Postischemic Cardiac Dysfunction Research Articles

Antispasmodic, cardioprotective and blood-pressure lowering properties of Gomphrena perennis L. and its mechanisms of action

BackgroundGomphrena perennis L. is a native plant of South America whose pharmacological properties have not been studied yet. AimTo evaluate the cardiovascular and intestinal pharmacological effects of Gomphrena perennis L. leaves tincture (GphT) and the mechanisms involved. Experimental procedureThe chromatographic profile of GphT was done. Its ex vivo effects were evaluated by contractile concentration-response curves (CRCs) obtained from the agonist carbachol or calcium found in isolated rat small intestine, as well as in the relaxant CRCs. Cardiac effects were evaluated on isolated rat hearts exposed to ischemia/reperfusion (I/R). Experiments in vivo were performed to evaluate the diuretic activity in conscious rats and the hypotensive effect in anaesthetised rats. ResultsFifteen flavonoids were identified in GphT by HPLC-UV, including diosmin. GphT induced a non-competitive inhibition in both carbachol and calcium CRCs on rat small intestine. The first was not affected by indomethacin. Moreover, GphT, unlike diosmin, relaxed the contracture produced by a high-potassium solution in a dose-dependently way. Neither propranolol nor l-NAME changed it. GphT did not show diuretic activity but induced hypotension insensitive to l-NAME. While GphT perfusion of isolated hearts increased injury consequent to I/R, oral administration was cardioprotective and reversed by l-NAME. However, diosmin did not improve the post-ischemic recovery. ConclusionsThis study supports the use of Gomphrena perennis L. tincture as an antispasmodic and hypotensive agent. Moreover, it has been demonstrated to be preventive of post-ischemic cardiac dysfunction. However, diosmin would not be responsible for these effects.

Efficacy of dexmedetomidine on myocardial ischemia/reperfusion injury in patients undergoing cardiac surgery with cardiopulmonary bypass: A protocol for systematic review and meta-analysis.

Cardiac surgery using cardiopulmonary bypass has been shown to cause reversible postischemic cardiac dysfunction and is associated with reperfusion injury and myocardial cell death. Therefore, it is very important to have a series of measures in place to reduce oxygen consumption and provide myocardial protection. We performed a protocol for systematic review and meta-analysis to evaluate the effect of dexmedetomidine administration on myocardial ischemia/reperfusion injury in patients undergoing cardiac surgery with cardiopulmonary bypass. This review protocol is registered in the PROSPERO International Prospective Register of systematic reviews, registration number CRD42023386749. A literature search is performed in January 2023 without restriction to regions, publication types or languages. The primary sources were the electronic databases of PubMed, Embase, Web of Science, the Cochrane Central Register of Controlled Trials, Chinese National Knowledge Infrastructure database, Chinese Biomedical Database, and Chinese Science and Technology Periodical database. Risk of bias will be assessed according to the Cochrane Risk of Bias Tool. The meta-analysis is performed using Reviewer Manager 5.4. The results of this meta-analysis will be submitted to a peer-reviewed journal for publication. This meta-analysis will evaluate the efficacy and safety of dexmedetomidine in patients undergoing cardiac surgery with cardiopulmonary bypass.

Abstract P2104: Adeno-associated Virus Serotype 5 Is A Suitable Vector For S100a1-based Gene Therapy Of Post-ischemic Chronic Cardiac Dysfunction

Introduction: For S100A1-based heart failure gene therapies, AAV9 and 6 have shown efficacy in pre-clinical large animal studies. As AAV9 has shown concerning signs of toxicity in clinical studies and AAV6 displays poor production yields, there is need for a novel safe and cardiac-specific AAV serotype. Hypothesis: We hypothesized that in a pig model the safety proven and scalably manufacturable AAV5 may be a suitable vector for S100A1-based gene therapy of post-ischemic cardiac dysfunction. Methods: AAV production, 2h balloon-occlusion of the LCX, retrograde cardiac gene delivery, cardiac MRI, late gadolinium enhancement (LGE), global T1 relaxation, qPCR, RNA-Seq, WGCNA, KEGG, Reactome, LAD-ligation mouse model Results: In a comparative study of AAV5-, 6- and 9-luciferase (luc) in healthy farm pigs (n=5 each; 1x10 13 vgc/pig), AAV5 achieved a more homogeneous cardiac apical-basal transduction pattern than AAV6 with a higher luc activity than AAV9. In a clinically relevant endpoint driven study, we demonstrated a significant improvement in EF (+19 ± 5 %) 12 weeks after retrograde AAV5- S100A1 gene delivery compared to AAV5-luc in infarcted pigs (n=4 each; 1x10 13 vgc/pig). Moreover, S100A1 -treated pigs showed significantly less infarct extension (-0.5 ± 0.3 g vs. 5 ± 1.3 g (luc)) measured by cardiac MRI. There were no unfavorable alterations in blood chemistry or ECG. S100A1 expression was predominantly contained to the heart. The WGCNA unveiled a significant correlation between the improved EF and a suppression of inflammatory and immunological pathways (r=-0.96, p < 0.01) and between the absent infarct extension and enhanced activity of cardioprotective signaling (r=-0.82, p < 0.05). With injections of 2х10 11 vgc of AAV5- S100A1 or AAV5-gfp (n=4 each) into the remote myocardium in the mouse model, we confirmed a significant improvement in FS (+43.8 ± 8.8 %, vs. gfp) and suppression of inflammatory gene expression including i.e., IL1b or TNFa by S100A1. Conclusion: We conclude that AAV5 is suitable for S100A1-based gene therapy of post-ischemic cardiac dysfunction and that this vector/target combination can help accelerating the way towards a clinical trial. We also found novel signaling pathways that may be involved in S100A1’s therapeutic actions.

