Karen M. O'Shea

Mitofusin-2 Maintains Mitochondrial Structure and Contributes to Stress-Induced Permeability Transition in Cardiac Myocytes

ABSTRACT Mitofusin-2 (Mfn-2) is a dynamin-like protein that is involved in the rearrangement of the outer mitochondrial membrane. Research using various experimental systems has shown that Mfn-2 is a mediator of mitochondrial fusion, an evolutionarily conserved process responsible for the surveillance of mitochondrial homeostasis. Here, we find that cardiac myocyte mitochondria lacking Mfn-2 are pleiomorphic and have the propensity to become enlarged. Consistent with an underlying mild mitochondrial dysfunction, Mfn-2-deficient mice display modest cardiac hypertrophy accompanied by slight functional deterioration. The absence of Mfn-2 is associated with a marked delay in mitochondrial permeability transition downstream of Ca2+ stimulation or due to local generation of reactive oxygen species (ROS). Consequently, Mfn-2-deficient adult cardiomyocytes are protected from a number of cell death-inducing stimuli and Mfn-2 knockout hearts display better recovery following reperfusion injury. We conclude that in cardiac myocytes, Mfn-2 controls mitochondrial morphogenesis and serves to predispose cells to mitochondrial permeability transition and to trigger cell death.

Fish oil, but not flaxseed oil, decreases inflammation and prevents pressure overload-induced cardiac dysfunction

AIMS Clinical studies suggest that intake of omega-3 polyunsaturated fatty acids (omega-3 PUFA) may lower the incidence of heart failure. Dietary supplementation with omega-3 PUFA exerts metabolic and anti-inflammatory effects that could prevent left ventricle (LV) pathology; however, it is unclear whether these effects occur at clinically relevant doses and whether there are differences between omega-3 PUFA from fish [eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA)] and vegetable sources [alpha-linolenic acid (ALA)]. METHODS AND RESULTS We assessed the development of LV remodelling and pathology in rats subjected to aortic banding treated with omega-3 PUFA over a dose range that spanned the intake of humans taking omega-3 PUFA supplements. Rats were fed a standard food or diets supplemented with EPA+DHA or ALA at 0.7, 2.3, or 7% of energy intake. Without supplementation, aortic banding increased LV mass and end-systolic and -diastolic volumes. ALA supplementation had little effect on LV remodelling and dysfunction. In contrast, EPA+DHA dose-dependently increased EPA and DHA, decreased arachidonic acid in cardiac membrane phospholipids, and prevented the increase in LV end-diastolic and -systolic volumes. EPA+DHA resulted in a dose-dependent increase in the anti-inflammatory adipokine adiponectin, and there was a strong correlation between the prevention of LV chamber enlargement and plasma levels of adiponectin (r = -0.78). Supplementation with EPA+DHA had anti-aggregatory and anti-inflammatory effects as evidenced by decreases in urinary thromboxane B(2) and serum tumour necrosis factor-alpha. CONCLUSION Dietary supplementation with omega-3 PUFA derived from fish, but not from vegetable sources, increased plasma adiponectin, suppressed inflammation, and prevented cardiac remodelling and dysfunction under pressure overload conditions.

Insulin Resistance: Metabolic Mechanisms and Consequences in the Heart

Insulin resistance is a characteristic feature of obesity and type 2 diabetes mellitus and impacts the heart in various ways. Impaired insulin-mediated glucose uptake is a uniformly observed characteristic of the heart in these states, although changes in upstream kinase signaling are variable and dependent on the severity and duration of the associated obesity or diabetes mellitus. The understanding of the physiological and pathophysiological role of insulin resistance in the heart is evolving. To maintain its high energy demands, the heart is capable of using many metabolic substrates. Although insulin signaling may directly regulate cardiac metabolism, its main role is likely the regulation of substrate delivery from the periphery to the heart. In addition to promoting glucose uptake, insulin regulates long-chain fatty acid uptake, protein synthesis, and vascular function in the normal cardiovascular system. Recent advances in understanding the role of metabolic, signaling, and inflammatory pathways in obesity have provided opportunities to better understand the pathophysiology of insulin resistance in the heart. This review will summarize our current understanding of metabolic mechanisms for and consequences of insulin resistance in the heart and will discuss potential new areas for investigating novel mechanisms that contribute to insulin resistance in the heart.

