Magnetic separation of peripheral nerve-resident cells underscores key molecular features of human Schwann cells and fibroblasts: an immunochemical and transcriptomics approach.
Magnetic separation of peripheral nerve-resident cells underscores key molecular features of human Schwann cells and fibroblasts: an immunochemical and transcriptomics approach.
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DOI：
10.1038/s41598-020-74128-3
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发表时间：
2020-10-28
影响因子：
4.6
通讯作者：
Monje PV
中科院分区：
文献类型：
--
作者：
Peng K;Sant D;Andersen N;Silvera R;Camarena V;Piñero G;Graham R;Khan A;Xu XM;Wang G;Monje PV
神经来源的人雪旺细胞(SC)培养是基础研究和翻译研究的不可替代的模型，但由于成纤维细胞过度生长的风险，它们的使用可能受到限制。成纤维细胞是一个定义不清的群体，由高度增殖的细胞组成，与人类干细胞相反，在培养中不会衰老。我们通过对成人神经来源的人干细胞和成纤维细胞进行详尽的免疫学和功能鉴定来揭示它们的特性，并优化磁激活细胞分选(MACs)方案来有效分离它们作为活细胞和生物活性细胞，从而启动了这项研究。接下来，我们使用免疫荧光显微镜成像、流式细胞仪分析和下一代RNA测序(RNA-SEQ)来明确表征后Mac细胞的产品。高分辨率转录组分析揭示了关键的谱系特异性转录本的身份，以及人类干细胞和成纤维细胞明显不同的神经脊和间充质起源。我们的分析强调了干细胞和成纤维细胞中的祖细胞或干细胞样分子表型以及成纤维细胞群体的异质性。此外，对RNA-SEQ数据的路径分析强调了成纤维细胞到干细胞信号的双向网络，这预示了干细胞和成纤维细胞对神经再生的互补但似乎独立的贡献。总之，将MACs与免疫化学和转录学方法相结合，提供了一种理想的工作流程，可以详尽地评估来自人类周围神经组织的高纯度细胞的身份、分化阶段和功能特征。
Nerve-derived human Schwann cell (SC) cultures are irreplaceable models for basic and translational research but their use can be limited due to the risk of fibroblast overgrowth. Fibroblasts are an ill-defined population consisting of highly proliferative cells that, contrary to human SCs, do not undergo senescence in culture. We initiated this study by performing an exhaustive immunological and functional characterization of adult nerve-derived human SCs and fibroblasts to reveal their properties and optimize a protocol of magnetic-activated cell sorting (MACS) to separate them effectively both as viable and biologically competent cells. We next used immunofluorescence microscopy imaging, flow cytometry analysis and next generation RNA sequencing (RNA-seq) to unambiguously characterize the post-MACS cell products. High resolution transcriptome profiling revealed the identity of key lineage-specific transcripts and the clearly distinct neural crest and mesenchymal origin of human SCs and fibroblasts, respectively. Our analysis underscored a progenitor- or stem cell-like molecular phenotype in SCs and fibroblasts and the heterogeneity of the fibroblast populations. In addition, pathway analysis of RNA-seq data highlighted putative bidirectional networks of fibroblast-to-SC signaling that predict a complementary, yet seemingly independent contribution of SCs and fibroblasts to nerve regeneration. In sum, combining MACS with immunochemical and transcriptomics approaches provides an ideal workflow to exhaustively assess the identity, the stage of differentiation and functional features of highly purified cells from human peripheral nerve tissues.
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影响因子：
16.2
作者：
Clements MP;Byrne E;Camarillo Guerrero LF;Cattin AL;Zakka L;Ashraf A;Burden JJ;Khadayate S;Lloyd AC;Marguerat S;Parrinello S
通讯作者：
Parrinello S
影响因子：
12.7
作者：
Garcia-Diaz, Beatriz;Bachelin, Corinne;Baron-Van Evercooren, Anne
通讯作者：
Baron-Van Evercooren, Anne
DOI：
10.1007/bf00305043
发表时间：
1988-01-01
期刊：
ANATOMY AND EMBRYOLOGY
影响因子：
--
作者：
HANINEC, P
通讯作者：
HANINEC, P
DOI：
10.1007/978-1-4939-7649-2_4
发表时间：
2018-01-01
期刊：
SCHWANN CELLS: METHODS AND PROTOCOLS
影响因子：
--
作者：
Andersen, Natalia D.;Monje, Paula V.
通讯作者：
Monje, Paula V.
影响因子：
4.6
作者：
Andersen ND;Srinivas S;Piñero G;Monje PV
通讯作者：
Monje PV