Anderson’s disease (AD) or chylomicron retention disease (CMRD) is a rare hereditary lipid malabsorption syndrome linked to SARA2 gene mutations. We report in this study a novel mutation in two ...sisters for which the Sar1b protein is predicted to be truncated by 32 amino acids at its carboxyl‐terminus. Because the SARA2 gene is also expressed in the muscle, heart, liver and placenta, extraintestinal clinical manifestations may exist. For the first time, we describe in this study in the two sisters muscular as well as cardiac abnormalities that could be related to the reported expression of SARA2 in these tissues. We also evaluated six other patients for potential manifestations of the SARA2 mutation. The creatine phosphokinase levels were increased in all patients 1.5–9.4 ×normal (N) and transaminases were moderately elevated in five of the eight patients (1.2–2.6 × N), probably related to muscle disease rather than to liver dysfunction. A decreased ejection fraction occurred in one patient (40%, N: 60%). The muscle, liver and placental tissues that were examined had no specific abnormalities and, in particular, no lipid accumulation. These results suggest that myolysis and other extraintestinal abnormalities can occur in AD/CMRD and that the clinical evaluation of patients should reflect this.
Summary
Exposure to fibrates leads to normalization of fatty acid oxidation (FAO) in fibroblasts from patients with myopathic forms of CPT2 deficiency or VLCAD deficiency. Correction of FAO is ...related to a drug‐induced increase of residual enzyme activity, and this could provide a new treatment strategy for these disorders.
The postnatal mammalian heart uses mitochondrial fatty acid oxidation (FAO) as the chief source of energy to meet the high energy demands necessary for pump function. Flux through the cardiac FAO ...pathway is tightly controlled in accordance with energy demands dictated by diverse physiologic and dietary conditions. In this report, we demonstrate that the lipid-activated nuclear receptor, peroxisome proliferator-activated receptor α (PPARα), regulates the expression of several key enzymes involved in cardiac mitochondrial FAO. In response to the metabolic stress imposed by pharmacologic inhibition of mitochondrial long-chain fatty acid import with etomoxir, PPARa serves as a molecular ‘lipostat’ factor by inducing the expression of target genes involved in fatty acid utilization including enzymes involved in mitochondrial and peroxisomal β-oxidation pathways. In mice lacking PPARα (PPARα-/- mice), etomoxir precipitates a cardiac phenotype characterized by myocyte lipid accumulation. Surprisingly, this metabolic regulatory response is influenced by gender as demonstrated by the observation that male PPARα-/- mice are more susceptible to the metabolic stress compared to female animals. These results identify an important role for PPARα in the control of cardiac lipid metabolism.
This study examines the ability of dietary lipids to regulate gene expression of mitochondrial and peroxisomal fatty acid beta-oxidation enzymes in the kidney cortex and medulla of 3-wk-old rats and ...evaluates the role of glucagon or of the alpha-isoform of peroxisome proliferator-activated receptor (PPARalpha) in mediating beta-oxidation enzyme gene regulation in the immature kidney. The long-chain (LCAD) and medium-chain acyl-CoA dehydrogenases (MCAD) and acyl-CoA oxidase (ACO) mRNA levels were found coordinately upregulated in renal cortex, but not in medulla, of pups weaned on a high-fat diet from day 16 to 21. Further results establish that switching pups from a low- to a high-fat diet for only 1 day was sufficient to induce large increases in cortical LCAD, MCAD, and ACO mRNA levels, and gavage experiments show that this upregulation of beta-oxidation gene expression is initiated within 6 h following lipid ingestion. Treatment of pups with clofibrate, a PPARalpha agonist, demonstrated that PPARalpha can mediate regulation of cortical beta-oxidation enzyme gene expression, whereas glucagon was found ineffective. Thus dietary lipids physiologically regulate gene expression of mitochondrial and peroxisomal beta-oxidation enzymes in the renal cortex of suckling pups, and this might involve PPARalpha-mediated mechanisms.
