Deep Tissue Second-Harmonics of Collagen Fibers in a Transparent Rat Heart from a Myocardial Infarction Model

The combination of tissue-clearing techniques with light-sheet microscopy has enabled detailed visualization of histological changes from the micrometer to millimeter scale, deepening understanding of various disease processes. However, these protocols are not fully optimized for animal species beyond mice or for organs outside the brain. Additionally, the lack of suitable fluorescent probes for target molecules limits their broader application. In this study, we present a protocol for whole-organ clearing of rat hearts in a myocardial infarction model, achieving complete transparency and enabling label-free imaging of collagen fibers in the myocardial wall up to a depth of [~]5 mm using Second Harmonic Generation (SHG) microscopy. For the first time, we successfully compared collagen fiber orientations between infarcted and healthy myocardial regions. Our approach facilitates high-resolution tissue remodeling analysis in cardiovascular research without the need for antibody staining, demonstrating that tissue-clearing techniques are feasible even in animal species with limited available antibodies. Summary statementWe present a novel method for visualizing collagen fibers deep within the heart using a combination of tissue clearing and advanced microscopy, providing valuable data for cardiac research.