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Toll-like receptors are expressed in immune cells and cardiac muscle. We examined whether the cardiac Toll-like receptor 4 (TLR4) is involved in the acute myocardial dysfunction caused by septic shock and myocardial ischemia (MI). We used wild type mice (WT), TLR4 deficient (TLR4-ko) mice and chimeras that underwent myeloablative bone marrow transplantation to dissociate between TLR4 expression in the heart (TLR4-ko/WT) and the immunohematopoietic system (WT/TLR4-ko). Mice were injected with lipopolysaccharide (LPS) (septic shock model) or subjected to coronary artery ligation (MI model) and tested in vivo and ex vivo, for function, histopathology proinflammatory cytokine and TLR4 expression. WT mice challenged with LPS or MI displayed reduced cardiac function, increased myocardial levels of IL-1 beta and TNF-alpha and upregulation of mRNA encoding TLR4 prior to myocardial leukocyte infiltration. TLR4 deficient mice sustained significantly smaller infarctions as compared to control mice at comparable areas at risk. The cardiac function of TLR4-ko mice was not affected by LPS and demonstrated reduced suppression by MI compared to WT. Chimeras deficient in myocardial TLR4 were resistant to suppression induced by LPS and the heart function was less depressed, compared to the TLR4-ko, following MI in the acute phase (4h). In contrast, hearts of chimeras deficient in immunohematopoietic TLR4 expression were suppressed both by LPS and MI, exhibiting increased myocardial cytokine levels, similar to WT mice. We concluded that cardiac function of TLR4-ko mice and chimeric mice expressing TLR4 in the immunohematopoietic system, but not in the heart, revealed resistance to LPS and reduced cardiac depression following MI, suggesting that TLR4 expressed by the cardiomyocytes themselves plays a key role in this acute phenomenon.

Previously we found that uridine-5'-triphosphate (UTP) significantly decreased cultured cardiomyocyte death, induced by hypoxia via activating P2Y(2) receptors, reduced infarct size and maintained higher ATP levels in an in vivo model. Mitochondrial contribution to the progression of cardiomyocyte injury in ischemia/hypoxia is well known. However, the protective effects of UTP in cardiac cells with a respiratory chain deficiency are poorly elucidated. The aim of our study was to further define the role of UTP on mitochondrial functional tolerance following chemical and/or ischemic stress in in vivo and in vitro models. Cardiac mitochondrial function was tested 24 h post left anterior descending (LAD) ligation in UTP (0.44 microg/kg)-treated rats. UTP's beneficial effect in LAD-ligated hearts was expressed by improved mitochondrial activity (Complexes I, II and IV). In the in vitro model, cultured cardiomyocytes were pretreated with 50 microM UTP prior to hypoxic and/or chemical stress with rotenone or sodium azide. Pretreatment with UTP maintained increased ATP levels as well as mitochondrial membrane potential and reduced lactate dehydrogenase (LDH) release. A modest reduction (12%) in the mitochondrial membrane potential was demonstrated when the cultured cardiomyocytes were subjected to UTP. This reduction was abolished by the P2Y receptor antagonist, reactive blue 2, but not with 5 hydroxydecanoate, a mitochondrial K(ATP) channel inhibitor, or by BAPTA-AM, the intracellular calcium chelator. We suggest that UTP may act as an uncoupling agent, which exerts a modest mitochondrial depolarization, resulting in a reduction of Ca(2+) uptake, preserving mitochondrial activity, thereby reducing cell damage during hypoxia.

The purpose of this study was to investigate whether vitamin D 3 deficiency affects the cardiac function of chick hearts directly or whether the influence is secondary through the hormone's effect on serum calcium levels. To this end, three experimental groups were studied: (a) the control group of vitamin D 3 supplemented chicks, D s , (b) vitamin D 3 deficient chicks, D d , and (c) vitamin D 3 supplemented hypocalcemic chicks raised on decreased calcium levels, D h . The three groups were compared by checking hemodynamic, metabolic and membrane parameters. Total and ionized serum calcium concentrations in the D h and D d groups were found to be lower than in the D s group. Perfusion of the isolated hearts with solutions containing various calcium concentrations (1, 1.5 and 2.5 mM) induced enhanced contractility levels, the magnitude of which was dependent on the difference between the in vivo and perfusate calcium levels. Thus, the inotropic effect was similar and more enhanced for the two hypocalcemic groups. The differences in hemodynamic behaviour could not be explained by variations in the levels of the high energy compounds and acidity, since similar ATP, creatine phosphate and intracellular pH levels were detected for both D s and D d groups. However, membrane studies revealed an increase in the number of slow calcium channels for the two hypocalcemic groups and this may be the possible mechanism for the differences in the contractile activity. In conclusion, our study strongly suggests that the effects of vitamin D 3 on the heart is mediated only indirectly through its effect on serum calcium levels.