Peroxisomal Beta-oxidation Activity Research Articles

Reduction of n-3 PUFAs, specifically DHA and EPA, and enhancement of peroxisomal beta-oxidation in type 2 diabetic rat heart

BackgroundThere is overwhelming evidence that dietary supplementation with n-3 polyunsaturated fatty acids (PUFAs), mainly EPA (C20:5n-3) and DHA (C22:6n-3), has cardiovascular protective effects on patients with type 2 diabetes mellitus (T2DM) but not on healthy people. Because the T2DM heart increases fatty acid oxidation (FAO) to compensate for the diminished utilization of glucose, we hypothesize that T2DM hearts consume more n-3 PUFAs and, therefore, need more n-3 PUFAs. In the present study, we investigated the changes in cardiac n-3 PUFAs and peroxisomal beta-oxidation, which are responsible for the degradation of PUFAs in a high-fat diet (HFD) and low-dose streptozotocin- (STZ) induced type 2 diabetic rat model.Methods and resultsThe capillary gas chromatography results showed that all the n-3 (or omega-3) PUFAs, especially DHA (~50%) and EPA (~100%), were significantly decreased, and the n-6/n-3 ratio (~115%) was significantly increased in the hearts of diabetic rats. The activity of peroxisomal beta-oxidation, which is crucial to very-long-chain and unsaturated FA metabolism (including DHA), was significantly elevated in DM hearts. Additionally, the real-time PCR results showed that the mRNA expression of most peroxisomal beta-oxidation key enzymes were up-regulated in T2DM rat hearts, which might contribute to the reduction of n-3 (or omega-3) PUFAs.ConclusionIn conclusion, our results indicate that T2DM hearts consume more n-3 PUFAs, especially DHA and EPA, due to exaggerated peroxisomal beta-oxidation.

Dicer ablation in oligodendrocytes provokes neuronal impairment in mice

MicroRNAs (miRNAs) regulate gene expression and have many roles in the brain, but a role in oligodendrocyte (OL) function has not been demonstrated. A Dicer floxed conditional allele was crossed with the proteolipid protein promoter-driven inducible Cre allele to generate inducible, OL-specific Dicer-floxed mice. OL-specific Dicer mutants show demyelination, oxidative damage, inflammatory astrocytosis and microgliosis in the brain, and eventually neuronal degeneration and shorter lifespan. miR-219 and its target ELOVL7 (elongation of very long chain fatty acids protein 7) were identified as the main molecular components that are involved in the development of the phenotype in these mice. Overexpressing ELOVL7 results in lipid accumulation, which is suppressed by miR-219 co-overexpression. In Dicer mutant brain, excess lipids accumulate in myelin-rich brain regions, and the peroxisomal beta-oxidation activity is dramatically reduced. Postnatal Dicer ablation in mature OLs results in inflammatory neuronal degeneration through increased demyelination, lipid accumulation, and peroxisomal and oxidative damage, and therefore indicates that miRNAs play an essential role in the maintenance of lipids and redox homeostasis in mature OLs that are necessary for supporting axonal integrity as well as the formation of compact myelin.

Effect of Dietary Omega 3 Phosphatidylcholine on Obesity-Related Disorders in Obese Otsuka Long-Evans Tokushima Fatty Rats

Recent reports have shown that dietary phosphatidylcholine (PC) has various beneficial biological effects. Omega 3 polyunsaturated fatty acids (omega3 PUFAs) have also been reported to have lipid-lowering effects in animal models and human studies. In the present study, we investigated the effect of omega3 PUFAs containing PC (omega3-PC) on obesity-related disorders in Otsuka Long-Evans Tokushima fatty (OLETF) rats. Rats were fed semisynthetic diets that contained either 5% corn oil and 2% egg-PC or 5% corn oil and 2% omega3-PC for 4 weeks. During this 4 week feeding of the omega3-PC, the OLEFT rats showed a decrease of omental white adipose tissue weight. In addition, the omega3-PC diet significantly decreased liver weight and hepatic lipid levels in OLETF rats. These changes were attributable to the significant suppression of fatty acid synthase activity and significant enhancement in the activities of carnitine palmitoyltransferase and peroxisomal beta-oxidation. Moreover, the omega3-PC diet reduced serum glucose levels concomitant with the increase of serum adiponectin levels. These results show that compared with egg-PC, omega3-PC can prevent or alleviate obesity-related disorders through the suppression of fatty acid synthesis, enhancement of fatty acid beta-oxidation, and increase of the serum adiponectin level in OLETF rats.

