Background: Revascularization after myocardial infarction (MI) induces a strong inflammatory response resulting in an increase of circulating neutrophils. Neutrophil quantities have found to be a good predictor for future adverse events. In this study, we hypothesized that the extent of morphological changes in circulating neutrophils also reflects the myocardial damage after MI and therefore relates to outcome. Methods: One hundred seven STEMI patients treated between 2009 and 2012 with at least one white blood cell count determined by an automated hematology analyzer within 24 hours after PCI were selected. This analyzer differentiates between leukocyte subsets based on morphological characteristics derived from light scatter patterns. Neutrophil morphology was compared with simultaneously measured creatine kinase (CK). Pigs (n=9) were subjected to left anterior descending artery (LAD) occlusion for 75 minutes followed by 3 days of reperfusion. Blood was collected at baseline, during ischemia and at multiple time-points during reperfusion followed by whole-blood analysis with the same hematology analyzer as above. Cardiac damage was determined by histological infarct size, 3D-echocardiography and Troponin measurements. Coronary blood sampling was performed to determine differences in neutrophil morphology between simultaneously sampled arterial and venous coronary blood. Results: In STEMI patients, a significant increase in neutrophil axial light loss (ALL) over time was seen (p<0.001) correlating with CK levels (R=0.314, p<0.001). In pigs, neutrophil ALL increased over time (p<0.001). Neutrophil ALL measured at 15 min after reperfusion correlated significantly with infarct size (R=0.745, p=0.021) and Troponin I levels (R=0.792, p=0.019). Neutrophil ALL also negatively correlated with LVEF measured by 3D-echocardiography (R=-0.760, p=0.017). Coronary sinus sampling revealed structural differences in neutrophil morphology between arterial and coronary venous blood (p=0.013), pinpointing the infarcted myocardium as the source of the observed changes. Conclusion: MI alters the morphology of circulating neutrophils in both patients and pigs in relation to the extent of damage reflected by CK and Troponin I levels. In pigs, neutrophil morphology early after reperfusion predicts infarct size and cardiac function after 3 days. Neutrophil scatter profiles might therefore prove valuable markers for the prediction of cardiac damage after MI.

Introduction: Sudden restoration of blood flow (reperfusion) after myocardial ischemia induces a strong inflammatory response that leads to additional tissue damage (reperfusion injury). In this study, we hypothesize that the extent to which the immune system is activated is not only reflected in the number of circulating leukocytes, but also in time-dependent morphological changes of subfractions, reflecting the severity of myocardial damage. Methods: We retrospectively selected 183 STEMI patients between 2009 and 2012 with at least one white blood cell count determined by an automated hematology analyzer within 24 hours after PCI. Morphological characteristics of different leukocyte subpopulations were compared with simultaneously measured cardiac enzymes. Furthermore, pigs (n=37) were subjected to left anterior descending artery occlusion for 90 minutes followed by 8 weeks of follow up. Blood was drawn at baseline, during ischemia and at multiple time points for up to 48 hours of reperfusion followed by whole-blood analysis with the same analyzer. Cardiac damage was assessed by troponin measurements and functional outcome by 3D-echocardiography. Results: In STEMI patients, a significant increase in neutrophil size (R=0.250, p<0.001) and a decrease in overall monocyte complexity (R=-0.329, p<0.001) over time was seen from the moment of PCI up to 24 hours of reperfusion. Neutrophil size significantly correlated with troponin (R=0.155, p=0.035) and CK measurements (R=0.186, p=0.005). Changes in monocyte complexity inversely correlated with troponin (R=-0.152, p=0.039) and CKMB (R=-0.217, p=0.014). In pigs, a significant correlation was seen between troponin measurements after 4.5 hours of reperfusion with neutrophil size after 90 minutes of reperfusion (R=0.468, p=0.014). Ejection fraction derived from 3D Echocardiography performed at 8 weeks of follow up was significantly inversely correlated with neutrophil size 24 hours after reperfusion (R=-0.542 p=0.011). Moreover, monocyte complexity after 90 minutes of reperfusion correlated significantly with ejection fraction measured directly post myocardial infarction (R=0.686, p<0.001). Conclusion: These results suggest that myocardial reperfusion injury not only activates the immune system but also affects the morphology of circulating cells. In a porcine model of ischemia reperfusion injury these morphological changes early after MI were predictive for impaired cardiac function at follow up.

