Similar results have been shown in by Wang et al. [12]. These difference in results of cardiotoxicity between clinical and animal studies may be attributed to difference in hemodynamics or the rate of formation of clozapine metabolites and free radicals. The cardiotoxic effects were confirmed by elevation in the activities of serum CK-MB and LDH, the two enzymes that are

considered important markers of early and late cardiac injury, especially during clinical follow-up of drug-induced cardiotoxicities [31]. Among various hypotheses of clozapine-induced cardiotoxicity, Killian et al. [7] proposed that clozapine-induced myocarditis may result from a type I IgE-mediated acute hypersensitivity reaction. This hypothesis is supported by the onset of clozapine-induced myocarditis, selleck compound which commonly includes peripheral eosinophilia and eosinophilic myocardial infiltrates [10]. These reports are consistent with our results showing Selleckchem BTK inhibitor an increase in cardiac MPO level, which is an index of neutrophil migration, in clozapine-treated animals. Activated eosinophils induce tissue injury and necrosis through the production and release of reactive oxygen metabolites and cytotoxic proteins (e.g., proteases and MPO) into the extracellular fluid

[32]. One possible explanation of this hypothesis comes from the fact that clozapine undergoes bioactivation in myocardial tissue to a chemically reactive nitrinium ion metabolite, which stimulates cellular injury, lipid peroxidation and free radical production [33]. These results are consistent with our results showing increased cardiac levels of the lipid peroxidation product (MDA) with clozapine treatment. This nitrinium ion also binds with Bay 11-7085 proteins in the myocardium, leading to formation of an antigenic complex that stimulates the immune response and macrophages [34]. This complex subsequently

leads to myocardial cell damage via the release of free radicals and the activation of a variety of proinflammatory cytokines such as TNF-α [35]. The increase of cardiac TNF-α by clozapine is dose-dependent [12]. These findings are consistent with the results of this study. TNF-α is known to be able to attract leukocytes to inflammatory sites, enhancing the generation of reactive species [36]. Moreover, TNF-α seems to be responsible for regulating the production of some mediators that stimulate inflammatory reaction, such as NF-κβ and COX-2 [37]. The results of this study clearly showed an increase in cardiac NF-κβ levels in CLZ-treated animals. Previous studies support the concept of the involvement of TNF-α in clozapine-induced cardiotoxicity, in which clozapine can stimulate in vivo release of TNF-α and various interleukins [12] and [13]. In addition, clozapine-induced myocarditis in humans is accompanied by the release of proinflammatory cytokines, including TNF-α [38].