Department of Physics and Astronomy, University of Florence, 50019, Sesto Fiorentino, Italy.
European Laboratory for Non-Linear Spectroscopy (LENS), 50019, Sesto Fiorentino, Italy.
Sci Rep. 2023 Mar 13;13(1):4160. doi: 10.1038/s41598-023-30953-w.
Fluorescence microscopy can be exploited for evaluating the brain's fiber architecture with unsurpassed spatial resolution in combination with different tissue preparation and staining protocols. Differently from state-of-the-art polarimetry-based neuroimaging modalities, the quantification of fiber tract orientations from fluorescence microscopy volume images entails the application of specific image processing techniques, such as Fourier or structure tensor analysis. These, however, may lead to unreliable outcomes as they do not isolate myelinated fibers from the surrounding tissue. In this work, we describe a novel image processing pipeline that enables the computation of accurate 3D fiber orientation maps from both grey and white matter regions, exploiting the selective multiscale enhancement of tubular structures of varying diameters provided by a 3D implementation of the Frangi filter. The developed software tool can efficiently generate orientation distribution function maps at arbitrary spatial scales which may support the histological validation of modern diffusion-weighted magnetic resonance imaging tractography. Despite being tested here on two-photon scanning fluorescence microscopy images, acquired from tissue samples treated with a label-free technique enhancing the autofluorescence of myelinated fibers, the presented pipeline was developed to be employed on all types of 3D fluorescence images and fiber staining.
荧光显微镜可以与不同的组织准备和染色方案结合使用，以无与伦比的空间分辨率来评估大脑的纤维结构。与最先进的基于偏光的神经影像学模式不同，从荧光显微镜体积图像中定量纤维束方向需要应用特定的图像处理技术，如傅立叶或结构张量分析。然而，这些方法可能会导致不可靠的结果，因为它们不能将髓鞘纤维与周围组织分离。在这项工作中，我们描述了一种新的图像处理管道，该管道可利用 Frangi 滤波器的 3D 实现提供的不同直径管状结构的选择性多尺度增强，从灰质和白质区域计算准确的 3D 纤维方向图。开发的软件工具可以在任意空间尺度上高效地生成方向分布函数图，这可能支持对现代扩散加权磁共振成像纤维追踪的组织学验证。尽管这里是在使用免标记技术增强髓鞘纤维自发荧光的组织样本的双光子扫描荧光显微镜图像上进行测试的，但所提出的管道旨在用于所有类型的 3D 荧光图像和纤维染色。
