Déficit de 3-hidroxiacil-CoA-deshidrogenasa de cadena larga: a propósito de un caso / Long-chain 3-hydroxyacyl-CoA dehydrogenase deficiency: A case report

CASO CLÍNICO: Paciente de 5 años de edad remitido para valoración oftalmológica con el diagnóstico de déficit de 3-hidroxiacil-CoA deshidrogenasa de cadena larga. Presentaba como antecedente la aparición de crisis metabólicas agudas precipitadas por infecciones banales y rabdomiólisis. El examen oftalmoscópico reveló una atrofia coriorretiniana peripapilar y una maculopatía granular difusa. La agudeza visual era de 6/6 en ambos ojos y las pruebas electrofisiológicas normales. DISCUSIÓN: Se realiza una revisión de la bibliografía y los conocimientos recientes de esta enfermedad mediante la descripción de un caso clínico documentando los hallazgos obtenidos mediante autofluorescencia y tomografía de coherencia óptica para mejorar el conocimiento existente sobre ella

CLINICAL CASE: A five-year-old patient, with a diagnosis of long-chain 3-hydroxyacyl-CoA dehydrogenase deficiency, was referred for an ophthalmological examination. He had a history of acute metabolic crises precipitated by intercurrent infections,as well as rhabdomyolysis. The fundoscopic examination revealed a peripapillary chorioretinal atrophy and a diffuse granular appearance of the macular retinal pigment epithelium. Best corrected visual acuity was 6/6 in both eyes, and he had a normal electroretinography test. DISCUSSION: We perform a review of the literature and recent findings in relation to this disease through the description of a clinical case in order to improve the knowledge of this uncommon disorder

The yeast eukaryotic translation initiation factor 2B translation initiation complex interacts with the fatty acid synthesis enzyme YBR159W and endoplasmic reticulum membranes.

Using affinity purifications coupled with mass spectrometry and yeast two-hybrid assays, we show the Saccharomyces cerevisiae translation initiation factor complex eukaryotic translation initiation factor 2B (eIF2B) and the very-long-chain fatty acid (VLCFA) synthesis keto-reductase enzyme YBR159W physically interact. The data show that the interaction is specifically between YBR159W and eIF2B and not between other members of the translation initiation or VLCFA pathways. A ybr159wΔ null strain has a slow-growth phenotype and a reduced translation rate but a normal GCN4 response to amino acid starvation. Although YBR159W localizes to the endoplasmic reticulum membrane, subcellular fractionation experiments show that a fraction of eIF2B cofractionates with lipid membranes in a YBR159W-independent manner. We show that a ybr159wΔ yeast strain and other strains with null mutations in the VLCFA pathway cause eIF2B to appear as numerous foci throughout the cytoplasm.

Dihydroartemisinin-induced inhibition of proliferation in BEL-7402 cells: an analysis of the mitochondrial proteome.

Artemisinin, the active ingredient of the Chinese medicinal herb Artemisia annua L., and its derivatives (ARTs) are currently widely used as anti-malarial drugs around the world. In this study, we found that dihydroartemisinin (DHA), one of the main active metabolites of ARTs, inhibited the proliferation of human hepatocarcinoma BEL-7402 cells in a concentration-dependent manner. To interpret the mechanisms involved, an analysis of the mitochondrial proteome was performed employing two-dimensional gel electrophoresis and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Seven mitochondrial proteins including fumarate hydratase, 60 kDa heat shock protein, enoyl-CoA hydratase, 3-hydroxyacyl-CoA dehydrogenase, two subunits of ATP synthase and NADPH:adrenodoxin oxidoreductase were identified to be differentially expressed between the control and DHA-treated groups. Our results indicate that the imbalance of energy metabolism induced by DHA may contribute, at least in part, to its anti-cancer potential in BEL-7402 cells.

Dietary soya protein intake and exercise training have an additive effect on skeletal muscle fatty acid oxidation enzyme activities and mRNA levels in rats.

