In vivo imaging of reactive oxygen species after myocardial ischemia-reperfusion injury: a large animal multimodal imaging and transcriptomic study

Background Reactive oxygen species (ROS) contribute to myocardial ischemia-reperfusion injury (IRI), but in-vivo data on the spatial myocardial distribution and systemic effects of ROS after IRI remain limited. This multimodal CMR and PET/CT study aimed to non-invasively image ROS activity in a clinically-relevant swine model of IRI using [18F]ROStrace, a fluorine-18-labeled analogue of dihydroethidium (DHE), and to investigate regional changes in ROS activity in the infarcted myocardium during the subacute post-IRI phase.

IRI was induced by percutaneous occlusion of the left anterior descending artery for 90 minutes in swine (N=9). CMR and whole-body PET/CT imaging with [18F]ROStrace were performed before myocardial infarction (MI) and 3-5 days post-MI to assess ROS in non-infarct myocardium, lungs, bone marrow, spleen and skeletal muscle. Late gadolinium enhanced CMR was performed to structurally characterize infarct regions. Post-MI, in vivo [18F]ROStrace signal in infarcted myocardium was compared with remote, non-infarcted myocardium and validated via ex vivo DHE fluorescent imaging. Bulk RNA-sequencing (RNA-seq) and Gene Ontology pathway analysis were conducted on biopsies from infarct and remote myocardial tissue to identify differentially expressed genes and pathways connected to oxidative stress.

During the subacute phase following MI, [18F]ROStrace fractional uptake rate (FUR; min-1) was significantly increased in skeletal muscle, compared to baseline (0.011±0.003 vs 0.016±0.005, p=0.04), with a trend toward increased FUR in bone marrow (0.046±0.009 vs 0.056±0.011, p=0.12) and the left ventricular free wall (0.067±0.007 vs 0.073±0.010, p=0.15). Within the myocardium, [18F]ROStrace FUR ((min-1)/(mL/min/g)) was significantly higher in infarcted compared to non-infarcted myocardium regions (0.110±0.034, vs 0.148±0.035, p=0.0005). DHE staining confirmed elevated ROS levels in the infarcted myocardium. RNA-seq identified 8,707 differentially expressed genes between infarct and remote myocardium, with downregulated pathways in the infarct associated with mitochondrial function, cellular respiration, and metabolic adaptation.

This study demonstrated MI ROS imaging using [18F]ROStrace using a whole-body PET/CT scanner and structural assessment with CMR. Systemic and myocardial increases in ROS activity were observed post-MI, accompanied by substantial molecular alterations in infarcted tissue. These findings show potential imaging strategies to evaluate therapeutic targets that can mitigate oxidative stress after MI. Competing Interest Statement The authors have declared no competing interest. - Non-Standard Abbreviations and Acronyms - MI - Myocardial infarction - ROS - Reactive oxygen species - IRI - Ischemic reperfusion injury - DHE - Dihydroethidium - PET - Positron emission tomography - DHE-analog PET radiotracer - ROStrace - CMR - Cardiac magnetic resonance - LAD - Left anterior descending coronary - 82Rb - 82-Rubidium - MBF - Myocardial blood flow - AFOV - Axial field of view