Dietary flavonoids increase plasma very long-chain (n-3) fatty acids in rats.

Flavonoids probably contribute to the health benefits associated with the consumption of fruit and vegetables. However, the mechanisms by which they exert their effects are not fully elucidated. PUFA of the (n-3) series also have health benefits. Epidemiological and clinical studies have suggested that wine flavonoids may interact with the metabolism of (n-3) PUFA and increase their blood and cell levels. The present studies in rats were designed to assess whether flavonoids actually increase plasma levels of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), the main very long-chain (n-3) PUFA. Rats were fed a corn-derived anthocyanin (ACN)-rich (ACN-rich) or ACN-free diet with constant intakes of plant and marine (n-3) PUFA for 8 wk (Expt. 1). Plasma fatty acids were measured by GC. The ACN-rich diet contained ~0.24 ± 0.01 mg of ACN/g pellets. There were no significant differences between groups in the main saturated, monounsaturated, and (n-6) fatty acids. In contrast, plasma EPA and DHA were greater in the ACN-rich diet group than in the ACN-free diet group (P < 0.05). We obtained similar results in 2 subsequent experiments in which rats were administered palm oil (80 µL/d) and consumed the ACN-rich or ACN-free diet (Expt. 2) or were supplemented with fish oil (60 mg/d, providing 35 mg DHA and 12 mg EPA) and consumed the ACN-rich or ACN-free diet (Expt. 3). In both experiments, plasma EPA and DHA were significantly greater in the ACN-rich diet group. These studies demonstrate that the consumption of flavonoids increases plasma very long-chain (n-3) PUFA levels. These data confirm previous clinical and epidemiological studies and provide new insights into the health benefits of flavonoids.

Chronic dietary intake of plant-derived anthocyanins protects the rat heart against ischemia-reperfusion injury.

Consumption of flavonoid-rich foods and beverages is thought to reduce the risk of cardiovascular diseases. Whereas the biological activities of flavonoids have been characterized in vitro, there are no clear experimental data demonstrating that chronic dietary intake and intestinal absorption of flavonoids actually protects the heart against ischemia-reperfusion injury. We tested whether long-term consumption of specific flavonoids (anthocyanins) included in normal food could render the heart of rats more resistant to myocardial infarction. Maize kernels that differed specifically in their accumulation of anthocyanins were used to prepare rodent food in which anthocyanins were either present or absent. Male Wistar rats were fed the anthocyanin-rich (ACN-rich) or the anthocyanin-free (ACN-free) diet for a period of 8 wk. Anthocyanins were significantly absorbed and detected in the blood and urine of only rats fed the ACN-rich diet. In Langendorff preparations, the hearts of rats fed the ACN-rich diet were more resistant to regional ischemia and reperfusion insult. Moreover, on an in vivo model of coronary occlusion and reperfusion, infarct size was reduced in rats that ate the ACN-rich diet than in those that consumed the ACN-free diet (P < 0.01). Cardioprotection was associated with increased myocardial glutathione levels, suggesting that dietary anthocyanins might modulate cardiac antioxidant defenses. Our findings suggest important potential health benefits of foods rich in anthocyanins and emphasize the need to develop anthocyanin-rich functional foods with protective activities for promoting human health.

A single intravenous sTNFR-Fc administration at the time of reperfusion limits infarct size--implications in reperfusion strategies in man.

