Phenotypic clustering: a novel method for microglial morphology analysis.
Phenotypic clustering: a novel method for microglial morphology analysis.
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DOI：
10.1186/s12974-016-0614-7
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发表时间：
2016-06-17
影响因子：
9.3
通讯作者：
Danckaert A
中科院分区：
文献类型：
--
作者：
Verdonk F;Roux P;Flamant P;Fiette L;Bozza FA;Simard S;Lemaire M;Plaud B;Shorte SL;Sharshar T;Chrétien F;Danckaert A
小胶质细胞是中枢神经系统的组织驻留巨噬细胞。它们非常动态，对其微环境非常敏感，并在脑组织中呈现出独特的复杂和异质的形态和分布。许多实验线索强调了它们的形态和它们在应对攻击时的功能之间的密切联系。然而，由于它们复杂的树突状方面构成了小鼠小胶质细胞的主要池和致密的网络，精确而强大的形态学研究并不容易实现，并且与分子或临床参数的相关性复杂。使用敲入小鼠模型CX3CR1GFP/+，我们开发了3D自动共聚焦组织成像系统，结合数千个小胶质细胞的形态建模，揭示了主要细胞特征的精确和定量评估：细胞密度、细胞体面积、细胞质面积和第一、第二和第三突起的数量。我们确定了两个形态标准，即复杂性指数(CI)和覆盖环境面积(CEA)，这使得创新的方法在于(I)准确和客观地研究健康或病理条件下的形态变化，(Ii)不同神经解剖区域的小胶质细胞分布的原位图谱，以及(Iii)大量细胞聚集的研究，使我们能够区分不同的亚群。我们对20,000多个细胞的研究结果证实，在系统注射脂多糖(LPS)诱导神经炎症后，小胶质细胞分布和表型的区域异质性持续存在。利用聚类分析，我们强调，在静息状态，小胶质细胞分布在由其CI和CEA定义的四个小胶质细胞亚群中，并具有区域模式和攻击后的特定行为。我们的结果反驳了经典的观点，即大脑内小胶质细胞处于同质的区域休眠状态。小胶质细胞分布在不同定义的亚群中，这些亚群在病理挑战后表现出特定的行为，从而可以假设细胞和功能的特化。此外，这种新的实验方法不仅将为神经病理学诊断提供支持，还将为研究各种疾病模型中的小胶质细胞功能提供支持，同时减少接近国际伦理声明所需的动物数量。本文的在线版本(doi：10.1186/s12974-0160614-7)包含补充材料，授权用户可以使用。
Microglial cells are tissue-resident macrophages of the central nervous system. They are extremely dynamic, sensitive to their microenvironment and present a characteristic complex and heterogeneous morphology and distribution within the brain tissue. Many experimental clues highlight a strong link between their morphology and their function in response to aggression. However, due to their complex “dendritic-like” aspect that constitutes the major pool of murine microglial cells and their dense network, precise and powerful morphological studies are not easy to realize and complicate correlation with molecular or clinical parameters. Using the knock-in mouse model CX3CR1GFP/+, we developed a 3D automated confocal tissue imaging system coupled with morphological modelling of many thousands of microglial cells revealing precise and quantitative assessment of major cell features: cell density, cell body area, cytoplasm area and number of primary, secondary and tertiary processes. We determined two morphological criteria that are the complexity index (CI) and the covered environment area (CEA) allowing an innovative approach lying in (i) an accurate and objective study of morphological changes in healthy or pathological condition, (ii) an in situ mapping of the microglial distribution in different neuroanatomical regions and (iii) a study of the clustering of numerous cells, allowing us to discriminate different sub-populations. Our results on more than 20,000 cells by condition confirm at baseline a regional heterogeneity of the microglial distribution and phenotype that persists after induction of neuroinflammation by systemic injection of lipopolysaccharide (LPS). Using clustering analysis, we highlight that, at resting state, microglial cells are distributed in four microglial sub-populations defined by their CI and CEA with a regional pattern and a specific behaviour after challenge. Our results counteract the classical view of a homogenous regional resting state of the microglial cells within the brain. Microglial cells are distributed in different defined sub-populations that present specific behaviour after pathological challenge, allowing postulating for a cellular and functional specialization. Moreover, this new experimental approach will provide a support not only to neuropathological diagnosis but also to study microglial function in various disease models while reducing the number of animals needed to approach the international ethical statements. The online version of this article (doi:10.1186/s12974-016-0614-7) contains supplementary material, which is available to authorized users.