ISO-1 reduced aerobic capacity. (A) Basal <i>V</i>O₂ and (B) <i>V</i>CO₂ measured by indirect calorimetry.

<p>(C) Maximal <i>V</i>O₂ (<i>V</i>O₂max) obtained from mice subjected to treadmill exercise. (D) Difference between <i>V</i>O₂ and <i>V</i>O₂max (Δ<i>V</i>O₂) in ND, HFD and ISO-1-treated HFD mice. n = 6–10, *p<0.05 vs. ND, ^†p<0.05 vs. HFD.</p

ISO-1 treatment did not modify metabolic syndrome-associated parameters.

<p>(A) Abdominal fat mass, (B) subcutaneous fat mass, (C) abdominal to subcutaneous fat mass ratio, (D) plasma true triglyceride levels, (E) plasma cholesterol concentration and (F) fasting blood glucose. Data are means ± SEM, n = 5; *p<0.05 vs. ND. (G) Oral glucose tolerance test and (H) insulin tolerance test performed on ND, HFD and HFD+ISO-1 mice. n = 5–10 in each group. *p<0.05 vs. ND.</p

HFD induced systemic and cardiac MIF expression.

<p>(A) Tautomerase activity measured on plasma sample prepared from ND and HFD mice. Consumption of L-dopachrome-methyl ester was followed by measuring optical density at 475 nm. (B) Cardiac <i>Mif</i> transcript expression detected by RT-qPCR normalized to <i>β-actin</i>. (C) MIF protein cardiac expression observed by western-blot. Left panel is a representative western-blot, right panel is a semi-quantitation calculated after densitometric analysis. Data are means ± SEM. n = 6–10; *p<0.05 vs. ND.</p

ISO-1 did not worsen mitochondrial function but blocked HFD-induced mitochondrial biogenesis.

<p>(A) Glutamate/malate (GM) and (B) palmitoylcarnitine/malate (PC) were used as substrates. Then, 2.5 mM ADP were added. Ratio was calculated as described in materials and methods. n = 6–10, *p<0.05 vs. ND. (C) Relative expression of mitochondrial-encoded genes <i>mtCOII</i> and <i>Nd1</i> compared to the nuclear-encoded gene <i>Ppia</i>. (D) Expression of <i>Pgc-1α</i> transcripts analyzed by real-time RT-PCR. n = 5, *p<0.05 vs. ND, ^†p<0.05 vs. HFD.</p

ISO-1 amplified HFD-induced cardiac dysfunction.

<p>(A) Coronary perfusion pressure (CPP) measured in presence of rising concentration of sodium nitroprusside (SNP) on isolated heart preparations obtained from ND, ISO-1-injected ND mice (ND+ISO-1), HFD and ISO-1-treated HFD mice (HFD+ISO-1). (B) Left ventricular developed pressure (LVDP), (C) +d<i>P</i>/d<i>t</i> max and (D) −d<i>P</i>/d<i>t</i> min measured on ND, ND+ISO-1, HFD, HFD+ISO-1 hearts, in presence of the β-adrenergic receptor agonist isoproterenol. Data are means ± SEM of 5 experiments. *p<0.05 vs. ND; ^†p<0.05 vs. HFD.</p

Carbon Monoxide Improves Cardiac Function and Mitochondrial Population Quality in a Mouse Model of Metabolic Syndrome

