Histological Evaluation of Skeletal Muscle Atrophy

All histological experiments were completed as previously described by us [36 (link),105 (link)]. To determine whether IRI had atrophic effects on skeletal muscle and, subsequently, whether BGP-15 adjuvant therapy could rescue any atrophy, we histologically assessed TA muscles which were cryopreserved in optimal cutting temperature compound (Sakura Finetek, Maumee, OH, USA). TAs were sectioned (10 µm, −20°C, Leica CM1950, Leica Biosystems, Mount Waverley, Australia) and mounted onto glass slides and then stained with hematoxylin and eosin (H&E) to evaluate muscle fiber size through CSA. All slides were imaged on a Zeiss Axio Imager Z2 microscope (Carl Zeiss MicroImaging GmbH, Oberkochen, Germany) at 20× magnification and analyzed as described previously [36 (link),105 (link)] using ImageJ software (NIH, Bethesda, MD, USA).