An unbiased, automated platform for scoring dopaminergic neurodegeneration in C. elegans.
An unbiased, automated platform for scoring dopaminergic neurodegeneration in C. elegans.
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DOI：
10.1371/journal.pone.0281797
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发表时间：
2023
期刊：
影响因子：
3.7
通讯作者：
San-Miguel, Adriana
中科院分区：
文献类型：
--
作者：
Clark, Andrew S.;Kalmanson, Zachary;Morton, Katherine;Hartman, Jessica;Meyer, Joel;San-Miguel, Adriana
秀丽隐杆线虫（Caenorhabditis elegans，C. elegans）作为研究多巴胺能神经变性的简单模式生物，因为它能够定量分析活体动物的细胞和亚细胞形态。这些等基因线虫具有快速的生命周期和透明的身体，使得荧光标记神经元的高通量成像和评估成为可能。然而，目前用于量化多巴胺能变性的最先进的方法需要研究人员手动检查图像并将树突分为不同神经变性严重程度的组，这是耗时的，易受偏见影响，并且数据敏感性有限。我们的目标是克服人工神经元评分的缺陷，通过开发一种自动化的，无偏的图像处理算法来量化C。优雅该算法可用于不同的显微镜设置获得的图像，只需要两个输入：在C的四个头神经元的最大投影图像。elegans头和用户相机的像素大小。我们通过检测和量化暴露于鱼藤酮，冷休克和6-羟基多巴胺的线虫中的神经变性，分别使用63倍荧光，63倍共聚焦和40倍荧光显微镜来验证该平台。对脂肪储存改变的矮胖突变蠕虫的分析表明，与我们的假设相反，肥胖的增加对应激诱导的神经退行性变不敏感。我们通过比较代码生成的分类退化结果与相同实验的手动评分树突，进一步验证了算法的准确性。该平台可以检测20种不同的神经退行性变指标，可以比较了解每种暴露如何影响多巴胺能神经退行性变模式。
Caenorhabditis elegans (C. elegans) has served as a simple model organism to study dopaminergic neurodegeneration, as it enables quantitative analysis of cellular and sub-cellular morphologies in live animals. These isogenic nematodes have a rapid life cycle and transparent body, making high-throughput imaging and evaluation of fluorescently tagged neurons possible. However, the current state-of-the-art method for quantifying dopaminergic degeneration requires researchers to manually examine images and score dendrites into groups of varying levels of neurodegeneration severity, which is time consuming, subject to bias, and limited in data sensitivity. We aim to overcome the pitfalls of manual neuron scoring by developing an automated, unbiased image processing algorithm to quantify dopaminergic neurodegeneration in C. elegans. The algorithm can be used on images acquired with different microscopy setups and only requires two inputs: a maximum projection image of the four cephalic neurons in the C. elegans head and the pixel size of the user’s camera. We validate the platform by detecting and quantifying neurodegeneration in nematodes exposed to rotenone, cold shock, and 6-hydroxydopamine using 63x epifluorescence, 63x confocal, and 40x epifluorescence microscopy, respectively. Analysis of tubby mutant worms with altered fat storage showed that, contrary to our hypothesis, increased adiposity did not sensitize to stressor-induced neurodegeneration. We further verify the accuracy of the algorithm by comparing code-generated, categorical degeneration results with manually scored dendrites of the same experiments. The platform, which detects 20 different metrics of neurodegeneration, can provide comparative insight into how each exposure affects dopaminergic neurodegeneration patterns.
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