<div><h3>Aims</h3><p>Metabolic syndrome induces cardiac dysfunction associated with mitochondria abnormalities. As low levels of carbon monoxide (CO) may improve myocardial and mitochondrial activities, we tested whether a CO-releasing molecule (CORM-3) reverses metabolic syndrome-induced cardiac alteration through changes in mitochondrial biogenesis, dynamics and autophagy.</p> <h3>Methods and Results</h3><p>Mice were fed with normal diet (ND) or high-fat diet (HFD) for twelve weeks. Then, mice received two intraperitoneal injections of CORM-3 (10 mg.kg⁻¹), with the second one given 16 hours after the first. Contractile function in isolated hearts and mitochondrial parameters were evaluated 24 hours after the last injection. Mitochondrial population was explored by electron microscopy. Changes in mitochondrial dynamics, biogenesis and autophagy were assessed by western-blot and RT-qPCR. Left ventricular developed pressure was reduced in HFD hearts. Mitochondria from HFD hearts presented reduced membrane potential and diminished ADP-coupled respiration. CORM-3 restored both cardiac and mitochondrial functions. Size and number of mitochondria increased in the HFD hearts but not in the CORM-3–treated HFD group. CORM-3 modulated HFD-activated mitochondrial fusion and biogenesis signalling. While autophagy was not activated in the HFD group, CORM-3 increased the autophagy marker LC3-II. Finally, <em>ex vivo</em> experiments demonstrated that autophagy inhibition by 3-methyladenine abolished the cardioprotective effects of CORM-3.</p> <h3>Conclusion</h3><p>CORM-3 may modulate pathways controlling mitochondrial quality, thus leading to improvements of mitochondrial efficiency and HFD-induced cardiac dysfunction.</p> </div

Effects of CORM-3 on metabolic parameters.

<p>(A) Oral glucose tolerance and (B) insulin tolerance tests were performed on ND (black circles), ND + CORM-3 (black triangles), HFD (black squares) and HFD + CORM-3 (white diamonds) mice. Data are means ± SEM. n  = 10 in each group. (a) p<0.05 between HFD and ND; (b) p<0.05 between HFD + CORM-3 and ND. (C) Plasma cholesterol and (D) true triglyceride measurements were performed on 6–10 different samples in each group. Data are means ± SEM. (E) Basal oxygen uptake (<i>V</i>O₂) and carbon dioxide rejection (<i>V</i>CO₂), (F) respiratory exchange ratio (RER). Data represent means ± SEM. n  = 5–7 in each group, * p<0.05 vs. ND. White bars  =  ND; vertically hatched bars  =  ND + CORM-3; black bars  =  HFD; crosshatched bars  =  HFD + CORM-3.</p

Effects of CORM-3 on high-fat-induced cardiac dysfunction.

<p>(A) Left Ventricular Developed Pressure (LVDP) of hearts isolated from normal diet (ND) or high-fat diet (HFD) mice injected twice with iCORM-3 (10 mg.kg⁻¹, intraperitoneally, studied 24 hours after the second injection) or CORM-3 (10 mg.kg⁻¹, intraperitoneally, studied 24, 48 or 72 hours after the second injection, or 24 hours after the fifth injection (x5); see Materials and Methods). (B) LVDP first derivatives ±d<i>P</i>/d<i>t</i> and (C) ratio (LVDP x Heart Rate)/MVO₂ (see Materials and Methods) obtained from ND, ND + CORM-3, HFD and CORM-3–treated HFD mouse hearts. Evaluation was performed 24 hours after the second injection. (D) Maximal oxygen uptake (<i>V</i>O₂) increase of mice subjected to exercise stress test. Data represent means ± SEM. n  = 5–8 in each group. White bars  =  ND; vertically hatched bars  =  ND + CORM-3; black bars  =  HFD; crosshatched bars  =  HFD + CORM-3. * p<0.05 vs. ND, † p<0.05 vs. HFD.</p

CORM-3 did not change UCP expression.

<p>(A) Results of quantitative RT-PCR detecting <i>Ucp2</i> and <i>Ucp3</i> mRNA expressions. Results were normalized to <i>â-actin</i>. (B) Representative western-blot targeting UCP3 and GAPDH proteins and (C) densitometric analysis of UCP3 to GAPDH ratio. White bars  =  ND; vertically hatched bars  =  ND + CORM-3; black bars  =  HFD; crosshatched bars  =  CORM-3–treated HFD mice. Data are means ± SEM. n  = 5 in each group. * p<0.05 vs. ND.</p