Underlying Mechanisms of Pharmacology and Toxicity of a Novel PPAR Agonist Revealed Using Rodent and Canine Hepatocytes

Marked species-specific responses to agonists of the peroxisome proliferator-activated alpha receptor (PPAR alpha) have been observed in rats and dogs, two species typically used to assess the potential human risk of pharmaceuticals in development. In this study, we used primary cultured rat and dog hepatocytes to investigate the underlying mechanisms of a novel PPAR alpha and -gamma coagonist, LY465608, relative to fenofibrate, a prototypical PPAR alpha agonist. As expected, rat hepatocytes incubated with these two agonists demonstrated an increase in peroxisome number as evaluated by electron microscopy, whereas the peroxisome number remained unchanged in dog hepatocytes. Biochemical analysis showed that rat hepatocytes responded to PPAR agonists with an induction of both peroxisomal and mitochondrial beta-oxidation (PBox and MBox) activities. Dog hepatocytes treated with both PPAR agonists, however, did not show increased PBox activity but did demonstrate increased MBox activity. Analysis of peroxisomal beta-oxidation gene expression markers by quantitative real-time PCR confirmed that PPAR agonists induced the peroxisomal enzymes, acyl-coenzyme A (CoA) oxidase (Acox), enoyl-CoA hydratase/L-3-hydroxyacyl-CoA dehydrogenase (Ehhadh), and 3-ketoacyl-CoA thiolase (Acaa1) at the transcriptional level in rat hepatocytes, but not dog hepatocytes. Expression of mRNA for the mitochondrial beta-oxidation gene hydroxyacyl-CoA dehydrogenase/3-ketoacyl-CoA thiolase (Hadhb), however, increased in both rat and dog hepatocytes, consistent with biochemical measurements of peroxisomal and mitochondrial beta-oxidation. Repeat-dose nonclinical safety studies of LY465608 revealed abnormities in mitochondrial morphology and evidence of single-cell necrosis following 30 days of dosing exclusively in dogs, but not in rats. Microarray analysis indicated that dog hepatocytes, but not rat hepatocytes, treated with LY465608 had an expression profile consistent with abnormalities in the regulation of cell renewal and death, oxidative stress, and mitochondrial bioenergetics, which may explain the canine-specific toxicity observed in vivo with this compound. This increased sensitivity to mitochondrial toxicity of canine hepatocytes relative to rat hepatocytes identified using gene expression was confirmed using the fluorescent indicator tetramethylrhodamine ethyl ester (TMRE) and flow cytometry. At doses of 0.1 microM LY465608, canine hepatocytes showed a greater shift in fluorescence indicative of mitochondrial damage than observed with rat hepatocytes treated at 10 microM. In summary, using rat and dog primary hepatocytes, we replicated the pharmacologic and toxicologic effects of LY465608 observed in vivo during preclinical development and propose an underlying mechanism for these species-specific effects.

Responses of the Liver to Perfluorinated Fatty Acids with Different Carbon Chain Length in Male and Female Mice:In Relation to Induction of Hepatomegaly, Peroxisomal .BETA.-Oxidation and Microsomal 1-Acylglycerophosphocholine Acyltransferase

