Modelling Toxoplasma gondii infection in a 3D cell culture system In Vitro: Comparison with infection in 2D cell monolayers
Modelling Toxoplasma gondii infection in a 3D cell culture system In Vitro: Comparison with infection in 2D cell monolayers
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DOI：
10.1371/journal.pone.0208558
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发表时间：
2018-12-06
期刊：
影响因子：
3.7
通讯作者：
Coppens, Isabelle
中科院分区：
文献类型：
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作者：
Danielson, Jeffrey J.;Perez, Nicolas;Coppens, Isabelle
三维(3D)细胞培养模型弥合了二维(2D)单层培养和动物模型之间的差距。与生理相关的3D培养模型极大地促进了基础细胞科学的发展，并提供了与细胞形态内在联系的宿主-病原体相互作用的独特见解。弓形虫是一种专性空泡内寄生虫，慢性感染全球很大一部分人口。我们目前对弓形虫感染的理解主要基于2D细胞培养，即哺乳动物细胞生长在平面上。然而，2D细胞培养可能不能概括体内感染的关键条件，因为它们引入了人为的压力和张力，这可能随后改变依赖于空间性的感染过程，例如入侵、复制和出口。在这项研究中，我们采用基于胶原的3D细胞培养系统来再现弓形虫自然感染的3D环境，以研究寄生虫从寄生虫的空泡中复制和排出的情况。弓形虫感染的Vero细胞悬浮在3D基质中，具有圆形的形态，而感染的Vero细胞则在2D单层中。弓形虫在Vero细胞基质中的倍增时间与在Vero细胞单层中培养的寄生虫相当。在2D培养中，在没有扁平寄主细胞压力的情况下，弓形虫的寄生液泡呈球形，液泡内寄生虫呈放射状分布，形成3D球状“玫瑰花结”结构。在3D基质中，寄生虫径向离开破裂的宿主细胞，而在2D单层培养中，寄生虫垂直从宿主细胞下方的平面逃逸。这些观察证明了胶原基质在研究寄生虫感染模式方面的实用性，因为这些3D分析更接近于体内的条件。
Three-dimensional (3D) cell culture models bridge the gap between two-dimensional (2D) monolayer cultures and animal models. Physiologically relevant, 3D culture models have significantly advanced basic cell science and provide unique insights into host-pathogen interactions intrinsically linked to cell morphology. Toxoplasma gondii is an obligate intravacuolar parasite that chronically infects a large portion of the global human population. Our current understanding of Toxoplasma infection is largely based on 2D cell cultures, in which mammalian cells are grown on flat surfaces. However, 2D cell cultures may not recapitulate key conditions of in vivo infections as they introduce artificial pressures and tensions, which may subsequently alter infectious processes that are dependent on spatiality, e.g., invasion, replication and egress. In this study, we adapted a collagen-based 3D cell culture system to reproduce the 3D environment of T. gondii natural infections for investigation of the replication and egress of the parasite from the parasitophorous vacuole. Suspended in the 3D matrix, Toxoplasma-infected VERO cells have round morphology, as opposed to infected VERO cells in 2D monolayers. The doubling time of Toxoplasma in VERO cells within the matrix is comparable to that of parasites cultivated in VERO cell monolayers. In the absence of the pressure of flattened host cells grown in 2D cultures, the parasitophorous vacuole of T. gondii has a globular shape, with intravacuolar parasites distributed radially, forming 3D spherical 'rosette' structures. Parasites egress radially away from the ruptured host cell in 3D matrices, in contrast to Toxoplasma cultivated in 2D monolayer cultures, where the parasites escape perpendicularly from the flat surface below the host cells. These observations demonstrate the utility of collagen matrices for studying parasite modes of infection as these 3D assays more closely mimic in vivo conditions.