Michael Courtois

PGC-1α deficiency causes multi-system energy metabolic derangements: Muscle dysfunction, abnormal weight control and hepatic steatosis

The gene encoding the transcriptional coactivator peroxisome proliferator-activated receptor-γ coactivator-1α (PGC-1α) was targeted in mice. PGC-1α null (PGC-1α−/−) mice were viable. However, extensive phenotyping revealed multi-system abnormalities indicative of an abnormal energy metabolic phenotype. The postnatal growth of heart and slow-twitch skeletal muscle, organs with high mitochondrial energy demands, is blunted in PGC-1α−/− mice. With age, the PGC-1α−/− mice develop abnormally increased body fat, a phenotype that is more severe in females. Mitochondrial number and respiratory capacity is diminished in slow-twitch skeletal muscle of PGC-1α−/− mice, leading to reduced muscle performance and exercise capacity. PGC-1α−/− mice exhibit a modest diminution in cardiac function related largely to abnormal control of heart rate. The PGC-1α−/− mice were unable to maintain core body temperature following exposure to cold, consistent with an altered thermogenic response. Following short-term starvation, PGC-1α−/− mice develop hepatic steatosis due to a combination of reduced mitochondrial respiratory capacity and an increased expression of lipogenic genes. Surprisingly, PGC-1α−/− mice were less susceptible to diet-induced insulin resistance than wild-type controls. Lastly, vacuolar lesions were detected in the central nervous system of PGC-1α−/− mice. These results demonstrate that PGC-1α is necessary for appropriate adaptation to the metabolic and physiologic stressors of postnatal life.

A critical role for PPARα-mediated lipotoxicity in the pathogenesis of diabetic cardiomyopathy: Modulation by dietary fat content

To explore the role of peroxisome proliferator-activated receptor α (PPARα)-mediated derangements in myocardial metabolism in the pathogenesis of diabetic cardiomyopathy, insulinopenic mice with PPARα deficiency (PPARα−/−) or cardiac-restricted overexpression [myosin heavy chain (MHC)-PPAR] were characterized. Whereas PPARα−/− mice were protected from the development of diabetes-induced cardiac hypertrophy, the combination of diabetes and the MHC-PPAR genotype resulted in a more severe cardiomyopathic phenotype than either did alone. Cardiomyopathy in diabetic MHC-PPAR mice was accompanied by myocardial long-chain triglyceride accumulation. The cardiomyopathic phenotype was exacerbated in MHC-PPAR mice fed a diet enriched in triglyceride containing long-chain fatty acid, an effect that was reversed by discontinuing the high-fat diet and absent in mice given a medium-chain triglyceride-enriched diet. Reactive oxygen intermediates were identified as candidate mediators of cardiomyopathic effects in MHC-PPAR mice. These results link dysregulation of the PPARα gene regulatory pathway to cardiac dysfunction in the diabetic and provide a rationale for serum lipid-lowering strategies in the treatment of diabetic cardiomyopathy.

Akt1 Is Required for Physiological Cardiac Growth

Background— Postnatal growth of the heart chiefly involves nonproliferative cardiomyocyte enlargement. Cardiac hypertrophy exists in a “physiological” form that is an adaptive response to long-term exercise training and as a “pathological” form that often is a maladaptive response to provocative stimuli such as hypertension and aortic valvular stenosis. A signaling cascade that includes the protein kinase Akt regulates the growth and survival of many cell types, but the precise role of Akt1 in either form of cardiac hypertrophy is unknown. Methods and Results— To evaluate the role of Akt1 in physiological cardiac growth, akt1−/− adult murine cardiac myocytes (AMCMs) were treated with IGF-1, and akt1−/− mice were subjected to exercise training. akt1−/− AMCMs were resistant to insulin-like growth factor-1–stimulated protein synthesis. The akt1−/− mice were found to be resistant to swimming training–induced cardiac hypertrophy. To evaluate the role of Akt in pathological cardiac growth, akt1−/− AMCMs were treated with endothelin-1, and akt1−/− mice were subjected to pressure overload by transverse aortic constriction. Surprisingly, akt1−/− AMCMs were sensitized to endothelin-1–induced protein synthesis, and akt1−/− mice developed an exacerbated form of cardiac hypertrophy in response to transverse aortic constriction. Conclusions— These results establish Akt1 as a pivotal regulatory switch that promotes physiological cardiac hypertrophy while antagonizing pathological hypertrophy.

