The broad objective of this program is to perform preclinical experimentation on animal models of MI and subsequent CHF to elucidate the mechanisms of its development and to evaluate the potential of different therapeutic modalities to limit the extent of MI and to prevent or attenuate the development of CHF. I. Beta-2 adrenergic receptors (2AR) in treatment of CHF The role of AR subtype signaling in the development of CHF is clearly important. It is widely accepted now that 1AR activation is associated with development of CHF. Recent data indicate that stimulation of 1AR is pro-apoptotic, thus, the use of 1AR antagonists became a recommended therapy for CHF. The possible role of 2AR agonists remains debatable; however, the consensus is that similar to 1AR, activation of 2AR during CHF is harmful. Recent research in LCS using single myocytes indicated that the 2AR agonist, fenoterol, possesses a unique cardioprotective property, and, in fact, is antiapoptotic. In several previously published studies we have reported the positive therapeutic effect of chronic treatment with a combination of the 2AR agonist, fenoterol and 1AR blocker, metaprolol in the rat model of post MI CHF. Proposed treatment was equal to a currently standard therapy for CHF (combination of 1AR blocker and ACE-inhibitor) with respect to mortality and exceeded the standard therapy with respect to cardiac remodeling. II. Fenoterol Stereoisomers Combination of two of enantiomers, RR and SS, are a racemic mixture which represents an actual drug that had been used clinically for treatment of asthma and for our experiments cited above. In order to refine the therapeutic effectiveness of fenoterol in the CHF model, researchers in LCS have synthesized a series of stereoisomers of fenoterol and its derivatives and characterized their receptor binding and pharmacological properties. The hypothesis had been tested that the stereochemistry of an agonist determines selectivity of receptor coupling to different G protein(s). It has been reported that in experiments in isolated rat cardiomyocytes the RR fenoterol exhibits more potent effects on cardiomyocyte contractility and better Gs protein selectivity than other stereoisomers, and had been proposed as a promising new drug. We used the rat experimental model of post-MI CHF to compare the efficacy of the RR and SS fenoterol with its racemic mixture, whose efficacy had been demonstrated in our previous experiments. Fenoterol and one of its enantiomers, RR or SS, were given to rats in drinking water as described above, starting 2 weeks after induction of MI. The LV remodeling and function were assessed during the next 5 months via serial Echo. All the effects of fenoterol described above, attenuation of LV remodeling and functional decline and arrest of MI expansion, were replicated in the present experiment, including the loss of effectiveness after 3 months as a monotherapy. Both groups treated with RR or SS were indistinguishable from the untreated group. Thus, the therapeutic potential of RR reported on the basis of experiments on single cardiomyocyte level was not confirmed in our whole animal experiments. III. Non-erythropoietic derivatives of Erythropoietin. A pyroglutamate helix B surface peptide (HBSP), a small peptide, had been engineered on the basis of erythropoietin molecule to effect tissue protection without erythropoiesis. In experiments in the rat model of MI induced by a permanent ligation of a coronary artery, we demonstrated strong cardioprotective effects of HBSP. Taking advantage of the fact that multiple injections of HBSP do not result in the elevation of hematocrit, we tested the idea that repeated injections of HBSP can attenuate the post-MI LV remodeling. We initiated a year-long experiment injecting post-MI rats twice a week with HBSP. Survival of HBSP-treated rats was significantly improved in comparison with untreated animals. For the 10 months, starting two weeks after left coronary artery ligation, rats received the i.p. injections of HBSP (60ug/kg; n=33) or saline (n=33) 2 times per week. Compared to control, saline-treated rats, HBSP treatment significantly reduced mortality. Repeated Echo demonstrated remarkable attenuation of LV chamber dilatation, LV functional deterioration and MI expansion in HPSP-treated compared to untreated rats. Blood hematocrit level measured monthly was not affected by HBSP treatment. An invasive hemodynamic assessment at the end of 10-mo treatment showed better LV systolic function and arterio-ventricular coupling in HBSP-treated compared to saline-treated rats. Histological analysis revealed less apoptosis, myocardial fibrosis, and myocyte hypertrophy in non-infarcted myocardium among HBSP-treated rats compared to saline-treated rats. Results suggest that HBSP could be considered as a safe alternative to EPO for further clinical testing in the patients with chronic heart failure. A further pathological investigation of myocardial tissue samples revealed that HBSP treatment significantly increased LV posterior wall thickness and myocyte cell density, reduced the myocyte cell hypertrophy. A classification of myocytes based on the cell diameter showed that there are significantly higher portion (43%) of small myocytes (< 20um) in LV posterior wall in HBSP treated group than in saline-treated group (23%). It indicates a possibility of myocyte regeneration by HBSP treatment in the failing heart. IV. The receptor for advanced glycation end-products (RAGE) RAGE and its soluble forms of RAGE (sRAGE) are the emerging mechanisms that involve in the pathobiology of a wide range of diseases including cardiovascular diseases. Experimental data suggest that sRAGE may neutralize the ligand-mediated damage by acting as a decoy. Our results showed a strong anti-inflammatory effect of sRAGE in rat carotid artery balloon injury model (Tae et al., J Mol Med. 2013; 91(12):1369-81). We tested whether sRAGE is cardio protective against myocardial acute ischemic injury. In an experimental model of myocardial infarction induced by permanent ligation of a coronary artery in rats, a single bolus injection of 1ug/kg of sRAGE immediately after coronary ligation, reduced myocardial infarction (MI) measured 24 h after coronary ligation by 50% compared to saline injection. V. Ivabradine Ivabradine reduces heart rate through HCN4 channel in sino-arterial nodal cells. A bradycardia and accompanied cardio-protective effects of ivabradine have been reported in preclinical animal models of heart failure and in patients with chronic heart failure. Thus, heart rate reduction was perceived as the main mechanism of cardio-protective effects of ivabradine. Our preliminary studies showed that ivabradine administration (2mg/kg) can reduce the core temperature (32.90.5 from 36.80.30C; p<0.05) as well as the heart rate (22921 from 51818 bpm; p<0.05) in normal conscious mice after i.p. administration. Based on the reports that ivabradine can suppress the activity of temperature sensory neurons isolated from hippocampus (Cai et al. Neuroscience Letters; 2012; 506:336-341), we hypothesized that ivabradine is more than a bradycardia agent, and hypothermia may play a role in its cardio-protective effects. Currently, we are investigating the roles of its bradycardia and hypothermia properties in the context of cardio-protection in a rat model of acute myocardial infarction.
