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首页> 外文期刊>Journal of International Medical Research >An integrated cell isolation and purification method for rat dorsal root ganglion neurons
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An integrated cell isolation and purification method for rat dorsal root ganglion neurons

机译：一种大鼠背根神经节神经元的细胞分离纯化综合方法

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Objective Neurobiology studies are increasingly focused on the dorsal root ganglion (DRG), which plays an important role in neuropathic pain. Existing DRG neuron primary culture methods have considerable limitations, including challenging cell isolation and poor cell yield, which cause difficulty in signaling pathway studies. The present study aimed to establish an integrated primary culture method for DRG neurons. Methods DRGs were obtained from fetal rats by microdissection, and then dissociated with trypsin. The dissociated neurons were treated with 5-fluorouracil to promote growth of neurons from the isolated cells. Then, reverse transcription polymerase chain reaction and immunofluorescence assays were used to identify and purify DRG neurons. Results Isolated DRGs were successfully dissociated and showed robust growth as individual DRG neurons in neurobasal medium. Both mRNA and protein assays confirmed that DRG neurons expressed neurofilament-200 and neuron-specific enolase. Conclusions Highly purified, stable DRG neurons could be easily harvested and grown for extended periods by using this integrated cell isolation and purification method, which may help to elucidate the mechanisms underlying neuropathic pain.
机译：客观的神经生物学研究越来越集中在背根神经节（DRG）上，背根神经节在神经性疼痛中起重要作用。现有的DRG神经元原代培养方法具有相当大的局限性，包括挑战性的细胞分离和较差的细胞产量，这导致了信号通路研究的困难。本研究旨在建立DRG神经元的综合原代培养方法。方法通过显微解剖从胎鼠中提取DRG，然后与胰蛋白酶解离。用5-氟尿嘧啶处理解离的神经元，以促进来自分离细胞的神经元的生长。然后，使用逆转录聚合酶链反应和免疫荧光法鉴定和纯化DRG神经元。结果分离的DRGs成功分离，并在神经基础培养基中作为单个DRG神经元显示出强劲的生长。 mRNA和蛋白质测定均证实DRG神经元表达了Neurofilament-200和神经元特异性烯醇化酶。结论采用这种整合的细胞分离和纯化方法可以很容易地收获高纯度，稳定的DRG神经元，并可以延长其生长时间，这可能有助于阐明神经性疼痛的潜在机制。

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