Transgenic Expression of Fatty Acid Transport Protein 1 in the Heart Causes Lipotoxic Cardiomyopathy

Evidence is emerging that systemic metabolic disturbances contribute to cardiac myocyte dysfunction and clinically apparent heart failure, independent of associated coronary artery disease. To test the hypothesis that perturbation of lipid homeostasis in cardiomyocytes contributes to cardiac dysfunction, we engineered transgenic mice with cardiac-specific overexpression of fatty acid transport protein 1 (FATP1) using the &agr;-myosin heavy chain gene promoter. Two independent transgenic lines demonstrate 4-fold increased myocardial free fatty acid (FFA) uptake that is consistent with the known function of FATP1. Increased FFA uptake in this model likely contributes to early cardiomyocyte FFA accumulation (2-fold increased) and subsequent increased cardiac FFA metabolism (2-fold). By 3 months of age, transgenic mice have echocardiographic evidence of impaired left ventricular filling and biatrial enlargement, but preserved systolic function. Doppler tissue imaging and hemodynamic studies confirm that these mice have predominantly diastolic dysfunction. Furthermore, ambulatory ECG monitoring reveals prolonged QTc intervals, reflecting reductions in the densities of repolarizing, voltage-gated K+ currents in ventricular myocytes. Our results show that in the absence of systemic metabolic disturbances, such as diabetes or hyperlipidemia, perturbation of cardiomyocyte lipid homeostasis leads to cardiac dysfunction with pathophysiological findings similar to those in diabetic cardiomyopathy. Moreover, the MHC-FATP model supports a role for FATPs in FFA import into the heart in vivo.

Transmitral pressure-flow velocity relation. Importance of regional pressure gradients in the left ventricle during diastole.

Effects of regional diastolic pressure differences within the left ventricle on the measured transmitral pressure-flow relation were determined by simultaneous micromanometric left atrial (LAP) and left ventricular pressure (LVP) measurements, and Doppler echocardiograms in 11 anesthetized, closed-chest dogs. Intraventricular pressure recordings at sites that were 2, 4, and 6 cm from the apex were obtained. Profound differences between these sites were noted in the transmitral pressure relation during early (preatrial) diastolic filling. In measurements from apex to base, minimum LVP increased (1.6 +/- 0.7 to 3.1 +/- 0.8 mm Hg, mean +/- SD); the time interval between the first crossover of transmitral pressures and minimum LVP increased (31 +/- 3 to 50 +/- 17 msec); the slope of the rapid-filling LVP wave decreased (74 +/- 13 to 26 +/- 5 mm Hg/sec); the maximum forward (i.e., LAP greater than LVP) transmitral pressure gradient decreased (3.6 +/- 1.3 to 2.1 +/- 0.7 mm Hg); the time interval between the first and second points of transmitral pressure crossover increased (71 +/- 9 to 96 +/- 13 msec); and the area of reversed (i.e., LVP greater than LAP) gradient between the second and third points of transmitral pressure crossover decreased (101 +/- 41 to 40 +/- 33 mm Hg.msec). During atrial contraction, significant regional ventricular apex-to-base gradients were also noted. The slope of the LV A wave decreased (26 +/- 10 to 16 +/- 4 mm Hg/sec); LV end-diastolic pressure decreased (8.1 +/- 2.0 to 7.4 +/- 2.0 mm Hg), and the upstroke of the LV A wave near the base was recorded earlier than near the apex. All differences were significant at the 0.05 level. Simultaneous transmitral Doppler velocity profiles and transmitral pressures were measured at the 4-cm intraventricular site. The average interval between the first and second points of pressure crossover and between the onset of early rapid filling and maximum E-wave velocity were statistically similar (81 +/- 13 vs. 85 +/- 12 msec; NS); and the average area of the forward transmitral pressure gradient associated with acceleration of early flow was significantly greater than the area of reversed gradient associated with deceleration of early flow (133 +/- 36 vs. 80 +/- 46 msec.mm Hg; p less than 0.025).(ABSTRACT TRUNCATED AT 400 WORDS)

Cardiac-Specific Induction of the Transcriptional Coactivator Peroxisome Proliferator-Activated Receptor γ Coactivator-1α Promotes Mitochondrial Biogenesis and Reversible Cardiomyopathy in a Developmental Stage-Dependent Manner

