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Cancer cell invasion is enhanced by applied mechanical stimulation.

Menon S, Beningo KA - PLoS ONE (2011)

Bottom Line: We have discovered a significant difference in the extent of invasion in mechanically stimulated verses non-stimulated cell culture environments.Finally, we have found that the protein cofilin is needed to sense the mechanical stimuli that enhances invasion.We conclude that other types of mechanical signals in the tumor microenvironment, besides the rigidity, can enhance the invasive abilities of cancer cells in vitro.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Sciences, Wayne State University, Detroit, Michigan, United States of America.

ABSTRACT
Metastatic cells migrate from the site of the primary tumor, through the stroma, into the blood and lymphatic vessels, finally colonizing various other tissues to form secondary tumors. Numerous studies have been done to identify the stimuli that drive the metastatic cascade. This has led to the identification of multiple biochemical signals that promote metastasis. However, information on the role of mechanical factors in cancer metastasis has been limited to the affect of compliance. Interestingly, the tumor microenvironment is rich in many cell types including highly contractile cells that are responsible for extensive remodeling and production of the dense extracellular matrix surrounding the cancerous tissue. We hypothesize that the mechanical forces produced by remodeling activities of cells in the tumor microenvironment contribute to the invasion efficiency of metastatic cells. We have discovered a significant difference in the extent of invasion in mechanically stimulated verses non-stimulated cell culture environments. Furthermore, this mechanically enhanced invasion is dependent upon substrate protein composition, and influenced by topography. Finally, we have found that the protein cofilin is needed to sense the mechanical stimuli that enhances invasion. We conclude that other types of mechanical signals in the tumor microenvironment, besides the rigidity, can enhance the invasive abilities of cancer cells in vitro. We further propose that in vivo, non-cancerous cells located within the tumor micro-environment may be capable of providing the necessary mechanical stimulus during the remodeling of the extracellular matrix surrounding the tumor.

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Stimulation of paramagnetic beads.A) A rare earth magnet placed 1.5 cm below a matrix produces a gradient field ranging from 500G to 80G within the matrix as it rotates in a 2 cm orbit. A paramagnetic bead at position X would receive a magnetic force of 500G, ∼300G and ∼200G when the magnet is orbiting at positions P1, P2 and P3 respectively. B) Series of four images depicting the displacement of beads by the magnet when held in stationary positions within the orbit. Clusters of beads responding to the mechanical stimulus and showing a positional shift have been demarcated using a circle, a square and an arrow. From left to right, image one is outside the magnetic field while the second and third images were taken with the magnet held in positions P1 and P2 respectively. The final image demonstrates the beads return to their original position after the magnet is removed. C) MEF cellular extensions cause fluorescent bead displacement. Four images (0, 15, 30 and 60 minutes) from a single focal plane were selected from a series of 30 phase images taken every 2 minutes of a MEF cell within a collagen/fibronectin matrix. Cell outlines and corresponding fluorescent bead images are shown. A bead undergoing displacement is outlined using a white rectangular box. The area within the box from all four images has been enlarged and displayed with an inset ruler to show the bead displacement more clearly. The contrast of the magnified images have been altered to better reflect the position of the bead in each case. Mag. Bar  = 10 µm.
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pone-0017277-g002: Stimulation of paramagnetic beads.A) A rare earth magnet placed 1.5 cm below a matrix produces a gradient field ranging from 500G to 80G within the matrix as it rotates in a 2 cm orbit. A paramagnetic bead at position X would receive a magnetic force of 500G, ∼300G and ∼200G when the magnet is orbiting at positions P1, P2 and P3 respectively. B) Series of four images depicting the displacement of beads by the magnet when held in stationary positions within the orbit. Clusters of beads responding to the mechanical stimulus and showing a positional shift have been demarcated using a circle, a square and an arrow. From left to right, image one is outside the magnetic field while the second and third images were taken with the magnet held in positions P1 and P2 respectively. The final image demonstrates the beads return to their original position after the magnet is removed. C) MEF cellular extensions cause fluorescent bead displacement. Four images (0, 15, 30 and 60 minutes) from a single focal plane were selected from a series of 30 phase images taken every 2 minutes of a MEF cell within a collagen/fibronectin matrix. Cell outlines and corresponding fluorescent bead images are shown. A bead undergoing displacement is outlined using a white rectangular box. The area within the box from all four images has been enlarged and displayed with an inset ruler to show the bead displacement more clearly. The contrast of the magnified images have been altered to better reflect the position of the bead in each case. Mag. Bar  = 10 µm.

