Department of Biomedical Engineering, University of Rochester, Rochester, New York.
Department of Biomedical Engineering, University of Rochester, Rochester, New York.
Biophys J. 2020 Apr 7;118(7):1564-1575. doi: 10.1016/j.bpj.2020.02.010. Epub 2020 Feb 15.
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.
内皮糖萼层（EGL）由从内皮突出的长蛋白聚糖组成，通过防止白细胞与内皮表面的粘附分子结合来充当炎症调节剂。EGL 提供的抗粘附事件的阻力大小取决于两个特性：EGL 厚度和刚性。为了确定这些特性，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由此产生的力-压痕曲线。在一种方法中，我们将 EGL 视为分子刷，而在另一种方法中，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更为稳健，得出的厚度为 110nm，模量为 0.025kPa。这两个值都没有显示出明显的依赖于压痕率。刷子模型表明了更大的层厚度（约 350nm），但往往导致拟合参数的不确定性更大。内皮细胞的模量被确定为 3.0-6.5kPa（刷子模型为 1.5-2.5kPa），随着压痕率的增加，模量显著增加。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 110nm 厚的层，结合范围内的分子数量应该减少一个数量级，对于更厚的糖萼，甚至会进一步减少。这些预测一致表明，由于缺乏流体切应力，内皮细胞 EGL 厚度减小会导致中性粒细胞黏附增加。这些研究建立了一个框架，用于理解具有不同厚度和刚性的糖萼层如何在稳态条件下限制粘附事件，以及糖萼层的损伤或去除如何在炎症期间增加白细胞的黏附潜力。
