Therapeutic potentials of cell death inhibitors in rats with cardiac ischaemia/reperfusion injury

Growing evidence demonstrated that cell death pathways including ferroptosis, apoptosis and necroptosis contribute to cardiac ischaemia/reperfusion (I/R) injury. We hypothesized that ferroptosis, apoptosis and necroptosis contribute differently to myocardial damage during acute cardiac I/R injury. Rats underwent cardiac I/R or sham operation. I/R‐operated rats were divided into 4 groups: vehicle, apoptosis (Z‐vad), ferroptosis (Fer‐1) and necroptosis (Nec‐1) inhibition. Rats in each cell death inhibitor group were subdivided into 3 different dose regimens: low, medium and high. Infarct size, left ventricular (LV) function, arrhythmias and molecular mechanism were investigated. Cardiac I/R caused myocardial infarction, LV dysfunction, arrhythmias, mitochondrial dysfunction, mitochondrial dynamic imbalance, inflammation, apoptosis and ferroptosis. Infarct size, LV dysfunction, mitochondrial dysfunction, apoptosis and ferroptosis were all reduced to a similar extent in rats treated with Z‐vad (low and medium doses) or Fer‐1 (medium and high doses). Fer‐1 treatment also reduced mitochondrial dynamic imbalance and inflammation. No evidence of necroptosis was found in association with acute I/R injury, therefore Nec‐1 treatment could not be assessed. Apoptosis and ferroptosis, not necroptosis, contributed to myocardial damage in acute I/R injury. Inhibitors of these 2 pathways provided effective cardioprotection in rats with I/R injury though modulation of mitochondrial function and attenuated apoptosis and ferroptosis.     

Putative therapeutic impacts of cardiac CTRP9 in ischaemia/reperfusion injury

Recently, cytokines belonging to C1q/tumour necrosis factor‐related proteins (CTRPs) superfamily have attracted increasing attention due to multiple metabolic functions and desirable anti‐inflammatory effects. These various molecular effectors exhibit key roles upon the onset of cardiovascular diseases, making them novel adipo/cardiokines. This review article aimed to highlight recent findings correlated with therapeutic effects and additional mechanisms specific to the CTRP9, particularly in cardiac ischaemia/reperfusion injury (IRI). Besides, the network of the CTPR9 signalling pathway and its possible relationship with IRI were discussed. Together, the discovery of all involved underlying mechanisms could shed light to alleviate the pathological sequelae after the occurrence of IRI.     

Roles of mitochondrial dynamics modulators in cardiac ischaemia/reperfusion injury

The current therapeutic strategy for the management of acute myocardial infarction (AMI) is to return blood flow into the occluded coronary artery of the heart, a process defined as reperfusion. However, reperfusion itself can increase mortality rates in AMI patients because of cardiac tissue damage and dysfunction, which is termed ‘ischaemia/reperfusion (I/R) injury’. Mitochondria play an important role in myocardial I/R injury as disturbance of mitochondrial dynamics, especially excessive mitochondrial fission, is a predominant cause of cardiac dysfunction. Therefore, pharmacological intervention and therapeutic strategies which modulate the mitochondrial dynamics balance during I/R injury could exert great beneficial effects to the I/R heart. This review comprehensively summarizes and discusses the effects of mitochondrial fission inhibitors as well as mitochondrial fusion promoters on cardiac and mitochondrial function during myocardial I/R injury. The comparison of the effects of both compounds given at different time‐points during the course of I/R injury (i.e. prior to ischaemia, during ischaemia and at the reperfusion period) are also summarized and discussed. Finally, this review also details important information which may contribute to clinical practices using these drugs to improve the quality of life in AMI patients.     

