TY - JOUR
T1 - Modulation of Corneal Fibroblast Contractility within Fibrillar Collagen Matrices
AU - Vishwanath, Mridula
AU - Ma, Lisha
AU - Otey, Carol A.
AU - Jester, James V.
AU - Petroll, W. Matthew
PY - 2003/11
Y1 - 2003/11
N2 - PURPOSE. To investigate the migratory and contractile behavior of isolated human corneal fibroblasts in fibrillar collagen matrices. METHODS. A telomerase-infected, extended-lifespan human corneal fibroblast cell line (HTK) was transfected by using a vector for enhanced green fluorescent protein (GFP)-α-actinin. Cells were plated at low density on top of or within 100-μm-thick fibrillar collagen lattices. After 18 hours to 7 days, time-lapse imaging was performed. At each 1- to 3-minute interval, GFP and Nomarski differential interference contrast (DIC) images were acquired in rapid succession. Serum-containing (S+) medium was used initially for perfusion. After 2 hours, perfusion was switched to either serum-free (S-) or S+ medium containing the Rho-kinase inhibitor Y-27632 for 1 to 2 hours. Finally, perfusion was changed back to S+ medium for 1 hour. RESULTS. Two to 4 days after plating, many cells underwent spontaneous contraction and/or relaxation in S+ medium. A decrease in the distance between consecutive α-actinin-dense bodies along stress fibers was measured during contraction, and focal adhesion and matrix displacements correlated significantly. Removal of serum or inhibition of Rho-kinase induced cell body elongation and relaxation of matrix stress, as confirmed using finite element modeling. Rapid formation and extension of pseudopodia and filopodia were also observed, and transient tractional forces were generated by these extending processes. CONCLUSIONS. Cultured human corneal fibroblasts can undergo rapid changes in the subcellular pattern of force generation that are mediated, in part, by Rho-kinase. Sarcomeric shortening of stress fibers in contracting corneal fibroblasts is also demonstrated for the first time.
AB - PURPOSE. To investigate the migratory and contractile behavior of isolated human corneal fibroblasts in fibrillar collagen matrices. METHODS. A telomerase-infected, extended-lifespan human corneal fibroblast cell line (HTK) was transfected by using a vector for enhanced green fluorescent protein (GFP)-α-actinin. Cells were plated at low density on top of or within 100-μm-thick fibrillar collagen lattices. After 18 hours to 7 days, time-lapse imaging was performed. At each 1- to 3-minute interval, GFP and Nomarski differential interference contrast (DIC) images were acquired in rapid succession. Serum-containing (S+) medium was used initially for perfusion. After 2 hours, perfusion was switched to either serum-free (S-) or S+ medium containing the Rho-kinase inhibitor Y-27632 for 1 to 2 hours. Finally, perfusion was changed back to S+ medium for 1 hour. RESULTS. Two to 4 days after plating, many cells underwent spontaneous contraction and/or relaxation in S+ medium. A decrease in the distance between consecutive α-actinin-dense bodies along stress fibers was measured during contraction, and focal adhesion and matrix displacements correlated significantly. Removal of serum or inhibition of Rho-kinase induced cell body elongation and relaxation of matrix stress, as confirmed using finite element modeling. Rapid formation and extension of pseudopodia and filopodia were also observed, and transient tractional forces were generated by these extending processes. CONCLUSIONS. Cultured human corneal fibroblasts can undergo rapid changes in the subcellular pattern of force generation that are mediated, in part, by Rho-kinase. Sarcomeric shortening of stress fibers in contracting corneal fibroblasts is also demonstrated for the first time.
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U2 - 10.1167/iovs.03-0513
DO - 10.1167/iovs.03-0513
M3 - Article
C2 - 14578392
AN - SCOPUS:0142231944
SN - 0146-0404
VL - 44
SP - 4724
EP - 4735
JO - Investigative Ophthalmology and Visual Science
JF - Investigative Ophthalmology and Visual Science
IS - 11
ER -