Curtis Olsen

PGC-1β Deficiency Accelerates the Transition to Heart Failure in Pressure Overload Hypertrophy

Rationale: Pressure overload cardiac hypertrophy, a risk factor for heart failure, is associated with reduced mitochondrial fatty acid oxidation (FAO) and oxidative phosphorylation (OXPHOS) proteins that correlate in rodents with reduced PGC-1&agr; expression. Objective: To determine the role of PGC-1&bgr; in maintaining mitochondrial energy metabolism and contractile function in pressure overload hypertrophy. Methods and Results: PGC-1&bgr; deficient (KO) mice and wildtype (WT) controls were subjected to transverse aortic constriction (TAC). Although LV function was modestly reduced in young KO hearts, there was no further decline with age so that LV function was similar between KO and WT when TAC was performed. WT-TAC mice developed relatively compensated LVH, despite reduced mitochondrial function and repression of OXPHOS and FAO genes. In nonstressed KO hearts, OXPHOS gene expression and palmitoyl-carnitine-supported mitochondrial function were reduced to the same extent as banded WT, but FAO gene expression was normal. Following TAC, KO mice progressed more rapidly to heart failure and developed more severe mitochondrial dysfunction, despite a similar overall pattern of repression of OXPHOS and FAO genes as WT-TAC. However, in relation to WT-TAC, PGC-1&bgr; deficient mice exhibited greater degrees of oxidative stress, decreased cardiac efficiency, lower rates of glucose metabolism, and repression of hexokinase II protein. Conclusions: PGC-1&bgr; plays an important role in maintaining baseline mitochondrial function and cardiac contractile function following pressure overload hypertrophy by preserving glucose metabolism and preventing oxidative stress.

Impaired insulin signaling accelerates cardiac mitochondrial dysfunction after myocardial infarction

Diabetes increases mortality and accelerates left ventricular (LV) dysfunction following myocardial infarction (MI). This study sought to determine the impact of impaired myocardial insulin signaling, in the absence of diabetes, on the development of LV dysfunction following MI. Mice with cardiomyocyte-restricted knock out of the insulin receptor (CIRKO) and wildtype (WT) mice were subjected to proximal left coronary artery ligation (MI) and followed for 14 days. Despite equivalent infarct size, mortality was increased in CIRKO-MI vs. WT-MI mice (68% vs. 40%, respectively). In surviving mice, LV ejection fraction and dP/dt were reduced by >40% in CIRKO-MI vs. WT-MI. Relative to shams, isometric developed tension in LV papillary muscles increased in WT-MI but not in CIRKO-MI. Time to peak tension and relaxation times were prolonged in CIRKO-MI vs. WT-MI suggesting impaired, load-independent myocardial contractile function. To elucidate mechanisms for impaired LV contractility, mitochondrial function was examined in permeabilized cardiac fibers. Whereas maximal ADP-stimulated mitochondrial O(2) consumption rates (V(ADP)) with palmitoyl carnitine were unchanged in WT-MI mice relative to sham-operated animals, V(ADP) was significantly reduced in CIRKO-MI (13.17+/-0.94 vs. 9.14+/-0.88 nmol O(2)/min/mgdw, p<0.05). Relative to WT-MI, expression levels of GLUT4, PPAR-alpha, SERCA2, and the FA-Oxidation genes MCAD, LCAD, CPT2 and the electron transfer flavoprotein ETFDH were repressed in CIRKO-MI. Thus reduced insulin action in cardiac myocytes accelerates post-MI LV dysfunction, due in part to a rapid decline in mitochondrial FA oxidative capacity, which combined with limited glucose transport capacity that may reduce substrate utilization and availability.

