“Neutrophil-Mediated Injury May Play a Role in the Increased Severity of Myocardial Infarction in the ZDF Model of Type 2 Diabetes” 

ABSTRACT
Introduction – Diabetic patients have significantly more severe and fatal heart attacks than non-diabetic patients. In fact, cardiovascular disease is the leading cause of death among people with diabetes, accounting for two-thirds to three-fourths of all diabetes-related deaths. Inflammation may exacerbate ischemic heart disease, yet the mechanisms are not clear. Our lab has previously demonstrated that neutrophil trapping and sequestration in the microcirculation plays a significant role in ischemia/reperfusion (I/R) injury. The effects of neutrophil accumulation are not solely confined to mechanical plugging of capillaries, the so-called “no-reflow” phenomenon. Neutrophils also produce oxidants and pro-inflammatory cytokines that can enhance tissue injury. The purpose of this study was to determine if the severity of myocardial infarction in the type 2 diabetic heart is associated with neutrophil accumulation. 
 
Methods – Non-diabetic Zucker Lean Control (ZLC) and Zucker Diabetic Fatty (ZDF) rats underwent left anterior descending (LAD) coronary artery occlusion for 30min followed by 120min of reperfusion. Blood was sampled from the femoral artery pre-ischemia and incubated with a FITC-conjugated anti-CD11b antibody for detection of activated neutrophils using a FACSCaliber flow cytometer. At the end of reperfusion, cardiac tissue samples were freeze-fixed, sectioned, and stained for neutrophils using Napthol AS-D Chloroacetate Esterase. 
 
Results – There was significantly greater CD11b expression on ZDF neutrophils compared to their lean controls (ZLC: 12.3 ± 2.1 TFI; ZDF: 17.1 ± 1.9 TFI; p<0.05), suggesting that neutrophils from diabetic blood are in a pre-activated or “primed” state. Indeed, there was significantly greater neutrophil accumulation in the post-ischemic left ventricle of the ZDF compared to the ZLC (ZLC: 53.0 ± 9.4; ZDF: 96.8 ± 16.4; p<0.05).  These results parallel infarct size measurements, a common index of myocardial injury, in which the “infarct-to-area at risk” (I/AAR) percentage of the ZDF was significantly greater than control (ZLC: 25.7% I/AAR; ZDF: 56.9% I/AAR; p<0.05). 
 
Conclusions – These findings indicate that increased neutrophil accumulation in the left ventricle of the ZDF heart following I/R injury may result from a chronic pre-activation of diabetic neutrophils and may contribute to the increased myocardial injury observed in type 2 diabetes.