To explore the gene regulatory mechanisms involved in the metabolic control of cardiac fatty acid oxidative flux, the expression of muscle-type carnitine palmitoyltransferase I (M-CPT I) was ...characterized in primary cardiac myocytes in culture following exposure to the long-chain mono-unsaturated fatty acid, oleate. Oleate induced steady-state levels of M-CPT I mRNA 4.5-fold. The transcription of a plasmid construct containing the human M-CPT I gene promoter region fused to a luciferase gene reporter transfected into cardiac myocytes, was induced over 20-fold by long-chain fatty acid in a concentration-dependent and fatty acyl-chain length-specific manner. The M-CPT I gene promoter fatty acid response element (FARE-1) was localized to a hexameric repeat sequence located between 775 and 763 base pairs upstream of the initiator codon. Cotransfection experiments with expression vectors for the peroxisome proliferator-activated receptor α (PPARα) demonstrated that FARE-1 is a PPARα response element capable of conferring oleate-mediated transcriptional activation to homologous or heterologous promoters. Electrophoretic mobility shift assays demonstrated that PPARα bound FARE-1 with the retinoid X receptor α. The expression of M-CPT I in hearts of mice null for PPARα was approximately 50% lower than levels in wild-type controls. Moreover, a PPARα activator did not induce cardiac expression of the M-CPT I gene in the PPARα null mice. These results demonstrate that long-chain fatty acids regulate the transcription of a gene encoding a pivotal enzyme in the mitochondrial fatty acid uptake pathway in cardiac myocytes and define a role for PPARα in the control of myocardial lipid metabolism.
Aims Arrhythmogenic right ventricular cardiomyopathy (ARVC) is characterized by major fibro-fatty replacement of the right ventricle (RV). We hypothesized that changes in peroxisome ...proliferator-activated receptor (PPAR) signalling contributed to myocardium fatty accumulation and contractile dysfunction in ARVC. Methods and results Real-time quantitative reverse transcriptase–polymerase chain reaction and western blotting were used to assess cardiac expression of PPARα and γ and two of their downstream target genes—medium-chain acyl-CoA dehydrogenase (MCAD) and phosphoenolpyruvate carboxykinase (PEPCK)—in both RV and left ventricle (LV) from five controls and five ARVC patients. In vitro motility assays were used to analyse functional properties of myosin. In the RV, sliding velocity was nearly two-fold lower in ARVC than in controls, whereas a 10% reduction in velocity values was noted between ARVC and non-failing myocardium in the LV. In controls, PPARα and MCAD mRNA and protein levels were higher in the RV compared with the LV. In ARVC, the expression of PPARα and MCAD mRNA and/or proteins was decreased in both RV and LV. RV from ARVC was also characterized by a dramatic activation of the PPARγ pathway, as attested by the increase in PPARγ mRNA and protein (500 and 270%, respectively, each P < 0.001) and by the induction of PEPCK gene. In contrast, the LV of ARVC heart exhibited no changes in the expression of the PPARγ regulatory pathway compared with control. Conclusion ARVC is associated with major disturbances in the PPARα and PPARγ signalling pathway in the RV that may contribute to intracellular lipid overload and severe myosin dysfunction.