Phosphoproteomic analysis identifies phospho-Threonine-17 site of phospholamban important in low molecular weight isoform of fibroblast growth factor 2-induced protection against post-ischemic cardiac dysfunction

RationaleAmong its many biological roles, fibroblast growth factor 2 (FGF2) protects the heart from dysfunction and damage associated with an ischemic attack. Our laboratory demonstrated that its protection against myocardial dysfunction occurs by the low molecular weight (LMW) isoform of FGF2, while the high molecular weight (HMW) isoforms are associated with a worsening in post-ischemic recovery of cardiac function. LMW FGF2-mediated cardioprotection is facilitated by activation of multiple kinases, including PKCalpha, PKCepsilon, and ERK, and inhibition of p38 and JNK. ObjectiveYet, the substrates of those kinases associated with LMW FGF2-induced cardioprotection against myocardial dysfunction remain to be elucidated. Methods and resultsTo identify substrates in LMW FGF2 improvement of post-ischemic cardiac function, mouse hearts expressing only LMW FGF2 were subjected to ischemia-reperfusion (I/R) injury and analyzed by a mass spectrometry (MS)-based quantitative phosphoproteomic strategy. MS analysis identified 50 phosphorylation sites from 7 sarcoendoplasmic reticulum (SR) proteins that were significantly altered in I/R-treated hearts only expressing LMW FGF2 compared to those hearts lacking FGF2. One of those phosphorylated SR proteins identified was phospholamban (PLB), which exhibited rapid, increased phosphorylation at Threonine-17 (Thr17) after I/R in hearts expressing only LMW FGF2; this was further validated using Selected Reaction Monitoring-based MS workflow. To demonstrate a mechanistic role of phospho-Thr17 PLB in LMW FGF2-mediated cardioprotection, hearts only expressing LMW FGF2 and those expressing only LMW FGF2 with a mutant PLB lacking phosphorylatable Thr17 (Thr17Ala PLB) were subjected to I/R. Hearts only expressing LMW FGF2 showed significantly improved recovery of cardiac function following I/R (p < 0.05), and this functional improvement was significantly abrogated in hearts expressing LMW FGF2 and Thr17Ala PLB (p < 0.05). ConclusionThe findings indicate that LMW FGF2 modulates intracellular calcium handling/cycling via regulatory changes in SR proteins essential for recovery from I/R injury, and thereby protects the heart from post-ischemic cardiac dysfunction.

Abstract 340: Myocyte-borne Bdnf is Essential to Limit Post-ischemic Cardiac Injury and Dysfunction

