Label-free evaluation of myocardial infarction and its repair by spontaneous Raman spectroscopy.

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.