Abstract
Under physiological conditions, the cornea is exposed to various enzymes, some of them have digestive actions, such as amylase and collagenase that may change the ultrastructure (collagen morphology) and sequentially change the mechanical response of the cornea and distort vision, such as in keratoconus. This study investigates the ultrastructure and nanomechanical properties of porcine cornea following incubation with α-amylase and collagenase. Atomic force microscopy (AFM) was used to capture nanoscale topographical details of stromal collagen fibrils (diameter and D-periodicity) and calculate their elastic modulus. Samples were incubated with varying concentrations of α-amylase and collagenase (crude and purified). Dimethylmethylene blue (DMMB) assay was utilised to detect depleted glycosaminoglycans (GAGs) following incubation with amylase. Collagen fibril diameters were decreased following incubation with amylase, but not D-periodicity. Elastic modulus was gradually decreased with enzyme concentration in amylase-treated samples. Elastic modulus, diameter, and D-periodicity were greatly reduced in collagenase-treated samples. The effect of crude collagenase on corneal samples was more pronounced than purified collagenase. Amylase was found to deplete GAGs from the samples. This enzymatic treatment may help in answering some questions related to keratoconus, and possibly be used to build an empirical animal model of keratoconic corneas with different progression levels.
Highlights
There is a need to investigate and develop a better understanding of corneal ultrastructure and biomechanics at the nano-level
Understanding corneal ultrastructure and its response to chemicals is important for a number of ocular disorders, such as keratoconus that is characterised by a significant deterioration in the collagenous network resulting in a cone-shape cornea
The results showed that Phosphate Buffered Saline (PBS) slightly decreased collagen fibril diameter
Summary
There is a need to investigate and develop a better understanding of corneal ultrastructure and biomechanics at the nano-level. Understanding corneal ultrastructure and its response to chemicals is important for a number of ocular disorders, such as keratoconus that is characterised by a significant deterioration in the collagenous network resulting in a cone-shape cornea. The human cornea consists of five layers, in which the stroma represents about 90%. The stroma is composed of many lamellae that are parallel to the corneal surface. The stroma mainly consists of Type I collagen fibrils that are arranged in a high degree of lateral order and run in the same direction as their lamellae. Collagen fibrils consist of several collagen molecules that are aligned together in staggered arrangement
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