Background

Astrocytes are the main cellular constituent in the central nervous system. Astrocyte cultures from rodent brains are most commonly used in the experimental practice. However, important differences between rodent and human astrocytes exist. The aim of this study was to develop an improved protocol for routine preparation of primary astrocyte culture from adult human brain, obtained after trauma.

New Method

Tissue obtained during neurotrauma operation was mechanically decomposed and centrifuged. The cell sediment was resuspended in cell culture medium, plated in T25 tissue flasks and incubated for one month at 37 °C in 5% CO_2. The medium was replaced twice weekly and microglia were removed. Once confluent, the purity of cultures was assessed. The culture was characterised immunocytochemically for specific astrocytic markers (GFAP, GLAST and S100B). Cell morphology was examined through the actin cytoskeleton labelling with fluorescent phalloidin.

Results

Under basal conditions, adult astrocytes exhibited astrocyte-specific morphology and expressed specific markers. Approximately 95% of cells were positive for the main glial markers (GFAP, GLAST, S100B).

Comparison with Existing Method

We established an easy and cost-effective method for a highly enriched primary astrocyte culture from adult human brain.

Conclusion

The isolation technique provides sufficient quantities of isolated cells. The culture obtained in this study exhibits the biochemical and physiological properties of astrocytes. It may be useful for elucidating the mechanisms related to the adult brain, exploring changes between neonatal and adult astrocytes, novel therapeutic targets, cell therapy experiments, as well as investigating compounds involved in cytotoxicity and cytoprotection.

中文翻译：

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优化了神经创伤患者的成人星形胶质细胞的分离和鉴定。

背景

星形胶质细胞是中枢神经系统的主要细胞成分。来自啮齿动物大脑的星形胶质细胞培养物最常用于实验实践中。但是，啮齿动物和人类星形胶质细胞之间存在重要差异。这项研究的目的是开发一种改进的方案，用于从成年人脑常规制备创伤后获得的原代星形胶质细胞培养物。

新方法

对神经外伤手术中获得的组织进行机械分解并离心。将细胞沉淀物重悬于细胞培养基中，铺在T25组织瓶中，并在37℃，5％CO ₂中温育一个月。每周更换培养基两次，并去除小胶质细胞。汇合后，评估培养物的纯度。用特定的星形细胞标记（GFAP，GLAST和S100B）对细胞进行免疫细胞化学鉴定。通过用荧光鬼笔环肽标记肌动蛋白细胞骨架来检查细胞形态。

结果

在基础条件下，成年星形胶质细胞表现出星形胶质细胞特异性形态并表达特异性标记。大约95％的细胞对主要神经胶质标记（GFAP，GLAST，S100B）呈阳性。

与现有方法的比较

我们建立了一种简单而经济高效的方法，用于从成年人脑中高度富集的原代星形胶质细胞培养。

结论

分离技术可提供足够数量的分离细胞。在这项研究中获得的培养物表现出星形胶质细胞的生化和生理特性。它对于阐明与成人大脑有关的机制，探索新生儿和成人星形胶质细胞之间的变化，新的治疗靶标，细胞疗法实验以及研究涉及细胞毒性和细胞保护作用的化合物可能是有用的。