Subtle variations in polymer chemistry modulate substrate stiffness and fibronectin activity
Subtle variations in polymer chemistry modulate substrate stiffness and fibronectin activity
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DOI：
10.1039/c0sm00074d
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发表时间：
2010-01-01
期刊：
影响因子：
3.4
通讯作者：
Salmeron-Sanchez, Manuel
中科院分区：
文献类型：
--
作者：
Brizuela Guerra, Nayrim;Gonzalez-Garcia, Cristina;Salmeron-Sanchez, Manuel
制备了以乙烯基为主链，侧基为-COO(CH2)(x)CH3 （x = 0,1,3,5）的聚合物底物家族。得到了硬度降低的底物，其特征是在37℃时的弹性模量，以及类似的化学基团。首先，我们研究了聚合物化学中的这些微小变化是否会导致纤维连接蛋白（FN）吸附的差异：在每个底物上获得相同的FN密度（450 ng cm(-2)），但在材料界面上的蛋白质的超分子组织，如用AFM获得的，对于x = 0和其他表面（x = 1,3,5）是不同的。因此，这允许人们使用一组底物（x = 1,3,5）作为唯一的物理参数来研究底物刚度对细胞行为的影响，即在排除了侧基长度对蛋白质构象的任何影响之后。此外，研究细胞-材料界面蛋白质中间层的重要性被强调：x = 0和x = 1对细胞行为的影响不能再归因于基质刚度的不同，因为蛋白质在材料表面的生物活性也不同。随后，利用MC3T3-E1成骨细胞样细胞研究了细胞间的初始相互作用，重点研究了肌动蛋白细胞骨架的发育、局灶黏附的形成以及细胞在不同基质上重组吸附FN层的能力。图像分析用于量化焦斑的频率分布，结果显示在较硬的基底上分布更广，形成较大的焦斑表明细胞在较硬的基底上施加更大的力。
A family of polymer substrates which consists of a vinyl backbone chain with the side groups -COO(CH2)(x)CH3, with x = 0, 1, 3, 5 was prepared. Substrates with decreasing stiffness, characterised by the elastic modulus at 37 degrees C, and similar chemical groups were obtained. Firstly, we have investigated whether these minute variations in polymer chemistry lead to differences in fibronectin (FN) adsorption: the same FN density was obtained on every substrate (450 ng cm(-2)) but the supramolecular organisation of the protein at the material interface, as obtained with AFM, was different for x = 0 and the other surfaces (x = 1, 3, 5). Consequently, this allows one to use a set of substrates (x = 1, 3, 5) to investigate the effect of substrate stiffness on cell behavior as the unique physical parameter, i.e. after ruling out any influence of the length of the side group on protein conformation. Moreover, the importance of investigating the intermediate layer of proteins at the cell-material interface is stressed: the effect of x = 0 and x = 1 on cell behavior cannot be ascribed to the different stiffness of the substrate anymore, since the biological activity of the protein on the material surface was also different. Afterwards, initial cellular interaction was investigated using MC3T3-E1 osteoblasts-like cells and focusing on actin cytoskeleton development, focal adhesion formation and the ability of cells to reorganize the adsorbed FN layer on the different substrates. Image analysis was used to quantify the frequency distribution of the focal plaques, which revealed broader distributions on the stiffer substrates, with formation of larger focal plaques revealing that cells exert higher forces on stiffer substrates.