Department of Pathology and Cell Regulation and ‡Department of Cardiovascular Surgery, Graduate School of Medical Science, Kyoto Prefectural University of Medicine , 465 Kajii-cho Kawaramachi-Hirokoji, Kamigyo-ku, Kyoto 602-8566, Japan.
Anal Chem. 2014 Jul 15;86(14):6903-10. doi: 10.1021/ac500592y. Epub 2014 Jun 23.
Raman spectroscopy, which provides information about molecular species and structures of biomolecules via intrinsic molecular vibrations, can analyze physiological and pathological states of tissues on the basis of molecular constituents without staining. In this study, we analyzed Raman spectra of myocardial infarction and its repair in rats using the hypothesis that the myocardium in the course of myocardial infarction and its repair could be recognized by spontaneous Raman spectroscopy on the basis of chemical changes in myocardial tissues. Raman spectra were acquired from unfixed frozen cross sections of normal and infarcted heart tissues upon excitation at 532 nm. Raman spectra of the infarcted tissues were successfully obtained at characteristic time points: days 2, 5, and 21 after coronary ligation, at which the main components of the infarcted region were coagulation necrosis, granulation tissue, and fibrotic tissue, respectively. The latent variable weights calculated by a multivariate classification method, partial least-squares-discriminant analysis (PLS-DA), revealed fundamental information about the spectral differences among the types of tissues on the basis of molecular constituents. A prediction model for the evaluation of these tissue types was established via PLS-DA. Cross-validated sensitivities of 99.3, 95.3, 96.4, and 91.3% and specificities of 99.4, 99.5, 96.5, and 98.3% were attained for the discrimination of normal, necrotic, granulation, and fibrotic tissue, respectively. A two-dimensional image of a marginal area of infarction was successfully visualized via PLS-DA. Our results demonstrated that spontaneous Raman spectroscopy combined with PLS-DA is a novel label-free method of evaluating myocardial infarction and its repair.
拉曼光谱通过分子固有振动提供生物分子的分子种类和结构信息，可以在不染色的情况下基于分子成分分析组织的生理和病理状态。在这项研究中，我们基于心肌组织的化学变化，假设通过自发拉曼光谱可以识别心肌梗死及其修复过程中的心肌，从而对大鼠的心肌梗死及其修复进行拉曼光谱分析。在 532nm 激发下，从正常和梗死心脏组织的未固定冷冻横切片中获取拉曼光谱。在结扎冠状动脉后的第 2、5 和 21 天等特征时间点成功获得了梗死组织的拉曼光谱，此时梗死区域的主要成分分别是凝固性坏死、肉芽组织和纤维化组织。通过多元分类方法（偏最小二乘判别分析，PLS-DA）计算的潜在变量权重揭示了基于分子成分的组织类型之间光谱差异的基本信息。通过 PLS-DA 建立了用于评估这些组织类型的预测模型。对正常、坏死、肉芽和纤维化组织的判别，交叉验证的灵敏度分别为 99.3%、95.3%、96.4%和 91.3%，特异性分别为 99.4%、99.5%、96.5%和 98.3%。通过 PLS-DA 成功可视化了梗死边缘区域的二维图像。我们的结果表明，自发拉曼光谱结合 PLS-DA 是一种评估心肌梗死及其修复的新型无标记方法。
