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Gah-Jone Won, Vivian Choh, Douglas S. Fudge; The Effects of Non-Muscle Inhibitors on the Biomechanics of Avian Crystalline Lenses. Invest. Ophthalmol. Vis. Sci. 2012;53(14):3037.
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Posterior fibre cells anchored at the lens capsule are associated with proteins such as actin, myosin and N-cadherins. Together, these proteins form a hexagonal lattice cradling the posterior surface of the lens. This lattice was disrupted using various inhibitors to examine their effects on lenticular biomechanics.
Lenses of 7-day-old White Leghorn chickens (Gallus gallus domesticus) were treated with either an inhibitor or its vehicle. Inhibitors were 10 µM and 100 µM 1-(5-iodonaphthalene-1-sulfonyl)-1H-hexahydro-1,4-diazepine hydrochloride (ML-7, a myosin light chain kinase (MLCK) inhibitor; n=14 for both groups), 10 µM 1-phenyl-1,2,3,4-tetrahydro-4-hydroxypyrrolo[2.3-b]-7-methylquinolin-4-one (blebbistatin, a myosin II inhibitor; n=16), 10 µM latrunculin (an actin polymerization inhibitor; n=18). Vehicle treatments were the equivalent concentrations of dimethyl sulfoxide. Structural integrity, assessed as the mechanical response of the lens to compression, was tested using an instrument that compresses the lens while measuring the responding force. Mechanical trials consisted of 0.75 mm compressions, repeated 3 times. The force vs. load curve for each lens was fitted with a growth exponential curve (y = yo + aebx). The b-coefficient was used to represent stiffness, with higher b-values indicating greater stiffness.
Inhibition of MLCK by 100 μM ML-7 led to an unexpected increase in stiffness, while 10 μM ML-7 led to a softening of the lens. The biphasic nature of ML-7 is presumably due to phosphorylation of proteins other than, and in addition to MLCK. 2D gel electrophoresis confirms this idea. Inhibition of myosin II led to a similar softening of the lens as the 10 μM ML-7 treatment, suggesting that disruption of myosin, either by prevention of phosphorylation or by disrupting its stability, leads to a loss of structural integrity. Softening, due to structural integrity loss, was also confirmed using an actin polymerization inhibitor.
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