BACKGROUND: Reperfusion of the ischemic myocardium is associated with increased inflammatory processes that can exert deleterious effects and therefore contribute to cardiac dysfunction. The aim of the present study was to verify whether the administration of sTNFR-Fc, a scavenger of the pro-inflammatory cytokine TNF-alpha, at the time of reperfusion would protect against myocardial infarction and reduce the severity of early mechanical dysfunction. METHODS: Male Wistar rats were subjected to 60 min coronary occlusion followed by reperfusion. A bolus of sTNFR-Fc (10 microg/kg, i.v.) (MI + sTNFR-Fc group) or a placebo (MI group) was injected prior to reperfusion. Cardiac geometry was assessed by echocardiography 1, 3 and 7 days after reperfusion. Eight days after reperfusion, left ventricular (LV) function was evaluated under basal conditions and during an experimental challenge of volume overload. Finally, infarct size was measured after euthanasia. RESULTS: sTNFR-Fc administration markedly reduced infarct size (P < 0.01) and decreased LV dilation as assessed by the echocardiographic measurement of the LV end diastolic area, 7 days post-MI (P < 0.01). Moreover, LV end-diastolic pressure was significantly preserved by sTNFR-Fc 1 week after myocardial infarction, under basal conditions (P < 0.05) as well as during cardiac overload (P < 0.05). CONCLUSION: A single administration of sTNFR-Fc at the time of reperfusion after myocardial infarction is able to limit infarct size and to reduce early LV diastolic dysfunction in rats. These findings suggest that intravenous neutralization of TNF-alpha during surgical cardiac reperfusion might improve the outcome of myocardial infarction in humans.

Topoisomerase Inhibition Accelerates Gene Expression after Adeno-associated Virus-mediated Gene Transfer to the Mammalian Heart.

Utility of adeno-associated virus 2 (AAV2) vectors for cardiac gene therapy is limited by the prolonged lag phase before maximal gene expression. Topoisomerase inhibition can induce AAV2-mediated gene expression in vivo, but with variable success in different tissues. In this study, we demonstrate that topoisomerase inhibition can accelerate AAV2-mediated gene expression in the mouse heart. We used an AAV2 vector expressing firefly luciferase and monitored expression kinetics using non-invasive bioluminescence imaging. In the group receiving vector alone, cardiac luciferase activity was evident from week 2 onward and increased progressively to reach a steady plateau by 9 weeks postinjection. In the group receiving vector and camptothecine (CPT), luciferase expression was evident from days 2 to 4 onward and increased rapidly to reach a steady plateau by 3-4 weeks postinjection, nearly three times faster than in the absence of CPT (P<0.05). Southern blot analysis of AAV2 genomes in cardiac tissue showed rapid conversion of the AAV2 genome from its single-stranded to double-stranded form in CPT-treated mice. Non-invasive determinations of luciferase expression correlated well with in vitro luciferase assays. Direct injection of the AAV2 vector and long-term luciferase gene expression had no detectable effects on normal cardiac function as assessed by magnetic resonance imaging.

Myocardial infarct-sparing effect of adenosine A2A receptor activation is due to its action on CD4+ T lymphocytes.

BACKGROUND: We previously used adenosine A2A receptor (A2AR) knockout (KO) mice and bone marrow transplantation to show that the infarct-sparing effect of A2AR activation at reperfusion is primarily due to effects on bone marrow-derived cells. In this study we show that CD4+ but not CD8+ T lymphocytes contribute to myocardial ischemia/reperfusion injury. METHOD AND RESULTS: After a 45-minute occlusion of the left anterior descending coronary artery and reperfusion, T cells accumulate in the infarct zone within 2 minutes. Addition of 10 microg/kg of the A2AR agonist ATL146e 5 minutes before reperfusion produces a significant reduction in T-cell accumulation and a significant reduction in infarct size (percentage of risk area) measured at 24 hours. In Rag1 KO mice lacking mature lymphocytes, infarct size is significantly smaller than in C57BL/6 mice. Infarct size in Rag1 KO mice is increased to the level of B6 mice by adoptive transfer of 50 million CD4+ T lymphocytes derived from C57BL/6 or A2AR KO but not interferon-gamma KO mice. ATL146e completely blocked the increase in infarct size in Rag1 KO mice reconstituted with B6 but not A2AR KO CD4+ T cells. The number of neutrophils in the reperfused heart at 24 hours after infarction correlated well with the number of lymphocytes and infarct size. CONCLUSIONS: These results strongly suggest that the infarct-sparing effect of A2AR activation is primarily due to inhibition of CD4+ T-cell accumulation and activation in the reperfused heart.