Exercise training and regular physical activity increase oxidation of fat. Enhanced oxidation of fat is important for preventing lifestyle diseases such as hypertension and obesity. The aim of the present study in rats was to determine whether intake of dietary soya protein and exercise training have an additive effect on the activity and mRNA expression of enzymes involved in skeletal muscle fatty acid oxidation. Male Sprague-Dawley rats (n 32) were assigned randomly into four groups (eight rats per group) and then divided further into sedentary or exercise-trained groups fed either casein or soya protein diets. Rats in the exercise groups were trained for 2 weeks by swimming for 120 min/d, 6 d/week. Exercise training decreased hepatic triacylglycerol levels and retroperitoneal adipose tissue weight and increased skeletal muscle carnitine palmitoyltransferase 1 (CPT1) activity and mRNA expression of CPT1, beta-hydroxyacyl-CoA dehydrogenase (HAD), acyl-CoA oxidase, PPARgamma coactivator 1alpha (PGC1alpha) and PPARalpha. Soya protein significantly decreased hepatic triacylglycerol levels and epididymal adipose tissue weight and increased skeletal muscle CPT1 activity and CPT1, HAD, acyl-CoA oxidase, medium-chain acyl-CoA dehydrogenase, PGC1alpha and PPARalpha mRNA levels. Furthermore, skeletal muscle HAD activity was the highest in exercise-trained rats fed soya protein. We conclude that exercise training and soya protein intake have an important additive role on induction of PPAR pathways, leading to increased activity and mRNA expression of enzymes involved in fatty acid oxidation in skeletal muscle and reduced accumulation of body fat.

Different susceptibility to peroxisome proliferator-induced hepatocarcinogenesis in rats with polymorphic glutathione transferase genes.

Although peroxisomal bifunctional enzyme (enoyl-CoA hydratase/L-3-hydroxyacyl-CoA dehydrogenase; BE) is a positive marker for peroxisome proliferation, it is completely absent or expressed very weakly in rat hepatic preneoplastic and neoplastic lesions induced by peroxisome proliferators (PP). After administration of PP for 8-15 weeks, some rats exhibit BE-negative preneoplastic foci but other rats do not. In the present study, to investigate the involvement of glutathione S-transferase (GST) M1 gene polymorphism in interindividual differences in susceptibility to PP, we developed a method to determine the genotypes of rats. We then examined whether rats with one type encoding 198Asn-199Cys (NC-type) or another encoding 198Lys-199Ser (KS-type) exhibit differences in clofibrate (CF) susceptibility. After administration of 0.3% CF for 6 weeks or more, BE-negative foci were found immunohistochemically in KS/KS-type rats, but not in NC/NC-type rats. The number of BE-negative foci in KS/KS rats was 15.3 +/- 9.0 foci/cm2 of liver section after 6 weeks of CF administration, and the values did not alter thereafter. The mean areas of BE-negative foci in KS/KS rat livers increased during the period from 6 to 60 weeks. At weeks 30 and 60, almost all BE-negative foci exhibited a clear cell phenotype, a type of preneoplastic hepatic lesion. BE-negative foci were devoid of peroxisome proliferator-activated receptor alpha, whereas surrounding tissues were positive for the receptor. These results indicate that rats that are polymorphic for the GST M1 gene exhibit different susceptibilities to CF in vivo.

Reye-like syndrome resulting from novel missense mutations in mitochondrial medium- and short-chain l-3-hydroxy-acyl-CoA dehydrogenase.