Brain-derived neurotrophic factor (BDNF)/ tyrosine receptor kinase B (TrkB) signaling is essential for normal cardiac contraction/relaxation. Alterations in this pathway, i.e., defective neuronal BDNF, account for post-ischemic cardiac injury. Less clear, however, is whether myocyte-borne BDNF has a role in this setting. We generated myocyte-selective BDNF knock-out (myoBDNF -/- ) mice, using Myh6-Cre mice crossed with BDNF floxed mice, confirming bdnf deletion via RT-PCR in isolated myocytes. Hearts from 12-wk old myoBDNF -/- mice and WT littermates underwent global ischemia (30 min) and reperfusion (2 h). At this age, the two strains had similar left ventricular (LV) sizes and fractional shortening [63±1.1 (WT) vs. 60±1.2% (myoBDNF -/- )]. At reperfusion, myoBDNF -/- hearts displayed larger infarct size compared to WT (38±3 vs. 14±2%, n=9, p&lt;.0001) and worsened LV functional recovery ( Fig. 1 ). For example, the rate-pressure product recuperated only by 14±1.5 in myoBDNF -/- mice vs. 36±3% in WT (p&lt;.0001). The two groups had similar heart rates at 2 h reperfusion, however myoBDNF -/- mice markedly lost their contractility [dP/dt max = 436±58 vs. 1407±142 mmHg/sec (WT), p&lt; .0001], likely due to the exacerbated myocyte loss. Accordingly, post-ischemic troponin I release was significantly higher in myoBDNF -/- than in WT mice (0.9±0.04 vs. 1.3±0.04 ng/ml, p&lt;.0002). Thus, deleting bdnf in myocytes severely limits recovery after ischemia, directly linking myocyte-borne BDNF to the heart response to injury. Therefore, preserving or enhancing autologous myocyte BDNF generation offers new avenues to counter cardiac ischemic injury and subsequent heart failure progression.

Relevant preclinical model of heart failure in rat: Impact of myocardial infarction and hypertension

In acute myocardial infarction (MI) setting, successful revascularization of the ischemic myocardium improves prognosis. However, MI remains a leading cause of heart failure (HF) associated with mortality [1] and the risk to develop HF is increased by co-morbidities especially hypertension [2], [3]. Relevant preclinical models of heart failure with comorbidities as in human are needed to improve the selection of new therapeutic strategies. to determine the best HF model by comparing left ventricular (LV) dysfunction and signs of HF in 3 preclinical models of MI in normotensive and hypertensive rats. In total, 60 minutes MI was induced by coronary ligation in male rats (8-week old) followed by reperfusion (I/R) or not (permanent, PMI) in normotensive Wistar rats (Wi) or in hypertensive rats (SHR PMI). Ejection fraction (EF, %) and LV end-diastolic volume (LVEDV, μl) were measured using echocardiography. Signs of heart failure were assessed by LV end-diastolic pressure (LVEDP, mmHg), right ventricle weight (RV, mg) and plasma NT-Pro-BNP (pg/ml). These different parameters were assessed post-MI at 3 months for the groups (Wi Sham, Wi I/R and SHR PMI) and at 6 months for the Wi PMI group. MI led to a clear LV dysfunction and adverse eccentric remodeling. As expected, these alterations were limited by reperfusion of the ischemic myocardium in normotensive rats. Hypertension exacerbated post ischemic cardiac dysfunction. Three months post-MI, signs of HF were observed only in SHR PMI while no sign were detected in Wi rats 6 months after MI. in line with human disease, our results show that hypertensive rats are more prone to develop signs of HF after MI than normotensive rats. SHR PMI should be considered as a relevant preclinical model for the efficacy assessment of new HFrEF therapeutic strategies.

(-)-Epigallocatechin-3-gallate attenuates myocardial injury induced by ischemia/reperfusion in diabetic rats and in H9c2 cells under hyperglycemic conditions.

(−)-Epigallocatechin gallate (EGCG) exerts multiple beneficial effects on cardiovascular performance. In this study, we aimed to examine the effects of EGCG on diabetic cardiomyopathy during myocardial ischemia/reperfusion (I/R) injury. EGCG (100 mg/kg/day) was administered at week 6 for 2 weeks to diabetic rats following the induction of type 1 diabetes by streptozotocin (STZ). At the end of week 8, the animals were subjected to myocardial I/R injury. The EGCG-elicited structural and functional effects were analyzed. Additionally, EGCG (20 μM) was administered for 24 h to cultured cardiac H9c2 cells under hyperglycemic conditions (30 mM glucose) prior to hypoxia/reoxygenation (H/R) challenge, and its effects on oxidative stress were compared to H9c2 cells transfecteed with silent information regulator 1 (SIRT1) small interfering RNA (siRNA). In rats with STZ-induced diabetes, EGCG treatment ameliorated post-ischemic cardiac dysfunction, decreased the myocardial infarct size, apoptosis and cardiac fibrosis, and reduced the elevated lactate dehydrogenase (LDH) and malonaldehyde (MDA) levels, and attenuated oxidative stress. Furthermore, EGCG significantly reduced H/R injury in cardiac H9c2 cells exposed to high glucose as evidenced by reduced apoptotic cell death and oxidative stress. The protein expression levels of SIRT1 and manganese superoxide dismutase (MnSOD) were reduced in the diabetic rats and the H9c2 cells under hyperglycemic conditions, compared with the control rats following I/R injury and H9c2 cells under normal glucose conditions. EGCG pre-treatment significantly upregulated the levels of htese proteins in vitro and in vivo. However, treatment with EX527 and SIRT1 siRNA blocked the EGCG-mediated cardioprotective effects. Taken together, our data indicate that SIRT1 plays a critical role in the EGCG-mediated amelioration of I/R injury in diabetic rats, which suggests that EGCG may be a promising dietary supplement for the prevention of diabetic cardiomyopathy.