PCSK9 inhibitor and atorvastatin reduce cardiac impairment in ovariectomized prediabetic rats via improved mitochondrial function and Ca2+ regulation

Post‐menopausal women have a higher risk of developing cardiometabolic dysfunction. Atorvastatin attenuates dyslipidaemia and cardiac dysfunction but it can have undesirable effects including increased risk of diabetes and myalgia. Currently, the proprotein convertase subtilisin/kexin type 9 (PCSK9) inhibitor efficiently reduces low‐density lipoprotein cholesterol (LDL‐C) levels more effectively than atorvastatin. We have been suggested that PCSK9 inhibitor attenuated cardiometabolic impairment more effectively than atorvastatin in ovariectomized prediabetic rats. Female Wistar rats (n = 48) were fed a normal diet (ND) or high‐fat diet (HFD) for 12 weeks. Then, HFD rats were assigned to a sham‐operated (Sham) or ovariectomized (OVX) group. Six weeks after surgery, the OVX group was subdivided into 4 treatment groups: vehicle (HFOV), atorvastatin (HFOA) (40 mg/kg/day; s.c.), PCSK9 inhibitor (HFOP) (4 mg/kg/day; s.c.) and oestrogen (HFOE₂) (50 µg/kg/day; s.c.) for an additional 3 weeks. Metabolic parameters, cardiac and mitochondrial function, and [Ca²⁺]_i transients were evaluated. All HFD rats became obese‐insulin resistant. HFS rats had significantly impaired left ventricular (LV) function, cardiac mitochondrial function and [Ca²⁺]_i transient dysregulation. Oestrogen deprivation (HFOV) aggravated all of these impairments. Our findings indicated that the atorvastatin, PCSK9 inhibitor and oestrogen shared similar efficacy in the attenuation in cardiometabolic impairment in ovariectomized prediabetic rats.     

Lin28a protects against cardiac ischaemia/reperfusion injury in diabetic mice through the insulin‐PI3K‐mTOR pathway

The insulin‐PI3K‐mTOR pathway exhibits a variety of cardiovascular activities including protection against I/R injury. Lin28a enhanced glucose uptake and insulin‐sensitivity via insulin‐PI3K‐mTOR signalling pathway. However, the role of lin28a on experimental cardiac I/R injury in diabetic mice are not well understood. Diabetic mice underwent 30 min. of ischaemia followed by 3 hrs of reperfusion. Animals were randomized to be treated with lentivirus carrying lin28a siRNA (siLin28a) or lin28a cDNA (Lin28a) 72 hrs before coronary artery ligation. Myocardial infarct size (IS), cardiac function, cardiomyocyte apoptosis and mitochondria morphology in diabetic mice who underwent cardiac I/R injury were compared between groups. The target proteins of lin28a were examined by western blot analysis. Lin28a overexpression significantly reduced myocardial IS, improved LV ejection fraction (LVEF), decreased myocardial apoptotic index and alleviated mitochondria cristae destruction in diabetic mice underwent cardiac I/R injury. Lin28a knockdown exacerbated cardiac I/R injury as demonstrated by increased IS, decreased LVEF, increased apoptotic index and aggravated mitochondria cristae destruction. Interestingly, pre‐treatment with rapamycin abolished the beneficial effects of lin28a overexpression. Lin28a overexpression increased, while Lin28a knockdown decreased the expression of IGF1R, p‐Akt, p‐mTOR and p‐p70s6k after cardiac I/R injury in diabetic mice. Rapamycin pre‐treatment abolished the effects of increased p‐mTOR and p‐p70s6k expression exerted by lin28a overexpression. This study indicates that lin28a overexpression reduces IS, improves cardiac function, decreases cardiomyocyte apoptosis index and alleviates cardiomyocyte mitochondria impairment after cardiac I/R injury in diabetic mice. The mechanism responsible for the effects of lin28a is associated with the insulin‐PI3K‐mTOR dependent pathway.     

