一种新的APP剪接变异体依赖性标记系统用于精确划分SH-SY5Y细胞衍生神经元的成熟度-动脉网

A novel APP splice variant-dependent marker system to precisely demarcate maturity in SH-SY5Y cell-derived neurons

Nature 等信源发布 2024-05-27 01:36

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AbstractSH-SY5Y, a neuroblastoma cell line, can be converted into mature neuronal phenotypes, characterized by the expression of mature neuronal and neurotransmitter markers. However, the mature phenotypes described across multiple studies appear inconsistent. As this cell line expresses common neuronal markers after a simple induction, there is a high chance of misinterpreting its maturity.

摘要SH-SY5Y是一种神经母细胞瘤细胞系，可以转化为成熟的神经元表型，其特征在于成熟神经元和神经递质标志物的表达。然而，多项研究中描述的成熟表型似乎不一致。由于该细胞系在简单诱导后表达常见的神经元标记，因此很有可能误解其成熟度。

Therefore, sole reliance on common neuronal markers is presumably inadequate. The Alzheimer's disease (AD) central gene, amyloid precursor protein (APP), has shown contrasting transcript variant dynamics in various cell types. We differentiated SH-SY5Y cells into mature neuron-like cells using a concise protocol and observed the upregulation of total APP throughout differentiation.

因此，仅依赖常见的神经元标记物可能是不够的。阿尔茨海默病（AD）中心基因淀粉样前体蛋白（APP）在各种细胞类型中显示出对比的转录物变异动态。我们使用简明的方案将SH-SY5Y细胞分化为成熟的神经元样细胞，并观察到整个分化过程中总APP的上调。

However, APP transcript variant-1 was upregulated only during the early to middle stages of differentiation and declined in later stages. We identified the maturity state where this post-transcriptional shift occurs, terming it 'true maturity.' At this stage, we observed a predominant expression of mature neuronal and cholinergic markers, along with a distinct APP variant pattern.

然而，APP转录本变体-1仅在分化的早期至中期上调，而在后期下降。我们确定了这种转录后转变发生的成熟状态，将其称为“真正的成熟”在此阶段，我们观察到成熟神经元和胆碱能标记物的主要表达，以及独特的APP变异模式。

Our findings emphasize the necessity of using a differentiation state-sensitive marker system to precisely characterize SH-SY5Y differentiation. Moreover, this study offers an APP-guided, alternative neuronal marker system to enhance the accuracy of the conventional markers..

我们的发现强调了使用分化状态敏感标记系统来精确表征SH-SY5Y分化的必要性。此外，这项研究提供了一个应用程序引导的替代神经元标记系统，以提高常规标记的准确性。。

IntroductionNeuronal cell lines play a crucial role in neuroscience research as in vitro model systems that mimic the nervous system and its associated disorders at the molecular, physiological, and cellular levels. Neuroblastoma (NB) cell lines are more versatile than other neural cell lines because they are neuroblast-like, undifferentiated neural precursors derived from the neural crest and could differentiate into mature neuronal cell types.

引言神经元细胞系作为在分子，生理和细胞水平上模拟神经系统及其相关疾病的体外模型系统，在神经科学研究中起着至关重要的作用。神经母细胞瘤（NB）细胞系比其他神经细胞系更通用，因为它们是源自神经c的成神经细胞样，未分化的神经前体，可以分化成成熟的神经元细胞类型。

Among NB cell lines, such as rat PC-12, mouse neuro-2a, and human SH-SY5Y, SH-SY5Y stands out as the most widely used cell model in neurodegenerative research. This cell line, derived from a sub-cloned metastatic bone marrow biopsy of a patient with neuroblastoma1, is a well-established model in neurological research, including investigating the molecular mechanisms underlying numerous disorders, such as Parkinson's disease (PD) and Alzheimer's disease (AD).

在NB细胞系中，例如大鼠PC-12，小鼠neuro-2a和人SH-SY5Y，SH-SY5Y是神经退行性研究中使用最广泛的细胞模型。该细胞系来源于神经母细胞瘤1患者的亚克隆转移性骨髓活检，是神经学研究中公认的模型，包括研究帕金森病（PD）和阿尔茨海默病（AD）等多种疾病的分子机制。

Moreover, the SH-SY5Y cell line can differentiate into multiple neuronal phenotypes, making it a more attractive tool for studying neurons. The SH-SY5Y cell line has been frequently used in both its undifferentiated and differentiated states2. Proliferating SH-SY5Y cells represent an undifferentiated state, which closely resembles that of neural progenitor cells or immature neurons.