Effects of Total Flavone from Rhododendron simsii Planch. Flower on Postischemic Cardiac Dysfunction and Cardiac Remodeling in Rats.

This study investigated the effect of total flavone from Rhododendron simsii Planch. flower (TFR) on postischemic cardiac dysfunction and ventricular remodeling and was to test the hypothesis that TFR has an antiventricular remodeling effect through inhibition of urotensin-II receptor- (UTR-) mediated activation of RhoA-ROCK pathways. Twenty-four hours after ligation of the left anterior descending coronary artery, male Sprague-Dawley rats were randomized to receive 4-week treatment with saline (model group) or TFR. Compared to the model group, TFR treatment restored cardiac function, attenuated cardiomyocyte hypertrophy, and reduced interstitial fibrosis. Expression levels of several fibrosis-related factors, including alpha-smooth muscle actin, transforming growth factor-beta 1, matrix metalloproteinase-2, and collagen type I, were increased after MI. TFR treatment attenuated the upregulation of these factors, downregulated UTR expression, and markedly diminished the expression of RhoA and ROCK1/2. These results suggested that TFR could improve cardiac function and ameliorate ventricular remodeling through blocking UTR-mediated activation of RhoA-ROCK pathways in myocardial infarction rats.

N-acetylcysteine attenuates myocardial dysfunction and postischemic injury by restoring caveolin-3/eNOS signaling in diabetic rats.

BackgroundPatients with diabetes are prone to develop cardiac hypertrophy and more susceptible to myocardial ischemia–reperfusion (I/R) injury, which are concomitant with hyperglycemia-induced oxidative stress and impaired endothelial nitric oxide (NO) synthase (eNOS)/NO signaling. Caveolae are critical in the transduction of eNOS/NO signaling in cardiovascular system. Caveolin (Cav)-3, the cardiomyocytes-specific caveolae structural protein, is decreased in the diabetic heart in which production of reactive oxygen species are increased. We hypothesized that treatment with antioxidant N-acetylcysteine (NAC) could enhance cardiac Cav-3 expression and attenuate caveolae dysfunction and the accompanying eNOS/NO signaling abnormalities in diabetes.MethodsControl or streptozotocin-induced diabetic rats were either untreated or treated with NAC (1.5 g/kg/day, NAC) by oral gavage for 4 weeks. Rats in subgroup were randomly assigned to receive 30 min of left anterior descending artery ligation followed by 2 h of reperfusion. Isolated rat cardiomyocytes or H9C2 cells were exposed to low glucose (LG, 5.5 mmol/L) or high glucose (HG, 25 mmol/L) for 36 h before being subjected to 4 h of hypoxia followed by 4 h of reoxygenation (H/R).ResultsNAC treatment ameliorated myocardial dysfunction and cardiac hypertrophy, and attenuated myocardial I/R injury and post-ischemic cardiac dysfunction in diabetic rats. NAC attenuated the reductions of NO, Cav-3 and phosphorylated eNOS and mitigated the augmentation of O2 −, nitrotyrosine and 15-F2t-isoprostane in diabetic myocardium. Immunofluorescence analysis demonstrated the colocalization of Cav-3 and eNOS in isolated cardiomyocytes. Immunoprecipitation analysis revealed that diabetic conditions decreased the association of Cav-3 and eNOS in isolated cardiomyocytes, which was enhanced by treatment with NAC. Disruption of caveolae by methyl-β-cyclodextrin or Cav-3 siRNA transfection reduced eNOS phosphorylation. NAC treatment attenuated the reductions of Cav-3 expression and eNOS phosphorylation in HG-treated cardiomyocytes or H9C2 cells. NAC treatment attenuated HG and H/R induced cell injury, which was abolished during concomitant treatment with Cav-3 siRNA or eNOS siRNA.ConclusionsHyperglycemia-induced inhibition of eNOS activity might be consequences of caveolae dysfunction and reduced Cav-3 expression. Antioxidant NAC attenuated myocardial dysfunction and myocardial I/R injury by improving Cav-3/eNOS signaling.

Attenuated recovery of contractile function in aging hearts following global ischemia/reperfusion: Role of extracellular HSP27 and TLR4.