Melatonin and its protective role in attenuating warm or cold hepatic ischaemia/reperfusion injury

Although the liver is the only organ with regenerative capacity, various injury factors induce irreversible liver dysfunction and end‐stage liver disease. Liver resection and liver transplantation (LT) are effective treatments for individuals with liver failure, liver cirrhosis and liver cancers. The remnant or transplanted liver tissues will undergo hepatic ischaemia/reperfusion (IR), which leads to oxidative stress, inflammation, immune injury and liver damage. Moreover, systemic ischaemia induced by trauma, stroke, myocardial ischaemia, haemorrhagic shock and other injury factors also induces liver ischaemia/reperfusion injury (IRI) in individuals. Hepatic IRI can be divided into warm IRI, which is induced by liver surgery and systemic ischaemia, and cold IRI, which is induced by LT. Multiple studies have shown that melatonin (MT) acts as an endogenous free radical scavenger with antioxidant capacity and is also able to attenuate hepatic IRI via its anti‐inflammatory and antiapoptotic capacities. In this review, we discuss the potential mechanisms and current strategies of MT administration in liver surgery for protecting against warm or cold hepatic IRI. We highlight strategies to improve the efficacy and safety of MT for attenuating hepatic IRI in different conditions. After the potential mechanisms underlying the interactions between MT and other important cellular processes during hepatic IR are clarified, more opportunities will be available to use MT to treat liver diseases in the future.     

Cardioprotective effects of melatonin against myocardial ischaemia/reperfusion injury: Activation of AMPK/Nrf2 pathway

Although reperfusion is the most effective therapy for patients with acute myocardial infarction, reperfusion injury limits the therapeutic effects of early reperfusion. Oxidative stress plays a crucial role in myocardial ischaemia/reperfusion (I/R) injury. Melatonin, a circulating hormone, is well-known as an antioxidant in cardiovascular diseases. In this short communication, we show that melatonin significantly improves post-ischaemic cardiac function, reduces infarct size and decreases oxidative stress. Furthermore, melatonin markedly increases AMPK activation and Nrf2 nuclear translocation. Nevertheless, these melatonin-induced changes are abrogated by compound C. In addition, ML-385, an Nrf2 inhibitor, also withdraws the antioxidative effects of melatonin but has little effect on AMPK activation. In conclusion, our results demonstrate that melatonin alleviates myocardial I/R injury by inhibiting oxidative stress via the AMPK/Nrf2 signalling pathway.     

miRNA‐182/Deptor/mTOR axis regulates autophagy to reduce intestinal ischaemia/reperfusion injury

It had been reported miR‐182 was down‐regulated after intestinal ischaemia/reperfusion (I/R) damage. However, its role and potential mechanisms are still unknown. This study was aimed to elucidate the function of miR‐182 in intestinal I/R injury and the underlying mechanisms. The model of intestinal injury was constructed in wild‐type and Deptor knockout (KO) mice. Haematoxylin‐eosin staining, Chiu's score and diamine oxidase were utilized to detect intestinal damage. RT‐qPCR assay was used to detected miR‐182 expression. Electronic microscopy was used to detect autophagosome. Western blot was applied to detect the expression of Deptor, S6/pS6, LC3‐II/LC3‐I and p62. Dual‐luciferase reporter assay was used to verify the relationship between miR‐182 and Deptor. The results showed miR‐182 was down‐regulated following intestinal I/R. Up‐regulation of miR‐182 reduced intestinal damage, autophagy, Deptor expression and enhanced mTOR activity following intestinal I/R. Moreover, suppression of autophagy reduced intestinal damage and inhibition of mTOR by rapamycin aggravated intestinal damage following intestinal I/R. Besides, damage of intestine was reduced and mTOR activity was enhanced in Deptor KO mice. In addition, Deptor was the target gene of miR‐182 and was indispensable for the protection of miR‐182 on intestine under I/R condition. Together, our research implicated up‐regulation of miR‐182 inhibited autophagy to alleviate intestinal I/R injury via mTOR by targeting Deptor.     