Cystic fibrosis (CF), a multisystem disease caused by CFTR (cystic fibrosis transmembrane conductance regulator) gene mutations, is associated with an abnormal inflammatory response and compromised ...redox homeostasis in the airways. Recent evidence suggests that dysfunctional CFTR leads to redox imbalance and to mitochondrial reduced glutathione (mtGSH) depletion in CF models. This study was designed to investigate the consequences of mtGSH depletion on mitochondrial function and inflammatory response. mtGSH depletion was confirmed in colonic epithelium of CFTR-null mice and in CFTR-mutated human epithelial cells. GSH uptake experiments performed on isolated mitochondria suggest that mtGSH depletion is not due to a defective GSH transport capacity by CF mitochondria, despite the decreased expression of two mtGSH carriers, oxoglutarate carrier and dicarboxylate carrier. CM-H2DCFDA 5 (and 6)-chloromethyl-2′,7′-dichlorodihydrofluorescein diacetate, acetyl ester fluorescence and aconitase activity showed an increase in reactive oxygen species levels in CFTR-defective cells and a pro-oxidative environment within CF mitochondria. The activities of respiratory chain complexes were further examined. Results showed a selective loss of Complex I (CI) function in CF models associated with an altered mitochondrial membrane potential (Δψ
m). CI analysis showed normal expression but an overoxidation of its NADH-ubiquinone oxidoreductase Fe-S protein 1 subunit. GSH monoethyl ester (GSH-EE) significantly enhanced mtGSH levels in the IB3-1/C38 model and reversed CI inhibition, suggesting that mtGSH depletion is responsible for the loss of CI activity. Furthermore, GSH-EE attenuated Δψ
m depolarization and restored normal IL-8 secretion by CFTR-defective cells. These studies provide evidence for a critical role of a mtGSH defect in mitochondrial dysfunction and abnormal IL-8 secretion in CF cells and reveal the therapeutic potential of mitochondria-targeted antioxidants in CF.
Type 2 carnitine palmitoyl transferase (CPT2) is involved in the transfer of long-chain fatty acid into the mitochondria. CPT2-deficient patients carry gene mutations associated with different ...clinical presentations, correlating with various levels of fatty acid oxidation (FAO) and residual CPT2 enzyme activity. We tested the hypothesis that pharmacological stimulation of peroxisome proliferator-activated receptors (PPAR) can stimulate FAO in CPT2-deficient muscle cells. Accordingly, we show that a 48-h treatment of CPT2-deficient myoblasts by bezafibrate restored FAO in patient cells. Specific agonists of PPARδ (GWδ 0742), and, to a lower extent, PPARα (GWα 7647) also stimulated FAO in control myoblasts. However, when tested in CPT2-deficient myoblasts, only the δ-agonist was able to restore FAO, whereas the α-agonist had no effect. GWδ 0742 increased CPT2 mRNA levels, whereas no change in CPT2 transcripts was found in response to GWα 7647. Bezafibrate and GWδ 0742 increased residual CPT2 activity and normalized long-chain acylcarnitine production by deficient cells. Finally, CPT1-B mRNA was also stimulated after PPAR agonist treatment, and this likely takes part in drug-induced increase of FAO in control muscle cells. In conclusion, this study clearly suggests that PPARs could be therapeutic targets for correction of inborn β-oxidation defects in human muscle. Furthermore, these data also illustrate a selective control of β-oxidation enzyme gene expression by PPARδ, with no contribution of PPARα.
We have previously shown that the activity of several mitochondrial oxidative enzymes increased greatly in the medullary thick ascending limb of Henle's loop (MTAL) of developing rat kidney between ...16 days after birth and the adult stage. These changes were triggered by the postnatal rise in circulating glucocorticoids. To determine whether these increases are related to a mitochondrial biogenesis we have studied the changes in mitochondrial density of MTAL cells and mitochondrial DNA content in the inner stripe of the outer medulla during the postnatal period. The activities of respiratory chain (RC) complexes II and IV, located in the inner mitochondrial membrane (IMM), were also assayed, and developmental changes in the ultrastructure of the IMM were quantified. Quantitative electron-microscopic data showed a doubling in mitochondrial density from day 16 to the adult, accompanied by twofold increase in mitochondrial DNA, as determined by slot-blot experiments. The surface density of IMM increased by 77%, and there was a concomitant large rise in the activity of both RC enzymes. A possible role of glucocorticoids on the regulation of mitochondrial biogenesis was examined in similar experiments performed in adrenalectomized rat pups. The data demonstrated that glucocorticoids are essential for the rise in RC enzymes and the development of IMM but do not regulate postnatal changes in mitochondrial density and mitochondrial DNA content. Finally, ontogenic changes in oxidative capacities of MTAL cells could be a critical factor in the development of kidney urine concentrating mechanisms in weanling rats.
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