Faculty of Chemistry, Jagiellonian University, Krakow, Poland.
Analyst. 2011 Dec 21;136(24):5247-55. doi: 10.1039/c1an15311k. Epub 2011 Oct 17.
Spectroscopy-based approaches can provide an insight into the biochemical composition of a tissue sample. In the present work Fourier transform infrared (FT-IR) spectroscopy was used to develop a reliable methodology to study the content of free fatty acids, triglycerides, cholesteryl esters as well as cholesterol in aorta from mice with atherosclerosis (ApoE/LDLR(-/-) mice). In particular, distribution and concentration of palmitic, oleic and linoleic acid derivatives were analyzed. Spectral analysis of pure compounds allowed for clear discrimination between free fatty acids and other similar moieties based on the carbonyl band position (1699-1710 cm(-1) range). In order to distinguish cholesteryl esters from triglycerides a ratio of carbonyl band to signal at 1010 cm(-1) was used. Imaging of lipids in atherosclerotic aortic lesions in ApoE/LDLR(-/-) mice was followed by Hierarchical Cluster Analysis (HCA). The aorta from C57Bl/6J control mice (fed with chow diet) was used for comparison. The measurements were completed with an FT-IR spectrometer equipped with a 128 × 128 FPA detector. In cross-section of aorta from ApoE/LDLR(-/-) mice a region of atherosclerotic plaque was clearly identified by HCA, which was later divided into 2 sub-regions, one characterized by the higher content of cholesterol, while the other by higher contents of cholesteryl esters. HCA of tissues deposited on normal microscopic glass, hence limited to the 2200-3800 cm(-1) spectral range, also identified a region of atherosclerotic plaque. Importantly, this region correlates with the area stained by standard histological staining for atherosclerotic plaque (Oil Red O). In conclusion, the use of FT-IR and HCA may provide a novel tool for qualitative and quantitative analysis of contents and distribution of lipids in atherosclerotic plaque.
基于光谱的方法可以深入了解组织样本的生化组成。在本工作中，傅里叶变换红外（FT-IR）光谱用于开发一种可靠的方法来研究动脉粥样硬化小鼠（ApoE/LDLR（-/-）小鼠）主动脉中游离脂肪酸、甘油三酯、胆固醇酯以及胆固醇的含量。特别分析了棕榈酸、油酸和亚油酸衍生物的分布和浓度。纯化合物的光谱分析可以根据羰基带位置（1699-1710 cm（-1）范围内）清楚地区分游离脂肪酸和其他类似基团。为了区分胆固醇酯和甘油三酯，使用羰基带与 1010 cm（-1）处信号的比值。对 ApoE/LDLR（-/-）小鼠动脉粥样硬化主动脉病变中的脂质进行成像后，进行了层次聚类分析（HCA）。C57Bl/6J 对照小鼠（用普通饮食喂养）的主动脉用于比较。使用配备 128×128 FPA 探测器的 FT-IR 光谱仪完成测量。在 ApoE/LDLR（-/-）小鼠的主动脉横截面上，HCA 清楚地识别出动脉粥样硬化斑块区域，随后将其分为 2 个亚区，一个区域胆固醇含量较高，另一个区域胆固醇酯含量较高。在普通显微镜玻璃上沉积的组织的 HCA，因此限制在 2200-3800 cm（-1）光谱范围内，也可以识别动脉粥样硬化斑块区域。重要的是，该区域与动脉粥样硬化斑块的标准组织学染色（油红 O）染色区域相关。总之，FT-IR 和 HCA 的使用可以为动脉粥样硬化斑块中脂质的含量和分布提供一种定性和定量分析的新工具。