Does nitric oxide contribute to progressive cardiac tissue damage and dysfunction after infarction?

Myocardial infarction induces contractile dysfunction and remodeling that can lead to heart failure. Nitric oxide has been proposed as one of the major actors of this pathophysiologic process. We note that N (G)-nitro-L-arginine methyl ester (L-NAME) administration from day 2 to day 7 after myocardial infarction in rats improves stroke volume, preserves cardiac compliance, and reduces infarct expansion. Our observations lead to the hypothesis that the mechanisms by which cytokines contribute to myocardial remodeling and dysfunction in the days after infarction might involve *NO signalling pathways.

Infarct-sparing effect of A2A-adenosine receptor activation is due primarily to its action on lymphocytes.

BACKGROUND: A2A-adenosine receptor (A2AAR) activation on reperfusion after ischemia reduces the size of myocardial infarction, but the mechanism of action has not been fully defined. METHODS AND RESULTS: We created chimeric mice by bone marrow transplantation from A2AAR-knockout or green fluorescent donor mice to irradiated congenic C57BL/6 (B6) recipients. In the GFP chimeras, we were unable to detect green fluorescent-producing cells in the vascular endothelium, indicating that bone marrow-derived cells were not recruited to endothelium at appreciable levels after bone marrow transplantation and/or acute myocardial infarction. Injection of 5 or 10 microg/kg of a potent and selective agonist of A2AAR, ATL146e, had no effect on hemodynamic parameters but reduced infarct size in B6 mice after 45 minutes of left anterior descending artery occlusion followed by 24 hours of reperfusion to 42.5+/-3.0% and 39.3+/-4.7% of risk region, respectively, compared with 61.0+/-2.3% in vehicle-treated B6 mice (P<0.05). Myocardial myeloperoxidase activity in the risk region measured at 4 hours after reperfusion was significantly reduced by ATL146e. The salutary effects of ATL146e were absent in A2AAR-knockout mice or in mice treated with a selective A2AAR antagonist, ZM241385. ATL146e also reduced infarct size and myeloperoxidase in B6/B6 (donor/recipient) chimeras (P<0.05) but not in A2AAR-knockout/B6 chimeras. In immunocompromised Rag-1-KO mice, infarct size was significantly reduced compared with B6 mice but was not further reduced by ATL146e. CONCLUSIONS: The results indicate that A2AAR activation on bone marrow-derived cells, specifically T or B lymphocytes, is responsible for the infarct-sparing and antiinflammatory effects of ATL146e administered at the time of reperfusion after coronary occlusion.

Simultaneous evaluation of infarct size and cardiac function in intact mice by contrast-enhanced cardiac magnetic resonance imaging reveals contractile dysfunction in noninfarcted regions early after myocardial infarction.

BACKGROUND: The objective of this study was to noninvasively determine the effects of reperfused myocardial infarction (MI) on regional and global left-ventricular (LV) function 24 hours after MI in intact mice with contrast-enhanced cardiac MRI and a single, gradient-echo pulse sequence. METHODS AND RESULTS: Twenty-three mice received baseline MRI scans followed by either 60 minutes of coronary occlusion (MI group, n=15) or thoracotomy without occlusion (sham group, n=8). Gadolinium-DTPA-enhanced magnetic resonance (MR) images were acquired 24 hours after surgery. Hearts were then excised for conventional infarct size determination via 2,3,5-triphenyl tetrazolium chloride (TTC) staining. In addition to infarct size, analysis of the MR images yielded left ventricular (LV) mass, LV end-systolic volume (LVESV), LV end-diastolic volume (LVEDV), LV ejection fraction (LVEF), cardiac output, and percent LV wall thickening (%WTh). Twenty-four hours after surgery, infarct size was 28.1+/-1.8% of LV mass by MRI and 27.5+/-1.7% by TTC (P=NS). Bland-Altman analysis revealed close agreement between the results obtained by the 2 methods. MI had little effect on LVEDV but caused a 98% increase in LVESV (from 11.3 to 22.4 microL, P<0.05), which resulted in a significant reduction in LVEF (from 70% to 37%, P<0.05). Compared with LV regional function at baseline, %WTh 24 hours after MI was significantly depressed, not only in infarcted myocardium but also in regions remote from the infarct zone. In contrast, sham-operated mice showed a small but significant increase in %WTh 24 hours after surgery (P<0.05). CONCLUSIONS: MRI can accurately assess both infarct size and cardiac function in intact mice early after large, reperfused MI, revealing the existence of contractile dysfunction in noninfarcted regions of the heart.