The potency of the induction of hepatomegaly, peroxisomal beta-oxidation and microsomal 1-acylglycerophoshocholine (1-acyl-GPC) acyltransferase was compared among perfluorinated fatty acids (PFCAs) with 6-9 carbon chain length in the liver of male and female mice. All PFCAs examined induced hepatomegaly and peroxisomal beta-oxidation and the potency was in the order of perfluorononanonic acid (PFNA), perfluorooctanoic acid (PFOA), perfluoroheptanoic acid (PFHA) and perfluorohexanoic acid (PFHeA) when compared with the relative doses to induce the two parameters. Microsomal 1-acyl-GPC acyltransferase was induced by PFHA, PFOA and PFNA, as was peroxisomal beta-oxidation. No significant sex-related difference was observed in the induction of peroxisomal beta-oxidation by any PFCAs examined. PFNA and PFOA accumulated in the liver of both male and female mice in a dose-dependent manner. PFHA accumulated in the liver to a lesser extent; little PFHeA accumulated in the liver. Hepatic concentrations of PFNA, PFOA and PFHA were higher in male mice than those in female mice. One linear regression line was confirmed between the activities of peroxisomal beta-oxidation and hepatic concentrations of PFHeA, PFHA, PFOA and PFNA in male mice regardless of their carbon chain lengths, and the activities were saturable at the concentrations over approximately 500 nmol/g liver. Similar linear regression line was obtained between the two parameters in female mice. These results suggest (i) that the longer the perfluoroalkyl chain becomes, the more PFCA accumulates in the liver of both male and female mice, (ii) that the accumulated PFCAs induce hepatomegaly, peroxisomal beta-oxidation and microsomal 1-acyl-GPC acyltransferase, and (iii) that the difference observed in the accumulation of PFHA, PFOA and PFNA in the liver between male and female mice is not enough to produce obvious sex-related difference in the induction of peroxisomal beta-oxidation.

Short-Term Feeding of Conjugated Linoleic Acid Does Not Induce Hepatic Steatosis in C57BL/6J Mice

We investigated the effect of short-term feeding of conjugated linoleic acid (CLA) on adipose tissue weights, liver weight, hepatic lipid metabolism, and serum lipoprotein profiles in C57BL/6J mice. Mice were fed semi-synthetic diets containing either 6%, high-linoleic safflower oil (HL-SAF) or 4% HL-SAF+2% CLA for 1 wk. Short-term feeding of CLA showed an anti-obesity effect without inducing hepatomegaly in mice. In addition to the decline of hepatic triglyceride concentration, significant inhibition of A9 desaturation of fatty acid in the total liver lipids was found in CLA-fed mice. The CLA diet significantly increased the activities of peroxisomal beta-oxidation and decreased the activities of diacylglycerol acyltransferase, a triglyceride synthesis-related enzyme, in the liver. Moreover, serum lipoprotein profiles of CLA-fed mice showed preferable changes in the atherogenic indices. However, serum leptin and adiponectin were drastically decreased by CLA feeding, suggesting that prolonged administration of CLA would induce further decrease of serum adipocytokine levels, which may be a cause of lipodystrophy in mice. These results show that short-term feeding of CLA does not induce adverse effect in C57BL/6J mice.

Biochemical effect evaluation of perfluorooctane sulfonic acid-contaminated wood mice (Apodemus sylvaticus).

Wood mice (Apodemus sylvaticus) were captured at Blokkersdijk, a nature reserve in the immediate vicinity of a fluorochemical plant in Antwerp, Belgium, and at Galgenweel, 3 kilometers farther away. The liver perfluorooctane sulfonic acid (PFOS) concentrations in the Blokkersdijk mice were extremely high (0.47-178.55 micro g/g wet weight). Perfluorononanoic, perfluorodecanoic, perfluoroundecanoic, and perfluorododecanoic acids were found sporadically in the liver tissue of the Blokkersdijk mice. The liver PFOS concentrations at Galgenweel were significantly lower than those at Blokkersdijk (0.14-1.11 micro g/g wet weight). Further results suggest sex independence of the liver PFOS levels, increased levels of PFOS bioaccumulation in older mice, and maternal PFOS transfer to the young. Several liver end points were significantly elevated in the Blokkersdijk mice: liver weight, relative liver weight, peroxisomal beta-oxidation activity, microsomal lipid peroxidation level, and mitochondrial fraction protein content. For the mitochondrial fraction catalase activity, no significant difference between locations was found. The liver weight, relative liver weight, and liver microsomal lipid peroxidation level increased significantly with the liver PFOS concentration. No indications for PFOS-mediated effects on the serum triglyceride, cholesterol, or potassium levels were obtained. The liver PFOS concentration was negatively related to the serum alanine aminotransferase activity.

Peroxisome proliferators do not activate the transcription factors AP-1, early growth response-1, or heat shock factors 1 and 2 in rats or hamsters.

