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Deformation gradients imprint the direction and speed of en masse fibroblast migration for fast healing.

Pan Z, Ghosh K, Hung V, Macri LK, Einhorn J, Bhatnagar D, Simon M, Clark RA, Rafailovich MH - J. Invest. Dermatol. (2013)

Bottom Line: In these situations, cells are influenced by the proximity of other cells including interactions facilitated by substrate mechanics.The mechanics of the hydrogel determined the magnitude of the gradient.The largest degree of alignment and migration velocity occurred on the gels with the largest gradients.

View Article: PubMed Central - PubMed

Affiliation: Department of Materials Science and Engineering, SUNY at Stony Brook, Stony Brook, New York, USA.

ABSTRACT
En masse cell migration is more relevant compared with single-cell migration in physiological processes of tissue formation, such as embryogenesis, morphogenesis, and wound healing. In these situations, cells are influenced by the proximity of other cells including interactions facilitated by substrate mechanics. Here, we found that when fibroblasts migrated en masse over a hydrogel, they established a well-defined deformation field by traction forces and migrated along a trajectory defined by field gradients. The mechanics of the hydrogel determined the magnitude of the gradient. For materials stiff enough to withstand deformation related to cellular traction forces, such patterns did not form. Furthermore, migration patterns functioned poorly on very soft matrices where only a minimal traction gradient could be established. The largest degree of alignment and migration velocity occurred on the gels with the largest gradients. Granulation tissue formation in punch wounds of juvenile pigs was correlated strongly with the modulus of the implanted gel, in agreement with in vitro en masse cell migration studies. These findings provide basic insight into the biomechanical influences on fibroblast movement in early wounds and relevant design criteria for the development of tissue-engineered constructs that aim to stimulate en masse cell recruitment for rapid wound healing.

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A) Masson Trichrome stained specimens showing granulation tissue regeneration in 4-day porcine excisional wounds as a function of hydrogel stiffness. Black lines encompass granulation tissue. No treatment: a partial rim of nascent pink granulation tissue appeared under the fibrin clot. 95 Pa hydrogel: a rim of granulation tissue underlies the fibrin clot. 550 Pa hydrogel: Increased granulation tissue appeared under the fibrin clot compared to no treatment control and 95 Pa hydrogel. 4270 Pa hydrogel: A remarkable increase in granulation tissue ingrowth (en masse fibroblast movement) appeared to be displacing the fibrin clot. B) Histogram of results. N=6. * indicates P < 0.05 by ANOVA and Tukey post-hoc analysis. Bars = 2mm
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Figure 5: A) Masson Trichrome stained specimens showing granulation tissue regeneration in 4-day porcine excisional wounds as a function of hydrogel stiffness. Black lines encompass granulation tissue. No treatment: a partial rim of nascent pink granulation tissue appeared under the fibrin clot. 95 Pa hydrogel: a rim of granulation tissue underlies the fibrin clot. 550 Pa hydrogel: Increased granulation tissue appeared under the fibrin clot compared to no treatment control and 95 Pa hydrogel. 4270 Pa hydrogel: A remarkable increase in granulation tissue ingrowth (en masse fibroblast movement) appeared to be displacing the fibrin clot. B) Histogram of results. N=6. * indicates P < 0.05 by ANOVA and Tukey post-hoc analysis. Bars = 2mm

Mentions: The in vitro model indicated that the modulus of the substrate can regulate both cell migration as well as cell orientation through the establishment of deformation gradient fields. Importantly, the substrate modulus had to be sufficiently high to maintain an adequate gradient, while simultaneously retaining the ability to deform in response to cell traction forces. In order to test this model in vivo, we used a porcine wound model. Full-thickness excisional wound sites were made on the back of healthy, juvenile Yorkshire pigs. HA/FNfds pre-gelling solutions with different crosslinking ratios were prepared and injected into the punch wound site where they gelled within 10 min after implantation. Four days later, the harvested wounds were processed and analyzed for the extent of granulation tissue formed (black line indicates boundary between granulation tissue and fibrin clot), as shown in Figure 5A. Morphometric analysis of cross sectional areas of such healing wounds demonstrated little to no difference in the degree of granulation tissue accumulation in wounds containing the softest gel relative to control wounds without gel (Figure 5B) However, a significant increase in granulation accumulation was observed for the stiffest gels, where we had previously demonstrated the optimal condition for cell migration in vitro.


