Péter Ferdinandy

Adaptation to myocardial stress in disease states: is preconditioning a healthy heart phenomenon?

Effective therapeutic strategies for protecting the ischaemic myocardium are much sought after. Ischaemic heart disease in humans is a complex disorder, often associated with other systemic diseases such as dyslipidaemia, hypertension and diabetes that exert multiple biochemical effects on the heart, independently of ischaemia. Ischaemic preconditioning of myocardium is a well-described adaptive response in which brief exposure to ischaemia markedly enhances the ability of the heart to withstand a subsequent ischaemic insult. The underlying molecular mechanisms of this phenomenon have been extensively investigated in the hope of identifying new rational approaches to therapeutic protection of the ischaemic myocardium. However, most studies have been undertaken in animal models in which ischaemia is imposed in the absence of other disease processes. In this article, Peter Ferdinandy, Zoltan Szilvassy and Gary Baxter review the ways in which systemic diseases might modify the preconditioning response and they emphasize the importance of further preclinical studies that specifically examine preconditioning in relation to complicating disease states.

Direct myocardial anti‐ischaemic effect of GTN in both nitrate‐tolerant and nontolerant rats: a cyclic GMP‐independent activation of KATP

We have recently demonstrated that glyceryl trinitrate (GTN) exerts a direct myocardial anti‐ischaemic effect in both GTN‐tolerant and nontolerant rats. Here we examined if this effect is mediated by GTN‐derived nitric oxide (NO) and involves guanosine 3′5′ cyclic monophosphate (cyclic GMP) and ATP‐sensitive K+ channels (KATP). Rats were treated with 100u2003mgu2003kg−1 GTN or vehicle s.c. three times a day for 3 days to induce vascular GTN‐tolerance or nontolerance. Isolated working hearts obtained from either GTN‐tolerant or nontolerant rats were subjected to 10u2003min coronary occlusion in the presence of 10−7u2003M GTN or its solvent. GTN improved myocardial function and reduced lactate dehydrogenase (LDH) release during coronary occlusion in both GTN‐tolerant and nontolerant hearts. Cardiac NO content significantly increased after GTN administration in both GTN‐tolerant and nontolerant hearts as assessed by electron spin resonance. However, cardiac cyclic GMP content measured by radioimmunoassay was not changed by GTN administration. When hearts from both GTN‐tolerant and nontolerant rats were subjected to coronary occlusion in the presence of the KATP‐blocker glibenclamide (10−7u2003M), the drug itself did not affect myocardial function and LDH release, however, it abolished the anti‐ischaemic effect of GTN. We conclude that GTN opens KATP via a cyclic GMP‐independent mechanism, thereby leading to an anti‐ischaemic effect in the heart in both GTN‐tolerant and nontolerant rats.

Lack of correlation between myocardial nitric oxide and cyclic guanosine monophosphate content in both nitrate-tolerant and -nontolerant rats

We studied the effect of nitroglycerin (NTG) on cardiac nitric oxide (NO) and cyclic guanosine monophosphate (cGMP) content in nitrate-tolerant/nontolerant rats in vivo. The effect of the pharmacological blockade of endogenous NO synthesis and the effect of exogenous NO on cardiac cGMP were also examined. Rats were treated with 100 mg/kg of NTG and corresponding vehicle s.c. three times a day for 2.5 days to induce NTG-tolerance/nontolerance. Rats were then administered a single dose of s.c. 100 mg/kg of NTG to test the effect of NTG in tolerant/nontolerant states, respectively. Nontolerant rats treated with vehicle were controls, and nontolerant rats treated with the NO synthesis inhibitor NG-nitro-L-arginine (LNNA, 20 mg/kg) were negative controls. Another group of nontolerant rats treated i.v. with the direct NO donor sodium nitroprusside (SNP, 3 mg/kg) were positive controls. Cardiac NO assessed by electron spin resonance after in vivo spin-trapping increased 100-fold (P < 0.05) in the positive control, 10-fold (P < 0.05) in the NTG-tolerant group, and 4-fold (P < 0.05) in the single NTG group, when compared to controls. In the negative control group, NO was reduced to near the detection limit (four-fold reduction, P < 0.05). Cardiac cGMP measured by radioimmunoassay was increased significantly (two-fold, P < 0.05) only in the positive control group, and there were no differences among the other groups. This shows that: 1) in vivo cardiac bioconversion of NTG to NO is not impaired in nitrate tolerance; and 2) changes in cardiac NO content are not reflected by changes in cGMP content in nitrate-tolerant and -nontolerant rats.