Pre‐conditions for eliminating mitochondrial dysfunction and maintaining liver function after hepatic ischaemia reperfusion

The liver, the largest organ with multiple synthesis and secretion functions in mammals, consists of hepatocytes and Kupffer, stem, endothelial, stellate and other parenchymal cells. Because of early and extensive contact with the external environment, hepatic ischaemia reperfusion (IR) may result in mitochondrial dysfunction, autophagy and apoptosis of cells and tissues under various pathological conditions. Because the liver requires a high oxygen supply to maintain normal detoxification and synthesis functions, it is extremely susceptible to ischaemia and subsequent reperfusion with blood. Consequently, hepatic IR leads to acute or chronic liver failure and significantly increases the total rate of morbidity and mortality through multiple regulatory mechanisms. An increasing number of studies indicate that mitochondrial structure and function are impaired after hepatic IR, but that the health of liver tissues or liver grafts can be effectively rescued by attenuation of mitochondrial dysfunction. In this review, we mainly focus on the subsequent therapeutic interventions related to the conservation of mitochondrial function involved in mitigating hepatic IR injury and the potential mechanisms of protection. Because mitochondria are abundant in liver tissue, clarification of the regulatory mechanisms between mitochondrial dysfunction and hepatic IR should shed light on clinical therapies for alleviating hepatic IR‐induced injury.     

Luteolin limits infarct size and improves cardiac function after myocardium ischemia/reperfusion injury in diabetic rats

Background

The present study was to investigate the effects and mechanism of Luteolin on myocardial infarct size, cardiac function and cardiomyocyte apoptosis in diabetic rats with myocardial ischemia/reperfusion (I/R) injury.

Methodology/Principal Findings

Diabetic rats underwent 30 minutes of ischemia followed by 3 h of reperfusion. Animals were pretreated with or without Luteolin before coronary artery ligation. The severity of myocardial I/R induced LDH release, arrhythmia, infarct size, cardiac function impairment, cardiomyocyte apoptosis were compared. Western blot analysis was performed to elucidate the target proteins of Luteolin. The inflammatory cytokine production were also examined in ischemic myocardium underwent I/R injury. Our results revealed that Luteolin administration significantly reduced LDH release, decreased the incidence of arrhythmia, attenuated myocardial infarct size, enhanced left ventricular ejection fraction and decreased myocardial apoptotic death compared with I/R group. Western blot analysis showed that Luteolin treatment up-regulated anti-apoptotic proteins FGFR2 and LIF expression, increased BAD phosphorylation while decreased the ratio of Bax to Bcl-2. Luteolin treatment also inhibited MPO expression and inflammatory cytokine production including IL-6, IL-1a and TNF-a. Moreover, co-administration of wortmannin and Luteolin abolished the beneficial effects of Luteolin.

Conclusions/Significance

This study indicates that Luteolin preserves cardiac function, reduces infarct size and cardiomyocyte apoptotic rate after I/R injury in diabetic rats. Luteolin exerts its action by up-regulating of anti-apoptotic proteins FGFR2 and LIF expression, activating PI3K/Akt pathway while increasing BAD phosphorylation and decreasing ratio of Bax to Bcl-2.     

Alteration in gut microbiota caused by time‐restricted feeding alleviate hepatic ischaemia reperfusion injury in mice

Time‐restricted feeding (TRF), that is, no caloric intake for 14‐16 hours each day leads to favourable nutritional outcomes. This study is the first to investigate TRF through a surgical perspective verifying its efficacy against liver ischaemia reperfusion (I/R) injury. We randomly assigned 100 10‐week‐old wild‐type male C57BL/6 mice into two feeding regimens: TRF and ad libitum access to food. Main outcomes were evaluated at 6, 12 and 24 hours post‐I/R surgery after 12 weeks of intervention. TRF group demonstrated minor liver injury via histological study; lower serum levels of liver enzymes, glucose and lipids; higher concentrations of free fatty acid and β‐hydroxybutyrate; decreased oxidative stress and inflammatory biomarkers; as well as less severe cell apoptosis and proliferation. Further exploration indicated better gut microenvironment and intestinal epithelial tight junction function. TRF employed its positive influence on a wide spectrum of biochemical pathways and ultimately revealed protective effect against hepatic I/R injury possibly through adjusting the gut microbiota. The results referred to a strong indication of adopting better feeding pattern for surgical patients.     