Nitro-oleic acid protects against endotoxin-induced endotoxemia and multiorgan injury in mice

Nitroalkene derivatives of nitro-oleic acid (OA-NO2) are endogenous lipid products with potent anti-inflammatory properties in vitro. The present study was undertaken to evaluate the in vivo anti-inflammatory effect of OA-NO2 in mice given LPS. Two days before LPS administration, C57BL/6J mice were chronically infused with vehicle (LPS vehicle) or OA-NO2 (LPS OA-NO2) at 200 microg x kg(-1) x day(-1) via osmotic minipumps; LPS was administered via a single intraperitoneal (ip) injection (10 mg/kg in saline). A third group received an ip injection of saline without LPS or OA-NO2 and served as controls. At 18 h of LPS administration, LPS vehicle mice displayed multiorgan dysfunction as evidenced by elevated plasma urea and creatinine (kidney), aspartate aminotransferase (AST) and alanine aminotransferase (ALT; liver), and lactate dehydrogenase (LDH) and reduced ejection fraction (heart). In contrast, the severity of multiorgan dysfunction was less in LPS OA-NO2 animals. The levels of circulating TNF-alpha and renal TNF-alpha mRNA expression, together with renal mRNA expression of monocyte chemoattractant protein-1, ICAM-1, and VCAM-1, and with renal mRNA and protein expression of inducible nitric oxide synthase and cyclooxygenase 2, and renal cGMP and PGE2 contents, were greater in LPS vehicle vs. control mice, but were attenuated in LPS OA-NO2 animals. Similar patterns of changes in the expression of inflammatory mediators were observed in the liver. Together, pretreatment with OA-NO2 ameliorated the inflammatory response and multiorgan injury in endotoxin-induced endotoxemia in mice.

Mechanistic Target of Rapamycin (Mtor) Is Essential for Murine Embryonic Heart Development and Growth

Mechanistic target of rapamycin (Mtor) is required for embryonic inner cell mass proliferation during early development. However, Mtor expression levels are very low in the mouse heart during embryogenesis. To determine if Mtor plays a role during mouse cardiac development, cardiomyocyte specific Mtor deletion was achieved using α myosin heavy chain (α-MHC) driven Cre recombinase. Initial mosaic expression of Cre between embryonic day (E) 10.5 and E11.5 eliminated a subset of cardiomyocytes with high Cre activity by apoptosis and reduced overall cardiac proliferative capacity. The remaining cardiomyocytes proliferated and expanded normally. However loss of 50% of cardiomyocytes defined a threshold that impairs the ability of the embryonic heart to sustain the embryos circulatory requirements. As a result 92% of embryos with cardiomyocyte Mtor deficiency died by the end of gestation. Thus Mtor is required for survival and proliferation of cardiomyocytes in the developing heart.

Inducible Overexpression of GLUT1 Prevents Mitochondrial Dysfunction and Attenuates Structural Remodeling in Pressure Overload but Does Not Prevent Left Ventricular Dysfunction

Background Increased glucose transporter 1 (GLUT1) expression and glucose utilization that accompany pressure overload‐induced hypertrophy (POH) are believed to be cardioprotective. Moreover, it has been shown that lifelong transgenic overexpression of GLUT1 in the heart prevents cardiac dysfunction after aortic constriction. The relevance of this model to clinical practice is unclear because of the life‐long duration of increased glucose metabolism. Therefore, we sought to determine if a short‐term increase in GLUT1‐mediated myocardial glucose uptake would still confer cardioprotection if overexpression occurred at the onset of POH. Methods and Results Mice with cardiomyocyte‐specific inducible overexpression of a hemagglutinin (HA)‐tagged GLUT1 transgene (G1HA) and their controls (Cont) were subjected to transverse aortic constriction (TAC) 2 days after transgene induction with doxycycline (DOX). Analysis was performed 4 weeks after TAC. Mitochondrial function, adenosine triphosphate (ATP) synthesis, and mRNA expression of oxidative phosphorylation (OXPHOS) genes were reduced in Cont mice, but were maintained in concert with increased glucose utilization in G1HA following TAC. Despite attenuated adverse remodeling in G1HA relative to control TAC mice, cardiac hypertrophy was exacerbated in these mice, and positive dP/dt (in vivo) and cardiac power (ex vivo) were equivalently decreased in Cont and G1HA TAC mice compared to shams, consistent with left ventricular dysfunction. O‐GlcNAcylation of Ca2+ cycling proteins was increased in G1HA TAC hearts. Conclusions Short‐term cardiac specific induction of GLUT1 at the onset of POH preserves mitochondrial function and attenuates pathological remodeling, but exacerbates the hypertrophic phenotype and is insufficient to prevent POH‐induced cardiac contractile dysfunction, possibly due to impaired calcium cycling.

