Biochemical basis of ischemic heart injury and of cardioprotective interventions.

Cardioprotective interventions are defined as interventions able to increase myocardial resistance to ischemia. The authors approach the issue of cardioprotection on the basis of the present knowledge about the biochemical mechanisms responsible for the injury produced by myocardial ischemia or ischemia-reperfusion. Reversible and irreversible injury are distinguished. The former is largely accounted for by the direct consequences of reduced ATP synthesis, which causes decreased ATP phosphorylation potential, acidosis and phosphate accumulation. The biochemical mechanisms leading to irreversible injury include osmotic overload, production of toxic lipid metabolites, cytosolic calcium overload, and generation of reactive oxygen species, which lead to membrane disruption, mitochondrial dysfunction and possibly to the activation of apoptotic pathways. The major effect of the classical cardioprotective agents (nitrates, beta adrenergic antagonists, calcium channel blockers) consists in affecting ATP demand/supply ratio in such a way as to delay the decrease in ATP phosphorylation potential. Other drugs have been introduced, which allegedly interfere directly with the mechanisms responsible for irreversible ischemic injury. These include 3-ketoacyl-CoA tiolase inhibitors, modulators of intracellular calcium channels, ionic exchanger inhibitors, free radical scavengers, caspase inhibitors, purinergic agonists, K(+)(ATP) channel openers, and modulators of mitochondrial permeability transition. The results obtained with these substances in experimental models and in the clinical setting are discussed. Special attention is devoted to angiotensin converting enzyme inhibitors, whose direct cardioprotective properties has recently been demonstrated.

Ghrelin tissue distribution: comparison between gene and protein expression.

Ghrelin, the natural ligand of the GH secretagogue (GHS) receptor, was originally isolated from the stomach and detected in several tissues, but a systematic study of its tissue distribution has not been performed. In the present investigation, we evaluated ghrelin gene expression (by RT-PCR technique) and ghrelin protein concentration (by enzyme immunoassay technique) in tissues obtained from control rats as well as in rats subjected to 48-h fasting. The ghrelin gene was expressed in stomach, small intestine, brain, cerebellum, pituitary, heart, pancreas, salivary gland, adrenal, ovary and testis, with maximum expression occurring in the stomach, while no significant expression was detected by standard RT-PCR in liver, lung, kidney and skeletal muscle. Ghrelin protein was detected in stomach, small intestine, brain, cerebellum, pituitary, lung, skeletal muscle pancreas, salivary gland, adrenal, ovary and testis, at concentrations ranging from 0.05 to 1.43 ng/mg of homogenate protein (the highest concentration occurred in the lung, followed by the brain). Ghrelin was not detectable in the heart, liver and kidney. Therefore, gene and protein expression were dissociated. Fasting did not produce significant changes in ghrelin gene expression, while the distribution of ghrelin between different tissues was significantly modified: protein concentration increased in the brain, cerebellum, lung and salivary gland, while it decreased in the stomach.

A3 adenosine receptor stimulation modulates sarcoplasmic reticulum Ca(2+) release in rat heart.

OBJECTIVE: Stimulation of A3 adenosine receptors has been shown to protect cardiac myocytes from ischemic injury, but the mechanism of this action is unknown. We evaluated the effect of adenosine agonists and antagonists on the sarcoplasmic reticulum (SR) Ca(2+) channels. METHODS: Isolated rat hearts were perfused with control buffer or different adenosine agonists and antagonists. Hearts were then homogenized and used to determine SR Ca(2+)-induced Ca(2+) release, assayed by quick filtration technique after loading with 45Ca(2+), and the binding of [3H]ryanodine, a specific ligand of the SR Ca(2+) release channel. In parallel experiments, hearts were challenged with 30 min of global ischemia and 120 min of reperfusion, and the extent of tissue necrosis was evaluated by triphenyltetrazolium chloride staining. RESULTS: Perfusion with the A1>A3 agonist R-PIA and the A3>A1 agonist IB-MECA was associated with reduced [3H]ryanodine binding, due to reduced B(max) (by about 20%), whereas K(d) and Ca(2+)-dependence of the binding reaction were unaffected. These actions were abolished by the A3 antagonist MRS 1191, while they were not affected by A1 and A2 antagonists. The rate constant of SR Ca(2+) release decreased by 25-30% in hearts perfused with R-PIA or IB-MECA. Tissue necrosis was significantly reduced in the presence of R-PIA or IB-MECA. Protection was removed by MRS 1191, and it was not affected by A1 and A2 antagonists. Hearts were also protected by administration of dantrolene, a ryanodine receptor antagonist. In the presence of dantrolene, no further protection was provided by IB-MECA. CONCLUSION: A3 adenosine receptor stimulation modulates the SR Ca(2+) channel. This action might account for the protective effect of adenosine.

Effect of MEN 10755, a new disaccharide analogue of doxorubicin, on sarcoplasmic reticulum Ca(2+) handling and contractile function in rat heart.

1. The use of anthraquinone antineoplastic agents is limited by their cardiac toxicity, which is largely due to activation of the sarcoplasmic reticulum (SR) Ca(2+) release channel (ryanodine receptor). MEN 10755 is a new disaccharide analogue of doxorubicin. We have evaluated its effects on SR function and its toxicity in isolated working rat hearts. 2. In rat SR vesicles, doxorubicin stimulated [(3)H]-ryanodine binding by increasing its Ca(2+)-sensitivity. At 1 microM Ca(2+), ryanodine binding increased by 15.3+/-2.5 fold, with EC(50)=20.6 microM. Epirubicin produced a similar effect, i.e. 9.7+/-0.6 fold stimulation with EC(50)=11.1 microM. MEN 10755 increased ryanodine binding by 1.9+/-0.3 fold (P:<0.01 vs doxorubicin and epirubicin), with EC(50)=38.9 microM. 3. Ca(2+)-induced Ca(2+) release experiments were performed by quick filtration technique, after SR loading with (45)Ca(2+). At 2 microM Ca(2+), doxorubicin (50 microM) increased the rate constant of Ca(2+) release to 82+/-5 s(-1) vs a control value of 22+/-2 s(-1) (P:<0.01), whereas 50 microM MEN 10755 did not produce any significant effect (24+/-3 s(-1)). 4. Ca(2+)-ATPase activity and (45)Ca(2+)-uptake were not significantly affected by doxorubicin, its 13-dihydro-derivative, epirubicin, MEN 10755 and the 13-dihydro-derivative of MEN 10755, at concentrations < or =100 microM. 5. In isolated heart experiments, administration of 30 microM doxorubicin or epirubicin caused serious contractile impairment, whereas 30 microM MEN 10755 produced only minor effects. 6. In conclusion, in acute experiments MEN 10755 was much less cardiotoxic than equimolar doxorubicin or epirubicin. This result might be accounted for by reduced activation of SR Ca(2+) release.