Medium- and short-chain l-3-hydroxy-acyl-CoA dehydrogenase (M/SCHAD) deficiency is a recessively inherited disorder of fatty acid oxidation. Currently, only four patients from three families have been reported in the literature. All these patients presented with hypoglycemia associated with hyperinsulinism (HI). This association suggests that there is a role for M/SCHAD in regulating the pancreatic secretion of insulin. We present a fifth patient whose presentation was similar to Reye syndrome, a feature in common with most of the previously recognized disorders of fatty acid oxidation but with no clinical evidence of HI. Sequencing of the HAD1 gene on chromosome 4 revealed compound heterozygosity for two novel missense mutations, 170A>G, resulting in D45G, and 676T>C, resulting in Y214H. The mutant enzymes were expressed and subjected to kinetic analysis. Y214H has no detectable activity, whilst D45G, which resides in the cofactor-binding pocket, has an altered K(m) for NADH (96 microM versus 24 microM for the wild-type). This represents the first kinetic M/SCHAD mutant, which explains the high residual activity in skin fibroblasts. The lack of obvious HI in this patient may be related to the high residual activity and indicates that HI associated with M/SCHAD deficiency may only be present with complete deficiency. The spectrum of M/SCHAD phenotype should be broadened to include acute liver disease.

Metabolic capacity of the diaphragm in patients with COPD.

Chronic obstructive pulmonary disease (COPD) is associated with an increased load on the diaphragm. Chronic loading on skeletal muscles results in metabolic changes and fiber-type shifts. Therefore, we investigated whether the load on the human diaphragm imposed by COPD altered oxidative enzyme activity, glycogenolytic enzyme activity and mitochondrial energy generating capacity and efficiency. Biopsies of the diaphragm from COPD patients and control subjects were obtained and activities of L(+)3-hydroxyacylCoA-dehydrogenase (HADH, marker for beta-oxidation capacity) and phosphorylase (marker for glycogenolytic capacity) were measured spectrophotometrically. Mitochondrial energy generating capacity was measured by spectrophotometrical and radiochemical methods. Fiber-type distribution was determined electrophoretically. We found that HADH activity was increased with increasing severity of COPD (P=0.05). No change in glycogenolytic enzyme activity was observed. The activity of the mitochondrial respiratory chain complexes III and IV and oxidation of pyruvate was increased with increasing airflow obstruction. These results suggest that in COPD the diaphragm adapts to a higher workload by increasing the oxidative capacity and mitochondrial function.

Different forms of obesity as a function of diet composition.

OBJECTIVE: To characterize the phenotype of obesity on a high-carbohydrate diet (HCD) as compared to a high-fat diet (HFD) or moderate-fat diet (MFD). METHODS AND PROCEDURES: In four experiments, adult Sprague-Dawley rats (275-300 g) were maintained for several weeks on a: (1) HFD with 50% fat; (2) balanced MFD with 25% fat; or (3) HCD with 10% fat/65% carbohydrate. Then, based on the amount of body fat accumulated in four dissected fat pads, the animals were subgrouped as lean (lowest tertile) or obese (highest tertile) and characterized with multiple measures. RESULTS: The obese rats of these diet groups, with 70-80% greater body fat than the lean animals, exhibited elevated levels of leptin and insulin and increased activity of lipoprotein lipase in adipose tissue (aLPL), with no change in muscle LPL. Characteristics common to the obese rats on the HFD or MFD, but not seen on the HCD, were hyperphagia, elevated circulating levels of triglycerides (TG), nonesterified fatty acids (NEFA) and glucose, and a significant increase in beta-hydroxyacyl-CoA dehydrogenase (HADH) activity in muscle, reflecting its greater capacity to metabolize fat. This was accompanied by a significant increase in expression of the peptide, galanin (GAL), in the paraventricular nucleus (PVN), as measured by in situ hybridization and real-time quantitative PCR, and also in GAL peptide immunoreactivity. These measures of GAL were consistently, positively correlated with circulating TG levels and also with HADH activity in muscle. In contrast to these fat-associated changes, rats that became obese on an HCD maintained normal caloric intake and levels of TG, NEFA, and glucose. They also showed no change in PVN GAL mRNA or peptide. Instead, they exhibited a significant reduction in HADH activity compared to the lean animals, along with increased activity of phosphofructokinase in muscle, a key enzyme in glycolysis. CONCLUSION: Specific characteristics of obesity, including expression of hypothalamic peptides, are dependent upon diet composition. Whereas obesity on an HFD is associated with hyperphagia and elevated lipids, fat metabolism in muscle, and fat-stimulated peptides such as GAL, obesity on an HCD with a similar increase in body fat shows none of these characteristics and instead exhibits a metabolic pattern in muscle that favors carbohydrate over fat oxidation. These results suggest the existence of multiple forms of obesity with different underlying mechanisms that are diet dependent.

