A major challenge to experimental studies and therapeutic uses of hematopoietic stem cells (HSC) is the limited options for analytical tools that can reliably resolve functional differences in heterogeneous HSC subpopulations at the single cell level. Currently available methods require the use of external labels and/or separate clonogenic and transplantation assays to identify bona fide stem cells, necessitating the harvest of bulk cell populations and long incubation times that obscure how individual HSCs dynamically respond to exogenous and endogenous stimuli. In this study, we employ Raman spectroscopy to noninvasively resolve the dynamics of individual differentiating hematopoietic progenitor cells during the course of neutrophilic differentiation. We collected Raman peaks of individual cells daily over the course of 14-day neutrophilic differentiation. Principal component analysis (PCA) of the Raman peaks revealed spectral differences between individual cells during differentiation that were strongly correlated with changes in the nucleus shape and surface antigen expression, the primary traditional means of monitoring neutrophilic differentiation. Additionally, our results were consistently reproducible in independent rounds of neutrophilic differentiation, as demonstrated by our partial least-squares discriminant analysis (PLS-DA) of the Raman spectral information that predicted the degree of neutrophilic differentiation with high sensitivity and specificity. Our findings highlight the utility and reliability of Raman spectroscopy as a robust molecular imaging tool to monitor the kinetics of HSC differentiation patterns.

中文翻译：

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通过拉曼光谱追踪造血祖细胞中性粒细胞分化

造血干细胞 (HSC) 实验研究和治疗用途的一个主要挑战是，能够在单细胞水平上可靠地解决异质 HSC 亚群功能差异的分析工具选择有限。目前可用的方法需要使用外部标记和/或单独的克隆形成和移植测定来鉴定真正的干细胞，需要收获大量细胞群和较长的孵育时间，从而掩盖了单个 HSC 如何动态响应外源和内源刺激。在本研究中，我们采用拉曼光谱来无创地解析中性粒细胞分化过程中个体分化造血祖细胞的动态。在 14 天的中性粒细胞分化过程中，我们每天收集单个细胞的拉曼峰。拉曼峰的主成分分析 (PCA) 揭示了分化过程中各个细胞之间的光谱差异，这些差异与细胞核形状和表面抗原表达的变化密切相关，这是监测中性粒细胞分化的主要传统方法。此外，我们的结果在独立轮次的中性粒细胞分化中始终如一，正如我们对拉曼光谱信息的偏最小二乘判别分析（PLS-DA）所证明的那样，该分析以高灵敏度和特异性预测中性粒细胞分化的程度。我们的研究结果强调了拉曼光谱作为一种强大的分子成像工具来监测 HSC 分化模式的动力学的实用性和可靠性。