ application/pdf Public Library of Science PLoS |
| spellingShingle | Murrell, Michael Kamm, Roger Dale Matsudaira, Paul T. Tension, Free Space, and Cell Damage in a Microfluidic Wound Healing Assay |
| title | Tension, Free Space, and Cell Damage in a Microfluidic Wound Healing Assay |
| title_full | Tension, Free Space, and Cell Damage in a Microfluidic Wound Healing Assay |
| title_fullStr | Tension, Free Space, and Cell Damage in a Microfluidic Wound Healing Assay |
| title_full_unstemmed | Tension, Free Space, and Cell Damage in a Microfluidic Wound Healing Assay |
| title_short | Tension, Free Space, and Cell Damage in a Microfluidic Wound Healing Assay |
| title_sort | tension free space and cell damage in a microfluidic wound healing assay |
| url | http://hdl.handle.net/1721.1/69108 https://orcid.org/0000-0002-7232-304X |
| work_keys_str_mv | AT murrellmichael tensionfreespaceandcelldamageinamicrofluidicwoundhealingassay AT kammrogerdale tensionfreespaceandcelldamageinamicrofluidicwoundhealingassay AT matsudairapault tensionfreespaceandcelldamageinamicrofluidicwoundhealingassay |