Determination of beta-hydroxyacyl CoA-dehydrogenase activity in meat by electrophoretically mediated microanalysis.

The combined use of an in-tube enzyme assay and capillary electrophoresis for determining beta-hydroxyacyl CoA-dehydrogenase (beta-HADH) activity in meat was investigated. Beta-HADH is a significant mitochondrial enzyme in food muscle; thus, the determination of its activity is important in food analysis. The enzymatic assay and the separation of the reaction products were carried out by electrophoretically mediated microanalysis (EMMA) using a plug-plug reaction mode at variable potential. For the quantification of beta-HADH activity, the rate of conversion of reduced beta-nicotinamide adenine dinucleotide (NADH) to beta-nicotinamide adenine dinucleotide (NAD+) was calculated by determining NAD+ at 260 nm. A calibration curve for NAD+ concentration versus normalized areas showed a highly significant (p < 0.001) linear relationship (R2 = 0.993). Accurate quantification of beta-HADH activity was achieved since on-line monitoring allowed us to account for the NAD+ produced from NADH degradation by applying a correction factor. An average reaction time of 0.66 +/- 0.06 sec was determined for a protein concentration in the range of 0.1-0.5 mg protein/mL. The assay was reproducible since coefficients of variation of less than 6.2% were calculated for triplicate analyses.

The early ontogeny of digestive and metabolic enzyme activities in two commercial strains of arctic charr (Salvelinus alpinus L.).

The extent to which growth performance is linked to digestive or energetic capacities in the early life stages of a salmonid species was investigated. We compared two strains of Arctic charr known to have different growth potentials during their early development (Fraser and Yukon gold). Trypsin, lipase, and amylase activities of whole alevins were measured at regular intervals from hatching through 65 days of development. To assess catabolic ability, we also measured five enzymes representing the following metabolic pathways: amino acid oxidation (amino aspartate transferase), fatty acid oxidation (beta-hydroxy acyl CoA-dehydrogenase), tricarboxylic acid cycle (citrate synthase), glycolysis (pyruvate kinase), and anaerobic glycolysis (lactate dehydrogenase). The measurement of these enzyme activities in individual fish allowed a clear evaluation of digestive capacity in relation to energetic demand. We also compared triploid and diploid individuals within the Yukon gold strain. For the whole experimental period, diploid Yukon gold fish exhibited the highest growth rate (1.08+/-0.18% length/day) followed by triploid Yukon gold fish (1.00+/-0.28% length/day) and finally Fraser strain fish (0.84+/-0.28% length/day). When differences in enzyme activities were observed, the Fraser strain showed higher enzyme activities at a given length than the Yukon gold strain (diploid and triploid). Higher growth performance appears to be linked to lower metabolic capacity. Our results suggest that fish may have to reach an important increase in the ratio of digestive to catabolic enzyme activities or a leveling off of metabolic enzyme activities before the onset of large increases in mass.

Skeletal muscle enzymes as predictors of 24-h energy metabolism in reduced-obese persons.

