Acute myocardial infarction (AMI) initiates an intense inflammatory response that promotes cardiac dysfunction, cell death, and ventricular remodeling. The molecular events underlying this inflammatory response, however, are incompletely understood. In experimental models of sterile inflammation, ATP released from dying cells triggers, through activation of the purinergic P2X7 receptor, the formation of the inflammasome, a multiprotein complex necessary for caspase-1 activation and amplification of the inflammatory response. Here we describe the presence of the inflammasome in the heart in an experimental mouse model of AMI as evidenced by increased caspase-1 activity and cytoplasmic aggregates of the three components of the inflammasome-apoptosis speck-like protein containing a caspase-recruitment domain (ASC), cryopyrin, and caspase-1, localized to the granulation tissue and cardiomyocytes bordering the infarct. Cultured adult murine cardiomyocytes also showed the inducible formation of the inflammasome associated with increased cell death. P2X7 and cryopyrin inhibition (using silencing RNA or a pharmacologic inhibitor) prevented the formation of the inflammasome and limited infarct size and cardiac enlargement after AMI. The formation of the inflammasome in the mouse heart during AMI causes additional loss of functional myocardium, leading to heart failure. Modulation of the inflammasome may therefore represent a unique therapeutic strategy to limit cell death and prevent heart failure after AMI.interleukin-1 | NALP3 | NLRP3 | pyroptosis

Background-Experimental interleukin-1 receptor antagonist gene overexpression has shown that interleukin-1 receptor antagonist is cardioprotective during global cardiac ischemia. The aim of the present study was to test the impact of an exogenous recombinant human interleukin-1 receptor antagonist (anakinra) in experimental acute myocardial infarction. Methods and Results-Two animal studies were conducted: one of immediate anakinra administration during ischemia in the mouse and one of delayed anakinra administration 24 hours after ischemia in the rat. Seventy-eight Institute of Cancer Research mice and 20 Wistar rats underwent surgical coronary artery ligation (or sham operation) and were treated with either anakinra 1 mg/kg or NaCl 0.9% (saline). Treatment was administered during surgery and then daily for 6 doses in the mice and starting on day 2 daily for 5 doses in the rats. Twenty-eight mice underwent infarct size assessment 24 hours after surgery, 6 saline-treated mice and 22 mice treated with increasing doses of anakinra (1 mg/kg [nϭ6], 10 mg/kg [nϭ6], and 100 mg/kg [nϭ10]); 6 mice were euthanized at 7 days for protein expression analysis. The remaining animals underwent transthoracic echocardiography before surgery and 7 days later just before death. Cardiomyocyte apoptosis was measured in the peri-infarct regions. The antiapoptotic effect of anakinra was tested in a primary rat cardiomyocyte culture during simulated ischemia and in vitro on caspase-1 and -9 activities. At 7 days, 15 of the 16 mice (94%) treated with anakinra were alive versus 11 of the 20 mice (55%) treated with saline (Pϭ0.013).No differences in infarct size at 24 hours compared with saline were observed with the 1-and 10-mg/kg doses, whereas a 13% reduction in infarct size was found with the 100-mg/kg dose (Pϭ0.015). Treatment with anakinra was associated with a significant reduction in cardiomyocyte apoptosis in both the immediate and delayed treatment groups (3.1Ϯ0.2% versus 0.5Ϯ0.3% [PϽ0.001] and 4.2Ϯ0.4% versus 1.1Ϯ0.2% [PϽ0.001], respectively). Compared with saline-treated animals, anakinra-treated mice and rats showed signs of more favorable ventricular remodeling. In vitro, anakinra significantly prevented apoptosis induced by simulated ischemia and inhibited caspase-1 and -9 activities. Conclusions-Administration of anakinra within 24 hours of acute myocardial infarction significantly ameliorates the remodeling process by inhibiting cardiomyocyte apoptosis in 2 different experimental animal models of AMI. This may open the door for using anakinra to prevent postischemic cardiac remodeling and heart failure.

Sildenafil citrate (Viagra) is the pharmacological agent used to treat erectile dysfunction in men. Because this drug has a vasodilatory effect, we hypothesized that such an action may induce a preconditioning-like cardioprotective effect via opening of mitochondrial ATP-sensitive K (K(ATP)) channels. Rabbits were treated with sildenafil citrate (0.7 mg/kg iv) either 30 min (acute phase) or 24 h (delayed phase) before 30 min of ischemia and 3 h of reperfusion. Mitochondrial K(ATP) channel blocker 5-hydroxydecanoate (5-HD, 5 mg/kg iv) was given 10 min before ischemia-reperfusion. Infarct size was measured by tetrazolium staining. Sildenafil caused reduction in arterial blood pressure within 2 min of treatment, which returned to nearly baseline levels 3 min later. The infarct size (% risk area, means +/- SE) reduced from 33.8 +/- 1.7 in control rabbits to 10.8 +/- 0.9 during the acute phase (68% reduction, P < 0.05) and 19.9 +/- 2.0 during the delayed phase (41% reduction, P < 0.05). 5-HD abolished protection with an increase in infarct size to 35.6 +/- 0.4% and 36.8 +/- 1.6% during the acute and delayed phase, respectively (P < 0.05). Similar acute and delayed cardioprotective effects were observed when sildenafil was administered orally. Systemic hemodynamics also decreased after oral administration of the drug. However, these changes were mild and occurred slowly. For the first time, we demonstrate that sildenafil induces acute and delayed protective effects against ischemia-reperfusion injury, which are mediated by opening of mitochondrial K(ATP) channels.