Abstract— Recent evidence has identified the peroxisome proliferator-activated receptor &ggr; coactivator-1&agr; (PGC-1&agr;) as a regulator of cardiac energy metabolism and mitochondrial biogenesis. We describe the development of a transgenic system that permits inducible, cardiac-specific overexpression of PGC-1&agr;. Expression of the PGC-1&agr; transgene in this system (tet-on PGC-1&agr;) is cardiac-specific in the presence of doxycycline (dox) and is not leaky in the absence of dox. Overexpression of PGC-1&agr; in tet-on PGC-1&agr; mice during the neonatal stages leads to a dramatic increase in cardiac mitochondrial number and size coincident with upregulation of gene markers associated with mitochondrial biogenesis. In contrast, overexpression of PGC-1&agr; in the hearts of adult mice leads to a modest increase in mitochondrial number, derangements of mitochondrial ultrastructure, and development of cardiomyopathy. The cardiomyopathy in adult tet-on PGC-1&agr; mice is characterized by an increase in ventricular mass and chamber dilatation. Surprisingly, removal of dox and cessation of PGC-1&agr; overexpression in adult mice results in complete reversal of cardiac dysfunction within 4 weeks. These results indicate that PGC-1&agr; drives mitochondrial biogenesis in a developmental stage-dependent manner permissive during the neonatal period. This unique murine model should prove useful for the study of the molecular regulatory programs governing mitochondrial biogenesis and characterization of the relationship between mitochondrial dysfunction and cardiomyopathy and as a general model of inducible, reversible cardiomyopathy.

Nuclear receptors PPARβ/δ and PPARα direct distinct metabolic regulatory programs in the mouse heart

In the diabetic heart, chronic activation of the PPARalpha pathway drives excessive fatty acid (FA) oxidation, lipid accumulation, reduced glucose utilization, and cardiomyopathy. The related nuclear receptor, PPARbeta/delta, is also highly expressed in the heart, yet its function has not been fully delineated. To address its role in myocardial metabolism, we generated transgenic mice with cardiac-specific expression of PPARbeta/delta, driven by the myosin heavy chain (MHC-PPARbeta/delta mice). In striking contrast to MHC-PPARalpha mice, MHC-PPARbeta/delta mice had increased myocardial glucose utilization, did not accumulate myocardial lipid, and had normal cardiac function. Consistent with these observed metabolic phenotypes, we found that expression of genes involved in cellular FA transport were activated by PPARalpha but not by PPARbeta/delta. Conversely, cardiac glucose transport and glycolytic genes were activated in MHC-PPARbeta/delta mice, but repressed in MHC-PPARalpha mice. In reporter assays, we showed that PPARbeta/delta and PPARalpha exerted differential transcriptional control of the GLUT4 promoter, which may explain the observed isotype-specific effects on glucose uptake. Furthermore, myocardial injury due to ischemia/reperfusion injury was significantly reduced in the MHC-PPARbeta/delta mice compared with control or MHC-PPARalpha mice, consistent with an increased capacity for myocardial glucose utilization. These results demonstrate that PPARalpha and PPARbeta/delta drive distinct cardiac metabolic regulatory programs and identify PPARbeta/delta as a potential target for metabolic modulation therapy aimed at cardiac dysfunction caused by diabetes and ischemia.

CD36 Deficiency Rescues Lipotoxic Cardiomyopathy

Obesity-related diabetes mellitus leads to increased myocardial uptake of fatty acids (FAs), resulting in a form of cardiac dysfunction referred to as lipotoxic cardiomyopathy. We have shown previously that chronic activation of the FA-activated nuclear receptor, peroxisome proliferator-activated receptor &agr; (PPAR&agr;), is sufficient to drive the metabolic and functional abnormalities of the diabetic heart. Mice with cardiac-restricted overexpression of PPAR&agr; (myosin heavy chain [MHC]-PPAR&agr;) exhibit myocyte lipid accumulation and cardiac dysfunction. We sought to define the role of the long-chain FA transporter CD36 in the pathophysiology of lipotoxic forms of cardiomyopathy. MHC-PPAR&agr; mice were crossed with CD36-deficient mice (MHC-PPAR&agr;/CD36−/− mice). The absence of CD36 prevented myocyte triacylglyceride accumulation and cardiac dysfunction in the MHC-PPAR&agr; mice under basal conditions and following administration of high-fat diet. Surprisingly, the rescue of the MHC-PPAR&agr; phenotype by CD36 deficiency was associated with increased glucose uptake and oxidation rather than changes in FA utilization. As predicted by the metabolic changes, the activation of PPAR&agr; target genes involved in myocardial FA-oxidation pathways in the hearts of the MHC-PPAR&agr; mice was unchanged in the CD36-deficient background. However, PPAR&agr;-mediated suppression of genes involved in glucose uptake and oxidation was reversed in the MHC-PPAR&agr;/ CD36−/− mice. We conclude that CD36 is necessary for the development of lipotoxic cardiomyopathy in MHC-PPAR&agr; mice and that novel therapeutic strategies aimed at reducing CD36-mediated FA uptake show promise for the prevention or treatment of cardiac dysfunction related to obesity and diabetes.