Selenium status as determinant of connexin-43 dephosphorylation in ex vivo ischemic/reperfused rat myocardium.

Recent studies have demonstrated that electrical uncoupling at gap junctions during ischemia is associated with cardiac Connexin-43 (Cx43) dephosphorylation. Whether oxidative stress is involved in this phenomenon still remains unclear. In the present study, we examined the influence of selenium intake on reperfusion-induced Cx43 dephosphorylation. Male Wistar rats were fed a diet containing either 0.05 mg/kg (Low-Se, n = 13) or 1.5 mg/kg (High-Se, n = 11) selenium for 8 weeks. At the end of this diet, hearts were isolated and subjected to 10 min regional ischemia followed by 10 min reperfusion. The level of dephosphorylated Cx43 was determined in tissue samples from ischemic/reperfused and non-ischemic regions of the hearts. At the end of the experiemental diet, the activity of the antioxidant enzyme glutathione peroxidase (GSH-Px) was increased in high-Se hearts compared with low-Se hearts (+ 13%; p < 0.05). After ischemia/reperfusion, in low-Se hearts, Cx43 dephosphorylation appeared significantly increased in the left ventricle compared to the non-ischemic right ventricle (+ 149%; p < 0.05). The high-Se diet significantly reduced Cx43 dephosphorylation in the left ventricle (p < 0.05 vs. low-Se diet). In conclusion, our results suggest that oxidative stress may be involved in Cx43 dephosphorylation during myocardial ischemia/reperfusion, thereby contributing to arrhythmogenesis.

Involvement of reactive oxygen species in cardiac preconditioning in rats.

To date, the involvement of reactive oxygen species in ischemic preconditioning in vivo in rats is not clearly demonstrated. The aim of the present study was to determine whether N-(2-mercaptopropionyl)glycine (MPG), a cell-diffusible hydroxyl radical scavenger, and carnosine, a potent singlet oxygen quencher, could block protection afforded by a single cycle of ischemic preconditioning in vivo in the rat. An ESR study was first performed to validate in vitro the specific antioxidant properties of carnosine and MPG. In a second set of experiments, open-chest rats were subjected to 30 min of left coronary occlusion followed by 60 min of reperfusion. Preconditioning was elicited by 5 min of ischemia and 5 min of reperfusion. Neither MPG (1-h infusion, 20 mg/kg) nor carnosine injection (bolus, 25 micro mol/rat) affected infarct size. The infarct size-limiting effect of preconditioning was completely blunted by MPG, whereas carnosine did not alter the cardioprotection. It is concluded that free radicals and especially hydroxyl radicals could be involved in the adaptive mechanisms induced by a single cycle of preconditioning in vivo in rats.

Preischemic selenium status as a major determinant of myocardial infarct size in vivo in rats.

