Role of sulfated GAGs in shear mechanical properties of human and porcine cornea.

Abstract

The corneal extracellular matrix is mainly composed of collagen fibers, proteoglycans (PGs), and glycosaminoglycans (GAGs). The present work was done to investigate the effect of GAGs on linear viscoelastic shear properties of human and porcine cornea. A clear understanding of structural functions of GAGs could result in the development of new intervention methods for diseased conditions that involve changes to the expression of GAGs/PGs. Here, we used keratanase II enzyme to deplete sulfated GAGs from porcine and human donor corneal disks. After quantifying the GAG content, collagen fiber diameter, and interfibrillar spacings of control and GAG-depleted specimens using the Blyscan assay and transmission electron microscopy, we performed torsional rheometry to determine their shear properties at different levels of axial strain. We found that the GAG content of control human (52.35 ± 3.40 μg/mg dry tissue) and porcine cornea (48.59 ± 7.79 μg/mg dry tissue) significantly reduced following keratanase II enzyme treatment. Moreover, we observed that the diameter of collagen fibers (28.78 ± 2.33 nm) and interfibrillar spacing (45.93 ± 2.33 nm) of human specimens were significantly smaller than the collagen fiber diameter (34.77 ± 21.90 nm) and interfibrillar spacing (54.28 ± 3.99 nm) of porcine corneal samples. Although GAG depletion did not have any significant effect on the collagen fiber diameter, it significantly increased the interfibrillar spacing in both porcine and human samples. Within the range of linear viscoelastic behavior, the shear stiffness of human and porcine corneal samples did not depend on the shear strain but significantly increased with increasing the applied axial strain. The average complex shear modulus was found to be between 1.0 KPa and 6.5 KPa and between 8.5 KPa and 31 KPa for control porcine and human corneal discs, respectively. The GAG removal caused significant reduction of shear stiffness in both human and porcine corneal samples. Based on these findings, we conclude that sulfated GAGs are important in defining shear properties of porcine and human corneas and significant GAG content variation adversely affects corneal shear modulus.