Mentions: To verify that the magnet was capable of producing enough magnetic force and that the embedded beads responded to the force in a transient manner, we used a magnometer to measure the magnetic force at defined experimental distances. We discovered a magnetic bead at a fixed point within the center of the culture could be subjected to a range of 500 to 80 Gauss as the rare earth magnet rotates 1.5 cm beneath the culture dish while completing an orbit of 2 cm at 160 rpm (2.6 Hz) (Figure 2A). Simulation at these distances under the microscope resulted in bead displacements of approximately 0.5–5 µm (Figure 2B, Movie S1, Movie S2). Beads were observed to spring back to their original position in the x-y plane after the magnet was removed, indicative of their attachment to the collagen matrix and maintenance of the integrity of the gel network. To determine the physiological significance of this displacement, we recognized that we could calculate the amount of force that was applied on the bead by the magnet, however a more tangible test would be to observe MEF cells extending and retracting extensions within our controlled culture system. We recorded bead displacements in the x-y plane from cellular extensions of MEF cells that range from 0.08–5.1 µm (Figure 2C, Movie S3). This is a conservative comparison to the types of displacements that could occur in the stroma given that the most contractile cell type found there, the myofibroblasts, produce considerably more force than a MEF [28], [29].


Cancer cell invasion is enhanced by applied mechanical stimulation.

Menon S, Beningo KA - PLoS ONE (2011)

Stimulation of paramagnetic beads.A) A rare earth magnet placed 1.5 cm below a matrix produces a gradient field ranging from 500G to 80G within the matrix as it rotates in a 2 cm orbit. A paramagnetic bead at position X would receive a magnetic force of 500G, ∼300G and ∼200G when the magnet is orbiting at positions P1, P2 and P3 respectively. B) Series of four images depicting the displacement of beads by the magnet when held in stationary positions within the orbit. Clusters of beads responding to the mechanical stimulus and showing a positional shift have been demarcated using a circle, a square and an arrow. From left to right, image one is outside the magnetic field while the second and third images were taken with the magnet held in positions P1 and P2 respectively. The final image demonstrates the beads return to their original position after the magnet is removed. C) MEF cellular extensions cause fluorescent bead displacement. Four images (0, 15, 30 and 60 minutes) from a single focal plane were selected from a series of 30 phase images taken every 2 minutes of a MEF cell within a collagen/fibronectin matrix. Cell outlines and corresponding fluorescent bead images are shown. A bead undergoing displacement is outlined using a white rectangular box. The area within the box from all four images has been enlarged and displayed with an inset ruler to show the bead displacement more clearly. The contrast of the magnified images have been altered to better reflect the position of the bead in each case. Mag. Bar  = 10 µm.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3040771&req=5

