Background

Astrocytes, one of the main glial cell types, play critical roles in the central nervous system (CNS) development and function, including support of neuronal survival and differentiation, blood brain barrier formation, synapse homeostasis and injury response. Cell isolation and culture techniques have been proved to be a powerful tool to study astrocyte physiology and function. Due to financial constraints and rigid biosafety and ethics rules to use animal models, freezing techniques and the creation of cell banks emerged as alternatives to optimize the use of experimental animals. One of the main challenges, however, of these techniques is to guarantee that conserved cells keep their biological properties.

New method

In this work, we characterized morphologically and functionally murine secondary astrocyte cultures that have been submitted to freezing/thawing procedures.

Results

Morphological characterization of SAC (secondary astrocyte culture) and SFAC (secondary frozen-astrocyte culture) did not reveal significant differences on astrocyte morphology, confluence time and cell number along culture period. Functionally, SAC and SFAC did not reveal differences in their potential to support neuronal survival, maturation, neuritogenesis and synapse formation.

Conclusions

Our results suggest that murine astrocytes that are submitted to freezing/thawing procedure maintain morphological and functional characteristics when compared with non-frozen astrocytes. Thus, this methodological approach is a valuable tool for in vitro research and might allow experimental optimization and reduction of animal use.

中文翻译：

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冷冻保存的星形胶质细胞保持生物学特性：支持神经元存活和分化。

背景

星形胶质细胞是神经胶质细胞的主要类型之一，在中枢神经系统（CNS）的发育和功能中起关键作用，包括支持神经元存活和分化，血脑屏障形成，突触稳态和损伤反应。细胞分离和培养技术已被证明是研究星形胶质细胞生理和功能的有力工具。由于使用动物模型的资金拮据和严格的生物安全和道德规范，出现了冷冻技术和细胞库的创建，作为优化实验动物使用的替代方法。然而，这些技术的主要挑战之一是确保保守的细胞保持其生物学特性。

新方法

在这项工作中，我们表征了形态和功能上的小鼠继发星形胶质细胞培养物，这些培养物已经过冷冻/解冻程序。

结果

SAC（次生星形胶质细胞培养物）和SFAC（次生冷冻星形胶质细胞培养物）的形态学特征没有显示出星形胶质细胞形态，融合时间和培养期间细胞数量的显着差异。在功能上，SAC和SFAC在支持神经元存活，成熟，神经形成和突触形成的潜力方面没有发现差异。

结论

我们的结果表明，与非冷冻星形胶质细胞相比，经受冷冻/解冻程序的鼠星形胶质细胞保持形态和功能特征。因此，这种方法学方法是用于体外研究的有价值的工具，并且可能允许实验优化和减少动物使用。