TY - JOUR
T1 - Substratum compliance modulates corneal fibroblast to myofibroblast transformation
AU - Dreier, Britta
AU - Thomasy, Sara M
AU - Mendonsa, Rima
AU - Raghunathan, Vijay Krishna
AU - Russell, Paul
AU - Murphy, Christopher J
PY - 2013
Y1 - 2013
N2 - Purpose. The transformation of fibroblasts to myofibroblasts is critical to corneal wound healing, stromal haze formation, and scarring. It has recently been demonstrated that the provision of biomimetic substratum topographic cues inhibits the progression toward the myofibroblast phenotype under the influence of transforming growth factor β1 (TGF-β1). The objective of this study was to determine the effect of another fundamental biophysical cue, substrate compliance, on TGF-β1-induced myofibroblast transformation of primary corneal cells isolated from human and rabbit corneas. Methods. Human and rabbit corneal fibroblasts were cultured on surfaces of varying substrate compliance (4-71 kPa) and tissue culture plastic (TCP) (>1 gigapascal [GPa]). Cells were cultured in media containing TGF-β1 at concentrations of 0, 1, or 10 ng/mL for 72 hours. RNA and protein were collected from cells cultured on polyacrylamide gels and TCP and were analyzed for the expression of α-smooth muscle actin (α-SMA), a key marker of myofibroblast transformation, using quantitative PCR, immunocytochemistry, and Western blot. Results. Cells grown on more compliant substrates demonstrated significantly reduced amounts of α-SMA mRNA compared with TCP. Immunocytochemistry and Western blot analysis determining the presence of α-SMA corroborated this finding, thus confirming a reduced transformation to the myofibroblast phenotype on more compliant substrates compared with cells on TCP in the presence of TGF-β1. Conclusions. These data indicate that substrate compliance modulates TGF-β1-induced expression of α-SMA and thus influences myofibroblast transformation in the corneal stroma. This provides further evidence that biomimetic biophysical cues inhibit myofibroblast transformation and participate in stabilizing the native cellular phenotype.
AB - Purpose. The transformation of fibroblasts to myofibroblasts is critical to corneal wound healing, stromal haze formation, and scarring. It has recently been demonstrated that the provision of biomimetic substratum topographic cues inhibits the progression toward the myofibroblast phenotype under the influence of transforming growth factor β1 (TGF-β1). The objective of this study was to determine the effect of another fundamental biophysical cue, substrate compliance, on TGF-β1-induced myofibroblast transformation of primary corneal cells isolated from human and rabbit corneas. Methods. Human and rabbit corneal fibroblasts were cultured on surfaces of varying substrate compliance (4-71 kPa) and tissue culture plastic (TCP) (>1 gigapascal [GPa]). Cells were cultured in media containing TGF-β1 at concentrations of 0, 1, or 10 ng/mL for 72 hours. RNA and protein were collected from cells cultured on polyacrylamide gels and TCP and were analyzed for the expression of α-smooth muscle actin (α-SMA), a key marker of myofibroblast transformation, using quantitative PCR, immunocytochemistry, and Western blot. Results. Cells grown on more compliant substrates demonstrated significantly reduced amounts of α-SMA mRNA compared with TCP. Immunocytochemistry and Western blot analysis determining the presence of α-SMA corroborated this finding, thus confirming a reduced transformation to the myofibroblast phenotype on more compliant substrates compared with cells on TCP in the presence of TGF-β1. Conclusions. These data indicate that substrate compliance modulates TGF-β1-induced expression of α-SMA and thus influences myofibroblast transformation in the corneal stroma. This provides further evidence that biomimetic biophysical cues inhibit myofibroblast transformation and participate in stabilizing the native cellular phenotype.
KW - α-smooth muscle actin
KW - Biophysical cues
KW - Corneal wound healing
KW - Myofibroblast transformation
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U2 - 10.1167/iovs.12-11575
DO - 10.1167/iovs.12-11575
M3 - Article
C2 - 23860754
AN - SCOPUS:84883245855
VL - 54
SP - 5901
EP - 5907
JO - Investigative Ophthalmology and Visual Science
JF - Investigative Ophthalmology and Visual Science
SN - 0146-0404
IS - 8
ER -