NAFLD and type 2 diabetes (T2D) share a bidirectional relationship and cardiovascular diseases (CVD) as leading death cause, but this relationship in prediabetes is still poorly known, especially for women which exhibit an exacerbated CVD risk. Here, we evaluated the effects of resveratrol supplementation (RSV) or diet intervention on the heart-liver axis of prediabetic female rats submitted to high-fat-high-sucrose diet (HFS). Forty Female Wistar rats were divided into 4 groups fed for 5 months with: a standard diet (CTRL), HFS diet (HFS), HFS diet supplemented for the last 2 months with RSV (1 mg/kg/day in water) (RSV) or HFS diet for 3 months followed by 2 months of standard diet (RSD). In vivo Magnetic Resonance Imaging was performed to study cardiac morphology/function. Then, tolerance to ischemia-reperfusion injury was investigated ex vivo by simultaneous measurement of cardiac function (RPP, EDP) and energy metabolism by 31P Magnetic Resonance Spectroscopy. Hepatic total lipid and triglycerides (TG) contents along expression of genes involved in lipid metabolism/inflammation were studied. HFS induced glucose intolerance (P < 0.01), increased diastolic left ventricular volume and thickness (P < 0.05) and heart weight to tibia length ratio (HTLR) (P < 0.01). HFS diet also altered myocardial tolerance to IR, with impaired RPP and EDP (P < 0.001) and PCr during reperfusion (P < 0.05). Both treatments improved glucose tolerance (P < 0.05). Importantly, despite maintained HFS diet, RSV restored the elevated diastolic volume and thickness in vivo (P < 0.01 CTRL, RSV vs. HFS, RSD) and HTLR level (P < 0.001 CTRL, RSV vs. HFS, RSD), conversely to RSD. Both treatments improved the tolerance to IR, with increased RPP (P < 0.05 vs. HFS) along with PCr during reperfusion (P < 0.001, P < 0.01 vs. HFS respectively). Only RSV decreased EDP during IR injury (P < 0.05). In the liver HFS diet induced total lipid and TG accumulation (P < 0.01) as well as elevated expression of lipid oxidation/peroxidation genes without effect on lipogenesis and inflammation cytokines genes. Both treatments normalized hepatic total lipid levels. RSD restored TG levels along elevated lipid metabolism gene expressions (P < 0.01) whereas RSV partially reduced the increase (P < 0.05). Despite maintained HFS and moderate hepatic lipid metabolism effects, RSV exhibited better cardioprotection than RSD, suggesting its potential to protect against early cardiac alterations associated with prediabetes and NAFLD, especially for women.

Cardiovascular disease (CVD) is known to be linked with nonalcoholic fatty liver disease (NAFLD) in type 2 diabetes, but few studies are interested in this relationship in prediabetes. This association needs to be explored especially among prediabetic women, who are at greater risk of CVD than men. We aimed to evaluate cardiac alterations and their association with hepatic inflammation and lipid metabolism in prediabetic female rats submitted to 5 months of high-fat-high-sucrose diet (HFS). Wistar female rats were divided in 2 groups fed for 5 months with standard or HFS diet (CTRL/HFS). We analyzed cardiac function, morphology, perfusion and fibrosis by magnetic resonance imaging, then rats underwent an IPGTT. The expression of genes involved in lipid metabolism and inflammation was afterwards studied in liver samples, as well as total lipid mass and triglyceride (TG) contents. Five months of HFS diet induced glucose intolerance (P < 0.05), and cardiac alterations with increased heart perfusion (P < 0.01), diastolic and systolic myocardial wall thickness (P < 0.05 and P < 0.001, respectively) but also increased diastolic left ventricular mass (P < 0.05). No significant differences were found in left ventricular ejection fraction, cardiac fibrosis, cardiac output and index between groups. HFS diet induced hepatic lipid accumulation with increased total lipid mass (P < 0.001) and TG contents, but also lipid metabolism adaptations with increased acyl CoA oxidase (P < 0.0001), CPT1a (P < 0.0001) and LCAD (P < 0.0001) expressions. No significant differences were found for FAS, MCAD and inflammatory cytokines (IL-6, TNF-α and MCP1) expressions. HFS diet in female rats induced cardiac alterations associated with hepatic lipid metabolism adaptations characterized by increased lipid peroxidation and β oxidation. Further analyses must be performed to study the relation between cardiac and hepatic alterations observed in female prediabetic rats.