Deformation gradients imprint the direction and speed of en masse fibroblast migration for fast healing.

Pan Z, Ghosh K, Hung V, Macri LK, Einhorn J, Bhatnagar D, Simon M, Clark RA, Rafailovich MH - J. Invest. Dermatol. (2013)

A) Masson Trichrome stained specimens showing granulation tissue regeneration in 4-day porcine excisional wounds as a function of hydrogel stiffness. Black lines encompass granulation tissue. No treatment: a partial rim of nascent pink granulation tissue appeared under the fibrin clot. 95 Pa hydrogel: a rim of granulation tissue underlies the fibrin clot. 550 Pa hydrogel: Increased granulation tissue appeared under the fibrin clot compared to no treatment control and 95 Pa hydrogel. 4270 Pa hydrogel: A remarkable increase in granulation tissue ingrowth (en masse fibroblast movement) appeared to be displacing the fibrin clot. B) Histogram of results. N=6. * indicates P < 0.05 by ANOVA and Tukey post-hoc analysis. Bars = 2mm
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC3755030&req=5

Figure 5: A) Masson Trichrome stained specimens showing granulation tissue regeneration in 4-day porcine excisional wounds as a function of hydrogel stiffness. Black lines encompass granulation tissue. No treatment: a partial rim of nascent pink granulation tissue appeared under the fibrin clot. 95 Pa hydrogel: a rim of granulation tissue underlies the fibrin clot. 550 Pa hydrogel: Increased granulation tissue appeared under the fibrin clot compared to no treatment control and 95 Pa hydrogel. 4270 Pa hydrogel: A remarkable increase in granulation tissue ingrowth (en masse fibroblast movement) appeared to be displacing the fibrin clot. B) Histogram of results. N=6. * indicates P < 0.05 by ANOVA and Tukey post-hoc analysis. Bars = 2mm
Mentions: The in vitro model indicated that the modulus of the substrate can regulate both cell migration as well as cell orientation through the establishment of deformation gradient fields. Importantly, the substrate modulus had to be sufficiently high to maintain an adequate gradient, while simultaneously retaining the ability to deform in response to cell traction forces. In order to test this model in vivo, we used a porcine wound model. Full-thickness excisional wound sites were made on the back of healthy, juvenile Yorkshire pigs. HA/FNfds pre-gelling solutions with different crosslinking ratios were prepared and injected into the punch wound site where they gelled within 10 min after implantation. Four days later, the harvested wounds were processed and analyzed for the extent of granulation tissue formed (black line indicates boundary between granulation tissue and fibrin clot), as shown in Figure 5A. Morphometric analysis of cross sectional areas of such healing wounds demonstrated little to no difference in the degree of granulation tissue accumulation in wounds containing the softest gel relative to control wounds without gel (Figure 5B) However, a significant increase in granulation accumulation was observed for the stiffest gels, where we had previously demonstrated the optimal condition for cell migration in vitro.

Bottom Line: In these situations, cells are influenced by the proximity of other cells including interactions facilitated by substrate mechanics.The mechanics of the hydrogel determined the magnitude of the gradient.The largest degree of alignment and migration velocity occurred on the gels with the largest gradients.

View Article: PubMed Central - PubMed

Affiliation: Department of Materials Science and Engineering, SUNY at Stony Brook, Stony Brook, New York, USA.

ABSTRACT
En masse cell migration is more relevant compared with single-cell migration in physiological processes of tissue formation, such as embryogenesis, morphogenesis, and wound healing. In these situations, cells are influenced by the proximity of other cells including interactions facilitated by substrate mechanics. Here, we found that when fibroblasts migrated en masse over a hydrogel, they established a well-defined deformation field by traction forces and migrated along a trajectory defined by field gradients. The mechanics of the hydrogel determined the magnitude of the gradient. For materials stiff enough to withstand deformation related to cellular traction forces, such patterns did not form. Furthermore, migration patterns functioned poorly on very soft matrices where only a minimal traction gradient could be established. The largest degree of alignment and migration velocity occurred on the gels with the largest gradients. Granulation tissue formation in punch wounds of juvenile pigs was correlated strongly with the modulus of the implanted gel, in agreement with in vitro en masse cell migration studies. These findings provide basic insight into the biomechanical influences on fibroblast movement in early wounds and relevant design criteria for the development of tissue-engineered constructs that aim to stimulate en masse cell recruitment for rapid wound healing.

Show MeSH
Related in: MedlinePlus