Craig R. Hampton

Inhibition of Toll-like Receptor 4 With Eritoran Attenuates Myocardial Ischemia-Reperfusion Injury

Background— We previously reported that the functional mutation of Toll-like receptor 4 (TLR4) in C3H/HeJ mice subjected to myocardial ischemia-reperfusion (MI/R) injury resulted in an attenuation of myocardial infarction size. To investigate the ligand-activating TLR4 during MI/R injury, we evaluated the effect of eritoran, a specific TLR4 antagonist, on MI/R injury, with the goal of defining better therapeutic options for MI/R injury. Methods and Results— C57BL/6 mice received eritoran (5 mg/kg) intravenously 10 minutes before 30 minutes of in situ of transient occlusion of the left anterior descending artery, followed by 120 minutes of reperfusion. Infarct size was measured using triphenyltetrazoliumchloride staining. A c-Jun NH2-terminal kinase (JNK) activation was determined by Western blotting, nuclear factor (NF)-&kgr;B activity was detected by gel-shift assay, and cytokine expression was measured by ribonuclease protection assay. Mice treated with eritoran developed significantly smaller infarcts when compared with mice treated with vehicle alone (21.0±6.4% versus 30.9±13.9%; P=0.041). Eritoran pretreatment resulted in a reduction in JNK phosphorylation (eritoran versus vehicle: 3.98±0.81 versus 7.01±2.21-fold increase; P=0.020), less nuclear NF-&kgr;B translocation (2.70±0.35 versus 7.75±0.60-fold increase; P=0.00007), and a decrease in cytokine expression (P<0.05). Conclusions— We conclude that inhibition of TLR4 with eritoran in an in situ murine model significantly reduces MI/R injury and markers of an inflammatory response.

Protease-Activated Receptor-1 Contributes to Cardiac Remodeling and Hypertrophy

Background— Protease-activated receptor-1 (PAR-1) is the high-affinity receptor for the coagulation protease thrombin. It is expressed by a variety of cell types in the heart, including cardiomyocytes and cardiac fibroblasts. We have shown that tissue factor (TF) and thrombin contribute to infarct size after cardiac ischemia-reperfusion (I/R) injury. Moreover, in vitro studies have shown that PAR-1 signaling induces hypertrophy of cardiomyocytes and proliferation of cardiac fibroblasts. The purpose of the present study was to investigate the role of PAR-1 in infarction, cardiac remodeling, and hypertrophy after I/R injury. In addition, we analyzed the effect of overexpression of PAR-1 on cardiomyocytes. Methods and Results— We found that PAR-1 deficiency reduced dilation of the left ventricle and reduced impairment of left ventricular function 2 weeks after I/R injury. Activation of ERK1/2 was increased in injured PAR-1−/− mice compared with wild-type mice; however, PAR-1 deficiency did not affect infarct size. Cardiomyocyte-specific overexpression of PAR-1 in mice induced eccentric hypertrophy (increased left ventricular dimension and normal left ventricular wall thickness) and dilated cardiomyopathy. Deletion of the TF gene in cardiomyocytes reduced the eccentric hypertrophy in mice overexpressing PAR-1. Conclusions— Our results demonstrate that PAR-1 contributes to cardiac remodeling and hypertrophy. Moreover, overexpression of PAR-1 on cardiomyocytes induced eccentric hypertrophy. Inhibition of PAR-1 after myocardial infarction may represent a novel therapy to reduce hypertrophy and heart failure in humans.

Tissue factor and thrombin mediate myocardial ischemia-reperfusion injury

Reperfusion of the ischemic heart is necessary to prevent irreversible injury of the myocardium, which leads to permanent organ dysfunction. However, reperfusion in itself leads to myocardial ischemia/reperfusion (I/R) injury, which is characterized by an acute inflammatory response mediated by activated inflammatory cells, chemokines, cytokines, and adhesion molecules. The molecular mechanisms of myocardial I/R injury are not completely known. Tissue factor (TF) and thrombin, two potent procoagulant and proinflammatory mediators, are recognized to play significant roles in myocardial I/R injury. To investigate the role of TF and thrombin in myocardial I/R injury, we used rabbit and murine in situ coronary artery ligation models. Increased TF mRNA, antigen, and activity were found in ischemic cardiomyocytes. Administration of an inhibitory antirabbit TF monoclonal antibody before or during the onset of ischemia resulted in a significant reduction in infarct size. Functional inhibition of thrombin with hirudin also reduced the infarct size. However, defibrinogenating rabbits with ancrod had no effect on infarct size, suggesting a requirement of thrombin generation but not fibrin deposition in myocardial I/R injury.

Microvascular Inflammatory Response in Cardiac Surgery

Cardiac surgical procedures, with or without cardiopulmonary bypass, elicit a systemic inflammatory response in patients that induces the elaboration of multiple cytokines, chemokines, adhesion molecules, and destructive enzymes. This inflammatory reaction involves multiple interdependent and redundant cell types and humoral cascades, which allows for amplification and positive feedback at numerous steps. This systemic inflammatory response ultimately results in a broad spectrum of clinical manifestations, with multiple organ failure being the most severe form. Investigative efforts have focused on understanding the mechanism of this systemic inflammatory response syndrome in order to develop potential therapeutic targets to inhibit it, thereby possibly decreasing postoperative morbidity and mortality. Multiple therapeutic methods have been investigated, including pharmacologic inhibitors and modifications of surgical technique and the cardiopulmonary bypass circuit. Although studies have demonstrated that the use of these therapies in experimental and clinical settings has attenuated the systemic inflammatory response, they have failed to conclusively show clinical benefit from these therapies. These therapies may be too specific to minimize the deleterious effects of a systemic inflammatory response that results from the activation of multiple, interdependent, and redundant inflammatory cascades and cell types. Hence, further studies that investigate the molecular and cellular events underlying the systemic inflammatory response syndrome and the resultant effects of anti-inflammatory therapies are warranted to ultimately achieve improvements in clinical outcome after cardiac surgical procedures.

Molecular Mechanisms of Neurologic Injury Following Cardiopulmonary Bypass

Neurologic injury is a potentially devastating consequence of heart surgery. Between 1% and 5% of patients undergoing cardiopulmonary bypass have postoperative strokes and 30% to 80% of patients demonstrate some neurologic dysfunction postoperatively. This review focuses on anatomic, molecular and clinical markers of neurologic injury following cardiopulmonary bypass. Attention is directed to the molecular mechanisms underlying neurologic injury and clinical biochemical markers of injury during heart surgery. Novel strategies to modulate injury are also discussed.

Endothelial Injury During Minimally Invasive Bypass Grafting

Over the last few decades, the vascular endothelium has emerged as a central mediator of the biochemical events that underlie the preoperative, operative, and postoperative course of nearly all patients who undergo cardiovascular interventional procedures and cardiovascular surgery. While initially felt to be a passive bystander of the whole-body response to cardiac surgery, it is now clear that the endothelium is a dynamic organ which is a central regulator of vascular tone, vasomotor function, coagulation, and cellular interactions. Although in the physiologically unstressed state the role of the endothelium is to maintain intravascular homeostasis, it also serves a critical role in the response to injury. Specifically, in terms of cardiovascular surgery, the endothelium modulates the systemic inflammatory response, changing vasomotor tone, impacting coagulation responses, initiating the intimal hyperplastic response, and mediating the chronic changes leading to atherosclerosis.