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Quantitative assessment of forward and backward second harmonic three dimensional images of collagen Type I matrix remodeling in a stimulated cellular environment

机译：向前和向后的二次谐波三维Ⅰ型胶原基质的重塑图像的定量评价刺激的细胞环境

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摘要

The structural remodeling of collagens is important in several biological processes including wound healing, tendon repair and adaptation, fibrosis and morphogenesis. Multiphoton microscopy is efficient in the induction of highly specific second harmonic generation (SHG) signal from non-centrosymmetric macromolecules such as fibrillar collagens. Although the detectors in the reflection geometry have been normally employed for capturing the backward scattered SHG considering the wide range of engineered thick tissue applications, there are still questions about the generated 3D collagen structures because of the directional pattern of SHG signals. The present study dealt with an in vitro collagen-fibroblast raft or bioartificial tendon model where the stimulation of fibroblast cells induced lateral orientation of collagen Type I fibers. The SHG signals originating from 3D collagen matrix were captured simultaneously in both forward and backward scattering directions. Our structural analysis indicates that collagen fibers formed in such in vitro model systems are predominantly of uniform sizes and are aligned preferentially in the lateral direction. The criss-cross arrangements of laterally oriented fibers are evident in the initial stages of contraction but eventually those laterally oriented collagen fibers are found to be aligned in parallel to each other as well as to the fibroblasts after an extended period of contraction. Our comprehensive quantitative assessment of simultaneously captured forward and backward 3D SHG image datasets, which includes the SHG signal decay, fiber diameter, cell dimensions, colocalization profiles, the 3D voxel volumes and Fourier analysis, indicates strong correlation of structural features identified in forward and backward directions.
机译：胶原蛋白的结构重塑在几种生物过程中是重要的，包括伤口愈合，肌腱修复和适应，纤维化和形态发生。多光子显微镜显微镜在诱导来自非亚苯二甲菌等纤维状胶原蛋白的高焦摩擦大分子的高度特异性二次谐波产生（SHG）信号中有效。尽管反射几何中的探测器通常用于捕获考虑到各种工程化厚组织应用的后向散射的SHG，但由于SHG信号的方向模式，仍然存在关于产生的3D胶原结构的问题。本研究涉及体外胶原成纤维细胞筏或生物肌腱模型，其中成纤维细胞诱导胶原型I纤维的横向取向。源自3D胶原矩阵的SHG信号同时捕获向前和向后散射方向。我们的结构分析表明，在这种体外模型系统中形成的胶原纤维主要是均匀的尺寸，并且在横向方向上优先对准。在收缩的初始阶段中，横向取向纤维的纵横交错是明显的，但是，发现横向取向的胶原纤维在延长的收缩期后彼此平行地与成纤维细胞平行排列。我们对同时捕获的前向和后向3D SHG图像数据集的综合定量评估，包括SHG信号衰减，光纤直径，小区尺寸，分层化配置文件，3D Voxel体积和傅立叶分析，表明在向前和向后识别的结构特征的强相关方向。

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    T Abraham; D Kayra; B McManus; A Scott;

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  • 年(卷),期 -1(180),1
  • 年度 -1
  • 页码 17–25
  • 总页数 19
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