While cardiac functional recovery is attenuated in the elderly following cardiac surgery with obligatory global myocardial ischemia/reperfusion (I/R), the underlying mechanism remains incompletely understood. We observed previously that human and mouse myocardium releases heat shock protein (HSP) 27 during global I/R. Extracellular HSP27 induces myocardial inflammatory response and plays a role in post-ischemic cardiac dysfunction in adult mouse hearts. This study was to determine the role of extracellular HSP27 and Toll-like receptor 4 (TLR4) in the attenuated functional recovery in aging mouse hearts following global I/R. Hearts isolated from aging (18-24 months) and adult (4-6 months) mice were subjected to ex vivo global I/R. Augmented release of HSP27 in aging hearts is associated with greater production of cytokines (TNF-α and IL-1β) and worse functional recovery. Anti-HSP27 suppressed the inflammatory response and markedly improved functional recovery in aging hearts. Perfusion of recombinant HSP27 to aging hearts resulted in greater cytokine production and more severe contractile depression in comparison to adult hearts. TLR4 deficiency abolished cytokine production and functional injury in aging hearts exposed to recombinant HSP27. Interestingly, aging hearts had higher TLR4 protein levels and displayed enhanced TLR4-mediated NF-κB activation following HSP27 stimulation or I/R. Extracellular HSP27 and TLR4 jointly enhance the inflammatory response and hamper functional recovery following I/R in aging hearts. The enhanced inflammatory response to global I/R and attenuated post-ischemic functional recovery in aging hearts is due, at least in part, to augmented myocardial release of HSP27 and elevated myocardial TLR4 levels.

Caveolin‐3/AdipoR1 dependent Mitochondrial STAT3 Activation in Ischemic Postconditioning Cardioprotection

Ischemic postconditioning (IPo) protects against myocardial ischemic reperfusion (IR) injury by activating signal transducer and activator of transcription 3 (STAT3) which requires adiponectin (APN) (FASEB, 2014, 28: 663.7), but how APN activates STAT3 during IPo is unclear. Caveolin-3 (Cav3), a protein of the lipid rafts/caveolae, is essential in APN receptor 1 (AdipoR1) transmembrane signaling. We postulated that IPo activates mitochondrial STAT3 (mitoSTAT3) via Cav3 AdipoR1 signaling. Wild type (WT) and APN knockout (KO) mice were subjected to sham operation or IR (30min of coronary artery occlusion and 2 hours of reperfusion) with or without IPo (3 cycles of 10s of reperfusion and 10s of reocclusion). KO mice exhibited more severe myocardial injury (increased post-ischemic infarction and poorer functional recovery; P<0.05 vs. WT), impaired mitochondrial function and reduced mitoSTAT3 phosphorylation (p-mitoSTAT3). IPo attenuated post-ischemic cardiac injury and mitochondrial dysfunction and increased p-mitoSTAT3 in WT but not in KO mice. IPo increased Cav3 in buoyant membrane fraction and increased colocolazation of Cav3 with AdipoR1 but not AdipoR2 in both WT and KO mice (P<0.05 vs. IR). In cultured cardiac H9C2 cell, hypoxic postconditioning (HPo) attenuated hypoxia/reoxygenation (HR) induced cell injury and restored mitochondrial transmembrane potential, concomitant with increased p-mitoSTAT3 and Cav3 (P<0.05, vs. HR). Cav3 or AdipoR1 but not AdipoR2 gene knockdown abolished the above HPo beneficial effects. APN supplementation restored HPo protection in cells with APN gene knockdown but not in cells with Cav3 or AdipoR1 gene knockdown. It is concluded that IPo conferred cardioprotection through Cav3/AdipoR1 dependent mitoSTAT3 activation.

Exercise Mitigates Aberrant Mitophagy and Cardiovascular Remodeling in Diabetes

Previously, we reported aberrant mitophagy contributed to cardiac failure and inhibition of mitochondrial fission was cardioprotective. Cardiac connexins (Cxs), play crucial role in ischemic preconditioning, myocyte coupling, and serve to regulate post ischemic cardiac dysfunction and apoptosis. Diabetic cardiomyopathy is associated with abnormal mitophagy and decreased Cxs expression. Although moderate exercise is reported to be beneficial in cardiovascular diseases, it is not known whether exercise can mitigate aberrant mitophagy and induce Cxs in diabetic cardiomyopathy. We hypothesize that exercise decreases the aberrant mitochondrial fragmentation (mitophagy), increases Cx-37, -43 expression and attenuates cardiovascular dysfunction in diabetes. To verify our hypothesis, we used db/db obese type 2 diabetic and control mice. Mice underwent 8 wks of moderate exercise on treadmill. Blood pressure (BP) by tail cuff method showed decreased mean BP and echocardiography demonstrated improved left ventricula...