BACKGROUND: Little is known about the effects of weight loss on the relation between skeletal muscle enzymes and energy metabolism. OBJECTIVE: This study was performed retrospectively to investigate the relation between skeletal muscle enzymes and 24-h energy metabolism in obese persons before and after weight loss. DESIGN: Ten women and 9 men [with body mass indexes (in kg/m(2)) > 30] underwent a 15-wk weight-loss program (-700 kcal/d). Body weight and composition, 24-h energy metabolism (whole-body indirect calorimetry), and maximal activities of phosphofructokinase (EC 2.7.1.11), creatine kinase (CK; EC 2.7.3.2), citrate synthase (CS; EC 4.1.3.7), 3-hydroxyacyl-CoA dehydrogenase (HADH; EC 1.1.1.35), and cytochrome-c oxidase (COX; EC 1.9.3.1) were determined from biopsy samples of the vastus lateralis taken before and after weight loss. RESULTS: Before weight loss, fat-free mass (FFM) was the only predictor of 24-h energy expenditure (R(2) = 0.70, P < 0.001), whereas the cumulative variance in sleeping metabolic rate explained by FFM and fat mass (FM) was 83% (P < 0.001). After weight loss, CS (r = 0.45, P = 0.05) and COX (r = 0.65, P < 0.01) were significantly associated with 24-h energy expenditure, whereas CK (r = 0.53, P < 0.05), CS (r = 0.45, P < 0.05), COX (r = 0.64, P < 0.01), and HADH (r = 0.45, P = 0.05) were all significant correlates of sleeping metabolic rate. After weight loss, FFM, FM, and COX explained 84% (P < 0.01) of the variance in 24-h energy expenditure, whereas FFM, FM, and CK all contributed to the cumulative variance in sleeping metabolic rate explained by this model (R(2) = 0.82, P < 0.05). CONCLUSION: Maximal activities of key skeletal muscle enzymes contribute to the variability in 24-h energy metabolism in reduced-obese persons.

Activity and mRNA levels of enzymes involved in hepatic fatty acid synthesis and oxidation in mice fed conjugated linoleic acid.

The effects of dietary conjugated linoleic acid (CLA) on the activity and mRNA levels of hepatic enzymes involved in fatty acid synthesis and oxidation were examined in mice. In the first experiment, male ICR and C57BL/6J mice were fed diets containing either a 1.5% fatty acid preparation rich in CLA or a preparation rich in linoleic acid. In the second experiment, male ICR mice were fed diets containing either 1.5% linoleic acid, palmitic acid or the CLA preparation. After 21 days, CLA relative to linoleic acid greatly decreased white adipose tissue mass but caused hepatomegaly accompanying an approximate 10-fold increase in the tissue triacylglycerol content irrespective of mouse strain. CLA compared to linoleic acid greatly increased the activity and mRNA levels of various lipogenic enzymes in both experiments. Moreover, CLA increased the mRNA expression of Delta6- and Delta5-desaturases, and sterol regulatory element binding protein-1 (SREBP-1). The mitochondrial and peroxisomal palmitoyl-CoA oxidation rate was about 2.5-fold higher in mice fed CLA than in those fed linoleic acid in both experiments. The increase was associated with the up-regulation of the activity and mRNA expression of various fatty acid oxidation enzymes. The palmitic acid diet compared to the linoleic acid diet was rather ineffective in modulating the hepatic lipid levels or activity and mRNA levels of enzymes in fatty acid metabolism. It is apparent that dietary CLA concomitantly increases the activity and mRNA levels of enzymes involved in fatty acid synthesis and oxidation, and desaturation of polyunsaturated fatty acid in the mouse liver. Both the activation of peroxisomal proliferator alpha and up-regulation of SREBP-1 may be responsible for this.

A novel HPLC-based method to diagnose peroxisomal D-bifunctional protein enoyl-CoA hydratase deficiency.

D-bifunctional protein (D-BP) plays an indispensable role in peroxisomal beta-oxidation, and its inherited deficiency in humans is associated with severe clinical abnormalities. Three different subtypes of D-BP deficiency can be distinguished: 1) a complete deficiency of D-BP (type I), 2) an isolated D-BP enoyl-CoA hydratase deficiency (type II), and 3) an isolated D-BP 3-hydroxyacyl-CoA dehydrogenase deficiency (type III). In this study, we developed a method to measure D-BP dehydrogenase activity independent of D-BP hydratase (D-BP HY) activity to distinguish between D-BP deficiency type I and type II, which until now was only possible by mutation analysis. For this assay, the hydratase domain of D-BP was expressed in the yeast Saccharomyces cerevisiae. After a coincubation of yeast homogenate expressing D-BP HY with fibroblast homogenate of patients using the enoyl-CoA ester of the bile acid intermediate trihydroxycholestanoic acid as substrate, D-BP dehydrogenase activity was measured. Fibroblasts of patients with a D-BP deficiency type II displayed D-BP dehydrogenase activity, whereas type I and type III patients did not. This newly developed assay to measure D-BP dehydrogenase activity in fibroblast homogenates provides a quick and reliable method to assign patients with deficient D-BP HY activity to the D-BP deficiency subgroups type I or type II.