Prediabetes is a strong predictor of type 2 diabetes and its associated cardiovascular complications, but few studies explore sexual dimorphism in this context. Here, we aim to determine whether sex influences physiological response to high-fat high-sucrose diet (HFS) and myocardial tolerance to ischemia-reperfusion injury. Male and female Wistar rats were subjected to standard (CTRL) or HFS diet for 5 months. Then, ex-vivo experiments on isolated perfused heart model were performed to evaluate tolerance to ischemia-reperfusion injury. HFS diet induced fasting hyperglycemia and increased body fat percent to a similar level in both sexes. However, glucose intolerance was more pronounced in female HFS. Cholesterol was increased only in female while male displayed higher level of plasmatic leptin. We observed increased heart weight to tibia length ratio only in males, but we showed a similar decrease in tolerance to ischemia-reperfusion injury in female and male HFS compared with respective controls, characterized by impaired cardiac function, energy metabolism and coronary flow during reperfusion. In conclusion, as soon as glucose intolerance and hyperglycemia develop, we observe higher sensitivity of hearts to ischemia-reperfusion injury without difference between males and females.

Prediabetes is an important risk factor for Type 2 diabetes and cardiovascular complications, such as myocardial infarction. However, few studies explore female sex in this context. Here, we aim to investigate the effects of high-fat high-sucrose diet on cardiac parameters and sensitivity to ischemia–reperfusion injury of female Wistar rats. Female Wistar rats received for 5 months normal diet (CTRL group) or high-fat high-sucrose diet (HFS group). Every month, MRI was performed to follow myocardial morphology, function and perfusion; cardiac and hepatic triglyceride content; and amount of sub-cutaneous and visceral adipose tissues. Then, ex vivo experiments were performed on isolated perfused hearts to evaluate tolerance to ischemia–reperfusion, with simultaneous measurement of energy metabolism by 31P MRS and contractile function. Coronary flow was measured before and after ischemia. At the end of the experiments, hearts were freeze-clamped for biochemical assays. Five months of high-fat high-sucrose diet induced a prediabetic condition in female Wistar rats, associated with an increase in myocardial perfusion, systolic and diastolic wall thickness. HFS rats also exhibited higher sensitivity to ischemia–reperfusion injury in comparison to controls, characterized by impaired cardiac function, energy metabolism and endothelial function. Biochemical analyses in hearts highlighted eNOS uncoupling, higher malondialdehyde level and lower S-Glutathionylation of proteins in HFS rats, indicating higher oxidative stress. Prediabetes induced by energy-dense diet was associated with modification of cardiac morphology and higher myocardial sensitivity to ischemia–reperfusion injury. These results may be related to the high risk of cardiovascular complications among Type 2 diabetic women.

Type 2 diabetic women have a high risk of mortality via myocardial infarction even with anti-diabetic treatments. Resveratrol (RSV) is a natural polyphenol, well-known for its antioxidant property, which has also shown interesting positive effects on mitochondrial function. Therefore, we aim to investigate the potential protective effect of 1 mg/kg/day of RSV on high energy compounds, during myocardial ischemia-reperfusion in type 2 diabetic female Goto-Kakizaki (GK) rats. For this purpose, we used 31P magnetic resonance spectroscopy in isolated perfused heart experiments, with a simultaneous measurement of myocardial function and coronary flow. RSV enhanced adenosine triphosphate (ATP) and phosphocreatine (PCr) contents in type 2 diabetic hearts during reperfusion, in combination with better functional recovery. Complementary biochemical analyses showed that RSV increased creatine, total adenine nucleotide heart contents and citrate synthase activity, which could be involved in better mitochondrial functioning. Moreover, improved coronary flow during reperfusion by RSV was associated with increased eNOS, SIRT1, and P-Akt protein expression in GK rat hearts. In conclusion, RSV induced cardioprotection against ischemia-reperfusion injury in type 2 diabetic female rats via increased high energy compound contents and expression of protein involved in NO pathway. Thus, RSV presents high potential to protect the heart of type 2 diabetic women from myocardial infarction.