Rapid pacing-induced preconditioning is recaptured by farnesol treatment in hearts of cholesterol-fed rats: Role of polyprenyl derivatives and nitric oxide

We have previously shown that hypercholesterolemia leads to the loss of pacing-induced preconditioning (PC), possibly due to the impairment of cardiac nitric oxide (NO) synthesis. It has been shown that excess exogenous cholesterol inhibits formation of several polyprenyl derivatives involved in signal transduction. In the present study, we examined whether PC and cardiac NO synthesis are restored by treatment with the key polyprenyl product, farnesol, in cholesterol-fed rats. Rats fed 2% cholesterol-enriched/control diet for 24 weeks were given i.p. 5 μM/kg farnesol/vehicle, respectively. An hour later, hearts were isolated and prepared for ‘working’ perfusion, then subjected to PC/non-PC protocols of 3 intermittent periods of pacing of 5 min duration at 10 Hz, followed by a 10 min coronary occlusion to test the effect of PC. PC increased ischemic aortic flow (AF) from its control value of 15.6 ± 1.5 to 27.3 ± 1.7 mL/min (p < 0.05). PC was not observed in hearts obtained from hypercholesterolemic rats (AF: 15.7 ± 1.2 mL/min), however, it reappeared in the farnesol-treated hypercholesterolemic group (AF: 31.8 ± 3.4 mL/min, p < 0.05). In tissue samples from the left ventricle, cholesterol-diet markedly decreased the intensity of the electron spin resonance spectra of NO obtained after in vivo spin trapping with Fe2+-diethyl-dithio-carbamate complex. Farnesol-treatment did not influence cardiac NO content in the cholesterol-fed or in the control group. These results show that the lost PC can be recaptured by farnesol-treatment in hypercholesterolemia, however, farnesol-treatment does not restore cardiac NO synthesis.

Loss of preconditioning in rabbits with vascular tolerance to nitroglycerin.

A preceding right ventricular overdrive pacing (VOP) of 500 b.p.m. for 5 min, markedly reduced the severity of global myocardial ischaemia produced by a subsequent 5‐min VOP in conscious rabbits. This VOP‐induced preconditioning developed in parallel with an increase in cardiac cyclic guanosine 3′: 5′‐monophosphate (cyclic GMP) content. VOP‐induced preconditioning was abolished when the animals had been made tolerant to the vasodilator effect of nitroglycerin (NG). In the heart of the NG‐tolerant rabbits, neither VOP nor preconditioning increased cyclic GMP content. This suggests that changes by NG tolerance of cyclic GMP metabolism may account for the loss of VOP‐induced preconditioning.

Cardiac Nitric Oxide Signalling in Metabolic Syndrome

It is well documented that metabolic syndrome (i.e. a group of risk factors, such as abdominal obesity, elevated blood pressure, elevated fasting plasma glucose, high serum triglycerides and low cholesterol level in high‐density lipoprotein), which raises the risk for heart disease and diabetes, is associated with increased reactive oxygen and nitrogen species (ROS/RNS) generation. ROS/RNS can modulate cardiac NO signalling and trigger various adaptive changes in NOS and antioxidant enzyme expressions/activities. While initially these changes may represent protective mechanisms in metabolic syndrome, later with more prolonged oxidative, nitrosative and nitrative stress, these are often exhausted, eventually favouring myocardial RNS generation and decreased NO bioavailability. The increased oxidative and nitrative stress also impairs the NO‐soluble guanylate cyclase (sGC) signalling pathway, limiting the ability of NO to exert its fundamental signalling roles in the heart. Enhanced ROS/RNS generation in the presence of risk factors also facilitates activation of redox‐dependent transcriptional factors such as NF‐κB, promoting myocardial expression of various pro‐inflammatory mediators, and eventually the development of cardiac dysfunction and remodelling. While the dysregulation of NO signalling may interfere with the therapeutic efficacy of conventional drugs used in the management of metabolic syndrome, the modulation of NO signalling may also be responsible for the therapeutic benefits of already proven or recently developed treatment approaches, such as ACE inhibitors, certain β‐blockers, and sGC activators. Better understanding of the above‐mentioned pathological processes may ultimately lead to more successful therapeutic approaches to overcome metabolic syndrome and its pathological consequences in cardiac NO signalling.

The effect of continuous versus intermittent treatment with transdermal nitroglycerin on pacing-induced preconditioning in conscious rabbits.