Leg and arm lactate and substrate kinetics during exercise.

To study the role of muscle mass and muscle activity on lactate and energy kinetics during exercise, whole body and limb lactate, glucose, and fatty acid fluxes were determined in six elite cross-country skiers during roller-skiing for 40 min with the diagonal stride (Continuous Arm + Leg) followed by 10 min of double poling and diagonal stride at 72-76% maximal O(2) uptake. A high lactate appearance rate (R(a), 184 +/- 17 micromol x kg(-1) x min(-1)) but a low arterial lactate concentration ( approximately 2.5 mmol/l) were observed during Continuous Arm + Leg despite a substantial net lactate release by the arm of approximately 2.1 mmol/min, which was balanced by a similar net lactate uptake by the leg. Whole body and limb lactate oxidation during Continuous Arm + Leg was approximately 45% at rest and approximately 95% of disappearance rate and limb lactate uptake, respectively. Limb lactate kinetics changed multiple times when exercise mode was changed. Whole body glucose and glycerol turnover was unchanged during the different skiing modes; however, limb net glucose uptake changed severalfold. In conclusion, the arterial lactate concentration can be maintained at a relatively low level despite high lactate R(a) during exercise with a large muscle mass because of the large capacity of active skeletal muscle to take up lactate, which is tightly correlated with lactate delivery. The limb lactate uptake during exercise is oxidized at rates far above resting oxygen consumption, implying that lactate uptake and subsequent oxidation are also dependent on an elevated metabolic rate. The relative contribution of whole body and limb lactate oxidation is between 20 and 30% of total carbohydrate oxidation at rest and during exercise under the various conditions. Skeletal muscle can change its limb net glucose uptake severalfold within minutes, causing a redistribution of the available glucose because whole body glucose turnover was unchanged.

Skeletal muscle mitochondrial function and exercise capacity in HIV-infected patients with lipodystrophy and elevated p-lactate levels.

OBJECTIVE: To investigate the skeletal muscle mitochondrial function in HIV-infected patients with lipodystrophy or elevated p-lactate levels. DESIGN: Eight HIV patients treated with highly active antiretroviral therapy, with lipodystrophy or elevated p-lactate, and eight healthy controls were exposed to incremental exercise until exhaustion. METHODS: Blood samples and gas analysis were performed at rest, during exercise and in recovery. Oxygen consumption, workload and blood lactate were assessed. Before and immediately after exercise muscle biopsies were obtained, in which citrate synthase (CS), hydroxyacyl-coenzyme A dehydrogenase (HD), glycogen and nucleotides were measured. RESULTS: Maximal workload was significantly lower in patients compared with controls [171 Watt (88-206) versus 235 Watt (118-294) P = 0.05]. A trend towards lower maximal oxygen consumption (VO(2max)) was detected in patients [2136 ml/min (1221-2598) versus 2985 ml/min (1506-3959) P = 0.11]. Patients had significantly elevated levels of blood lactate at rest [1.55 mmol/l (1-2.5) versus 0.8 mmo/l (0.37-1.1) P < 0.01), but no significant difference in maximal blood-lactate values was found. The decline in blood lactate in the recovery period was similar between groups. There was no significant difference in CS, HD, glycogen or nucleotides. CONCLUSION: The significantly lower working capacity and the trend towards reduced VO(2max) in patients could be caused by mitochondrial dysfunction, but may also be caused by impaired physical fitness. The similar levels of nucleotides, CS, HD, and glycogen and the normal increase in blood lactate during exercise indicates a normal oxidative phosphorylation. No evidence of serious damage to skeletal muscle mitochondrial function was found.