Prospective epidemiological studies have shown that the incidence of numerous cardiovascular pathologies is correlated with body selenium status. However, it remains unclear whether selenium status also influences the outcome of myocardial infarction. The aim of the present study was to test whether dietary selenium intake affects myocardial necrosis induced by transient regional ischemia in vivo in rats. For this purpose, male Wistar rats received either a high-selenium (High-Se: 1.5 mg of Se/kg) or a low-selenium (Low-Se: 0.05 mg of Se/kg) diet for 10 weeks. Animals were subjected to 30 min of myocardial ischemia induced by coronary artery ligation followed by 60 min of reperfusion. Pre- and postischemic blood samples were collected for glutathione (GSH and GSSG) determination and for glutathione peroxidase (GSH-Px) assessment. Our results show that high-selenium intake reduces myocardial infarct size (High-Se: 25.16 +/- 1.19% versus Low-Se: 36.51 +/- 4.14%, p < 0.05), preserves postischemic GSH/GSSG ratio (High-Se: 1.37 +/- 0.37 versus Low-Se: 0.47 +/- 0.10, p < 0.05), increases plasma GSH-Px activity, and improves postischemic mean arterial pressure. In conclusion, preischemic body selenium status is a major determinant of the outcome of myocardial ischemia in vivo in rats probably because it influences the cellular redox status.

Effects of increased lipid concentration and hyperemic blood flow on the intrinsic myocardial washout kinetics of (99m)TcN-NOET.

UNLABELLED: Bis(N-ethoxy,N-ethyldithiocarbamato)nitrido technetium (V) ((99m)Tc) ((99m)TcN-NOET) is a myocardial perfusion imaging agent demonstrating significant redistribution and currently in phase III clinical trials. Previous studies have suggested that (99m)TcN-NOET is bound intravascularly. Therefore, we sought to determine whether modifications in the vascular compartment would provide further insights into the mechanisms of (99m)TcN-NOET myocardial washout and redistribution. METHODS: (99m)TcN-NOET cardiac washout was studied ex vivo in 15 isolated perfused rat hearts after bolus injection (1.5 MBq) in the absence (n = 6) or presence of bovine serum albumin ([BSA] 0.03%) with (n = 5) or without (n = 4) bound lipids. The intrinsic myocardial washout of the tracer was also studied in vivo in 6 dogs after intracoronary bolus injection of the tracer (0.75 MBq) before and after hyperlipidemia induced by intravenous administration of 300 mL of 20% intralipids (n = 3) or hyperemia induced by intravenous infusion of the adenosine A(2A) receptor agonist ATL-146e (0.3 micro g/kg/min; n = 6). RESULTS: On isolated hearts, there was no significant myocardial washout of (99m)TcN-NOET with Krebs-Henseleit buffer. Addition of BSA without bound lipids resulted in a significant cardiac washout of the tracer (P < 0.001 by repeated measures ANOVA). The presence of lipids bound to BSA further accelerated the washout rate of (99m)TcN-NOET (half-life [t(1/2)], 431.5 +/- 23.2 min vs. 242.9 +/- 63.2 min; P < 0.05). In vivo in dogs, intralipid administration significantly increased the intrinsic washout rate of (99m)TcN-NOET (t(1/2), 108.0 +/- 23.9 min vs. 51.8 +/- 11.8 min; P < 0.05). In addition, vasodilatation with ATL-146e resulted in a 4.9-fold increase in coronary flow (P < 0.05 vs. baseline) and a significantly faster intrinsic (99m)TcN-NOET myocardial washout (t(1/2), 81.1 +/- 12.1 min vs. 40.7 +/- 7.3 min; P < 0.05). CONCLUSION: The myocardial washout kinetics of (99m)TcN-NOET are affected by a variety of intravascular factors, supporting the hypothesis that the tracer is most likely localized on the vascular endothelium. The potential impact of variations in circulating lipid levels among patients on clinical imaging with (99m)TcN-NOET requires further investigation.

Cardiac dysfunction in rats with dietary-induced insulin resistance associated with pharmacologically-induced dyslipidemia.