The transmitral pressure-flow velocity relation. Effect of abrupt preload reduction.

Although recent animal and clinical studies suggest that Doppler-derived indexes may be useful for the characterization of ventricular diastolic behavior, the hemodynamic basis for the preload dependency of these indexes has not previously been fully elucidated. Accordingly, effects of reduction of left atrial load on the pressure-flow velocity relation were characterized in 10 anesthetized, closed-chest dogs during transient inferior vena caval occlusion by means of simultaneously recorded left atrial and left ventricular micromanometric pressure measurement and transesophageal Doppler echocardiograms. Within four or five beats after inferior vena caval balloon occlusion, left atrial loading was reduced as evidenced by a decrease in the slope of the left atrial v wave from 21 +/- 4 to 13 +/- 4 mm Hg/sec (p less than 0.001) and by a decrease in the first crossover point of left atrial and left ventricular pressures from 5.6 +/- 1.1 to 2.9 +/- 1.5 mm Hg (p less than 0.001). This decrease in left atrial loading resulted in reductions during early diastole of minimum left ventricular pressure (from 1.0 +/- 0.8 to -0.4 +/- 1.2 mm Hg, p less than 0.001), the maximum early forward (i.e., left atrial pressure greater than left ventricular pressure) transmitral pressure gradient (from 2.8 +/- 0.8 to 2.4 +/- 0.5 mm Hg, p less than 0.01); the slope of the rapid filling pressure wave (from 44 +/- 11 to 38 +/- 10 mm Hg/sec, p less than 0.025); and the area of the reversed (i.e., left ventricular pressure greater than left atrial pressure) transmitral pressure gradient (from 79 +/- 42 to 53 +/- 33 mm Hg.msec, p less than 0.05). During late diastole, both the heights and slopes of the left atrial and left ventricular a waves fell, resulting in a decrease in the maximum late transmitral pressure gradient (from 1.2 +/- 0.7 to 0.9 +/- 0.5 mm Hg, p less than 0.05). Vena caval occlusion also altered Doppler transmitral velocity profiles during both the early and late phases of diastole. Peak velocity of the E wave decreased (from 50 +/- 11 to 41 +/- 7 cm/sec, p less than 0.01) as did acceleration (from 880 +/- 222 to 757 +/- 258 cm/sec2, p less than 0.025) and deceleration (from 597 +/- 260 to 429 +/- 197 cm/sec2, p less than 0.025). Peak velocity of the A wave also fell (from 29 +/- 9 to 22 +/- 5 cm/sec, p less than 0.005). Abrupt inferior vena caval occlusion did not significantly change heart rate or mean aortic pressure.(ABSTRACT TRUNCATED AT 400 WORDS)

Akt2 Regulates Cardiac Metabolism and Cardiomyocyte Survival

The Akt family of serine-threonine kinases participates in diverse cellular processes, including the promotion of cell survival, glucose metabolism, and cellular protein synthesis. All three known Akt family members, Akt1, Akt2 and Akt3, are expressed in the myocardium, although Akt1 and Akt2 are most abundant. Previous studies demonstrated that Akt1 and Akt3 overexpression results in enhanced myocardial size and function. Yet, little is known about the role of Akt2 in modulating cardiac metabolism, survival, and growth. Here, we utilize murine models with targeted disruption of the akt2 or the akt1 genes to demonstrate that Akt2, but not Akt1, is required for insulin-stimulated 2-[3H]deoxyglucose uptake and metabolism. In contrast, akt2-/- mice displayed normal cardiac growth responses to provocative stimulation, including ligand stimulation of cultured cardiomyocytes, pressure overload by transverse aortic constriction, and myocardial infarction. However, akt2-/- mice were found to be sensitized to cardiomyocyte apoptosis in response to ischemic injury, and apoptosis was significantly increased in the peri-infarct zone of akt2-/- hearts 7 days after occlusion of the left coronary artery. These results implicate Akt2 in the regulation of cardiomyocyte metabolism and survival.