Polarized Raman Anisotropic Response of Collagen in Tendon: Towards 3D Orientation Mapping of Collagen in Tissues
Polarized Raman Anisotropic Response of Collagen in Tendon: Towards 3D Orientation Mapping of Collagen in Tissues
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DOI：
10.1371/journal.pone.0063518
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发表时间：
2013-05-15
期刊：
影响因子：
3.7
通讯作者：
Masic, Admir
中科院分区：
文献类型：
--
作者：
Galvis, Leonardo;Dunlop, John W. C.;Masic, Admir
在这项研究中，偏振拉曼光谱(PRS)被用来表征胶原酰胺I带的各向异性响应，作为评估三维胶原纤维在组织中取向的基础。首先，将经典的拉曼理论应用于类胶原蛋白多肽晶体结构，从理论上研究了类胶原多肽晶体结构的响应。然后，通过测量大鼠尾部的酰胺I强度各向异性作为入射激光偏振方向的函数，对理论方法进行了实验验证。在理论研究中，从蛋白质数据库中获得的几个胶原样三螺旋多肽晶体结构被“平面内”和“平面外”旋转，以评估分子取向对酰胺I带强度的作用。类胶原蛋白多肽在相对于偏振入射激光“平面”旋转时表现出正弦的各向异性响应。入射光垂直于分子的偏振时强度最大，平行时最小。在分子结构的“离面”旋转的情况下，观察到各向异性响应减弱，当结构垂直于观察平面时，变得完全各向同性。比较了胶原蛋白和α螺旋蛋白片段的理论拉曼响应。与胶原蛋白相反，当入射光平行于分子时，α螺旋的信号最大，垂直时最小。对于平面外的分子取向，α-螺旋结构的平均强度降低。在大鼠尾肌腱上的实验结果与理论预测非常吻合，从而证明了PRS在生物组织中胶原纤维三维取向的实验评估中的巨大潜力。
In this study, polarized Raman spectroscopy (PRS) was used to characterize the anisotropic response of the amide I band of collagen as a basis for evaluating three-dimensional collagen fibril orientation in tissues. Firstly, the response was investigated theoretically by applying classical Raman theory to collagen-like peptide crystal structures. The theoretical methodology was then tested experimentally, by measuring amide I intensity anisotropy in rat tail as a function of the orientation of the incident laser polarization. For the theoretical study, several collagen-like triple-helical peptide crystal structures obtained from the Protein Data Bank were rotated "in plane'' and "out of plane'' to evaluate the role of molecular orientation on the intensity of the amide I band. Collagen-like peptides exhibit a sinusoidal anisotropic response when rotated "in plane'' with respect to the polarized incident laser. Maximal intensity was obtained when the polarization of the incident light is perpendicular to the molecule and minimal when parallel. In the case of "out of plane'' rotation of the molecular structure a decreased anisotropic response was observed, becoming completely isotropic when the structure was perpendicular to the plane of observation. The theoretical Raman response of collagen was compared to that of alpha helical protein fragments. In contrast to collagen, alpha helices have a maximal signal when incident light is parallel to the molecule and minimal when perpendicular. For out-of-plane molecular orientations alpha-helix structures display a decreased average intensity. Results obtained from experiments on rat tail tendon are in excellent agreement with the theoretical predictions, thus demonstrating the high potential of PRS for experimental evaluation of the three-dimensional orientation of collagen fibers in biological tissues.