ω-3 polyunsaturated fatty acid supplementation for the treatment of heart failure: mechanisms and clinical potential

Heart failure (HF) is a complex clinical syndrome with multiple aetiologies. Current treatment options can slow the progression to HF, but overall the prognosis remains poor. Clinical studies suggest that high dietary intake of the omega-3 polyunsaturated fatty acids (omega-3PUFA) found in fish oils (eicosapentaenoic and docosahexaenoic acids) may lower the incidence of HF, and that supplementation with pharmacological doses prolongs event-free survival in patients with established HF. The mechanisms for these potential benefits are complex and not well defined. It is well established that fish oil supplementation lowers plasma triglyceride levels, and more recent work demonstrates anti-inflammatory effects, including reduced circulating levels of inflammatory cytokines and arachidonic acid-derived eicosanoids, and elevated plasma adiponectin. In animal studies, fish oil favourably alters cardiac mitochondrial function. All of these effects may work to prevent the development and progression of HF. The omega-3PUFA found in plant sources, alpha-linolenic acid, may also be protective in HF; however, the evidence is not as compelling as for fish oil. This review summarizes the evidence related to use of omega-3PUFA supplementation as a potential treatment for HF and discusses possible mechanisms of action. In general, there is growing evidence that supplementation with omega-3PUFA positively impacts established pathophysiological targets in HF and has potential therapeutic utility for HF patients.

Dietary ω-3 fatty acids alter cardiac mitochondrial phospholipid composition and delay Ca2+-induced permeability transition

Consumption of omega-3 fatty acids from fish oil, specifically eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), decreases risk for heart failure and attenuates pathologic cardiac remodeling in response to pressure overload. Dietary supplementation with EPA + DHA may also impact cardiac mitochondrial function and energetics through alteration of membrane phospholipids. We assessed the role of EPA + DHA supplementation on left ventricular (LV) function, cardiac mitochondrial membrane phospholipid composition, respiration, and sensitivity to mitochondrial permeability transition pore (MPTP) opening in normal and infarcted myocardium. Rats were subjected to sham surgery or myocardial infarction by coronary artery ligation (n=10-14), and fed a standard diet, or supplemented with EPA + DHA (2.3% of energy intake) for 12 weeks. EPA + DHA altered fatty acid composition of total mitochondrial phospholipids and cardiolipin by reducing arachidonic acid content and increasing DHA incorporation. EPA + DHA significantly increased calcium uptake capacity in both subsarcolemmal and intrafibrillar mitochondria from sham rats. This treatment effect persisted with the addition of cyclosporin A, and was not accompanied by changes in mitochondrial respiration or coupling, or cyclophilin D protein expression. Myocardial infarction resulted in heart failure as evidenced by LV dilation and contractile dysfunction. Infarcted LV myocardium had decreased mitochondrial protein yield and activity of mitochondrial marker enzymes, however respiratory function of isolated mitochondria was normal. EPA + DHA had no effect on LV function, mitochondrial respiration, or MPTP opening in rats with heart failure. In conclusion, dietary supplementation with EPA + DHA altered mitochondrial membrane phospholipid fatty acid composition in normal and infarcted hearts, but delayed MPTP opening only in normal hearts.

