School of Dentistry, Faculty of Medicine, Pontificia Universidad Católica de Chile, Santiago, Chile.
Institute for Biological and Medical Engineering, Schools of Engineering, Medicine and Biological Sciences, Pontificia Universidad Católica de Chile, Santiago, Chile.
J Dent Res. 2023 Jul;102(8):957-964. doi: 10.1177/00220345231166294. Epub 2023 May 18.
The adhesion of initial colonizers such as to collagen is critical for dentinal and root caries progression. One of the most described pathological and aging-associated changes in collagen-including dentinal collagen-is the generation of advanced glycation end-products (AGEs) such as methylglyoxal (MGO)-derived AGEs. Despite previous reports suggesting that AGEs alter bacterial adhesion to collagen, the biophysics driving oral streptococcal attachment to MGO-modified collagen remains largely understudied. Thus, the aim of this work was to unravel the dynamics of the initial adhesion of to type I collagen in the presence and absence of MGO-derived AGEs by employing bacterial cell force spectroscopy with atomic force microscopy (AFM). Type I collagen gels were treated with 10 mM MGO to induce AGE formation, which was characterized with microscopy and enzyme-linked immunosorbent assay. Subsequently, AFM cantilevers were functionalized with living UA 159 or SK 36 cells and probed against collagen surfaces to obtain force curves displaying bacterial attachment in real time, from which the adhesion force, number of events, Poisson analysis, and contour and rupture lengths for each individual detachment event were computed. Furthermore, in silico computer simulation docking studies between the relevant UA 159 collagen-binding protein SpaP and collagen were computed, in the presence and absence of MGO. Overall, results showed that MGO modification increased both the number and adhesion force of single-unbinding events between and collagen, without altering the contour or rupture lengths. Both experimental and in silico simulations suggest that this effect is due to increased specific and nonspecific forces and interactions between UA 159 and MGO-modified collagen substrates. In summary, these results suggest that collagen alterations due to aging and glycation may play a role in early bacterial adherence to oral tissues, associated with conditions such as aging or chronic hyperglycemia, among others.
初始定植菌（如 ）与胶原蛋白的黏附对于牙本质和根面龋的进展至关重要。在包括牙本质胶原蛋白在内的胶原蛋白中，最常描述的一种病理性和与衰老相关的变化是生成晚期糖基化终产物（AGEs），如甲基乙二醛（MGO）衍生的 AGEs。尽管先前有报道表明 AGEs 会改变细菌对胶原蛋白的黏附，但口腔链球菌黏附到 MGO 修饰的胶原蛋白的生物物理机制在很大程度上仍未得到充分研究。因此，本研究旨在通过原子力显微镜下的细菌细胞力谱法（AFM）揭示 MGO 衍生的 AGE 存在和不存在时， 与 I 型胶原蛋白初始黏附的动力学。使用 10 mM MGO 处理 I 型胶原蛋白凝胶以诱导 AGE 形成，并用显微镜和酶联免疫吸附测定法对其进行表征。随后，将 AFM 悬臂用活的 UA 159 或 SK 36 细胞功能化，并探测胶原蛋白表面以实时获得显示细菌附着的力曲线，从中计算出粘附力、事件数、泊松分析以及每个单独脱离事件的轮廓和断裂长度。此外，还计算了存在和不存在 MGO 时，相关的 UA 159 胶原蛋白结合蛋白 SpaP 与胶原蛋白之间的计算机模拟对接研究。总的来说，结果表明，MGO 修饰增加了 与胶原蛋白之间单键解吸事件的数量和粘附力，而不改变轮廓或断裂长度。实验和计算机模拟均表明，这种效应是由于 UA 159 与 MGO 修饰的胶原蛋白底物之间的特异性和非特异性力和相互作用增加所致。总之，这些结果表明，由于衰老和糖化导致的胶原蛋白改变可能在与衰老或慢性高血糖等相关的条件下，在早期细菌黏附到口腔组织中发挥作用。
