利用拉曼光谱评估骨髓间充质干细胞向神经胶质细胞表型的分化。
Raman spectroscopy to assess the differentiation of bone marrow mesenchymal stem cells into a glial phenotype.
作者信息
Bautista-González Sulei, Carrillo González Nidia Jannette, Campos-Ordoñez Tania, Acosta Elías Mónica Alessandra, Pedroza-Montero Martín Rafael, Beas-Zárate Carlos, Gudiño-Cabrera Graciela
机构信息
Laboratorio de Desarrollo y Regeneración Neural, Departamento de Biología Celular y Molecular, Centro Universitario de Ciencias Biológicas y Agropecuarias, Universidad de Guadalajara, Zapopan, Jalisco, Mexico.
Laboratorio de Biofísica Médica, Departamento de Investigación en Física, Universidad de Sonora, Hermosillo, Sonora, México.
出版信息
Regen Ther. 2023 Oct 7;24:528-535. doi: 10.1016/j.reth.2023.09.016. eCollection 2023 Dec.
BACKGROUND
Mesenchymal stem cells (MSCs) are multipotent precursor cells with the ability to self-renew and differentiate into multiple cell linage, including the Schwann-like fate that promotes regeneration after lesion. Raman spectroscopy provides a precise characterization of the osteogenic, adipogenic, hepatogenic and myogenic differentiation of MSCs. However, the differentiation of bone marrow mesenchymal stem cells (BMSCs) towards a glial phenotype (Schwann-like cells) has not been characterized before using Raman spectroscopy.
METHOD
We evaluated three conditions: 1) cell culture from rat bone marrow undifferentiated (uBMSCs), and two conditions of differentiation; 2) cells exposed to olfactory ensheathing cells-conditioned medium (dBMSCs) and 3) cells obtained from olfactory bulb (OECs). uBMSCs phenotyping was confirmed by morphology, immunocytochemistry and flow cytometry using antibodies of cell surface: CD90 and CD73. Glial phenotype of dBMSCs and OECs were verified by morphology and immunocytochemistry using markers of Schwann-like cells and OECs such as GFAP, p75 NTR and O4. Then, the Principal Component Analysis (PCA) of Raman spectroscopy was performed to discriminate components from the high wavenumber region between undifferentiated and glial-differentiated cells. Raman bands at the fingerprint region also were used to analyze the differentiation between conditions.
RESULTS
Differences between Raman spectra from uBMSC and glial phenotype groups were noted at multiple Raman shift values. A significant decrease in the concentration of all major cellular components, including nucleic acids, proteins, and lipids were found in the glial phenotype groups. PCA analysis confirmed that the highest spectral variations between groups came from the high wavenumber region observed in undifferentiated cells and contributed with the discrimination between glial phenotype groups.
CONCLUSION
These findings support the use of Raman spectroscopy for the characterization of uBMSCs and its differentiation in the glial phenotype.
背景
间充质干细胞（MSCs）是多能前体细胞，具有自我更新和分化为多种细胞谱系的能力，包括促进损伤后再生的雪旺氏样命运。拉曼光谱可对MSCs的成骨、成脂、成肝和成肌分化进行精确表征。然而，此前尚未使用拉曼光谱对骨髓间充质干细胞（BMSCs）向神经胶质表型（雪旺氏样细胞）的分化进行表征。
方法
我们评估了三种情况：1）大鼠骨髓未分化细胞（uBMSCs）的细胞培养，以及两种分化情况；2）暴露于嗅鞘细胞条件培养基的细胞（dBMSCs）和3）从嗅球获得的细胞（OECs）。通过使用细胞表面抗体CD90和CD73的形态学、免疫细胞化学和流式细胞术对uBMSCs进行表型鉴定。使用雪旺氏样细胞和OECs的标志物如GFAP、p75 NTR和O4，通过形态学和免疫细胞化学验证dBMSCs和OECs的神经胶质表型。然后，进行拉曼光谱的主成分分析（PCA），以区分未分化细胞和神经胶质分化细胞在高波数区域的成分。指纹区域的拉曼谱带也用于分析不同情况之间的分化。
结果
在多个拉曼位移值处，注意到uBMSC和神经胶质表型组的拉曼光谱之间存在差异。在神经胶质表型组中发现所有主要细胞成分（包括核酸、蛋白质和脂质）的浓度显著降低。PCA分析证实，组间最高的光谱变化来自未分化细胞中观察到的高波数区域，并有助于区分神经胶质表型组。
结论
这些发现支持使用拉曼光谱对uBMSCs及其向神经胶质表型的分化进行表征。
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