Improved Mitochondrial Function with Diet-Induced Increase in Either Docosahexaenoic Acid or Arachidonic Acid in Membrane Phospholipids

Mitochondria can depolarize and trigger cell death through the opening of the mitochondrial permeability transition pore (MPTP). We recently showed that an increase in the long chain n3 polyunsaturated fatty acids (PUFA) docosahexaenoic acid (DHA; 22:6n3) and depletion of the n6 PUFA arachidonic acid (ARA; 20:4n6) in mitochondrial membranes is associated with a greater Ca2+ load required to induce MPTP opening. Here we manipulated mitochondrial phospholipid composition by supplementing the diet with DHA, ARA or combined DHA+ARA in rats for 10 weeks. There were no effects on cardiac function, or respiration of isolated mitochondria. Analysis of mitochondrial phospholipids showed DHA supplementation increased DHA and displaced ARA in mitochondrial membranes, while supplementation with ARA or DHA+ARA increased ARA and depleted linoleic acid (18:2n6). Phospholipid analysis revealed a similar pattern, particularly in cardiolipin. Tetralinoleoyl cardiolipin was depleted by 80% with ARA or DHA+ARA supplementation, with linoleic acid side chains replaced by ARA. Both the DHA and ARA groups had delayed Ca2+-induced MPTP opening, but the DHA+ARA group was similar to the control diet. In conclusion, alterations in mitochondria membrane phospholipid fatty acid composition caused by dietary DHA or ARA was associated with a greater cumulative Ca2+ load required to induced MPTP opening. Further, high levels of tetralinoleoyl cardiolipin were not essential for normal mitochondrial function if replaced with very-long chain n3 or n6 PUFAs.

Dietary supplementation with docosahexaenoic acid, but not eicosapentaenoic acid, dramatically alters cardiac mitochondrial phospholipid fatty acid composition and prevents permeability transition.

Treatment with the omega-3 polyunsaturated fatty acids (PUFAs) docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA) exerts cardioprotective effects, and suppresses Ca2+-induced opening of the mitochondrial permeability transition pore (MPTP). These effects are associated with increased DHA and EPA, and lower arachidonic acid (ARA) in cardiac phospholipids. While clinical studies suggest the triglyceride lowering effects of DHA and EPA are equivalent, little is known about the independent effects of DHA and EPA on mitochondria function. We compared the effects of dietary supplementation with the omega-3 PUFAs DHA and EPA on cardiac mitochondrial phospholipid fatty acid composition and Ca2+-induced MPTP opening. Rats were fed a standard lab diet with either normal low levels of omega-3 PUFA, or DHA or EPA at 2.5% of energy intake for 8 weeks, and cardiac mitochondria were isolated and analyzed for Ca2+-induced MPTP opening and phospholipid fatty acyl composition. DHA supplementation increased both DHA and EPA and decreased ARA in mitochondrial phospholipid, and significantly delayed MPTP opening as assessed by increased Ca2+ retention capacity and decreased Ca2+-induced mitochondria swelling. EPA supplementation increased EPA in mitochondrial phospholipids, but did not affect DHA, only modestly lowered ARA, and did not affect MPTP opening. In summary, dietary supplementation with DHA but not EPA, profoundly altered mitochondrial phospholipid fatty acid composition and delayed Ca2+-induced MPTP opening.

A high-fat diet increases adiposity but maintains mitochondrial oxidative enzymes without affecting development of heart failure with pressure overload

A high-fat diet can increase adiposity, leptin secretion, and plasma fatty acid concentration. In hypertension, this scenario may accelerate cardiac hypertrophy and development of heart failure but could be protective by activating peroxisome proliferator-activated receptors and expression of mitochondrial oxidative enzymes. We assessed the effects of a high-fat diet on the development of left ventricular hypertrophy, remodeling, contractile dysfunction, and the activity of mitochondrial oxidative enzymes. Mice (n = 10-12/group) underwent transverse aortic constriction (TAC) or sham surgery and were fed either a low-fat diet (10% of energy intake as fat) or a high-fat diet (45% fat) for 6 wk. The high-fat diet increased adipose tissue mass and plasma leptin and insulin. Left ventricular mass and chamber size were unaffected by diet in sham animals. TAC increased left ventricular mass (approximately 70%) and end-systolic and end-diastolic areas (approximately 100% and approximately 45%, respectively) to the same extent in both dietary groups. The high-fat diet increased plasma free fatty acid concentration and prevented the decline in the activity of the mitochondrial enzymes medium chain acyl-coenzyme A dehydrogenase (MCAD) and citrate synthase that was observed with TAC animals on a low-fat diet. In conclusion, a high-fat diet did not worsen cardiac hypertrophy or left ventricular chamber enlargement despite increases in fat mass and insulin and leptin concentrations. Furthermore, a high-fat diet preserved MCAD and citrate synthase activities during pressure overload, suggesting that it may help maintain mitochondrial oxidative capacity in failing myocardium.