The functional and bioenergetics impact of regular physical activity upon type-2 diabetic skeletal muscle independently of confounding factors of overweight remains undocumented. Here, gastrocnemius muscle energy fluxes, mitochondrial capacity and mechanical performance were assessed noninvasively and longitudinally in non-obese diabetic Goto-Kakizaki rats using magnetic resonance (MR) imaging and dynamic 31-phosphorus MR spectroscopy (31P-MRS) throughout a 6-min fatiguing bout of exercise performed before, in the middle (4-week) and at the end of an 8-week training protocol consisting in 60-min daily run on a treadmill. The training protocol reduced plasmatic insulin level (-61%) whereas blood glucose and non-esterified fatty acids levels remained unaffected, thereby indicating an improvement of insulin sensitivity. It also increased muscle mitochondrial citrate synthase activity (+45%) but this increase did not enhance oxidative ATP synthesis capacity in working muscle in vivo while glycolytic ATP production was increased (+33%). On the other hand, the training protocol impaired maximal force-generating capacity (-9%), total amount of force produced (-12%) and increased ATP cost of contraction (+32%) during the fatiguing exercise. Importantly, these deleterious effects were transiently worsened in the middle of the 8-week period, in association with reduced oxidative capacity and increased basal [P-i]/[PCr] ratio (an in vivo biomarker of muscle damage). These data demonstrate that the beneficial effect of regular training on insulin sensitivity in non-obese diabetic rat occurs separately from any improvement in muscle mitochondrial function and might be linked to an increased capacity for metabolizing glucose through anaerobic process in exercising muscle. (c) 2018 Elsevier B.V. and Societe Francaise de Biochimie et Biologie Moleculaire (SFBBM). All rights reserved.

Objectives: To investigate any gender effect of the beta-1 adrenergic blocker, landiolol, on cardiac performance and energy metabolism in septic rats, and to explore the expression of genes and proteins involved in this process. Design: Randomized animal study. Setting: University research laboratory. Subjects: Male and female Wistar rats. Interventions: One hour after cecal ligation and puncture, male and female rats were randomly allocated to the following groups: sham male, cecal ligation and puncture male, cecal ligation and puncture + landiolol male, sham female, cecal ligation and puncture female, and cecal ligation and puncture + landiolol female. Cardiac MRI was carried out 18 hours after cecal ligation and puncture to assess in vivo cardiac function. Ex vivo cardiac function measurement and 31P magnetic resonance spectroscopy were subsequently performed using an isovolumic isolated heart preparation. Finally, we assessed cardiac gene and protein expression. Measurements and Main Results: In males, landiolol increased indexed stroke volume by reversing the indexed end-diastolic volume reduction without affecting left ventricle ejection fraction. In females, landiolol did not increase indexed stroke volume and indexed end-diastolic volume but decreased left ventricle ejection fraction. Landiolol had no effect on ex vivo cardiac function and on high-energy phosphate compounds. The effect of landiolol on the gene expression of natriuretic peptide receptor 3 and on protein expression of phosphorylated-AKT:AKT ratio and endothelial nitric oxide synthase was different in males and females. Conclusions: Landiolol improved the in vivo cardiac performance of septic male rats while deleterious effects were reported in females. Expression of natriuretic peptide receptor 3, phosphorylated-AKT:AKT, and endothelial nitric oxide synthase are signaling pathways to investigate to better understand the sex differences in sepsis.

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Type 2 diabetes is associated with greater relative risk of cardiovascular diseases in women than in men, which is not well understood. Consequently, we have investigated if male and female displayed differences in cardiac function, energy metabolism, and endothelial function which could contribute to increased cardiovascular complications in type 2 diabetic female.

Docosahexaenoic acid (DHA) might prevent heart failure or optimise drug treatments by improving cardiac contraction. We investigated whether DHA-enriched avian glycerophospholipids (GPL-DHA) exert cardioprotection in ouabain-treated rats after 4 weeks of dietary supplementation with 10, 35 or 60 mg DHA per kg body weight versus none (DHA10, DHA35, DHA60 and control groups, respectively). The contractile responsiveness to different doses of ouabain (10(-7) to 10(-4) M), ouabain intoxication (at 3 × 10(-4) M), and relative variations in cardiac energy metabolism were determined using (31)P NMR in isolated perfused rat hearts. The fatty acid composition of cardiac membranes was analysed by gas chromatography. DHA accretion in the heart was dose-dependent (+8%, +30% and +45% for DHA10, DHA35 and DHA60, respectively). The cardiac phosphocreatine content significantly increased at the baseline in DHA35 (+45%) and DHA60 groups (+85%), and at the different doses of ouabain in the DHA60 group (+73% to 98%). The maximum positive inotropy achieved at 10(-4) M ouabain was significantly increased in all DHA groups versus control (+150%, +122.5% and +135% for DHA10, DHA35 and DHA60, respectively), and ouabain intoxication was delayed. The increase in myocardial phosphocreatine content and the improved efficacy of ouabain on myocardial contraction without toxicity suggest the potential of GPL-DHA as a dietary supplement or ingredient for functional food, and possibly as a co-treatment with digitalis drugs in humans.

