The human innate immune system is able to recognize pathogen-associated molecular patterns like lipopolysaccharides (LPS) leading to the activation of signal cascades and the release of different cytokines. Activation of the immune cells can be assessed in different ways which are either indirect (ELISA of cytokine release), require staining protocols (flow cytometry) or lysis of the cells (mRNA analysis). Here, Raman spectroscopy as a non-destructive spectroscopic method is presented to enable direct and label-free monitoring of changes in cellular metabolism, biomolecular composition as well as morphology. Exemplarily, the potential of Raman spectroscopy is presented for the characterization of LPS-stimulation of monocytic THP-1 cells over a time course of 16 h. The cell culture stimulation model is characterized using gene transcription and expression of the two cytokines TNFα and IL-1β. After 1 h, 3 h, 8 h and 16 h specific Raman spectroscopic fingerprints are generated which encode cell activation pattern after TLR4 stimulation. Most prevalent changes in the spectra occur after 8 h, but slight differences are already detectable after 1 h. Spatially highly resolved Raman scans are used to generate false-color Raman images which provide spatial information of the biochemical state of the cells and changes over time. One of the most significant observed differences is an increase in Raman signal from DNA/RNA content in LPS-stimulated cells when compared to unstimulated cells. The systematic assignment of Raman spectroscopic profiles of LPS-activated cells to cellular activation assessed by cytokine gene transcription and expression opens new ways for label-free and direct immunological studies of specific pathogen recognizing receptors and their downstream signaling pathways.

中文翻译：

---

拉曼光谱以无标记方式揭示 LPS 诱导单核 THP-1 细胞中生物分子组成的变化


人类先天免疫系统能够识别脂多糖（LPS）等病原体相关分子模式，从而激活信号级联并释放不同的细胞因子。免疫细胞的激活可以通过不同的方式进行评估，这些方式可以是间接的（细胞因子释放的 ELISA），需要染色方案（流式细胞术）或细胞裂解（mRNA 分析）。在这里，拉曼光谱作为一种无损光谱方法，可以直接、无标记地监测细胞代谢、生物分子组成以及形态的变化。例如，拉曼光谱在表征 16 小时内单核 THP-1 细胞的 LPS 刺激方面具有潜力。使用两种细胞因子 TNFα 和 IL-1β 的基因转录和表达来表征细胞培养刺激模型。 1小时、3小时、8小时和16小时后生成特定拉曼光谱指纹，其编码TLR4刺激后的细胞激活模式。最普遍的光谱变化发生在 8 小时后，但 1 小时后就已经可以检测到细微的差异。空间高分辨率拉曼扫描用于生成假彩色拉曼图像，提供细胞生化状态和随时间变化的空间信息。观察到的最显着的差异之一是与未刺激的细胞相比，LPS 刺激的细胞中 DNA/RNA 含量的拉曼信号增加。 将 LPS 激活细胞的拉曼光谱图谱系统地分配给通过细胞因子基因转录和表达评估的细胞激活，为特定病原体识别受体及其下游信号通路的无标记和直接免疫学研究开辟了新途径。