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Ischaemic heart disease remains a leading cause of premature mortality and morbidity. Understanding the associated pathophysiological mechanisms of cardiac dysfunction arising from ischaemic heart disease and the identification of sites of novel therapeutic intervention requires a preclinical model that reproduces the key clinical characteristics of myocardial ischaemia, reperfusion and infarction. Here we describe and validate a refined and minimally invasive translationally relevant approach to induce ischaemia, reperfusion and infarction in the sheep. The protocol uses clinical cardiology devices and approaches and would be readily adopted by researchers with access to standard fluoroscopic instrumentation. In addition to being minimally invasive, the major refinements associated with the described methodology are the implantation of an intracardiac defibrillator prior to coronary engagement and use of an antiarrhythmic medication protocol during the procedure. These refinements lead to a reduction of intraoperative mortality to 6.7 %. The model produces key characteristics associated with the 4thUniversal Definition of Myocardial Infarction including electrocardiographic changes, elevated cardiac biomarkers and cardiac wall motility defects. In conclusion, the model closely replicates the clinical paradigm of myocardial ischaemia, reperfusion and infarction in a translationally relevant large-animal setting and the applied refinements reduce the incidence of intraoperative mortality typically associated with preclinical myocardial infarction models.

The highly organized transverse tubule (t-tubule) network facilitates cardiac excitation–contraction coupling and synchronous cardiac myocyte contraction. In cardiac failure secondary to myocardial infarction (MI), changes in the structure and organization of t-tubules result in impaired cardiac contractility. However, there is still little knowledge on the regional variation of t-tubule remodelling in cardiac failure post-MI. Here, we investigate post-MI t-tubule remodelling in infarct border and remote regions, using serial block face scanning electron microscopy (SBF-SEM) applied to a translationally relevant sheep ischaemia reperfusion MI model and matched controls. We performed minimally invasive coronary angioplasty of the left anterior descending artery, followed by reperfusion after 90 min to establish the MI model. Left ventricular tissues obtained from control and MI hearts eight weeks post-MI were imaged using SBF-SEM. Image analysis generated three-dimensional reconstructions of the t-tubular network in control, MI border and remote regions. Quantitative analysis revealed that the MI border region was characterized by t-tubule depletion and fragmentation, dilation of surviving t-tubules and t-tubule elongation. This study highlights region-dependent remodelling of the tubular network post-MI and may provide novel localized therapeutic targets aimed at preservation or restoration of the t-tubules to manage cardiac contractility post-MI. This article is part of the theme issue ‘The cardiomyocyte: new revelations on the interplay between architecture and function in growth, health, and disease’.

The highly organised transverse tubule (t-tubule) network consisting of invaginations of the cell sarcolemma facilitates synchronous cardiac myocyte contraction. This study aimed to investigate post-myocardial infarction (MI) t-tubule remodelling in infarct border and remote regions in a translationally relevant ischaemia reperfusion injury MI model. Six adult sheep were used in this study (n=3 MI, n=3 control). Eight weeks after MI, left ventricular tissue was collected from the remote and border MI regions and from control sheep, processed and imaged using 3D scanning electron microscopy. The t-tubule network was manually segmented using 3dmod. One-way ANOVA with Tukey’s post-hoc correction, unpaired t-tests or Mann-Whitney U test were used where appropriate. Marked disorganisation of the t-tubule network was observed in the border region following MI. Quantitative analysis revealed that in comparison to the control sheep myocardium, the MI border zone had a decreased t-tubule count (0.07 ± 0.007 tubules per μm3 in control vs 0.05 ± 0.004 tubules per μm3 in border; p = 0.02) and showed t-tubule dilation (405 ± 22 nm in control vs 533 ± 30 nm in border; p = 0.02). Whilst there was minimal disorganisation and loss of t-tubules in the MI remote region, we observed increased t-tubule length as a fraction of the cell diameter (0.41 ± 0.04 in control vs 0.56 ± 0.04 in remote; p = 0.045). In addition to gross t-tubule remodelling, we also noted post-MI fragmentation of t-tubules, particularly in the border region. In comparison to control, the number of t-tubule fragments per μm3 was increased in the post MI heart (control, 0.17 ± 0.1 fragments per μm3; border, 2.21 ± 0.7 fragments per μm3; remote, 1.20 ± 0.4 fragments per μm3; p = 0.04 border vs control; p = 0.02 remote vs control). The volume occupied by fragments as a percentage of the cell volume was also higher following MI (control, 0.003 ± 0.002 %; border, 0.071 ± 0.023 %; remote 0.014 ± 0.005 %; p = 0.003 border vs control; p = 0.013 border vs remote). Whilst there was no difference in fragments density between the remote and border regions, there was an increase in the volume of cell occupied by fragments in the MI border region compared to remote. This is explained by a larger average fragment volume in the border region (0.04 ± 0.006 μm3 in border vs 0.01 ± 0.002 μm3 in remote; p Our research shows remodelling of the t-tubule network in the post-MI sheep myocardium. We noted reduced t-tubule count, t-tubule fragmentation, and dilation of remaining t-tubules. Importantly our work shows that these changes occur in a regional manner, being most pronounced in the border region. These changes may reflect regional wall stresses post-MI, and we speculate that our observations may result in region-specific changes to systolic calcium and contractility post-MI. Conflict of Interest Authors declare that there is no conflict of interest.