UCP3 Regulates Cardiac Efficiency and Mitochondrial Coupling in High Fat–Fed Mice but Not in Leptin-Deficient Mice

These studies investigate the role of uncoupling protein 3 (UCP3) in cardiac energy metabolism, cardiac O2 consumption (MVO2), cardiac efficiency (CE), and mitochondrial uncoupling in high fat (HF)–fed or leptin-deficient mice. UCP3KO and wild-type (WT) mice were fed normal chow or HF diets for 10 weeks. Substrate utilization rates, MVO2, CE, and mitochondrial uncoupling were measured in perfused working hearts and saponin-permeabilized cardiac fibers, respectively. Similar analyses were performed in hearts of ob/ob mice lacking UCP3 (U3OB mice). HF increased cardiac UCP3 protein. However, fatty acid (FA) oxidation rates were similarly increased by HF diet in WT and UCP3KO mice. By contrast, MVO2 increased in WT, but not in UCP3KO with HF, leading to increased CE in UCP3KO mice. Consistent with increased CE, mitochondrial coupling was increased in the hearts of HF-fed UCP3KO mice. Unexpectedly, UCP3 deletion in ob/ob mice reduced FA oxidation but had no effect on MVO2 or CE. In addition, FA-induced mitochondrial uncoupling was similarly enhanced in U3OB compared with ob/ob hearts and was associated with elevated mitochondrial thioesterase-1 protein content. These studies show that although UCP3 may mediate mitochondrial uncoupling and reduced CE after HF feeding, it does not mediate uncoupling in leptin-deficient states.

GLUT1 Deficiency in Cardiomyocytes Does not Accelerate the Transition from Compensated Hypertrophy to Heart Failure

The aim of this study was to determine whether endogenous GLUT1 induction and the increased glucose utilization that accompanies pressure overload hypertrophy (POH) are required to maintain cardiac function during hemodynamic stress, and to test the hypothesis that lack of GLUT1 will accelerate the transition to heart failure. To determine the contribution of endogenous GLUT1 to the cardiac adaptation to POH, male mice with cardiomyocyte-restricted deletion of the GLUT1 gene (G1KO) and their littermate controls (Cont) were subjected to transverse aortic constriction (TAC). GLUT1 deficiency reduced glycolysis and glucose oxidation by 50%, which was associated with a reciprocal increase in fatty acid oxidation (FAO) relative to controls. Four weeks after TAC, glycolysis increased and FAO decreased by 50% in controls, but were unchanged in G1KO hearts relative to shams. G1KO and controls exhibited equivalent degrees of cardiac hypertrophy, fibrosis, and capillary density loss after TAC. Following TAC, in vivo left ventricular developed pressure was decreased in G1KO hearts relative to controls, but+dP/dt was equivalently reduced in Cont and G1KO mice. Mitochondrial function was equivalently impaired following TAC in both Cont and G1KO hearts. GLUT1 deficiency in cardiomyocytes alters myocardial substrate utilization, but does not substantially exacerbate pressure-overload induced contractile dysfunction or accelerate the progression to heart failure.

Maintaining PGC-1α expression following pressure overload-induced cardiac hypertrophy preserves angiogenesis but not contractile or mitochondrial function

