Anterior and posterior corneal stroma elasticity assessed using nanoindentation

Abstract

Corneal biomechanics is an essential parameter for developing diagnostic and treatment methods of corneal-related diseases. It is widely accepted that corneal mechanical strength stems from the stroma's collagenous composition. However, more comprehensive insight into the mechanical properties within the stroma is needed to improve current corneal diagnostic and treatment techniques. The purpose of this study was to perform elasticity characterization of anterior and posterior stromal regions of human corneas using atomic force microscopy (AFM). Nine pairs of human whole globes were placed in 20% Dextran solution, cornea side down, to restore the corneal thickness to physiological levels (400-600 μm). The epithelium and Bowman's membrane were removed from all eyes. Anterior stromal AFM elasticity testing was then performed on left (OS) eyes. Additional stroma was removed from right (OD) eyes to allow posterior stromal measurements at a depth of 50% of the original thickness. All experiments were performed with corneas submerged in 15% Dextran to maintain corneal hydration. The results of the study showed that the Young's modulus of elasticity of the anterior stroma (average: 281 ± 214 kPa; range: 59-764 kPa) was significantly higher than that of the posterior stroma (average: 89.5 ± 46.1 kPa; range: 29-179 kPa) (p = 0.014). In addition, a linear relationship was found between the posterior stromal elasticity and anterior stromal elasticity (p = 0.0428). On average, the elasticity of the posterior stroma is 39.3% of the anterior stroma. In summary, there appears to be an elasticity gradient within the corneal stroma, which should be considered in the design and development of corneal diagnostic and treatment methods to enhance efficacy.

Keywords: atomic force microscopy; cornea; mechanical properties.

Figure 1

Custom Cornea Holder. Side View (Top) and Top View (Bottom) of the Human Cornea in Custom Corneal Holder for AFM Testing. A human cornea (stained with 0.4% trypan blue for illustration purposes) is placed in the custom corneal holder during AFM testing. A human cornea (stained with 0.4% trypan blue for illustration purposes) was excised within the sclera beyond the limbus and was mounted onto the base of a custom cornea holder. The holder is comprised of a spherical protrusion that has a radius of curvature matching that of the cornea. The cornea was secured onto the spherical base with the holder's anchoring top that contains a hole to expose the central region of cornea for mechanical testing. This holder enables sturdy anchoring of an intact cornea, maintenance of corneal curvature, and the filling of 15% Dextran solution for proper corneal hydration during AFM testing.

Figure 2

Bar Graph of Anterior Stromal Elasticity and Posterior Stromal Elasticity. A bar graph comparing the Young's modulus of elasticity of the anterior stroma and posterior stroma for each respective eye pair is shown. Right eyes were subjected to anterior stromal elasticity testing and left eyes were subjected to posterior stromal elasticity testing using atomic force microscopy.

Figure 3

Linear Regression of Posterior Stromal Young's Modulus of Elasticity vs. Anterior Stromal Young's Modulus of Elasticity. This graph demonstrates a significant linear relationship between the elasticity of the posterior stroma and anterior stroma.