Peroxisome proliferators (PPs) cause hepatomegaly, peroxisome proliferation, and hepatocarcinogenesis in rats and mice, whereas hamsters are less responsive to these compounds. PPs increase peroxisomal beta-oxidation and P4504A subfamily activity, which have been hypothesized to result in oxidative stress. Work in our laboratory indicated that differential modulation of the redox-sensitive transcription factor NF-kappaB may contribute to the resulting difference in species susceptibility following PP administration. Therefore, we hypothesized that other redox-sensitive transcription factors such as AP-1, early growth response gene 1 (Egr-1), and heat-shock factors 1 and 2 (HSF1/2) may also be alternatively activated in differentially susceptible species. Accordingly, we measured the activation of these transcription factors using gel mobility shift assays, with hepatic nuclear extracts derived from rats and Syrian hamsters fed two doses of three peroxisome proliferators (dibutyl-phthalate [DBP], gemfibrozil and Wy-14,643) for 6, 34, or 90 days. Although changes were observed at various time points, no consistent, dose-responsive changes were observed in the DNA binding activities of these transcription factors following PP treatment. The lack of increased binding of AP-1, Egr-1, and HSFs suggests that these factors are not involved in increased cell proliferation following PP administration, although we cannot rule out that these factors are activated at earlier time points than those examined in this study.

Differential induction of peroxisomal beta-oxidation enzymes by clofibric acid and aspirin in piglet tissues.

Peroxisomal beta-oxidation (POX) of fatty acids is important in lipid catabolism and thermogenesis. To investigate the effects of peroxisome proliferators on peroxisomal and mitochondrial beta-oxidation in piglet tissues, newborn pigs (1-2 days old) were allowed ad libitum access to milk replacer supplemented with 0.5% clofibric acid (CA) or 1% aspirin for 14 days. CA increased ratios of liver weight to body weight (P < 0.07), kidney weight to body weight (P < 0.05), and heart weight to body weight (P < 0.001). Aspirin decreased daily food intake and final body weight but increased the ratio of heart weight to body weight (P < 0.01). In liver, activities of POX, fatty acyl-CoA oxidase (FAO), total carnitine palmitoyltransferase (CPT), and catalase were 2.7-, 2.2-, 1.5-fold, and 33% greater, respectively, for pigs given CA than for control pigs. In heart, these variables were 2.2-, 4.1-, 1.9-, and 1.8-fold greater, respectively, for pigs given CA than for control pigs. CA did not change these variables in either kidney or muscle, except that CPT activity was increased approximately 110% (P < 0.01) in kidney. Aspirin increased only hepatic FAO and CPT activities. Northern blot analysis revealed that CA increased the abundance of catalase mRNA in heart by approximately 2.2-fold. We conclude that 1) POX and CPT in newborn pigs can be induced by peroxisomal proliferators with tissue specificity and 2) the relatively smaller induction of POX in piglets (compared with that in young or adult rodents) may be related to either age or species differences.

Metabolism of n-3 and n-6 fatty acids in Atlantic salmon liver: stimulation by essential fatty acid deficiency.

Oxidation, esterification, desaturation, and elongation of [1-14C]18:2n-6 and [1-14C]18:3n-3 were studied using hepatocytes from Atlantic salmon (Salmo salar L.) maintained on diets deficient in n-3 and n-6 polyunsaturated fatty acids (PUFA) or supplemented with n-3 PUFA. For both dietary groups, radioactivity from 18:3n-3 was incorporated into lipid fractions two to three times faster than from 18:2n-6, and essential fatty acids (EFA) deficiency doubled the incorporation. Oxidation to CO2 was very low and was independent of substrate or diet, whereas oxidation to acid-soluble products was stimulated by EFA deficiency. Products from 18:2n-6 were mainly 18:3n-6, 20:3n-6, and 20:4n-6, with minor amounts of 20:2n-6 and 22:5n-6. Products from 18:3n-3 were mainly 18:4n-3, 20:5n-3, and 22:6n-3, with small amounts of 20:3n-3. The percentage of 22:6n-3 in the polar lipid fraction of EFA-deficient hepatocytes was fourfold higher than in n-3 PUFA-supplemented cells. This correlated well with our other results obtained after abdominal injection of [1-14C]18:3n-3 and [1-14C]18:2n-6. In hepatocytes incubated with [4,5-3H]-22:6n-3, 20:5n-3 was the main product. This retroconversion was increased by EFA deficiency, as was peroxisomal betaoxidation activity. This study shows that 18:2n-6 and 18:3n-3 can be elongated and desaturated in Atlantic salmon liver, and that this conversion and the activity of retroconversion of very long chain PUFA is markedly enhanced by EFA deficiency.

