Heart tissues from a normal rat, myocardial infarcted rat, and mesenchymal stem cell (MSC)-treated rat after myocardial infarction (MI) were studied using ToF-SIMS, with the assistance of principal component analysis (PCA). To our knowledge, this is the first such published study. In PCA results obtained from both positive and negative ion spectra of normal (control), MI, and MSC-treated MI tissues, the normal and MSC-treated tissues could be clearly distinguished from the MI tissue in terms of chemical composition. For the MI tissue, fatty acid signals from inside the cell and phosphocholine signals from the cell membrane decreased while glucose signals increased. In the MSC-treated MI tissue, the signal levels of phosphocholine and fatty acids returned to those for normal tissue. Our observations for myocardium tissues are consistent with previous biological studies, which show that the loss of reversibility of cell injury associated with MI could cause degradation to the cell membrane, and that MI could slow fatty acid uptake and oxidation and increase glucose uptake and utilization for ATP synthesis in myocardium. This is, of course, because fatty acids need oxygen for β-oxidation. We anticipate that this label-free ToF-SIMS method partnered with PCA will be helpful to the investigation of myocardial disease and MSC therapy since it is based on chemical information at the tissue level.