Metabolic disorders such as insulin resistance (IR) and dyslipidemia (DL) might contribute to the induction of diabetic cardiomyopathy (DCM). However, few relevant animal models are currently available for studying the time-course of DCM and evaluating experimental therapeutics. The present study proposes a rodent model of dietary-induced IR combined or not with DL in order to investigate the impact of chronic IR and DL on in vivo myocardial function. Male rats were fed a western-type diet (65% fat; 15% fructose; WD). DL was induced by combining the western diet with i.p. injections of a nonionic surface-active agent (P-407; 0.2 mg/kg, 3 times/wk; P-407). A chow diet was used as control. At 11 and 14 weeks, cardiac function was assessed by echocardiography. Fasting blood glucose increased in WD group while plasma lipids markedly accumulated in P-407 treated rats. Echocardiographic data showed no significant difference in cardiac geometry under basal conditions. Diastolic dysfunction was evidenced at 14 weeks by a significant decrease in E/A ratio in the P-407 group. Moreover, fractional shortening was significantly depressed under dobutamine stress in WD group at 14 weeks whereas systolic dysfunction appeared as early as 11 weeks and worsened at 14 weeks in P-407 animals. Finally, myocardial TNF-alpha tissue content increased in P-407 group. In conclusion, DL exacerbated cardiac lipotoxicity and functional complications associated with IR. This experimental model of combined IR and DL closely mimics the main clinical manifestations of DCM and might therefore constitute a useful tool for the evaluation of pharmacological treatments.

Analysis of the occupational, consumer and environmental exposure to engineered nanomaterials used in 10 technology sectors.

Humans and the environment can come into contact with nanomaterials through a wide range of applications during all stages of the life cycle of nanoproducts. The aim of this commentary is to present an assessment of the potential for exposure and thus identify possible environmental, health and safety (EHS) issues for nanomaterials used in 10 technology sectors. We analysed all life cycle stages with regard to potential for exposure of workers, consumers/patients, and the environment. A wide variety of nanomaterials are used of which many have negligible potential for exposure, while others have medium or even high potential for exposure. Based on the likelihood of exposure, it appears that in general most attention should be paid to the agrifood, chemistry/materials, textiles and health sectors; and less to the information and communication technology (ICT), security and energy sectors. Toxicity and exposure are both important; however, the EHS impact of nanomaterials is always dependent on their particular use.

Topoisomerase inhibition accelerates gene expression after adeno-associated virus-mediated gene transfer to the mammalian heart.

Utility of adeno-associated virus 2 (AAV2) vectors for cardiac gene therapy is limited by the prolonged lag phase before maximal gene expression. Topoisomerase inhibition can induce AAV2-mediated gene expression in vivo, but with variable success in different tissues. In this study, we demonstrate that topoisomerase inhibition can accelerate AAV2-mediated gene expression in the mouse heart. We used an AAV2 vector expressing firefly luciferase and monitored expression kinetics using non-invasive bioluminescence imaging. In the group receiving vector alone, cardiac luciferase activity was evident from week 2 onward and increased progressively to reach a steady plateau by 9 weeks postinjection. In the group receiving vector and camptothecine (CPT), luciferase expression was evident from days 2 to 4 onward and increased rapidly to reach a steady plateau by 3-4 weeks postinjection, nearly three times faster than in the absence of CPT (P<0.05). Southern blot analysis of AAV2 genomes in cardiac tissue showed rapid conversion of the AAV2 genome from its single-stranded to double-stranded form in CPT-treated mice. Non-invasive determinations of luciferase expression correlated well with in vitro luciferase assays. Direct injection of the AAV2 vector and long-term luciferase gene expression had no detectable effects on normal cardiac function as assessed by magnetic resonance imaging.

Delayed expression of cytokines after reperfused myocardial infarction: possible trigger for cardiac dysfunction and ventricular remodeling.