The role of mitochondrial reactive oxygen species,NO and H2S in ischaemia/reperfusion injury and cardioprotection

Redox signalling in mitochondria plays an important role in myocardial ischaemia/reperfusion (I/R) injury and in cardioprotection. Reactive oxygen and nitrogen species (ROS/RNS) modify cellular structures and functions by means of covalent changes in proteins including among others S‐nitros(yl)ation by nitric oxide (NO) and its derivatives, and S‐sulphydration by hydrogen sulphide (H₂S). Many enzymes are involved in the mitochondrial formation and handling of ROS, NO and H₂S under physiological and pathological conditions. In particular, the balance between formation and removal of reactive species is impaired during I/R favouring their accumulation. Therefore, various interventions aimed at decreasing mitochondrial ROS accumulation have been developed and have shown cardioprotective effects in experimental settings. However, ROS, NO and H₂S play also a role in endogenous cardioprotection, as in the case of ischaemic pre‐conditioning, so that preventing their increase might hamper self‐defence mechanisms. The aim of the present review was to provide a critical analysis of formation and role of reactive species, NO and H₂S in mitochondria, with a special emphasis on mechanisms of injury and protection that determine the fate of hearts subjected to I/R. The elucidation of the signalling pathways of ROS, NO and H₂S is likely to reveal novel molecular targets for cardioprotection that could be modulated by pharmacological agents to prevent I/R injury.     

Hepatic 31P‐magnetic resonance spectroscopy identified the impact of melatonin‐pretreated mitochondria in acute liver ischaemia‐reperfusion injury

Acute liver ischaemia‐reperfusion injury (IRI), commonly encountered during liver resection and transplantation surgery, is strongly associated with unfavourable clinical outcome. However, a prompt and accurate diagnosis and the treatment of this entity remain formidable challenges. This study tested the hypothesis that ³¹P‐magnetic resonance spectroscopy (³¹P‐MRS) findings could provide reliable living images to accurately identify the degree of acute liver IRI and melatonin‐pretreated mitochondria was an innovative treatment for protecting the liver from IRI in rat. Adult male SD rats were categorized into group 1 (sham‐operated control), group 2 (IRI only) and group 3 (IRI + melatonin [ie mitochondrial donor rat received intraperitoneal administration of melatonin] pretreated mitochondria [10 mg/per rat by portal vein]). By the end of study period at 72 hours, ³¹P‐MRS showed that, as compared with group 1, the hepatic levels of ATP and NADH were significantly lower in group 2 than in groups 1 and 3, and significantly lower in group 3 than in group 1. The liver protein expressions of mitochondrial‐electron‐transport‐chain complexes and mitochondrial integrity exhibited an identical pattern to ³¹P‐MRS finding. The protein expressions of oxidative stress, inflammatory, cellular stress signalling and mitochondrial‐damaged biomarkers displayed an opposite finding of ³¹P‐MRS, whereas the protein expressions of antioxidants were significantly progressively increased from groups 1 to 3. Microscopic findings showed that the fibrotic area/liver injury score and inflammatory and DNA‐damaged biomarkers exhibited an identical pattern of cellular stress signalling. Melatonin‐pretreated mitochondria effectively protected liver against IRI and ³¹P‐MRS was a reliable tool for measuring the mitochondrial/ATP consumption in living animals.     