Abstract 13879: Ischemic Postconditioning Confers Cardioprotection through Adiponectin-dependent Mitochondrial STAT3 Activation in Mice

Introduction: Signal transducer and activator of transcription 3 (STAT3) plays a key role in postconditioning (IPo) mediated protection against myocardial ischemia reperfusion injury, but the mechanism by which IPo activates STAT3 is unknown. Adiponectin (APN), a protein with anti-ischemic properties, activates STAT3. We hypothesized that IPo activates mitochondrial STAT3 (MitoSTAT3) via APN signaling. Methods and Results: Wild type (WT) and APN knockout (KO) mice were either sham operated or subjected to 30 min of coronary artery occlusion followed by 2 hours of reperfusion with or without IPo (3 cycles of 10 seconds reperfusion and 10 seconds reocclusion; n=8/group). At the end of reperfusion, KO mice exhibited more severe myocardial injury evidenced as increased infarct size (% of area at risk) 49.2±2.0 vs WT 39.4±3.5, P &lt;0.01; plasma troponin I (ng/ml): KO 72.8±7.6 vs WT 45.7±4.0, P &lt;0.01; worse cardiac function (lower dP/dt max and end-systolic pressure-volume relation, P &lt;0.05); more severely impaired mitochondrial function (reductions in complex IV and complex V protein expression) and more severe reduction of MitoSTAT3 phosphorylation (activation) at site Ser727, P &lt;0.01. IPo significantly attenuated post-ischemic cardiac injury and dysfunction with a concomitant increase in phosphorylated MitoSTAT3 and attenuation of mitochondrial dysfunction in WT (all P &lt;0.05) but not in KO mice. In cultured cardiac H9C2 cells, hypoxic postconditioning (HPo, 3 cycles of 5 min hypoxia and 5 min reoxygenation) significantly attenuated hypoxia/reoxygenation (HR, 3 hours hypoxia/3 hours reoxygenation) induced cell injury (increased apoptotic cell death as % of HR): HR 100.2±0.4 vs HPo 78.2±4.8, P &lt;0.05) and reduced mitochondrial transmembrane potential (% total cells, HR 37.2±4.9 vs HPo 23.5±3.7, P &lt;0.01). APN, adiponectin receptor 1 (AdipoR1), or STAT3 gene knockdown but not AdipoR2 gene knockdown, respectively, abolished HPo cellular protection (all P &lt;0.05 vs. HPo). APN supplementation (10μg/ml) restored HPo protection in cells with APN knockdown but not in cells with AdipoR1or STAT3 gene knockdown. Conclusion: Adiponectin and AdipoR1 signaling are required for IPo to activate myocardial mitochondrial STAT3 to confer cardioprotection.

Mitigation of postischemic cardiac contractile dysfunction by CaMKII inhibition: effects on programmed necrotic and apoptotic cell death

While Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) has been suggested to be an important protein regulating heart function upon ischemia/reperfusion (I/R), the mechanisms responsible are not fully known. Furthermore, it is not known whether CaMKII activation can modulate necroptosis, a recently described form of programmed cell death. In order to investigate these issues, Langendroff-perfused rat hearts were subjected to global ischemia and reperfusion, and CaMKII inhibition was achieved by adding the CaMKII inhibitor KN-93 (0.5μmol/dm(3)) to the perfusion solution before the induction of ischemia. Immunoblotting was used to detect changes in expression of proteins modulating both necroptotic and apoptotic cell death. CaMKII inhibition normalized I/R induced increases in expression of necroptotic RIP1 and caspase-8 along with proteins of the intrinsic apoptotic pathway, namely cytochrome c and caspase-9. In addition, it increased the Bcl-2/Bax ratio and reduced caspase-3 and cleaved PARP1 content suggesting reduction of cell death. These changes coexisted with improvement of postischemic contractile function. On the other hand, there was no correlation between levels of pT287-CaMKIIδ and LVDP recovery after I/R. These results demonstrate for the first time that CaMKII inhibition may mitigate cardiac contractile dysfunction, at least partially, by limiting the contents of not only apoptotic, but also necroptotic proteins. Phosphorylation of CaMKII seems unlikely to determine the degree of postischemic recovery of contractile function.