Tolerance to the hypotensive effect of nitroglycerin (NG) blocks preconditioning induced by rapid ventricular pacing (RVP) in rabbits. In the present work the effect of continuous versus intermittent treatment with transdermal nitroglycerin on the pacing‐induced preconditioning phenomenon was studied in conscious rabbits. RVP (500 beats min−1 over 5u2003min) increased left ventricular end‐diastolic pressure (LVEDP) from baseline 4.1±0.9 to postpacing 13.8±2.9u2003mmHg (P<0.001) with a right intraventricular ST‐segment elevation of 1.25±0.13u2003mV, two indicators of myocardial ischaemia. These changes were significantly attenuated when the RVP period was preceded by a preconditioning pacing of the same rate and duration with an interpacing interval of 5u2003min. Protection by preconditioning was abolished when the animals had been made tolerant to the vasodilator effect of 30u2003μgu2003kg−1 NG by the application of transdermal NG (approx. 0.07u2003mgu2003kg−1u2003h−1) over 7 days. Furthermore, transdermal NG per se attenuated both RVP‐induced ST‐segment elevation and LVEDP‐increase over the 7 day period. With intermittent transdermal NG treatment (12u2003h ‘patch on’ vs ‘patch off’), neither development of vascular tolerance nor attenuation of the NG‐ or preconditioning‐induced anti‐ischaemic effects were observed. However, the severity of pacing‐induced myocardial ischaemia was significantly increased during the ‘patch off’ periods. In a second set of experiments, postpacing changes in cardiac cyclic GMP and cyclic AMP levels were determined by means of radioimmunoassay in chronically instrumented anaesthetized open‐chest rabbits with the same NG‐treatment protocols. Preconditioning reduced postpacing increase in cyclic AMP with an increase in cyclic GMP concentrations in hearts of the untreated animals and in those given patches intermittently during both ‘patch on’ and ‘patch off’ periods. However, the preconditioning effect on either cyclic nucleotide was blocked in the tolerant animals. Transdermal NG increased resting levels of both cardiac cyclic nucleotides in the non‐tolerant but not in the tolerant state. The postpacing increase in cyclic AMP content was inhibited by transdermal NG, independent of vascular tolerance development, whereas, an increase in cyclic GMP content was exclusively seen in the non‐tolerant animals. We conclude that the anti‐ischaemic effect of NG is independent of the cyclic GMP mechanism in the tolerant state. While intermittent NG therapy prevents development of vascular tolerance and preserves preconditioning, the nitrate‐free periods yield an increased susceptibility of the heart to ischaemic challenges.

In vivo and in vitro acute cardiovascular effects of bimoclomol.

Effects of bimoclomol, the novel heat shock protein (HSP) coinducer, was studied in various mammalian cardiac and rabbit aortic preparations. Bimoclomol decreased the ST-segment elevation induced by coronary occlusion in anesthetized dogs (56% and 80% reduction with 1 and 5 mg/kg, respectively). In isolated working rat hearts, bimoclomol increased coronary flow (CF), decreased the reduction of cardiac output (CO) and left ventricular developed pressure (LVDP) developing after coronary occlusion, and prevented ventricular fibrillation (VF) during reperfusion. In rabbit aortic preparations, precontracted with phenylephrine, bimoclomol induced relaxation (EC(50)=214 microM). Bimoclomol produced partial relaxation against 20 mM KCl, however, bimoclomol failed to relax preparations precontracted with serotonin, PGF(2) or angiotensin II. All these effects were evident within a few minutes after application of bimoclomol. A rapid bimoclomol-induced compartmental translocation of the already preformed HSPs may explain the protective action of the compound.

Long-Term but not Short-Term Cardioprotection can be Induced by Preconditioning in Hypercholesterolemia

Preconditioning of the heart by single or multiple noninjurious ischemic stimuli may induce both short-term (up to 2 hours) and long-term (3 to 24–48 hours) cardiac protection against the consequences of a subsequent severe stress. Up to now, however, both short-term and long-term protection has been demonstrated only in normal, metabolically healthy animals.

Involvement of glibenclamide-sensitive potassium channels in vasorelaxation by cochlear nerve stimulation.

Rabbit aortic rings relaxed with an increase in cyclic guanosine monophosphate and cyclic adenosine monophosphate content in response to exposure to organ fluid of isolated cochleas of the guinea pig following field stimulation (50 Hz, 80 V, 0.2 ms). Relaxations were blocked by 30 μMNG-nitro-l-arginine methyl ester added to the vessel rings. This inhibitory effect was reversed by 3 MMl-arginine. Removal of the vascular endothelium also blocked the relaxation response. Glibenclamide attenuated vasorelaxation in a concentration-dependent manner. We conclude that cochlear nerve stimulation induces an endothelium-dependent vasorelaxation involving activation of adenosine triphosphate-sensitive potassium channels.