Maternal diabetes up‐regulates NOX2 and enhances myocardial ischaemia/reperfusion injury in adult offspring

Offspring of diabetic mothers are at risk of cardiovascular diseases in adulthood. However, the underlying molecular mechanisms are not clear. We hypothesize that prenatal exposure to maternal diabetes up‐regulates myocardial NOX2 expression and enhances ischaemia/reperfusion (I/R) injury in the adult offspring. Maternal diabetes was induced in C57BL/6 mice by streptozotocin. Glucose‐tolerant adult offspring of diabetic mothers and normal controls were subjected to myocardial I/R injury. Vascular endothelial growth factor (VEGF) expression, ROS generation, myocardial apoptosis and infarct size were assessed. The VEGF‐Akt (protein kinase B)‐mammalian target of rapamycin (mTOR)‐NOX2 signalling pathway was also studied in cultured cardiomyocytes in response to high glucose level. In the hearts of adult offspring from diabetic mothers, increases were observed in VEGF expression, NOX2 protein levels and both Akt and mTOR phosphorylation levels as compared to the offspring of control mothers. After I/R, ROS generation, myocardial apoptosis and infarct size were all significantly higher in the offspring of diabetic mothers relative to offspring of control mothers, and these differences were diminished by in vivo treatment with the NADPH oxidase inhibitor apocynin. In cultured cardiomyocytes, high glucose increased mTOR phosphorylation, which was inhibited by the PI3 kinase inhibitor LY294002. Notably, high glucose‐induced NOX2 protein expression and ROS production were inhibited by rapamycin. In conclusion, maternal diabetes promotes VEGF‐Akt‐mTOR‐NOX2 signalling and enhances myocardial I/R injury in the adult offspring. Increased ROS production from NOX2 is a possible molecular mechanism responsible for developmental origins of cardiovascular disease in offspring of diabetic mothers.     

Diabetes aggravates myocardial ischaemia reperfusion injury via activating Nox2‐related programmed cell death in an AMPK‐dependent manner

Cardiovascular diseases such as myocardial ischaemia have a high fatality rate in patients with diabetes. This study was designed to expose the crosstalk between oxidative stress and AMPK, a vital molecule that controls biological energy metabolism, in myocardial ischaemia reperfusion injury (I/RI) in diabetic rats. Diabetes was stimulated in rats using streptozotocin injection. Rats were separated on random into control, control + I/R, Diabetes, Diabetes + I/R, Diabetes + I/R + N‐acetylcysteine and Diabetes + I/R + Vas2870 groups. Myocardial infarct size was determined, and the predominant Nox family isoforms were analysed. In vitro, the H9C2 cells were administered excess glucose and exposed to hypoxia/reoxygenation to mimic diabetes and I/R. The AMPK siRNA or AICAR was used to inhibit or activate AMPK expression in H9C2 cells, respectively. Then, myocardial oxidative stress and programmed cell death were measured. Diabetes or high glucose levels were found to aggravate myocardial I/RI or hypoxia/reoxygenation in H9C2 cells, as demonstrated by an increase in myocardial infarct size or lactate dehydrogenase levels, oxidative stress generation and induction of programmed cell death. In diabetic rat hearts, cardiac Nox1, Nox2 and Nox4 were all heightened. The suppression of Nox2 expression using Vas2870 or Nox2‐siRNA treatment in vivo or in vitro, respectively, protected diabetic rats from myocardial I/RI. AMPK gene knockout increased Nox2 protein expression while AMPK agonist decreased Nox2 expression. Therefore, diabetes aggravates myocardial I/RI by generating of Nox2‐associated oxidative stress in an AMPK‐dependent manner, which led to the induction of programmed cell death such as apoptosis, pyroptosis and ferroptosis.     

