The endothelial glycocalyx layer (EGL), which consists of long proteoglycans protruding from the endothelium, acts as a regulator of inflammation by preventing leukocyte engagement with adhesion molecules on the endothelial surface. The amount of resistance to adhesive events the EGL provides is the result of two properties: EGL thickness and stiffness. To determine these, we used an atomic force microscope to indent the surfaces of cultured endothelial cells with a glass bead and evaluated two different approaches for interpreting the resulting force-indentation curves. In one, we treat the EGL as a molecular brush, and in the other, we treat it as a thin elastic layer on an elastic half-space. The latter approach proved more robust in our hands and yielded a thickness of 110 nm and a modulus of 0.025 kPa. Neither value showed significant dependence on indentation rate. The brush model indicated a larger layer thickness (∼350 nm) but tended to result in larger uncertainties in the fitted parameters. The modulus of the endothelial cell was determined to be 3.0-6.5 kPa (1.5-2.5 kPa for the brush model), with a significant increase in modulus with increasing indentation rates. For forces and leukocyte properties in the physiological range, a model of a leukocyte interacting with the endothelium predicts that the number of molecules within bonding range should decrease by an order of magnitude because of the presence of a 110-nm-thick layer and even further for a glycocalyx with larger thickness. Consistent with these predictions, neutrophil adhesion increased for endothelial cells with reduced EGL thickness because they were grown in the absence of fluid shear stress. These studies establish a framework for understanding how glycocalyx layers with different thickness and stiffness limit adhesive events under homeostatic conditions and how glycocalyx damage or removal will increase leukocyte adhesion potential during inflammation.

中文翻译：

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内皮糖萼层特性及其限制白细胞粘附的能力

内皮糖萼层 (EGL) 由从内皮突出的长蛋白多糖组成，通过阻止白细胞与内皮表面上的粘附分子结合，起到炎症调节剂的作用。EGL 提供的对粘附事件的抵抗力是两个特性的结果：EGL 厚度和刚度。为了确定这些，我们使用原子力显微镜用玻璃珠压痕培养的内皮细胞表面，并评估了两种不同的方法来解释所产生的力-压痕曲线。一方面，我们将 EGL 视为分子刷，另一方面，我们将其视为弹性半空间上的薄弹性层。后一种方法在我们的手中被证明更可靠，产生了 110 nm 的厚度和 0.025 kPa 的模量。这两个值均未显示出对压痕率的显着依赖性。刷子模型表明更大的层厚度（~350 nm），但往往会导致拟合参数的不确定性更大。内皮细胞的模量被确定为 3.0-6.5 kPa（刷模型为 1.5-2.5 kPa），随着压痕率的增加，模量显着增加。对于生理范围内的力和白细胞特性，白细胞与内皮相互作用的模型预测，由于存在 110 nm 厚的层，结合范围内的分子数量应减少一个数量级，甚至更远对于更厚的糖萼。与这些预测一致，EGL 厚度降低的内皮细胞的中性粒细胞粘附增加，因为它们在没有流体剪切应力的情况下生长。这些研究建立了一个框架，用于了解具有不同厚度和刚度的糖萼层如何限制稳态条件下的粘附事件，以及糖萼损伤或去除如何在炎症期间增加白细胞粘附潜力。