Enhanced potential for oxidative stress in hyperinsulinemic rats: imbalance between hepatic peroxisomal hydrogen peroxide production and decomposition due to hyperinsulinemia.

Oxidative stress is involved in aging and age-related diseases. Several metabolic alterations similar to those encountered with aging and age-related diseases have been observed in response to hyperinsulinemia. Surprisingly, this metabolic derangement diminished hepatic peroxisomal beta-oxidation which is a major source of H2O2 production in the liver, suggesting a protective effect against oxidative stress. However, the impact of hyperinsulinemia on the balance between H2O2 production and elimination in the liver is not known. Consequently, this study was undertaken to evaluate the effect of sustained high serum insulin levels on the activity of hepatic catalase, a peroxisomal antioxidant enzyme involved in the decomposition of H2O2. Male Sprague-Dawley rats received intravenous infusion of either 30% glucose, 30% galactose or normal saline for seven days. Activity of hepatic peroxisomal beta-oxidation and catalase decreased 58% and 74%, respectively, in glucose-infused rats compared with galactose- or saline-infused animals. When infused simultaneously with glucose, diazoxide blocked glucose-enhanced insulin secretion and prevented the decrease in peroxisomal enzyme activities, without altering blood glucose concentration. Neither diazoxide alone nor galactose, which did not alter serum insulin levels, had any effect on enzyme activities. These results suggest that hyperinsulinemia is responsible for the decreased enzyme activities observed in glucose-infused rats. Indeed, a strong negative correlation between serum insulin levels and hepatic peroxisomal enzyme activities was found. To investigate the mechanism by which insulin modulates catalase activity, we studied rates of synthesis and degradation of catalase in saline- and glucose-infused rats. Data show that insulin diminishes rates of catalase synthesis, while exhibiting no effect on its degradation. Upsetting the balance between the cellular capacity to produce and eliminate H2O2 may be a contributing factor to the known deleterious effects of hyperinsulinemia.

Cytokine-induced accumulation of very long-chain fatty acids in rat C6 glial cells: implication for X-adrenoleukodystrophy.

X-Adrenoleukodystrophy (X-ALD) is an inherited metabolic disorder of very long-chain fatty acids (VLCFA) with subsequent manifestation of neuroinflammatory disease. To investigate the possible role of proinflammatory cytokines in the X-ALD disease process, we examined the effect of cytokines on the metabolism of VLCFA in C6 glial cells expressing oligodendrocyte-like properties. C6 glial cells under serum-free conditions were treated with different combinations of cytokines (tumor necrosis factor-alpha, interleukin-1beta, interferon-gamma) or cytokine with bacterial lipopolysaccharide (LPS). Cytokine-treated C6 cells had higher concentrations of VLCFA, measured as percent weight and also as C26:0/C22:0 ratio, which were 300-400% as compared with the controls. We also found increased levels of C26:1 in cytokine-treated cells. The accumulation of VLCFA paralleled the decrease (35-55%) in peroxisomal beta-oxidation activity and a 12- to 14-fold increase in the production of nitric oxide (NO). Individual cytokines were unable either to produce NO or to increase the levels of VLCFA in C6 cells. Inhibition of cytokine-induced NO production by L-N-methylarginine, an inhibitor of NO synthase (NOS), and N-acetylcysteine, an inhibitor of cytokine-mediated induction of inducible NOS, normalized the peroxisomal beta-oxidation activity and the levels of VLCFA, suggesting a role for the proinflammatory cytokines and NO toxicity in the neuropathological changes associated with abnormal VLCFA metabolism (e.g., X-ALD). X-ALD is a peroxisomal disease having deficient oxidation of VLCFA, resulting in the excessive accumulation of VLCFA in all tissues but especially in brain. We observed greater increase in levels of VLCFA in the inflammatory region of ALD brain (in the demyelinating plaque and the area around the plaque) than in the normal-looking area away from the plaque; this also indicates that cytokines in the proinflammatory region may augment the VLCFA defect caused by the inherited abnormality in X-ALD brain. Although C6 glial cultured cells do not reflect the X-ALD model precisely, the observed relationship between the cytokine-induced inhibition of the oxidation of VLCFA, excessive accumulation of VLCFA, and excessive production of NO and their normalization by inhibitors of NOS in C6 glial cells suggests that NO-mediated toxicity may play a role in VLCFA-associated neuroinflammatory diseases (e.g., X-ALD).

