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.

中文翻译：

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使用Peakforce-QNM AFM进行酶降解后抢夺角膜

在生理条件下，角膜暴露于各种酶中，其中一些具有消化作用，例如淀粉酶和胶原酶，它们可能会改变超微结构（胶原蛋白的形态）并依次改变角膜的机械反应并扭曲视力，例如在圆锥角膜中。这项研究调查了猪角膜与α-淀粉酶和胶原酶孵育后的超微结构和纳米力学性能。原子力显微镜（AFM）用于捕获基质胶原纤维的纳米级地形学细节（直径和D周期），并计算其弹性模量。将样品与不同浓度的α-淀粉酶和胶原酶（粗品和纯化品）一起孵育。与淀粉酶孵育后，利用二甲基亚甲基蓝（DMMB）测定来检测耗尽的糖胺聚糖（GAG）。与淀粉酶一起孵育后，胶原原纤维直径减小，但D-周期性不降低。随着淀粉酶处理样品中酶浓度的增加，弹性模量逐渐降低。在胶原酶处理的样品中，弹性模量，直径和D周期大大降低。粗胶原酶对角膜样品的影响比纯化的胶原酶更明显。发现淀粉酶会耗尽样品中的GAG。这种酶促治疗可能有助于回答与圆锥角膜有关的一些问题，并可能用于建立具有不同进展水平的圆锥角膜角膜的经验动物模型。胶原酶处理过的样品中，D和周期明显降低。粗胶原酶对角膜样品的影响比纯化的胶原酶更明显。发现淀粉酶会耗尽样品中的GAG。这种酶促治疗可能有助于回答与圆锥角膜有关的一些问题，并可能用于建立具有不同进展水平的圆锥角膜角膜的经验动物模型。胶原酶处理过的样品中，D和周期明显降低。粗胶原酶对角膜样品的影响比纯化的胶原酶更明显。发现淀粉酶会耗尽样品中的GAG。这种酶促治疗可能有助于回答与圆锥角膜有关的一些问题，并可能用于建立具有不同进展水平的圆锥角膜角膜的经验动物模型。