Multi-tracer molecular imaging reveals critical role of macrophages for cardiac repair after acute myocardial infarction

Objectives: Myocardial infarction (MI) triggers a dynamic and organized infiltration of the damaged region by inflammatory leukocytes. Macrophages contribute to wound healing through various mechanisms, including interaction with fibroblasts to generate a stable scar. We sought to obtain further insights into the role of macrophages for healing after MI, by use of multi-tracer molecular imaging.

Methods: C57Bl6 mice received clodronate-loaded liposomes for peripheral macrophage depletion outside the bone marrow (n=30) or PBS-loaded liposomes as vehicle control (n=12). After 24h, mice underwent permanent left coronary artery ligation (MI) or sham surgery. Perfusion ^99mTc-sestamibi SPECT/CT calculated infarct sizes and cardiac magnetic resonance (CMR) assessed left ventricular function at MI+1wk and 6wks. Within the first 7d after MI, CXCR4-positive leukocyte recruitment was evaluated by ⁶⁸Ga-pentixafor and macrophage infiltration was assessed by translocator protein (TSPO) targeted ¹⁸F-GE180. At MI+4wks, microcalcification was evaluated by ¹⁸F-NaF PET/CT. Immunohistochemistry characterized the cellular substrate of the imaging signals.

Results: Macrophage depletion resulted in increased incidence of ventricular rupture compared to control MI (42% vs 16%), and in modest reduction of chronic contractile function among survivors (ejection fraction: 31.6±4.5 vs 37.3±5.9, p=0.029). Infarct sizes were comparable (38±8 vs 32±8, p=0.30). ¹⁸F-GE180 uptake in the infarct territory was lower compared to control MI at 7d (percent injected dose (ID)/g: 4.9±1.0 vs 10.5±2.9, p<0.001), while CXCR4 expression was steadily elevated (%ID/g, d1: 1.6±0.2 for depletion vs 1.2±0.1 for control MI; d3: 1.3±0.2 vs 1.0±0.2; d7: 1.1±0.4 vs 0.7±0.1; p<0.05). Immunostaining identified lower CD68+ macrophage content, but elevated and sustained recruitment of Ly6G+ neutrophils in the infarct territory. CMR identified a dense intra-cavity thrombus adherent to the infarct wall from 7d post-MI. ¹⁸F-NaF PET identified active calcification associated with the intraluminal thrombus at 4wks, which was absent in vehicle-treated MI and macrophage-depleted sham. Scar region calcification was visualized by CT in macrophage-depleted mice at MI+6wks.

Conclusions: Complete suppression of non-bone marrow macrophages results in increased ventricle rupture, thrombus formation, and tissue calcification, which is readily identified by multiparametric molecular imaging. The altered immune response is reflected by specific changes of inflammation imaging targets. These findings underscore the requirement of macrophages for effective healing. They may explain adverse response to broad anti-inflammatory therapy after acute MI, and support a role for molecular imaging to identify the individual healing response for targeted support of repair.