Treatment with Docosahexaenoic Acid, but Not Eicosapentaenoic Acid, Delays Ca2+-Induced Mitochondria Permeability Transition in Normal and Hypertrophied Myocardium

Intake of fish oil containing docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA) prevents heart failure; however, the mechanisms are unclear. Mitochondrial permeability transition pore (MPTP) opening contributes to myocardial pathology in cardiac hypertrophy and heart failure, and treatment with DHA + EPA delays MPTP opening. Here, we assessed: 1) whether supplementation with both DHA and EPA is needed for optimal prevention of MPTP opening, and 2) whether this benefit occurs in hypertrophied myocardium. Rats with either normal myocardium or cardiac hypertrophy induced by 8 weeks of abdominal aortic banding were fed one of four diets: control diet without DHA or EPA or diets enriched with either DHA, EPA, or DHA + EPA (1:1 ratio) at 2.5% of energy intake for 17 weeks. Aortic banding caused a 27% increase in left ventricular mass and 25% depletion in DHA in mitochondrial phosopholipids in rats fed the control diet. DHA supplementation raised DHA in phospholipids ∼2-fold in both normal and hypertrophied hearts and increased EPA. DHA + EPA supplementation also increased DHA, but to a lesser extent than DHA alone. EPA supplementation increased EPA, but did not affect DHA compared with the control diet. Ca2+-induced MPTP opening was delayed by DHA and DHA + EPA supplementation in both normal and hypertrophied hearts, but EPA had no effect on MPTP opening. These results show that supplementation with DHA alone effectively increases both DHA and EPA in cardiac mitochondrial phospholipids and delays MPTP and suggest that treatment with DHA + EPA offers no advantage over DHA alone.

The antioxidant tempol attenuates pressure overload-induced cardiac hypertrophy and contractile dysfunction in mice fed a high-fructose diet

We have previously shown that high-sugar diets increase mortality and left ventricular (LV) dysfunction during pressure overload. The mechanisms behind these diet-induced alterations are unclear but may involve increased oxidative stress in the myocardium. The present study examined whether high-fructose feeding increased myocardial oxidative damage and exacerbated systolic dysfunction after transverse aortic constriction (TAC) and if this effect could be attenuated by treatment with the antioxidant tempol. Immediately after surgery, TAC and sham mice were assigned to a high-starch diet (58% of total energy intake as cornstarch and 10% fat) or high-fructose diet (61% fructose and 10% fat) with or without the addition of tempol [0.1% (wt/wt) in the chow] and maintained on the treatment for 8 wk. In response to TAC, fructose-fed mice had greater cardiac hypertrophy (55.1% increase in the heart weight-to-tibia length ratio) than starch-fed mice (22.3% increase in the heart weight-to-tibia length ratio). Treatment with tempol significantly attenuated cardiac hypertrophy in fructose-fed TAC mice (18.3% increase in the heart weight-to-tibia ratio). Similarly, fructose-fed TAC mice had a decreased LV area of fractional shortening (from 38+/-2% in sham to 22+/-4% in TAC), which was prevented by tempol treatment (33+/-3%). Markers of lipid peroxidation in fructose-fed TAC hearts were also blunted by tempol. In conclusion, tempol significantly blunted markers of cardiac hypertrophy, LV remodeling, contractile dysfunction, and oxidative stress in fructose-fed TAC mice.