Utilization of an auxotrophic strain of the yeast Yarrowia lipolytica to improve gamma-decalactone production yields.

gamma-Decalactone is an aroma compound with a pleasant peachy odour. Most industrial processes use the bioconversion of ricinoleic acid by yeasts to produce gamma-decalactone. Peroxisomal beta-oxidation activity is responsible for the bioconversion. Some yeasts, Yarrowia lipolytica in particular, grow during the bioconversion, yielding a low bioconversion rate. Auxotrophy for uracil of a genetically engineered Y. lipolytica strain was used to prevent growth in the bioconversion medium. beta-Oxidation activities and gamma-decalactone production of the auxotrophic strain were measured and compared with a wild-type strain in media supplemented or not. Induction of beta-oxidation was observed in the non-supplemented medium, although to a lesser extent than in supplemented medium. Aroma productivity of the auxotrophic strain in the supplemented medium was similar to that observed for the wild-type strain in both media. However, in the non-supplemented medium the productivity of the auxotrophic strain was 10-20-fold higher.

Effects of ubiquinone and mevalonic acid on hepatic peroxisomal enzymes induced by dehydroepiandrosterone.

The adrenal steroid, dehydroepiandrosterone (DHEA) has been identified as a peroxisome proliferator. We examined the effects of the cellular antioxidant ubiquinone and its precursor mevalonic acid on the induction of enzymes associated with DHEA-mediated peroxisome proliferation in male F-344 rats. Upon treatment with DHEA (300 mg/kg orally for 14 days), there was a significant increase in hepatic activities of peroxisomal beta-oxidation (3 fold), 3-ketoacyl-CoA thiolase (4 fold) and catalase (1.8 fold). Co-administration of either mevalonic acid (100 mg/kg intraperitoneally) or ubiquinone (50 mg/kg orally) with DHEA significantly attenuated the DHEA-mediated induction of these enzymes. However, neither ubiquinone nor mevalonic acid alone significantly altered peroxisomal enzyme activities in rat liver. These data suggest that exogenous administration of ubiquinone or mevalonic acid can modulate the induction of the enzymes involved in peroxisome proliferation.

Induction of peroxisomal enzymes in rat liver by dehydroepiandrosterone sulfate.

Male F-344 rats, when treated with either 150 mg/kg or 300 mg/kg body weight of DHEAS for 14 days, produced a dose-dependent increase in liver weight and peroxisomal beta-oxidation activity, characteristic of peroxisomal proliferation. Contrary to previous observations in vitro, we also found a significant increase in catalase activity in rat liver with the higher dose of the steroid. Furthermore, the in vivo induction of peroxisomal beta-oxidation by DHEAS observed in our study was significantly less than reported in vitro, and also unlike previously reported in vitro results, was approximately equivalent to DHEA administered in vivo.

Biochemical and Immunocytochemical Properties of Peroxisomes and Mitochondria in Bovine Chromaffin Cells.

The biochemical and immunocytochemical properties of peroxisomal and mitochondrial beta-oxidation enzymes in bovine adrenal chromaffin cells were investigated. Peroxisomes were detectable by immunofluorescence staining using antibodies against acyl-CoA oxidase, peroxisomal 3-ketoacyl-CoA thiolase and catalase. The mitochondria were abundantly stained with antibody against mitochondrial 3-ketoacyl-CoA thiolase. The biosynthesis and intracellular processing of acyl-CoA oxidase and the peroxisomal 3-ketoacyl-CoA thiolase was slower than that in fibroblasts. The peroxisomal beta-oxidation activities shown by [1-14C] lignoceric acid oxidation were slightly lower than those in fibroblasts, whereas the mitochondrial beta-oxidation activities shown by [1-14C] palmitic acid oxidation were almost identical to those in fibroblasts. Adrenal chromaffin cells are useful materials for investigating the peroxisomal and mitochondrial metabolism of autonomic neurons and may contribute to the clarification of neuronal dysfunction in peroxisomal and mitochondrial disorders.

