Tsai Tsung-Hua, Short Michael A, McLean David I, Zeng Haishan, McElwee Kevin, Lui Harvey
Photomedicine Institute, Department of Dermatology and Skin Science, Vancouver Coastal Health Research Institute and University of British Columbia, Vancouver, Canada.
Analyst. 2014 Jun 7;139(11):2799-805. doi: 10.1039/c4an00155a.
Stem cells offer tremendous opportunities for regenerative medicine. Over the past decade considerable research has taken place to identify and characterize the differentiation states of stem cells in culture. Raman micro-spectroscopy has emerged as an ideal technology since it is fast, nondestructive, and does not require potentially toxic dyes. Raman spectroscopy systems can also be incorporated into confocal microscope imaging systems allowing spectra to be obtained from below the tissue surface. Thus there is significant potential for monitoring stem cells in living tissue. Stem cells that reside in hair follicles are suitable for testing this possibility since they are close to the skin surface, and typically clustered around the bulge area. One of the first steps needed would be to obtain Raman micro-spectra from stem cells located in thin sections of tissue, and then see whether these spectra are clearly different from those of the surrounding differentiated cells. To facilitate this test, standard 5 μm thick sections of murine skin tissue were stained to identify the location of hair follicle stem cells and their progeny. Raman spectra were then obtained from adjacent cells in a subsequent unstained 10 μm thick section. The spectra revealed significant differences in peak intensities associated with nucleic acids, proteins, lipids and amino acids. Statistical analyses of the Raman micro-spectra identified stem cells with 98% sensitivity and 94% specificity, as compared with a CD34 immunostaining gold standard. Furthermore analyses of the spectral variance indicated differences in cellular dynamics between the two cell groups. This study shows that Raman micro-spectroscopy has a potential role in identifying adult follicle stem cells, laying the groundwork for future applications of hair follicle stem cells and other somatic stem cells in situ.
干细胞为再生医学提供了巨大的机遇。在过去十年中，人们进行了大量研究以识别和表征培养中的干细胞分化状态。拉曼显微光谱技术已成为一种理想的技术，因为它快速、无损，且不需要使用可能有毒的染料。拉曼光谱系统还可以整合到共聚焦显微镜成像系统中，从而能够从组织表面下方获取光谱。因此，在活体组织中监测干细胞具有巨大潜力。毛囊中的干细胞适合用于测试这种可能性，因为它们靠近皮肤表面，通常聚集在隆突区周围。首先需要采取的步骤之一是从位于组织薄片中的干细胞获取拉曼显微光谱，然后观察这些光谱是否与周围分化细胞的光谱明显不同。为便于进行此测试，对小鼠皮肤组织的标准5微米厚切片进行染色，以确定毛囊干细胞及其子代的位置。然后在随后未染色的10微米厚切片中从相邻细胞获取拉曼光谱。光谱显示出与核酸、蛋白质、脂质和氨基酸相关的峰强度存在显著差异。与CD34免疫染色金标准相比，对拉曼显微光谱的统计分析识别干细胞的灵敏度为98%，特异性为94%。此外，对光谱方差的分析表明两个细胞组之间细胞动态存在差异。这项研究表明，拉曼显微光谱在识别成年毛囊干细胞方面具有潜在作用，为毛囊干细胞和其他体干细胞的原位未来应用奠定了基础。