Previous studies have shown that 1 wk after permanent coronary artery ligation in rats, some cellular mechanisms involving TNF-alpha occur and contribute to the development of cardiac dysfunction and subsequent heart failure. The aim of the present study was to determine whether similar phenomena also occur after ischemia-reperfusion and whether cytokines other than TNF-alpha can also be involved. Anesthetized male Wistar rats were subjected to 1 h coronary occlusion followed by reperfusion. Cardiac geometry and function were assessed by echocardiography at days 5, 7, 8, and 10 postligation. Before death, heart function was assessed in vivo under basal conditions, as well as after volume overload. Finally, hearts were frozen for histoenzymologic assessment of infarct size and remodeling. The profile of cardiac cytokines was determined by ELISA and ChemiArray on heart tissue extracts. As expected, ischemia-reperfusion induced a progressive remodeling of the heart, characterized by left ventricular free-wall thinning and cavity dilation. Heart function was also decreased in ischemic rats during the first week after surgery. Interestingly, a transient and marked increase in TNF-alpha, IL-1beta, IL-6, cytokine-induced neutrophil chemoattractant (CINC) 2, CINC3, and macrophage inflammatory protein-3alpha was also observed in the myocardium of myocardial ischemia (MI) animals at day 8, whereas the expression of anti-inflammatory interleukins IL-4 and IL-10 remained unchanged. These results suggest that overexpression of proinflammatory cytokines occurring during the first week after ischemia-reperfusion may play a role in the adaptative process in the myocardium and contribute to early dysfunction and remodeling.

Borderzone contractile dysfunction is transiently attenuated and left ventricular structural remodeling is markedly reduced following reperfused myocardial infarction in inducible nitric oxide synthase knockout mice.

OBJECTIVES: We sought to determine the effect of inducible nitric oxide synthase (iNOS) expression on regional contractile function and left ventricular (LV) remodeling after reperfused myocardial infarction (MI). BACKGROUND: Inducible nitric oxide synthase is known to contribute to global LV dysfunction after a large MI, but the mechanisms underlying this dysfunction remain unclear. METHODS: We used immunohistochemistry to investigate the distribution of iNOS expression in wild-type (WT) and iNOS knockout (KO) mice early (day 1) and late (day 28) after reperfused MI. We also used serial cardiac magnetic resonance imaging at baseline and at 1, 7, and 28 days after MI to assess LV volumes, ejection fraction (EF), regional circumferential strain (E(cc)), and day 1 infarct size. RESULTS: At baseline, LV volumes and EF were similar between groups. Day 1 infarct size was also similar between groups. Immunohistochemistry revealed that iNOS expression was abundant throughout the heart in WT mice on day 1 after MI, particularly near the infarct borderzone. On day 7 after MI, E(cc) in KO mice was significantly improved in some borderzone sectors compared with WT. The LV volumes were significantly lower in KO mice at days 7 and 28 compared with WT. The EF on days 7 and 28 was significantly higher in KO mice compared with WT. The circumferential extent of wall thinning was also significantly reduced in KO versus WT mice at days 7 and 28. CONCLUSIONS: Expression of iNOS contributes importantly to post-infarction contractile dysfunction and subsequent LV remodeling, suggesting new strategies to combat heart failure resulting from large MI.

Stimulation of A2A-adenosine receptors after myocardial infarction suppresses inflammatory activation and attenuates contractile dysfunction in the remote left ventricle.

Following myocardial infarction (MI), contractile dysfunction develops not only in the infarct zone but also in noninfarcted regions of the left ventricle remote from the infarct zone. Inflammatory activation secondary to MI stimulates inducible nitric oxide synthase (iNOS) induction with excess production of nitric oxide. We hypothesized that the anti-inflammatory effects of selective A(2A)-adenosine receptor (A(2A)AR) stimulation would suppress inflammation and preserve cardiac function in the remote zone early after MI. A total of 53 mice underwent 60 min of coronary occlusion followed by 24 h of reperfusion. The A(2A)AR agonist (ATL146e, 2.4 microg/kg) was administered intraperitoneally 1, 3, and 6 h postreperfusion. Because of the 1-h delay in treatment after MI, ATL146e had no effect on infarct size, as demonstrated by contrast-enhanced cardiac MRI (n = 18) performed 24 h post-MI. ATL146e did however preserve global cardiac function at that time by limiting contractile dysfunction in remote regions [left ventricle wall thickening: 51 +/- 4% in treated (n = 9) vs. 29 +/- 3% in nontreated groups (n = 9), P < 0.01]. RT-PCR, immunohistochemistry, and Western blot analysis indicated that iNOS mRNA and protein expression were significantly reduced by ATL146e treatment in both infarcted and noninfarcted zones. Similarly, elevations in plasma nitrate-nitrite after MI were substantially blunted by ATL146e (P < 0.01). Finally, treatment with ATL146e reduced NF-kappaB activation in the myocardium by over 50%, not only in the infarct zone but also in noninfarcted regions (P < 0.05). In conclusion, A(2A)AR stimulation after MI suppresses inflammatory activation and preserves cardiac function, suggesting the potential utility of A(2A)AR agonists against acute heart failure in the immediate post-MI period.

