β淀粉样蛋白斑块的高分辨全脑三维定量研究

Three-dimensional quantitative analysis of amyloid plaques in the whole brain with high voxel resolution

中枢神经系统中β淀粉样蛋白斑块是阿尔兹海默症的主要病理特征之一,其负荷和数目的变化是病程发展的重要标志.已有研究主要是对局部脑组织进行二维切片成像,尚缺少在全脑三维空间对斑块进行高分辨率定量分析的研究方法.本文建立了适用于哺乳动物三维完整脑内β淀粉样蛋白斑块定量分析策略,包括全脑斑块快速荧光染色方法、基于荧光显微光学切片断层成像技术的高分辨全脑数据获取,以及斑块自动定位、统计数目等.与免疫组化染色比较,证明本方法对直径大于10μm的斑块检出率为97.71%±0.18%.并以0.32μm×0.32μm×2μm的成像分辨率,获取了5XFAD转基因小鼠全脑Aβ斑块分布数据集,首次以脑区/核团的三维轮廓划分出立体区域,定量统计了90个亚区内Aβ斑块的数量及分布密度.本文建立的快速、精准、价廉的方法将有助于全面高效地研究阿尔兹海默症致病机理和药效评估.

Amyloidosis of the central nervous system is one of the main pathological hallmarks of Alzheimer’s disease(AD). Changes in the number and the load of amyloid plaque(Aβ) deposition are important indicators of the progression of AD. However, the quantitative analysis of amyloidosis at the subcellular level in the whole brain remains a substantial challenge. In this study, an automatic analysis method was established for assessing amyloid plaques in the whole brain, including convenient whole brain labeling, imaging with a fluorescence micro-optical sectioning tomography system(fMOST) and an automated three-dimensional(3 D) analysis method.Validation using immunostaining showed that the detection rate of amyloid plaques larger than 10 μm was 97.71%±0.18%. We achieved a 3 D dataset of amyloid plaques in the whole brain of a 5 XFAD transgenic mouse at a resolution of 0.32 μm×0.32 μm×2 μm and quantitatively analyzed the 3 D distribution of amyloid plaques in different brain regions and nuclei through automated registration, segmentation and counting. The results indicated that the combination of the fMOST system with a whole brain staining method enables the acquisition and quantitative analysis of a brain-wide dataset of amyloid plaques, which may contribute to elucidating the underlying pathogenesis of AD and therapeutic drug discovery.