Loss of ectonucleotidases from the coronary vascular bed after ischemia-reperfusion in isolated rat heart

BackgroundEctonucleotidase plays an important role in the regulation of cardiac function by controlling extracellular levels of adenine nucleotides and adenosine. To determine the influence of ischemia-reperfusion injury on ectonucleotidase activity in coronary vascular bed, we compared the metabolic profile of adenine nucleotides during the coronary circulation in pre- and post-ischemic heart.MethodsLangendorff-perfused rat hearts were used to assess the intracoronary metabolism of adenine nucleotides. The effects of ischemia on the adenine nucleotide metabolism were examined after 30 min of ischemia and 30 min of reperfusion. Adenine nucleotide metabolites were measured by high performance liquid chromatography.ResultsATP, ADP and AMP were rapidly metabolized to adenosine and inosine during the coronary circulation. After ischemia, ectonucleotidase activity of the coronary vascular bed was significantly decreased. In addition, the perfusate from the ischemic heart contained a considerable amount of enzymes degrading ATP, AMP and adenosine. Immunoblot analysis revealed that the perfusate from the ischemic heart dominantly contained ectonucleoside triphosphate diphosphohydrolase 1, and, to a lesser extent, ecto-5’-nucleotidase. The leakage of nucleotide metabolizing enzymes from the coronary vascular bed by ischemia-reperfusion was more remarkable in aged rats, in which post-ischemic cardiac dysfunction was more serious.ConclusionEctonucleotidases were liberated from the coronary vascular bed by ischemia-reperfusion, resulting in an overall decrease in ectonucleotidase activity in the post-ischemic coronary vascular bed. These results suggest that decreased ectonucleotidase activity by ischemia may exacerbate subsequent reperfusion injury, and that levels of circulating ectonucleotidase may reflect the severity of ischemic vascular injury.

Role of superoxide production in post-ischemic cardiac dysfunction and norepinephrine overflow in rat hearts

Reactive oxygen species and norepinephrine are known as physiological active substances which cause cell damage and cardiac dysfunction in myocardial ischemia/reperfusion injury. We investigated the role of reactive oxygen species, especially superoxide (O2−), in ischemia-induced norepinephrine overflow and cardiac dysfunction using superoxide scavengers tempol and tiron. According to the Langendorff technique, isolated rat hearts were subjected to 40-min global ischemia followed by 30-min reperfusion. Tempol (10 and 100µM) and tiron (100 and 500µM) were perfused 15min before ischemia and during reperfusion. Cardiac levels of oxidative stress markers such as O2− and malondialdehyde were notably increased during ischemia and following reperfusion, which were suppressed by the administration of tempol or tiron. These agents significantly improved ischemia/reperfusion-induced cardiac dysfunction such as decreased left ventricular developed pressure and the maximum and minimum value of the first derivative of left ventricular pressure and increased left ventricular end-diastolic pressure. Furthermore, norepinephrine overflow in the coronary effluent after ischemia/reperfusion was significantly suppressed by the administration of each agent. These results suggest that endogenously increased O2− is involved in norepinephrine overflow and cardiac dysfunction after myocardial ischemia/reperfusion.

Low molecular weight fibroblast growth factor-2 signals via protein kinase C and myofibrillar proteins to protect against postischemic cardiac dysfunction

Among its many biological roles, fibroblast growth factor-2 (FGF2) acutely protects the heart from dysfunction associated with ischemia/reperfusion (I/R) injury. Our laboratory has demonstrated that this is due to the activity of the low molecular weight (LMW) isoform of FGF2 and that FGF2-mediated cardioprotection relies on the activity of protein kinase C (PKC); however, which PKC isoforms are responsible for LMW FGF2-mediated cardioprotection, and their downstream targets, remain to be elucidated. To identify the PKC pathway(s) that contributes to postischemic cardiac recovery by LMW FGF2, mouse hearts expressing only LMW FGF2 (HMWKO) were bred to mouse hearts not expressing PKCα (PKCαKO) or subjected to a selective PKCε inhibitor (εV(1-2)) before and during I/R. Hearts only expressing LMW FGF2 showed significantly improved postischemic recovery of cardiac function following I/R (P < 0.05), which was significantly abrogated in the absence of PKCα (P < 0.05) or presence of PKCε inhibition (P < 0.05). Hearts only expressing LMW FGF2 demonstrated differences in actomyosin ATPase activity as well as increases in the phosphorylation of troponin I and T during I/R compared with wild-type hearts; several of these effects were dependent on PKCα activity. This evidence indicates that both PKCα and PKCε play a role in LMW FGF2-mediated protection from cardiac dysfunction and that PKCα signaling to the contractile apparatus is a key step in the mechanism of LMW FGF2-mediated protection against myocardial dysfunction.

