A.M. Perna

Novel drug development opportunity for relaxin in acute myocardial infarction: evidences from a swine model

The hormone relaxin has been shown to cause coronary vasodilation and to prevent ischemia/reperfusion‐induced cardiac injury in rodents. This study provides evidence that relaxin, used as an adjunctive drug to coronary reperfusion, reduces the functional, biochemical, and histopathological signs of myocardial injury in an in vivo swine model of heart ischemia/reperfusion, currently used to test cardiotropic drugs for myocardial infarction. Human recombinant relaxin, given at reperfusion at doses of 1.25, 2.5, and 5 µg/kg b.wt. after a 30‐min ischemia, caused a dose‐related reduction of key markers of myocardial damage (serum myoglobin, CK‐MB, troponin T) and cardiomyocyte apoptosis (caspase 3, TUNEL assay), as well as of cardiomyocyte contractile dysfunction (myofibril hypercontraction). Compared with the controls, relaxin also increased the uptake of the viability tracer 201Thallium and improved ventricular performance (cardiac index). Relaxin likely acts by reducing oxygen free radical‐induced myocardial injury (malondialdehyde, tissue calcium overload) and inflammatory leukocyte recruitment (myeloperoxidase). The present findings show that human relaxin, given as a drug to counteract reperfusion‐induced cardiac injury, affords a clear‐cut protection to the heart of swine with induced myocardial infarction. The findings also provide background to future clinical trials with relaxin as adjunctive therapy to catheter‐based coronary angioplasty in patients with acute myocardial infarction.

Paracrine effects of transplanted myoblasts and relaxin on post-infarction heart remodelling.

In the post‐infarcted heart, grafting of precursor cells may partially restore heart function but the improvement is modest and the mechanisms involved remain to be elucidated. Here, we explored this issue by transplanting C2C12 myoblasts, genetically engineered to express enhanced green fluorescent protein (eGFP) or eGFP and the cardiotropic hormone relaxin (RLX) through coronary venous route to swine with experimental chronic myocardial infarction. The rationale was to deliver constant, biologically effective levels of RLX at the site of cell engraftment. One month after engraftment, histological analysis showed that C2C12 myoblasts selectively settled in the ischaemic scar and were located around blood vessels showing an activated endothelium (ICAM‐1‐,VCAM‐positive). C2C12 myoblasts did not trans‐differentiate towards a cardiac phenotype, but did induce extracellular matrix remodelling by the secretion of matrix metalloproteases (MMP) and increase microvessel density through the expression of vascular endothelial growth factor (VEGF). Relaxin‐producing C2C12 myoblasts displayed greater efficacy to engraft the post‐ischaemic scar and to induce extracellular matrix re‐modelling and angiogenesis as compared with the control cells. By echocardio‐graphy, C2C12‐engrafted swine showed improved heart contractility compared with the ungrafted controls, especially those producing RLX. We suggest that the beneficial effects of myoblast grafting on cardiac function are primarily dependent on the paracrine effects of transplanted cells on extracellular matrix remodelling and vascularization. The combined treatment with myoblast transplantation and local RLX production may be helpful in preventing deleterious cardiac remodelling and may hold therapeutic possibility for post‐infarcted patients.

Human Recombinant Relaxin Reduces Heart Injury and Improves Ventricular Performance in a Swine Model of Acute Myocardial Infarction

Abstract: This study shows that relaxin can be effective in the treatment of acute myocardial infarction. In a swine model of heart ischemia‐reperfusion currently used to test cardiotropic drugs because of its similarities with human myocardial infarction, human recombinant relaxin (2.5 and 5 μg/kg body weight), given at reperfusion after a 30‐min ischemia, markedly reduced the main serum markers of myocardial damage (myoglobin, CK‐MB, and troponin T) and the metabolic and histopathologic parameters of myocardial inflammation and cardiomyocyte injury, resulting in overall improvement of ventricular performance (increased cardiac index) compared to the controls. These results provide a background for future clinical trials with human relaxin as adjunctive therapy to catheter‐based coronary angioplasty in patients with acute myocardial infarction.

Mast cell inhibition and reduced ventricular arrhythmias in a swine model of acute myocardial infarction upon therapeutic administration of relaxin

Resident cardiac mast cells, located mainly around coronary vessels and in the right atrium close to the sinoatrial node, are the main repository of histamine in the heart [1]. Infl ammatory activation of cardiac mast cells, as occurs upon acute myocardial infarction, causes the release of histamine and prostanoids [2]. These substances lead to severe tachyarrhythmias, cardiodepression and coronary spasm, thus contributing to myocardial injury and early, lethal outcome [2]. The peptide relaxin (RLX), known to inhibit mast cell and basophil activation [3,4], has been recently validated as a cardiotropic hormone [5], being produced by the heart and acting on specifi c cardiac receptors [6]. The present study was aimed at evaluating whether the cardioprotective effect of RLX on myocardial ischemia/reperfusion-induced damage described in previous studies [7] also involves the inhibition of cardiac mast cell activation.

Possible role of acylphosphatase, Bcl-2 and Fas/Fas-L system in the early changes of cardiac remodeling induced by volume overload.

To identify early adaptive processes of cardiac remodeling (CR) in response to volume overload, we investigated the molecular events that may link intracellular Ca(2+) homeostasis alterations and cardiomyocyte apoptosis. In swine heart subjected to aorto-cava shunt for 6, 12, 24, 48 and 96 h sarcoplasmic reticulum (SR) Ca(2+) pump activity was reduced until 48 h (-30%), but a recovery of control values was found at 96 h. The decrease in SR Ca(2+)-ATPase (SERCA2a) expression at 48 h, was more marked (-60%) and not relieved by a subsequent recovery, while phospholamban (PLB) concentration and phosphorylation were unchanged at all the considered times. Conversely, acylphosphatase activity and expression significantly increased from 48 to 96 h (+40%). Bcl-2 expression increased significantly from 6 to 24 h, but at 48 h, returned to control values. At 48 h, microscopic observations showed that overloaded myocardium underwent substantial damage and apoptotic cell death in concomitance with an enhanced Fas/Fas-L expression. At 96 h, apoptosis appeared attenuated, while Fas/Fas-L expression was still higher than control values and cardiomyocyte hypertrophy became to develop. These data suggest that in our experimental model, acylphosphatase could be involved in the recovery of SERCA2a activity, while cardiomyocyte apoptosis might be triggered by a decline in Bcl-2 expression and a concomitant activation of Fas.