Growth substrate induced functional changes elucidated by FTIR and Raman spectroscopy in in-vitro cultured human keratinocytes
Growth substrate induced functional changes elucidated by FTIR and Raman spectroscopy in in-vitro cultured human keratinocytes
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DOI：
10.1007/s00216-006-0876-5
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发表时间：
2007-03-01
影响因子：
4.3
通讯作者：
Byrne, Hugh J.
中科院分区：
文献类型：
--
作者：
Meade, Aidan D.;Lyng, Fiona M.;Byrne, Hugh J.
在体外培养的细胞系中，使用振动光谱对细胞功能进行无创测量，需要使用光谱底物，如石英、ZnSe和MirrIR等。这些底物通常与给定细胞系的原始体内细胞外环境不同，并且通常对培养的细胞系耐受性差，导致细胞的形态和功能变化。本研究证明了振动光谱分析和生化分析在评估正常人类上皮角质形成细胞细胞系（HaCaT）与MirrIR和涂有纤维连接蛋白、层粘连蛋白和明胶的石英底物的相互作用中的各种相关性。本研究的结果表明，在FTIR光谱中，细胞增殖能力和活力的定量测量与峰面积比之间存在相关性，并且在观察到的拉曼光谱的类似区域存在复制差异。在纤维连接蛋白和层粘连蛋白包被的两种光谱底物之间观察到细胞的生理差异，但在明胶包被的石英和MirrIR载玻片上观察到细胞的差异不大。这些相关性证明了光谱技术在评价系统生理特性方面的敏感性。此外，该研究表明，明胶是一种适合用于人体角质形成细胞细胞功能光谱测量的涂层，因为它提供了一种使底物附着在细胞生理学上的影响正常化的材料。这种效应可能是细胞系依赖的，建议对光谱底物和细胞系的组合进行类似的评估，这些组合将用于单独的实验。
Non-invasive measurements of cellular function in in vitro cultured cell lines using vibrational spectroscopy require the use of spectroscopic substrates such as quartz, ZnSe and MirrIR etc. These substrates are generally dissimilar to the original in vivo extracellular environment of a given cell line and are often tolerated poorly by cultured cell lines resulting in morphological and functional changes in the cell. The present study demonstrates various correlations between vibrational spectroscopic analyses and biochemical analyses in the evaluation of the interaction of a normal human epithelial keratinocyte cell line (HaCaT) with MirrIR and quartz substrates coated with fibronectin, laminin and gelatin. The findings of this study suggest that there is a correlation between quantitative measurements of cellular proliferative capacity and viability and peak area ratios in FTIR spectra, with replicated differences in similar areas of the observed Raman spectra. Differences in the physiology of cells were observed between the two spectroscopic substrates coated in fibronectin and laminin, but little differences were observed when the cells were attached to gelatin-coated quartz and MirrIR slides. The correlations demonstrate the sensitivity of the spectroscopic techniques to evaluate the physiology of the system. Furthermore the study suggests that gelatin is a suitable coating for use in spectroscopic measurements of cellular function in human keratinocytes, as it provides a material that normalises the effect of substrate attachment on cellular physiology. This effect is likely to be cell-line dependent, and it is recommended that similar evaluations of this effect are performed for those combinations of spectroscopic substrate and cell lines that are to be used in individual experiments.