Evaluating leukocyte subpopulation contributions to inflammation in the heart-brain axis after myocardial infarction in mice: a multimodal molecular imaging approach

Abstract Introduction Following myocardial infarction (MI), orchestrated infiltration of diverse leukocyte subtypes contributes to local repair. Systemic and concurrent neuroinflammation is thought to contribute to cognitive impairment after MI. We have demonstrated that macrophage depletion impairs ventricle healing and exacerbates neuroinflammation early after MI, with persistence of cardiac neutrophils. Purpose We aimed to explore the role of neutrophils in neuroinflammation post-MI in macrophage-depleted mice using timed anti-neutrophil antibody dosing and multiparametric imaging. Methods Male C57Bl6N mice received two distinct treatments: initially clodronate-loaded liposomes intravenously (n=27) to deplete peripheral macrophages. Subsequently, 24h later all mice underwent permanent ligation of the left coronary artery. These mice also received treatment at 24h after MI– with anti-Ly6G therapeutic antibody (n=15) for neutrophil depletion or with an isotype control antibody (n=12). Longitudinal TSPO PET/CT with 18F-GE180 at 3d, 7d, and 8w after MI measured cardiac and brain inflammation. SPECT/CT defined infarct size and CMR calculated contractile function at 8w. Correlative analyses including flow cytometry, immunostaining, and behavioural experiments validated PET data. Results Neutrophil inhibition 24h post-MI reduced incidence of LV rupture compared to clodronate alone (33% vs. 66%). However, isotype and Ly6G treated survivors at 8w showed similar contractile functional impairment (%EF 25±9 vs 16±7%, p=0.071) and perfusion defects (%LV 37±8 vs 40±9, p=0.64). Fluorescence associated cell sorting demonstrated successful neutrophil depletion in the infarct territory (cells x103 39±11 vs 9±4, p0.01), and a higher proportion of anti-inflammatory Ly6Clow monocytes (cells x103 32±19 vs 70±10, p0.05). In anti-Ly6G and isotype control treatment groups, the 18F-GE180 signal was lower in the infarct region 3d after MI (%ID/g 5.3±1.5 vs 5.8±2.4, p=0.55), reflecting selectivity to macrophages. Despite successful neutrophil inhibition in the myocardium, neuroinflammation was not significantly lower in anti-Ly6G treated compared to isotype control macrophage depleted mice post-MI (%ID/g 2.3±0.5 vs 2.7±1.0, p=0.27). This may suggest severity of functional impairment, rather than persistent neutrophil activity, is a more prominent contributor to microglial activation after MI. Conclusion Unrestricted neutrophil activity impedes the acute healing process of the infarct, resulting in increased ventricle rupture- a phenomenon reminiscent of infarct expansion observed in patients. The healing process after MI and the associated neuroinflammation constitute a multifaceted phenomenon that cannot be solely attributed to a single proinflammatory leukocyte subpopulation i.e. neutrophils. Effective therapeutic interventions may need to target entire systems rather than focusing on individual leukocyte cell types for the prevention of cardiac and cognitive dysfunction.Figure 1.Schematic timeline Figure 2.Supporting results