Corneal Biomechanical Properties from Two-Dimensional Corneal Flap Extensiometry: Application to UV-Riboflavin Cross-Linking

Abstract

Corneal biomechanical properties are usually measured by strip extensiometry or inflation methods. We developed a two-dimensional (2D) flap extensiometry technique, combining the advantages of both methods, and applied it to measure the effect of UV-Riboflavin cross-linking (CXL). Corneal flaps (13 pig/8 rabbit) from the de-epithelialized anterior stroma (96 μm) were mounted on a custom chamber, consisting of a BK7 lens, a reflective retina, and two reservoirs (filled with Riboflavin and silicone oil). Stretching the corneal flap during five pressure increase/decrease cycles (0-30 mm Hg) changed the refractive power of the system, whose Zernike aberrations were monitored with a ray-tracing aberrometer. Porcine flaps were used to test the system. Rabbits were treated with CXL unilaterally in vivo following standard clinical procedures. Flaps were measured 1 month postoperatively. An analytical model allowed estimating Young's modulus from the change in surface (strain) and pressure (stress). Confocal microscopy examination was performed before, and at different times after CXL. Flap curvature changed with increased function of IOP in pig flaps (23.4 × 10⁻³ D/mm Hg). In rabbit flaps curvature changed significantly less in 1 month post CXL (P = 0.026) than in untreated corneas [17.0 vs. 6.36 millidiopter (mD)/mm Hg]. Young's modulus was 2.29 megapascals (MPa) in porcine corneas, 1.98 MPa in untreated rabbit corneas, and 4.83 MPa in 1 month post CXL rabbit corneas. At the same time, highly reflective structures were observed in the rabbit midstroma after treatment. 2D flap extensiometry allows estimating corneal elasticity in vitro. The measurements are spatially resolved in depth, minimize the effects of corneal hydration, and preserve the integrity of the cornea. The method proved the efficacy of CXL in increasing corneal rigidity after 1 month in rabbits.