During pathological hypertrophy, peroxisome proliferator‐activated receptor coactivator 1α (PGC‐1α) is repressed in concert with reduced mitochondrial oxidative capacity and fatty acid oxidation (FAO). We therefore sought to determine if maintaining or increasing PGC‐1α levels in the context of pressure overload hypertrophy (POH) would preserve mitochondrial function and prevent contractile dysfunction. Pathological cardiac hypertrophy was induced using 4 wk of transverse aortic constriction (TAC) in mice overexpressing the human PGC‐1α genomic locus via a bacterial artificial chromosome (TG) and non‐transgenic controls (Cont). PGC‐1α levels were increased by 40% in TG mice and were sustained following TAC. Although TAC‐induced repression of FAO genes and oxidative phosphorylation (oxphos) genes was prevented in TG mice, mitochondrial function and ATP synthesis were equivalently impaired in Cont and TG mice after TAC. Contractile function was also equally impaired in Cont and TG mice following TAC, as demonstrated by decreased +dP/dt and ejection fraction and increased left ventricular developed pressure and end diastolic pressure. Conversely, capillary density was preserved, in concert with increased VEGF expression, while apoptosis and fibrosis were reduced in TG relative to Cont mice after TAC. Hence, sustaining physiological levels of PGC‐1α expression following POH, while preserving myocardial vascularity, does not prevent mitochondrial and contractile dysfunction.—Pereira, R. O., Wende, A. R., Crum, A., Hunter, D., Olsen, C. D., Rawlings, T., Riehle, C., Ward, W. F., Abel, E. D. Maintaining PGC‐1α expression following pressure overload‐induced cardiac hypertrophy preserves angiogenesis but not contractile or mitochondrial function. FASEB J. 28, 3691–3702 (2014). www.fasebj.org

Human erythropoietin gene delivery for cardiac remodeling of myocardial infarction in rats.

Considerable efforts have been made to exploit cardioprotective drugs and gene delivery systems for myocardial infarction (MI). The promising cardioprotective effects of recombinant human erythropoietin (rHuEPO) protein in animal experiments have not been consistently reproduced in clinical human trials of acute MI; however, the mechanisms underlying the inconsistent discrepancies are not yet fully understood. We hypothesized that the plasmid human erythropoietin gene (phEPO) delivered by our bioreducible polymer might produce cardioprotective effects on post-infarct cardiac remodeling. We demonstrated that intramyocardial delivery of phEPO by an arginine-grafted poly(disulfide amine) (ABP) polymer in infarcted rats preserves cardiac geometry and systolic function. The reduced infarct size of phEPO/ABP delivery was followed by decrease in fibrosis, protection from cardiomyocyte loss, and down-regulation of apoptotic activity. In addition, the increased angiogenesis and decreased myofibroblast density in the border zone of the infarct support the beneficial effects of phEPO/ABP administration. Furthermore, phEPO/ABP delivery induced prominent suppression on Ang II and TGF-β activity in all subdivisions of cardiac tissues except for the central zone of infarct. These results of phEPO gene therapy delivered by a bioreducible ABP polymer provide insight into the lack of phEPO gene therapy translation in the treatment of acute MI to human trials.

Antioxidant treatment normalizes mitochondrial energetics and myocardial insulin sensitivity independently of changes in systemic metabolic homeostasis in a mouse model of the metabolic syndrome

Cardiac dysfunction in obesity is associated with mitochondrial dysfunction, oxidative stress and altered insulin sensitivity. Whether oxidative stress directly contributes to myocardial insulin resistance remains to be determined. This study tested the hypothesis that ROS scavenging will improve mitochondrial function and insulin sensitivity in the hearts of rodent models with varying degrees of insulin resistance and hyperglycemia. The catalytic antioxidant MnTBAP was administered to the uncoupling protein-diphtheria toxin A (UCP-DTA) mouse model of insulin resistance (IR) and obesity, at early and late time points in the evolution of IR, and to db/db mice with severe obesity and type-two diabetes. Mitochondrial function was measured in saponin-permeabilized cardiac fibers. Aconitase activity and hydrogen peroxide emission were measured in isolated mitochondria. Insulin-stimulated glucose oxidation, glycolysis and fatty acid oxidation rates were measured in isolated working hearts, and 2-deoxyglucose uptake was measured in isolated cardiomyocytes. Four weeks of MnTBAP attenuated glucose intolerance in 13-week-old UCP-DTA mice but was without effect in 24-week-old UCP-DTA mice and in db/db mice. Despite the absence of improvement in the systemic metabolic milieu, MnTBAP reversed cardiac mitochondrial oxidative stress and improved mitochondrial bioenergetics by increasing ATP generation and reducing mitochondrial uncoupling in all models. MnTBAP also improved myocardial insulin mediated glucose metabolism in 13 and 24-week-old UCP-DTA mice. Pharmacological ROS scavenging improves myocardial energy metabolism and insulin responsiveness in obesity and type 2 diabetes via direct effects that might be independent of changes in systemic metabolism.