Background-Sildenafil, a phosphodiesterase-5 inhibitor, induces cardioprotection against ischemia/reperfusion injury via opening of mitochondrial K ATP channels. It is unclear whether sildenafil would provide similar protection from doxorubicin-induced cardiotoxicity. Methods and Results-Male ICR mice were randomized to 1 of 4 treatments: saline, sildenafil, doxorubicin (5 mg/kg IP), and sildenafil (0.7 mg/kg IP) plus doxorubicin (nϭ6 per group). Apoptosis was assessed with the use of terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling and in situ oligo ligation methods. Desmin distribution was determined via immunofluorescence. Bcl-2 expression was analyzed by Western blot. Left ventricular function was assessed by measuring developed pressure and rate pressure product in Langendorff mode. ECG changes indicative of doxorubicin cardiotoxicity were also measured. For in vitro studies, adult ventricular cardiomyocytes were exposed to doxorubicin (1 mol/L), sildenafil (1 mol/L) with or without N G -nitro-L-arginine methyl ester (L-NAME) (100 mol/L), or 5-hydroxydecanoate (100 mol/L) 1 hour before doxorubicin and incubated for 18 hours. Doxorubicintreated mice demonstrated increased apoptosis and desmin disruption, which was attenuated in the sildenafilϩdoxorubicin group. Bcl-2 was decreased in the doxorubicin group but was maintained at basal levels in the sildenafilϩdoxorubicin group. Left ventricular developed pressure and rate pressure product were significantly depressed in the doxorubicin group but were attenuated in the sildenafilϩdoxorubicin group. ST interval was significantly increased in the doxorubicin group over 8 weeks. In the sildenafilϩdoxorubicin group, ST interval remained unchanged from baseline. Doxorubicin caused a significant increase in apoptosis, caspase-3 activation, and disruption of mitochondrial membrane potential in vitro. In contrast, sildenafil significantly protected against doxorubicin cardiotoxicity; however, this protection was abolished by both L-NAME and 5-hydroxydecanoate. Conclusions-Prophylactic

Expression of the STAT3 transcription factor in the heart is cardioprotective and decreases the levels of reactive oxygen species. Recent studies indicate that a pool of STAT3 resides in the mitochondria where it is necessary for the maximal activity of complexes I and II of the electron transport chain. However, it has not been explored whether mitochondrial STAT3 modulates cardiac function under conditions of stress. Transgenic mice with cardiomyocyte-specific overexpression of mitochondria-targeted STAT3 with a mutation in the DNA-binding domain (MLS-STAT3E) were generated. We evaluated the role of mitochondrial STAT3 in the preservation of mitochondrial function during ischemia. Under conditions of ischemia heart mitochondria expressing MLS-STAT3E exhibited modest decreases in basal activities of complexes I and II of the electron transport chain. In contrast to WT hearts, complex I-dependent respiratory rates were protected against ischemic damage in MLS-STAT3E hearts. MLS-STAT3E prevented the release of cytochrome c into the cytosol during ischemia. In contrast to WT mitochondria, ischemia did not augment reactive oxygen species production in MLS-STAT3E mitochondria likely due to an MLS-STAT3E-mediated partial blockade of electron transport through complex I. Given the caveat of STAT3 overexpression, these results suggest a novel protective mechanism mediated by mitochondrial STAT3 that is independent of its canonical activity as a nuclear transcription factor. STAT3 was originally identified as an IL-6-induced transcriptional activator of acute phase genes (1). However, other members of the IL-6 family, which utilize gp-130 receptor, as well as leptin, IL-12, IFN␣/␤, IL-10, GM-CSF, several growth factors, oncogenes, and stress such as hypoxia, also activate STAT3 (1). STAT3 is vital to embryonic development and STAT3-null mice are embryonic lethal (2). Analysis of tissuespecific conditional STAT3 knock-out mice has provided strong evidence that transcriptional activity of STAT3 plays a central role in the control of cell growth and host responses to inflammation and cellular stress (1). STAT3 positively regulates expression of anti-apoptotic (Bcl-2 and Bcl-xL) (1) and antioxidative proteins (MnSOD and metallothionein-1 and -2) (3, 4).Expression of STAT3 in the heart is associated with cardiac survival (5). When STAT3 is selectively deleted in cardiomyocytes, mice develop enhanced cardiac inflammation with fibrosis, dilated cardiomyopathy, and die prematurely due to congestive heart failure (5). Female mice, where STAT3 is not expressed in cardiomyocytes, develop post-partum cardiomyopathy, which is also seen in humans with reduced STAT3 expression in the myocardium (6). Ventricles from STAT3-null hearts show elevated levels of reactive oxygen species (ROS) 2 (6). Ischemic and pharmacologic preconditioning protected the viability of wild type but not STAT3 Ϫ/Ϫ cardiomyocytes (5). When STAT3 is overexpressed in cardiomyocytes, mice are less sensitive to the cardiotoxic effects of doxorubicin, which exerts i...