pone-0017277-g002: Stimulation of paramagnetic beads.A) A rare earth magnet placed 1.5 cm below a matrix produces a gradient field ranging from 500G to 80G within the matrix as it rotates in a 2 cm orbit. A paramagnetic bead at position X would receive a magnetic force of 500G, ∼300G and ∼200G when the magnet is orbiting at positions P1, P2 and P3 respectively. B) Series of four images depicting the displacement of beads by the magnet when held in stationary positions within the orbit. Clusters of beads responding to the mechanical stimulus and showing a positional shift have been demarcated using a circle, a square and an arrow. From left to right, image one is outside the magnetic field while the second and third images were taken with the magnet held in positions P1 and P2 respectively. The final image demonstrates the beads return to their original position after the magnet is removed. C) MEF cellular extensions cause fluorescent bead displacement. Four images (0, 15, 30 and 60 minutes) from a single focal plane were selected from a series of 30 phase images taken every 2 minutes of a MEF cell within a collagen/fibronectin matrix. Cell outlines and corresponding fluorescent bead images are shown. A bead undergoing displacement is outlined using a white rectangular box. The area within the box from all four images has been enlarged and displayed with an inset ruler to show the bead displacement more clearly. The contrast of the magnified images have been altered to better reflect the position of the bead in each case. Mag. Bar  = 10 µm.
Mentions: To verify that the magnet was capable of producing enough magnetic force and that the embedded beads responded to the force in a transient manner, we used a magnometer to measure the magnetic force at defined experimental distances. We discovered a magnetic bead at a fixed point within the center of the culture could be subjected to a range of 500 to 80 Gauss as the rare earth magnet rotates 1.5 cm beneath the culture dish while completing an orbit of 2 cm at 160 rpm (2.6 Hz) (Figure 2A). Simulation at these distances under the microscope resulted in bead displacements of approximately 0.5–5 µm (Figure 2B, Movie S1, Movie S2). Beads were observed to spring back to their original position in the x-y plane after the magnet was removed, indicative of their attachment to the collagen matrix and maintenance of the integrity of the gel network. To determine the physiological significance of this displacement, we recognized that we could calculate the amount of force that was applied on the bead by the magnet, however a more tangible test would be to observe MEF cells extending and retracting extensions within our controlled culture system. We recorded bead displacements in the x-y plane from cellular extensions of MEF cells that range from 0.08–5.1 µm (Figure 2C, Movie S3). This is a conservative comparison to the types of displacements that could occur in the stroma given that the most contractile cell type found there, the myofibroblasts, produce considerably more force than a MEF [28], [29].

Bottom Line: We have discovered a significant difference in the extent of invasion in mechanically stimulated verses non-stimulated cell culture environments.Finally, we have found that the protein cofilin is needed to sense the mechanical stimuli that enhances invasion.We conclude that other types of mechanical signals in the tumor microenvironment, besides the rigidity, can enhance the invasive abilities of cancer cells in vitro.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Sciences, Wayne State University, Detroit, Michigan, United States of America.

ABSTRACT
Metastatic cells migrate from the site of the primary tumor, through the stroma, into the blood and lymphatic vessels, finally colonizing various other tissues to form secondary tumors. Numerous studies have been done to identify the stimuli that drive the metastatic cascade. This has led to the identification of multiple biochemical signals that promote metastasis. However, information on the role of mechanical factors in cancer metastasis has been limited to the affect of compliance. Interestingly, the tumor microenvironment is rich in many cell types including highly contractile cells that are responsible for extensive remodeling and production of the dense extracellular matrix surrounding the cancerous tissue. We hypothesize that the mechanical forces produced by remodeling activities of cells in the tumor microenvironment contribute to the invasion efficiency of metastatic cells. We have discovered a significant difference in the extent of invasion in mechanically stimulated verses non-stimulated cell culture environments. Furthermore, this mechanically enhanced invasion is dependent upon substrate protein composition, and influenced by topography. Finally, we have found that the protein cofilin is needed to sense the mechanical stimuli that enhances invasion. We conclude that other types of mechanical signals in the tumor microenvironment, besides the rigidity, can enhance the invasive abilities of cancer cells in vitro. We further propose that in vivo, non-cancerous cells located within the tumor micro-environment may be capable of providing the necessary mechanical stimulus during the remodeling of the extracellular matrix surrounding the tumor.

Show MeSH
Related in: MedlinePlus