Muscle glycogen stores and meat quality as affected by strategic finishing feeding of slaughter pigs.

The aim of the present study was to investigate whether muscle glycogen stores in slaughter pigs could be decreased through strategic finishing feeding before slaughter. Moreover, preliminary meat quality traits were measured to see whether such a regulation of muscle glycogen stores affected ultimate pH, color, and tenderness in the meat. The strategic finishing feeding was carried out the last 3 wk prior to slaughter. Seven experimental groups with eight animals per group were fed diets low in digestible carbohydrates. A control group with four animals was fed a traditional grower-finishing diet. The muscle glycogen stores were reduced in longissimus muscle (LM) 11 to 26% at the time of slaughter in pigs that were fed the experimental diets compared with the control group. Meat quality measured as ultimate pH and color on LM muscle in half the pigs 24 h postmortem showed that ultimate pH in LM was not affected by the reduction in glycogen stores in the muscles from pigs fed any of the experimental diets. However, the meat from pigs fed the experimental diets was darker than the meat from pigs that were fed the control diet with two of the experimental diets, resulting in significantly lower L* values. Activities of key enzymes in the glycolytic pathway, glycogen phoshorylase a and b, phosphofructokinase, and the fatty acid oxidative pathway, beta-hydrozyacyl-CoA-dehydrogenase, were not affected by the strategic feeding. In contrast, the activity of the proteolytic enzyme calpain as well as its inhibitor calpastatin was influenced by the strategic feeding. Lower activity of mu-calpain and greater activity of calpastatin in the muscle samples from the strategically fed pigs indicate a lesser muscle protein degradation in the muscles compared with muscles of control animals. The present study showed that the muscle glycogen stores in slaughter pigs can be reduced at the time of slaughter through strategic finishing feeding with diets low in digestible carbohydrate without compromising growth rate.

Effects of age and sex on the structural, chemical and technological characteristics of mule duck meat.

1. The aim of the study was to analyse the effect of age and sex on the chemical, structural and technological characteristics of mule duck meat. 2. Ten males and 10 females were weighed and slaughtered at 8, 10, 11, 12 and 13 weeks of age. Weight, pH value, colour, tenderness and juice loss of breast muscle were determined. 3. The activities of 3 enzymes (citrate synthase, beta-hydroxyacyl CoA dehydrogenase, lactate dehydrogenase) which indicate muscular metabolic activity were assayed. 4. Chemical composition (moisture, lipids, proteins, minerals, lipid and phospholipid classes, fatty acid composition) of breast muscle was analysed. 5. Fibre type, fibre type percentage and cross-sectional areas were determined using histochemistry and an image analysis system. 6. For growth performance and muscular structure, the ideal slaughter age of mule ducks is 10 weeks of age. Chemical and technological analysis indicated that muscular maturity in Pectoralis major was reached at 11 weeks of age, but, at this age, breast lipid content is high. Moreover, after 10 weeks of age, food costs rapidly increased. 7. Lastly, sexual dimorphism for body weight is minor. In this study, at any given age, no significant differences between males and females were shown. Thus, it is possible to rear both sexes together and to slaughter them at the same age.

Early and long-term changes of equine skeletal muscle in response to endurance training and detraining.