Insulin resistance, altered lipid metabolism and mitochondrial dysfunction in skeletal muscle would play a major role in type 2 diabetes mellitus (T2DM) development, but the causal relationships between these events remain conflicting. To clarify this issue, gastrocnemius muscle function and energetics were investigated throughout a multidisciplinary approach combining in vivo and in vitro measurements in Goto-Kakizaki (GK) rats, a non-obese T2DM model developing peripheral insulin resistant without abnormal level of plasma non-esterified fatty acids (NEFA). Wistar rats were used as controls. Mechanical performance and energy metabolism were assessed strictly non-invasively using magnetic resonance (MR) imaging and 31-phosphorus MR spectroscopy (31P-MRS). Compared with control group, plasma insulin and glucose were respectively lower and higher in GK rats, but plasma NEFA level was normal. In resting GK muscle, phosphocreatine content was reduced whereas glucose content and intracellular pH were both higher. However, there were not differences between both groups for basal oxidative ATP synthesis rate, citrate synthase activity, and intramyocellular contents for lipids, glycogen, ATP and ADP (an important in vivo mitochondrial regulator). During a standardized fatiguing protocol (6 min of maximal repeated isometric contractions electrically induced at a frequency of 1.7 Hz), mechanical performance and glycolytic ATP production rate were reduced in diabetic animals whereas oxidative ATP production rate, maximal mitochondrial capacity and ATP cost of contraction were not changed. These findings provide in vivo evidence that insulin resistance is not caused by an impairment of mitochondrial function in this diabetic model.

Informations about the effects of exercice training on diabetesinduced myocardial dysfunctions are lacking. Consequently, we investigated the effect of exercice training on the sensitivity of the type 2 diabetic Goto-Kakizaki (GK) rat heart to ischaemia-reperfusion injury by using a multiparametric approach combining 31P magnetic resonance spectroscopy (MRS) with simultaneous measurement of contractile function and biochemical assays. 8-month-old male GK rat (n=11, trained GK) rats ran 60 min on a treadmill 5 days/week at a speed of 20 m/min for 7 weeks and compared with age-matched male Control (n=6) and untrained GK (n=11) rats. Then, isolated rat hearts were perfused with a physiological buffer containing 0.4 mM palmitate, for 24 min before switching to 1.2mM palmitate during 32min low-flow (0.5 ml/min-1.g wet wt-1) ischaemia. Next, flow was restored with 0.4 mM palmitate buffer for 32 minutes. High energy compounds and intracellular pH were followed using 31P MRS with simultaneous measurement of contractile function. Total adenine nucleotides (TAN) and energy charge (EC) were determined in freeze-clamped tissues by H.P.L.C. Heart to body weight ratios were significantly higher in both untrained and trained diabetic groups (p<0.02) versus Control. During reperfusion, rate pressure product and recoveries of PCr and ATP were significantly lower (p<0.05) in trained GK versus untrained GK and Control rat hearts. TAN and EC were significantly decreased in trained GK compared with Control (p=0.0052 and p=0.0156) and untrained GK (p=0.0024 and p=0.0005) rat hearts. The intensity of exercice training exacerbed the sensitivity of the type 2 diabetic 8-month-old Goto-Kakizaki rat to myocardial ischaemiareperfusion by impairing energy metabolism and myocardial performance. Other exercice protocols and/or therapeutic approaches need to be explored to limit myocardial ischaemia-reperfusion injury in type 2 diabetes.

The chronic administration of the β-adrenoreceptor agonist isoproterenol (IsoP) is used in animals to study the mechanisms of cardiac hypertrophy and failure associated with a sustained increase in circulating catecholamines. Time-dependent changes in myocardial blood flow (MBF), morphological and functional parameters were assessed in rats in vivo using multimodal cardiac MRI. Energy metabolism, oxidative stress and the nitric oxide (NO) pathway were evaluated in isolated perfused rat hearts following 7 days of treatment. Male Wistar rats were infused for 7 days with IsoP or vehicle using osmotic pumps. Cine-MRI and arterial spin labeling were used to determine left ventricular morphology, function and MBF at days 1, 2 and 7 after pump implantation. Isolated hearts were then perfused, and high-energy phosphate compounds and intracellular pH were followed using ³¹P MRS with simultaneous measurement of contractile function. Total creatine and malondialdehyde (MDA) contents were measured by high-performance liquid chromatography. The NO pathway was evaluated by NO synthase isoform expression and total nitrate concentration (NO(x)). In IsoP-treated rats, left ventricular mass was increased at day 1 and maintained. Wall thickness was increased with a peak at day 2 and a tendency to return to baseline values at day 7. MBF was markedly increased at day 1 and returned to normal values between days 1 and 2. The rate-pressure product and phosphocreatine/adenosine triphosphate ratio in perfused hearts were reduced. MDA, endothelial NO synthase expression and NO(x) were increased. Sustained high cardiac function and normal MBF after 24 h of IsoP infusion indicate imbalance between functional demand and blood flow, leading to morphological changes. After 1 week, cardiac hypertrophy and decreased function were associated with impaired phosphocreatine, increased oxidative stress and up-regulation of the NO pathway. These results provide supplemental information on the evolution of the different contributing factors leading to morphological and functional changes in this model of cardiac hypertrophy and failure.