Tongxinluo Improves Cardiac Function and Ameliorates Ventricular Remodeling in Mice Model of Myocardial Infarction through Enhancing Angiogenesis

Background. Myocardial infarction (MI) is a major cause of morbidity and mortality in the world. Tongxinluo (TXL) is a traditional Chinese compound prescription which has cardioprotective functions. The present study was aimed to determine the effect of TXL on postischemic cardiac dysfunction and cardiac remodeling and to elucidate the underlying mechanisms. Methods and Results. MI was performed by ligation of left anterior descending coronary artery (LAD) in male adult mice. Mice were randomly divided into four groups: (1) sham group (Sham); (2) MI-control group (Control); (3) MI-low dose TXL group (TXL-L); and (4) MI-high dose TXL (TXL-H) group. Compared with the control group, TXL treatment restored cardiac function, increased revascularization, attenuated cardiomyocyte apoptosis, and reduced interstitial fibrosis. TXL treatment increased the phosphorylation of Akt, extracellular signal regulated kinase (ERK), and endothelial nitric oxide synthase (eNOS); the expression of phosphatidylinositol3-kinase (PI3K), hypoxia-inducible factors 1α (HIF-1α), and vascular endothelial growth factor (VEGF); and the DNA binding activity of HIF-1α after MI. Conclusion. TXL may improve cardiac function and ameliorate cardiac remodeling by increasing neovascularization through enhancing the phosphorylation of Akt and ERK, the expression and activity of HIF-1α, and the protein level of VEGF and p-eNOS.

Protective effects of 17beta-estradiol on post-ischemic cardiac dysfunction and norepinephrine overflow through the non-genomic estrogen receptor/nitric oxide-mediated pathway in the rat heart

The present study was undertaken to examine the effect of acute treatment with 17β-estradiol on post-ischemic cardiac dysfunction and norepinephrine overflow and its possible mechanisms. Male rat hearts were perfused with the Langendorff method and subjected to 40min of global ischemia followed by 30min of reperfusion. Each drug was perfused from 15min before ischemia to 5min after reperfusion. During reperfusion, 17β-estradiol treatment showed significantly greater functional recovery of left ventricular developed pressure (LVDP), left ventricular end diastolic pressure (LVEDP), and dP/dtmax. Excessive norepinephrine release in coronary effluent from the post-ischemic heart was notably suppressed by treatment with 17β-estradiol. These beneficial effects of 17β-estradiol were not observed in the presence of the nitric oxide synthase inhibitor NG-nitro-l-arginine and estrogen receptor antagonist ICI 182,780 ((7α, 17β)-7-[9-[(4,4,5,5,5-pentafluoropentyl)sulfinyl]nonyl]estra-1,3,5(10)-triene-3,17-diol), respectively. When NO2/NO3 levels in coronary effluents after the onset of reperfusion were measured, reverse-correlation relationships between NO2/NO3 production and ischemia/reperfusion-induced cardiac dysfunction, as well as norepinephrine overflow were observed. These findings suggest that 17β-estradiol exerts cardioprotective effects against ischemia/reperfusion-induced cardiac dysfunction, at least in part, by suppressing norepinephrine overflow, and that nitric oxide production via estrogen receptor activation plays a key role in this process.

Sex Differences in Postischemic Cardiac Dysfunction and Norepinephrine Overflow in Rat Heart

The purpose of this study is to elucidate the relationship between sex difference and norepinephrine (NE) release in the pathogenesis of myocardial ischemia/reperfusion (I/R) injury. Isolated male and female rat hearts were subjected to 40-minute global ischemia followed by 30-minute reperfusion. Compared with male hearts, I/R-induced cardiac dysfunction, such as decreased left ventricular developed pressure and dP/dtmax and increased left ventricular end diastolic pressure, was significantly attenuated in female hearts. An excessive NE overflow in the coronary effluent from the postischemic heart in females was much less than that in males. These sex differences were abolished by ovariectomy, but in vivo treatment with 17β-estradiol recovered it. This ameliorating effect of 17β-estradiol was not observed in the presence of nitric oxide (NO) synthase inhibitor N-nitro-L-arginine. When NOx (NO2/NO3) levels in the coronary effluent after onset of reperfusion were measured, reversed correlated relationships between NOx production and I/R-induced cardiac dysfunction, and NE overflow, were observed. These findings suggest that sex differences in the postischemic cardiac dysfunction are closely related to the NE overflow from the postischemic heart and that estrogen plays a key role in the cardioprotective effect against I/R injury in female rats, by suppressing NE release via the enhancement of NO production.