Twenty-four 4-year-old Andalusian (Spanish breed) stallions were used to examine the plasticity of myosin heavy chain (MHC) phenotype and the metabolic profile in horse skeletal muscle with long-term endurance-exercise training and detraining. Sixteen horses underwent a training programme based on aerobic exercises for 8 months. Afterwards, they were kept in paddocks for 3 months. The remaining eight horses were used as controls. Three gluteus medius muscle biopsy samples were removed at depths of 20, 40 and 60 mm from each horse before (month 0), during (month 3) and after (month 8) training, and again after 3 months of detraining (month 11). MHC composition was analysed by electrophoresis and immunohistochemistry with anti-MHC monoclonal antibodies. Fibre areas, oxidative capacity and capillaries were studied histochemically. The activities of key muscle enzymes of aerobic (citrate synthase and 3-hydroxy-acyl-CoA-dehydrogenase) and anaerobic (phosphofructokinase and lactic dehydrogenase) metabolism and the intramuscular glycogen and triglyceride contents were also biochemically analysed. Early changes with training (3 months) included hypertrophy of type IIA fibres, a reduction of MHC-IIX with a concomitant increase of MHC-IIA, a rise in the number of high-oxidative fibres and in the activities of aerobic muscle enzymes and glycogen content. Long-term changes with training (8 months) were a further decline in the expression of MHC-IIX, an increase of slow MHC-I, additional increases of high-oxidative fibres, capillary density, activities of aerobic enzymes and endogenous glycogen; intramuscular lipid deposits also increased after 8 months of training whereas the activities of anaerobic enzymes declined. Most of exercise-induced alterations reverted after 3 months of detraining. These results indicate that endurance-exercise training induces a reversible transition of MHC composition in equine muscle in the order IIX-->IIA-->I, which is coordinated with changes in the metabolic properties of the muscle. Furthermore, a dose-response relationship was evident between the duration (in total) of training and the magnitude of muscle adaptations.

A novel peroxisomal enzyme, D-3-hydroxyacyl-CoA dehydratase/D-3-hydroxyacyl-CoA dehydrogenase bifunctional protein: its expression in the developing human brain.

D-bifunctional protein, which is a newly recognized peroxisomal enzyme (D-3-hydroxyacyl-CoA dehydratase/D-3-hydroxyacyl-CoA dehydrogenase), demonstrates a characteristic development of pattern in the human brain. At 13 gestational weeks (GW), D-bifunctional protein immunoreactivity first appeared in the brain. Each neuron exhibited increased immunoreactivity along with growth in size as age increased and size with age. Glial cells in the white matter showed immunoreactivity after 30 GW. On the other hand, the L-bifunctional protein immunoreactivity was reported in neurons from 23 or 25 GW and in the white matter from 12 or 14 GW. Because of polymicrogyria in conditions such as infantile Refsum disease and infantile adrenoleukodystrophy, peroxisomal enzymes are thought to play an important role in neuronal migration and possibly myelination. D-bifunctional protein may be relevant to neuronal migration and L-bifunctional protein may be involved in axonal growth and synaptic development. This study is designed to access the ontogeny of D-bifunctional protein in the human brain.

Angiotensin-converting enzyme gene expression in skeletal muscle in patients with chronic heart failure.

BACKGROUND: Skeletal muscle factors may influence functional limitation in patients with heart failure. The renin-angiotensin system is activated in chronic heart failure. Treatment with angiotensin-converting enzyme (ACE) inhibitors improve symptoms and prognosis. The goal of this study was to quantify and localize skeletal muscle ACE-mRNA in patients with chronic heart failure and in control subjects, and to elucidate skeletal muscle fiber area and capillary density. METHODS AND RESULTS: Biopsies from the lateral vastus muscle were taken from 9 patients before and after treatment with enalapril and in 10 control subjects. ACE-mRNA was quantified with reverse transcription polymerase chain reaction. Immunohistochemistry was used to localize ACE within skeletal muscle. No difference in ACE-mRNA transcripts between patients and control subjects was detected, nor did ACE gene expression change after treatment with enalapril. The number of ACE-mRNA transcripts was related to muscle fiber area, whereas an inverse relationship between the number of ACE transcripts and capillary density was found. ACE was detected in the endothelial cells of capillaries in skeletal muscle. CONCLUSION: ACE is expressed in skeletal muscle and is confined to endothelial cells. The close relationship between capillary density and number of ACE transcripts indicate that activation of the renin-angiotensin system has an impact on capillary growth.