Activated platelets contribute importantly to myocardial reperfusion injury.

Platelets become activated during myocardial infarction (MI), but the direct contribution of activated platelets to myocardial reperfusion injury in vivo has yet to be reported. We tested the hypothesis that activated platelets contribute importantly to reperfusion injury during MI in mice. After 30 min of ischemia and 60 min of reperfusion, P-selectin knockout mice had a significantly smaller infarct size than that of wild-type mice (P < 0.05). Platelets were detected by P-selectin antibody in the previously ischemic region of wild-type mice as early as 2 min postreperfusion after 45 min, but not 20 min, of ischemia. The appearance of neutrophils in the heart was delayed when compared with platelets. Flow cytometry showed that the number of activated platelets more than doubled after 45 min of ischemia when compared with 20 min of ischemia or sham treatment (P < 0.05). Platelet-rich or platelet-poor plasma was then transfused from either sham-operated or infarcted mice after 45 and 10 min of ischemia-reperfusion to mice undergoing 20 and 60 min of ischemia-reperfusion. Infarct size was increased by threefold and platelet accumulation was remarkably enhanced in mice treated with wild-type, MI-activated platelet-rich plasma but not in mice receiving either platelet-poor plasma from wild types or MI-activated platelet-rich plasma from P-selectin knockout mice. In conclusion, circulating platelets become activated early during reperfusion and their activation depends on the duration of the preceding coronary occlusion and is proportional to the extent of myocardial injury. Activated platelets play an important role in the process of myocardial ischemia-reperfusion injury, and platelet-derived P-selectin is a critical mediator.

Early pre-diabetic state alters adaptation of myocardial glucose metabolism during ischemia in rats.

Pre-diabetic subjects with high insulin secretory capacity have double risk of cardiovascular disease compared with subjects who do not develop insulin-resistance. It is well established that the ability of the myocardium to increase its glycolytic ATP production plays a crucial role in determining cell survival under conditions of ischemia. Up to now, whether the pre-diabetic state reduces the tolerance of the heart to ischemia by affecting its ability to increase its energy production through glycolysis remains unknown. The aim of the present study was to assess whether insulin resistance affects the ability of the myocardium to increase glycolysis under ischemic conditions. Male Wistar rats were fed for 8 weeks a fructose-enriched (33%) diet to induce a pre-diabetic state. Hearts were isolated and subjected to ex-vivo low-flow (2%) ischemia for 30 min. The fructose diet increased sarcolemmal GLUT4 localisation in myocardial cells under basal conditions compared with controls. This effect was not accompanied by increased glucose utilisation. Ischemia induced the translocation of GLUT4 to the plasma membrane in controls but did not significantly modify the distribution of these transporters in pre-diabetic hearts. Glycolytic flux under ischemic conditions was significantly lower in fructose-fed rat hearts compared with controls. The reduction of glycolytic flux during ischemia in fructose-fed rat hearts was not due to metabolic inhibition downstream hexokinase II since no cardiac accumulation of glucose-6-phosphate was detected. In conclusion, our results suggest that the pre-diabetic state reduces the tolerance of the myocardium to ischemia by decreasing glycolytic flux adaptation.