缺血预处理对缺血/再灌注离体心脏的保护作用

目的：探讨连续多次短暂缺血预处理对缺血/再灌注损伤心肌的保护作用及机制。方法：采用大鼠离体心脏Lan-gendorff灌流模型,观察缺血预处理对心肌缺血/再灌注后不同时间点冠脉流出液中AST、CPK、UDH及冠脉流量,心肌组织中SOD、LPO以及再灌注性心律失常的影响。结果：缺血预处理可以减少缺血/再灌注损伤的心肌冠脉流出液中AST、CPK、LDH的含量,提高心肌SOD活性,降低LPO水平,并且抑制再灌注性心律失常的发生,提高再灌注期间的冠脉流量。结论：缺血预处理对心肌缺血/再灌注损伤具有一定保护作用。     

Long noncoding RNA TUG1 contributes to cerebral ischaemia/reperfusion injury by sponging mir‐145 to up‐regulate AQP4 expression

Emerging studies have shown that long noncoding RNA (lncRNA) TUG1 (taurine‐up‐regulated gene 1) plays critical roles in multiple biological processes. However, the expression and function of lncRNA TUG1 in cerebral ischaemia/reperfusion injury have not been reported yet. In this study, we found that LncRNA TUG1 expression was significantly up‐regulated in cultured MA‐C cells exposed to OGD/R injury, while similar results were also observed in MCAO model. Mechanistically, knockdown of TUG1 decreased lactate dehydrogenase levels and the ratio of apoptotic cells and promoted cell survival in vitro. Moreover, knockdown of TUG1 decreased AQP4 (encoding aquaporin 4) expression to attenuate OGD/R injury. TUG1 could interact directly with miR‐145, and down‐regulation of miR‐145 could efficiently reverse the function of TUG1 siRNA on AQP4 expression. Finally, the TUG1 shRNA reduced the infarction area and cell apoptosis in I/R mouse brains in vivo. In summary, our results suggested that lncRNA TUG1 may function as a competing endogenous RNA (ceRNA) for miR‐145 to induce cell damage, possibly providing a new therapeutic target in cerebral ischaemia/reperfusion injury.     

Omega 3 chronic supplementation attenuates myocardial ischaemia‐reperfusion injury through reinforcement of antioxidant defense system in rats

Currently, controversial clinical data about the protective effects in the consumption of n‐3 polyunsaturated fatty acids (PUFAs) in ischaemic heart diseases exist. Improved myocardial resistance to ischaemia‐reperfusion (IR) injury results in non‐lethal myocardial infarction, which is a relevant factor in the myocardial function. We hypothesized that chronic supplementation with PUFAs reduced infarct size (IS) and induced an improvement on oxidative stress‐related parameters in IR model. Rats were supplemented with two doses of PUFAs D1 (n = 7) (0.6 g kg^?1 d^?1) and D2 (n = 7) (1.2 g kg^?1 d^?1) for 8 weeks. Control group (n = 7) received only standard diet. In ex vivo model, all rat hearts were subjected to 30 min of global ischaemia followed by 120 min of reperfusion. The IS and left ventricular function were assessed. Lipid peroxidation, reduced glutathione (GSH)/oxidized glutathione (GSSG) ratio and antioxidant enzyme activity were measured in the whole heart. The results show a reduction in IS in a dose‐dependent manner with PUFAs D1 (30.6%) and D2 (48.5%) and higher values of left ventricular developed pressure, at the end of the reperfusion, for each dose, respectively (p < 0.05). The two PUFAs groups showed higher values of GSH/GSSG ratio and lipid peroxidation, and higher values of activity of antioxidant enzymes catalase, superoxide dismutase and glutathione peroxidase at basal condition (p < 0.05). At the end of reperfusion, the GSH/GSSG ratio and antioxidants enzyme activity did not show a significant drop in their values (p > 0.05). These findings suggested that the supplementation with PUFAs induces cardioprotection against IR injury, associated with reinforcement of the antioxidant defense system. Copyright © 2013 John Wiley & Sons, Ltd.