此外，SH-SY5Y细胞系可以分化为多种神经元表型，使其成为研究神经元的更有吸引力的工具。SH-SY5Y细胞系经常用于其未分化和分化状态2。增殖的SH-SY5Y细胞代表未分化状态，与神经祖细胞或未成熟神经元非常相似。

Although they have certain merits, including ease of culture and reproducibility, the validity of experiments conducted using undifferentiated neuronal cell lines is often disputed as these cell lines fail to reflect the complexity of mature neurons and their stimuli-response mechanisms2.To develop a more realistic neuronal model, it must be converted to a mature neuronal phenotype by inducing differentiation.

尽管它们具有某些优点，包括易于培养和可重复性，但使用未分化神经元细胞系进行的实验的有效性经常受到争议，因为这些细胞系不能反映成熟神经元及其刺激反应机制的复杂性2。为了开发更现实的神经元模型，必须通过诱导分化将其转化为成熟的神经元表型。

The differentiated state mimics mature neurons, thus exhibiting mature neuronal markers.

分化状态模仿成熟神经元，因此表现出成熟的神经元标记。

Data availability

数据可用性

The data of this is available from the corresponding author on request.

相关数据可应要求从通讯作者处获得。

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Download referencesAcknowledgementsWe acknowledge funding support from the Korea Health Technology R&D Project, administered through the Korea Health Industry Development Institute (KHIDI), funded by the Ministry of Health and Welfare, Republic of Korea (Grant Number: HI22C1754).Author informationAuthors and AffiliationsLaboratory of Animal Reproduction and Physiology, College of Agriculture and Life Sciences, Chungnam National University, Yuseong-gu, Daejeon, 34134, Republic of KoreaD.

下载参考文献致谢我们感谢韩国卫生技术研发项目的资金支持，该项目由韩国卫生与福利部资助的韩国卫生产业发展研究所（KHIDI）管理（资助号：HI22C1754）。作者信息作者和附属机构忠南国立大学农业与生命科学学院动物繁殖与生理学实验室，大田玉城，34134，科里德共和国。

Chanuka M. Kulatunga, Umanthi Ranaraja, Dong Ern Kim, Kuk Bin Ji & Min Kyu KimMK Biotech Inc., Daejeon, Republic of KoreaEun Young Kim, Ryoung Eun Kim & Min Kyu KimAuthorsD. Chanuka M. KulatungaView author publicationsYou can also search for this author in.

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PubMed Google ScholarContributionsD.C.M.K. and M.K.K. designed and conceived the study. D.C.M.K., U.R., E.Y.K., R.E.K., D.E.K., and K.B.J., performed the experiments. D.C.M.K. and M.K.K. made transgenic vectors, gene delivery, and transgenic cell lines. D.C.M.K., U.R., and E.Y.K.

PubMed谷歌学术贡献SD。C、 M.K.和M.K.K.设计并构思了这项研究。D、 C.M.K.，U.R.，E.Y.K.，R.E.K.，D.E.K.和K.B.J.进行了实验。D、 C.M.K.和M.K.K.制造了转基因载体，基因传递和转基因细胞系。D、 C.M.K.，U.R。和E.Y.K。

did microscopic imaging and image analysis. D.C.M.K., U.R., and R.E.K. did the data handling and statistical analysis. D.C.M.K. and M.K.K. wrote and edited the manuscript. All authors read, finalized, and approved the manuscript.Corresponding authorCorrespondence to.

进行了显微成像和图像分析。D、 C.M.K.，U.R。和R.E.K.进行了数据处理和统计分析。D、 C.M.K.和M.K.K.撰写并编辑了手稿。所有作者都阅读，定稿并批准了手稿。对应作者对应。

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Reprints and permissionsAbout this articleCite this articleKulatunga, D.C.M., Ranaraja, U., Kim, E.Y. et al. A novel APP splice variant-dependent marker system to precisely demarcate maturity in SH-SY5Y cell-derived neurons.

转载和许可本文引用本文Kulatunga，D.C.M.，Ranaraja，U.，Kim，E.Y。等人。一种新的APP剪接变体依赖性标记系统，用于精确划分SH-SY5Y细胞衍生神经元的成熟度。

Sci Rep 14, 12113 (2024). https://doi.org/10.1038/s41598-024-63005-yDownload citationReceived: 23 October 2023Accepted: 23 May 2024Published: 27 May 2024DOI: https://doi.org/10.1038/s41598-024-63005-yShare this articleAnyone you share the following link with will be able to read this content:Get shareable linkSorry, a shareable link is not currently available for this article.Copy to clipboard.

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