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器官型海马切片培养作为研究肿瘤细胞神经保护和侵袭性的模型

Organotypic Hippocampal Slice Cultures As a Model to Study Neuroprotection and Invasiveness of Tumor Cells.

作者信息

Grabiec Urszula, Hohmann Tim, Hammer Niels, Dehghani Faramarz

机构信息

Department of Anatomy and Cell Biology, Martin Luther University Halle-Wittenberg;

Department of Anatomy and Cell Biology, Martin Luther University Halle-Wittenberg.

出版信息

J Vis Exp. 2017 Aug 27(126):55359. doi: 10.3791/55359.

Abstract

In organotypic hippocampal slice cultures (OHSC), the morphological and functional characteristics of both neurons and glial cells are well preserved. This model is suitable for addressing different research questions that involve studies on neuroprotection, electrophysiological experiments on neurons, neuronal networks or tumor invasion. The hippocampal architecture and neuronal activity in multisynaptic circuits are well conserved in OHSC, even though the slicing procedure itself initially lesions and leads to formation of a glial scar. The scar formation alters presumably the mechanical properties and diffusive behavior of small molecules, etc. Slices allow the monitoring of time dependent processes after brain injury without animal surgery, and studies on interactions between various brain-derived cell types, namely astrocytes, microglia and neurons under both physiological and pathological conditions. An ambivalent aspect of this model is the absence of blood flow and immune blood cells. During the progression of the neuronal injury, migrating immune cells from the blood play an important role. As those cells are missing in slices, the intrinsic processes in the culture may be observed without external interference. Moreover, in OHSC the composition of the medium-external environment is precisely controlled. A further advantage of this method is the lower number of sacrificed animals compared to standard preparations. Several OHSC can be obtained from one animal making simultaneous studies with multiple treatments in one animal possible. For these reasons, OHSC are well suited to analyze the effects of new protective therapeutics after tissue damage or during tumor invasion. The protocol presented here describes a preparation method of OHSC that allows generating highly reproducible, well preserved slices that can be used for a variety of experimental research, like neuroprotection or tumor invasion studies.

摘要

在器官型海马切片培养物(OHSC)中,神经元和胶质细胞的形态及功能特征均得到良好保留。该模型适用于解决不同的研究问题,包括神经保护研究、神经元电生理实验、神经元网络研究或肿瘤侵袭研究。尽管切片过程本身最初会造成损伤并导致胶质瘢痕形成,但OHSC中多突触回路的海马结构和神经元活动仍得到很好的保留。瘢痕形成可能会改变小分子的机械性能和扩散行为等。切片能够在不进行动物手术的情况下监测脑损伤后的时间依赖性过程,还能研究各种脑源性细胞类型(即星形胶质细胞、小胶质细胞和神经元)在生理和病理条件下的相互作用。该模型的一个矛盾之处在于缺乏血流和免疫血细胞。在神经元损伤进展过程中,从血液中迁移的免疫细胞发挥着重要作用。由于切片中缺少这些细胞,因此可以在不受外部干扰的情况下观察培养物中的内在过程。此外,在OHSC中,培养基 - 外部环境的成分得到精确控制。与标准制剂相比,该方法的另一个优点是牺牲的动物数量较少。从一只动物身上可以获得多个OHSC,从而有可能在一只动物身上同时进行多种处理的研究。基于这些原因,OHSC非常适合分析组织损伤后或肿瘤侵袭期间新的保护性治疗方法的效果。此处介绍的方案描述了一种OHSC的制备方法,该方法能够生成高度可重复、保存良好的切片,可用于各种实验研究,如神经保护或肿瘤侵袭研究。

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