New Findings What is the central question of this study?Are admittance‐based pressure–volume measurements useful for the assessment of cardiac function in a chronic myocardial infarction model compared with three‐dimensional echocardiography and with classical conductance measurements? What is the main finding and its importance?Baseline admittance‐based measurements show good agreement with echocardiography and classical conductance in human‐sized porcine hearts. Myocardial infarction significantly alters ventricular dimensions and pressure–volume loop measurements compared with three‐dimensional echocardiography. The novel admittance system reliably monitors changes in cardiac function after myocardial infarction. The aim of this study was to validate admittance‐based pressure–volume (PV) loop measurements for the assessment of cardiac function in a porcine model of chronic myocardial infarction. The traditional PV loop measurement technique requires hypertonic saline injections for parallel conductance correction prior to signal conversion into volume. Furthermore, it assumes a linear relationship between conductance and volume. More recently, an admittance‐based technique has been developed, which continuously measures parallel conductance and uses a non‐linear equation for volume calculation. This technique has not yet been evaluated in a large‐animal model of myocardial ischaemia. Eleven pigs underwent invasive PV measurements with the admittance system (AS) and the traditional conductance system followed by three‐dimensional echocardiography (3DE). After baseline measurements, pigs were subjected to 90 min left anterior descending coronary artery occlusion, followed by the same measurements at 8 weeks follow‐up. In the healthy heart, the AS showed good agreement with 3DE for left ventricular volumes and a reasonable correlation for ejection fraction (r= 0.756, P= 0.007). At follow‐up, an increase in end‐systolic volume was observed with 3DE (+15.4 ± 14.4 ml, P= 0.005) and the AS (+34.6 ± 36.1 ml, P= 0.010). The ejection fraction measured with 3DE (−13.2 ± 5.2%, P < 0.001) and the AS (−20.3 ± 11.2%, P < 0.001) significantly decreased. We conclude that the AS can be used for quantitative monitoring of changes in cardiac function induced by myocardial infarction and provides comparable results to 3DE, rendering it a useful tool for functional testing in large‐animal cardiac models.

Background— Toll-like receptor (TLR)-2 is an important mediator of innate immunity and ischemia/reperfusion-induced cardiac injury. We have previously shown that TLR2 inhibition reduces infarct size and improves cardiac function in mice. However, the therapeutic efficacy of a clinical grade humanized anti-TLR2 antibody, OPN-305, in a large-animal model remained to be addressed. Methods and Results— Pigs (n=38) underwent 75 minutes ischemia followed by 24 hours of reperfusion. Saline or OPN-305 (12.5, 6.25, or 1.56 mg/kg) was infused intravenously 15 minutes before reperfusion. Cardiac function and geometry were assessed by echocardiography. Infarct size was calculated as the percentage of the area at risk and by serum Troponin-I levels. Flow cytometry analysis revealed specific binding of OPN-305 to porcine TLR2. In vivo, OPN-305 exhibited a secondary half-life of 8±2 days. Intravenous administration of OPN-305 before reperfusion significantly reduced infarct size (45% reduction, P=0.041) in a dose-dependent manner. In addition, pigs treated with OPN-305 exhibited a significant preservation of systolic performance in a dose-dependent fashion, whereas saline treatment completely diminished the contractile performance of the ischemic/reperfused myocardium. Conclusions— OPN-305 significantly reduces infarct size and preserves cardiac function in pigs after ischemia/reperfusion injury. Hence, OPN-305 is a promising adjunctive therapeutic for patients with acute myocardial infarction.