实验和有限元议定书调查中性和带电溶质的跨关节软骨运输
Summary
我们提出了一个协议，以调查带电和不带电分子的横跨关节软骨与最近开发的实验和数值方法的帮助下运输。
Abstract
骨关节炎（OA）是与关节软骨和软骨下骨的退化相关联的令人衰弱的疾病。关节软骨的退化基本上损害其承载功能，因为它经历了巨大的化学降解， 即 ，蛋白多糖的损失和胶原纤维的破坏。 OA期间，调查化学损伤机制中的一种有前途的方法是将试样软骨暴露到外部溶质和监控分子的扩散。软骨损伤（ 即浓度和基本大分子的构型）的程度与外部溶质的碰撞能量损失，同时跨越关节软骨移动相比健康软骨产生不同的扩散特性相关联。在这项研究中，我们引入一个协议，该协议包括几个步骤，并且基于先前开发的试验微-C14px的;“> omputedŤomography（微CT）和有限元建模使用微CT，随后是通过施加双相-溶质和多相有限元模型，以获得扩散系数首先被记录带电和不带电碘化分子的运输。固定跨软骨区的电荷密度。
Introduction
分子运输在铰接接头的内稳态中起着至关重要的作用，治疗剂递送至关节软骨和对比度增强软骨成像1，2，3。因素，如软骨整合和完整性，溶质电荷和大小以及重量渗透摩尔浓度，并与软骨可能影响传输速率4，5，6接触浴的浓度。溶质，无论是中性的或带电荷的传输，可以是关节软骨区域之间不同，因为每个区由不同浓度的和主要的细胞外基质分子的取向，即蛋白聚糖素（PGs）和II型胶原蛋白1，7，8，9，小姑娘=“外部参照”> 10,11。更重要的是，带电荷的溶质运输可以是高度依赖于包括横跨关节软骨8,9增加了的细胞外基质内负的固定电荷的蛋白聚糖的浓度。这些参数尤其是固定电荷密度（FCD），胶原原纤维和穿过软骨含水量变化的取向可经历改变为骨关节炎（OA）的进行，从而表示跨越软骨研究扩散的重要性。
在目前的研究中，协议基于先前建立的实验和计算研究6，8，9，提出了准确调查使用中性和带电溶质扩散的有限浴模型中的各种边界条件下扩散。 Ť他提出的方法是由包括软骨和由先进的双相 – 溶质和多相有限元模型支持的有限浴的系统的微型计算机断层扫描成像（微CT）的。这些模型使跨关节软骨的不同区域获得中性和带电分子以及FCDS的扩散系数。利用这些模型，可以更好地了解可用于研究软骨和覆盖有限浴之间的相互作用的扩散中性和带电分子的行为。
Protocol
注意：这里介绍的协议是从最近的研究论文6，8，9，实验和计算程序通过。该协议是在图1中示出。 该尸体材料与乌得勒支大学的兽医学院的许可收集。 1.样品和浴的制备使用定制的钻头（ 图1），同时喷涂凉爽磷酸盐缓冲盐水（PBS?…
Representative Results
这里提供的代表性的结果是从先前的研究论文6，8，9，16日通过。 在OA中，关节软骨发生显著变化最重要的GAG损失，以及胶原纤维17的损坏，18个 ，19。这些变?…
Discussion
我们提出了一个实验方案用有限元建模程序组合来研究跨关节软骨中性和带电溶质的扩散。根据我们最近的研究，所提出的模型可以准确地描述这两种中性（双相溶质）的跨关节软骨8,9的不同区域的运输和带负电荷（多相）溶质。人们普遍认为，关节软骨变得通过其重要部件的损耗功能的限制，诸如OA进展<sup class="xref"…
Disclosures
The authors have nothing to disclose.
Acknowledgements
作者想表达他们在乌得勒支UMC从开发力学组感谢的Jeroen先生范登贝尔赫和Matthijs Wassink先生为他们的包裹骨软骨插头的过程中提供帮助。这项工作是由来自荷兰关节炎基金会的资助。
Materials
| Hexabrix | Guerbet | 15HX005D | Negatively charged contrast agent |
| Visipaque | GE healthcare | 12570511 | Nuetral contrast agent |
| PBS | Life technologies | 10010023 | Medium |
| micro-CT | Perkin Elmer | Monitoring diffusion | |
| Freezing-point osmometer | Advanced instruments | Measuring solution osmolality |
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Cite This Article
Arbabi, V., Pouran, B., Zadpoor, A. A., Weinans, H. An Experimental and Finite Element Protocol to Investigate the Transport of Neutral and Charged Solutes across Articular Cartilage. J. Vis. Exp. (122), e54984, doi:10.3791/54984 (2017).