Calcium overload induced by chronic administration of the β-adrenoreceptor agonist, isoproterenol (IP), is used in animal to study the mechanisms of cardiac hypertrophy and failure associated with a sustained increase in circulating catecholamines.

We investigated the tolerance of the insulin-resistant diabetic heart to ischemic injury in the male Goto-Kakizaki (GK) rat, a model of type 2 diabetes. Changes in energy metabolism, nitric oxide (NO) pathway, and cardiac function were assessed in the presence of physiological substrates. Age-matched control Wistar (n = 19) and GK (n = 18) isolated rat hearts were perfused with 0.4 mM palmitate, 3% albumin, 11 mM glucose, 3 U/l insulin, 0.2 mM pyruvate, and 0.8 mM lactate for 24 min before switching to 1.2 mM palmitate (11 rats/group) during 32 min low-flow (0.5 ml·min(-1)·g wet wt(-1)) ischemia. Next, flow was restored with 0.4 mM palmitate buffer for 32 min. A subset of hearts from each group (n = 8 for control and n = 7 for GK groups) were freeze-clamped for determining baseline values after the initial perfusion of 24 min. ATP, phosphocreatine (PCr), and intracellular pH (pH(i)) were followed using (31)P magnetic resonance spectroscopy with simultaneous measurement of contractile function. The NO pathway was determined by nitric oxide synthase (NOS) isoform expression and total nitrate concentration (NOx) in hearts. We found that coronary flow was 26% lower (P < 0.05) during baseline conditions and 61% lower (P < 0.05) during reperfusion in GK vs. control rat hearts. Rate pressure product was lower during reperfusion in GK vs. control rat hearts (P < 0.05). ATP, PCr, and pH(i) during ischemia-reperfusion were similar in both groups. Endothelial NOS expression was increased in GK rat hearts during baseline conditions (P < 0.05). NOx was increased during baseline conditions (P < 0.05) and after reperfusion (P < 0.05) in GK rat hearts. We report increased susceptibility of type 2 diabetic GK rat heart to ischemic injury that is not associated with impaired energy metabolism. Reduced coronary flow, upregulation of eNOS expression, and increased total NOx levels confirm NO pathway modifications in this model, presumably related to increased oxidative stress. Modifications in the NO pathway may play a major role in ischemia-reperfusion injury of the type 2 diabetic GK rat heart.

Myocardial injury caused by prolonged storage compromises post-transplantation contractile performance and induces endothelial injury. The aim of this study was to compare a solution developed in our laboratory [Centre de Résonance Magnétique Biologique et Médicale (CRMBM) solution] with a widely used solution (Celsior, Genzyme, Saint Germain en Laye, France). Metabolic and contractile parameters as well as indexes of endothelial injury were measured in a heterotopic rat heart transplantation model with a 3-h ischaemia and a 1-h reperfusion. The two solutions were randomly used for cardioplegia and graft preservation in six experiments each. During reperfusion, developed pressure and rate pressure product were higher with CRMBM compared with Celsior (P = 0.0002 and P = 0.0135, respectively). Phosphocreatine and adenosine triphosphate (ATP) concentrations after reperfusion were significantly higher with CRMBM (P = 0.0069 and P = 0.0053, respectively). Endothelial nitric oxide synthase (eNOS) and neuronal nitric oxide synthase (nNOS) protein expression were decreased to the same extent after reperfusion compared with baseline with CRMBM (P = 0.0001 and P < 0.0001, respectively) and Celsior (P = 0.0007 and P < 0.0001, respectively). Total nitrate concentration (NOx) was significantly increased after reperfusion with CRMBM (P < 0.0001 versus baseline and P < 0.0001 versus Celsior). Na,K-ATPase activity was decreased in both groups versus baseline after reperfusion (P < 0.0001 for CRMBM and P < 0.0001 for Celsior). We showed limitation of both myocardial and endothelial damage with CRMBM compared with Celsior during heterotopic rat heart transplantation in vivo.