PRENATAL DIAGNOSIS OF PEROXISOMAL DISORDERS. † 869

Peroxisomal disorders are genetic disease caused either by a defect in peroxisome biogenesis resulting in deficiencies of multiple peroxisomal enzymes or by a deficiency of one peroxisomal enzyme. Seventeen different peroxisomal disorders are recognized nowadays, with progressive neurological deficits and early death in most. Both in peroxisome deficiency disorders(PDD) like Zellweger syndrome, neonatal Adrenoleukodystrophy (n-ALD) and infantile Refsum disease, in classic rhizomelic Chondrodysplasia punctata(RCDP) and disorders with a defect in peroxisomal beta-oxidation of fatty acids (including X-linked ALD) no effective treatment is available and early prenatal detection indicated. For PDD this is done by us by in vitro measurement of acylCoA:dihydroxyacetonephosphate acyltransferase(DHAPAT) activity and immuno(western)blotting with antisera raised against catalase, acylCoA oxidase and peroxisomal thiolase(Thio) resp. in uncultured chorionic villi(CV) homogenates. Since 1992 seventeen affected PDS fetusses were recognized by us by this procedure in sixty one at risk pregnancies in the 10th or 11th gestational week. Moreover, an additional thirteen affected PDS fetusses were found by analyzing de novo plasmalogen biosynthesis and very longchain fatty acids(VLCFA) in cultured CV-fibroblasts or amniocytes. Seven RCDP fetusses were diagnosed by the finding of abnormal immunoblots with anti-thiolase (absence of 41kDa band for mature thiolase protein) and deficient plasmalogens in CV. We also diagnosed isolated DHAPAT deficiency in a fetus by finding a deficiency of plasmalogens and DHAPAT, but normal Thio-blot in CV. Fifteen fetusses with a deficiency of one of the enzymes of peroxisomal beta-oxidation, including X-linked ALD, were diagnosed prenatally in 41 at risk pregnancies by the finding of elevated VLCFA and deficient activity of peroxisomal beta-oxidation in cultured CV-fibroblasts or amniocytes. No false positive or negative cases were found with these procedures.

Effect of nicardipine, a calcium antagonist, on induction of peroxisomal enzymes by dehydroepiandrosterone sulfate in cultured rat hepatocytes.

We examined the effect of nicardipine, a calcium antagonist, on the induction of peroxisomal enzymes, such as acyl-CoA oxidase and carnitine acetyltransferase, by dehydroepiandrosterone sulfate (DHEAS) and clofibric acid (CPIB), in primary cultured rat hepatocytes. Peroxisomal beta-oxidation and carnitine acetyltransferase activities were increased 11- and 20-fold, respectively, after 5 days of treatment with DHEAS (40 microM). However, 60 microM nicardipine significantly suppressed the induction of both of these activities by DHEAS to about 2-fold that of the control. This suppression was found to be both dose- and time-dependent. Immunoblot and Northern blot analyses of acyl-CoA oxidase revealed that suppression by nicardipine of the induction of peroxisomal beta-oxidation activity would be responsible for an increase in the amount of mRNA. In addition, the manner in which nicardipine suppressed the induction of peroxisomal beta-oxidation and carnitine acetyltransferase activity, was similar to that of clofibric acid. These findings suggest that in the calcium-dependent pathway, the mechanism for the induction of peroxisomal enzymes by DHEAS is basically the same as that by clofibric acid, a typical peroxisome proliferator. The present results also support our previous hypothesis that calcium may play an important role in the induction of these enzymes by peroxisome proliferators.

Effects of clofibric and beclobric acid in rat and monkey hepatocyte primary culture: influence on peroxisomal and mitochondrial beta-oxidation and the activity of catalase, glutathione S-transferase and glutathione peroxidase.

The effect of hypolipidaemic compounds on peroxisomal fatty acid beta-oxidation and on peroxisome morphology in the liver differs widely between rodent and primate species. We studied the relative importance of peroxisomal and mitochondrial beta-oxidation of palmitate in primary cultures of hepatocytes isolated from rat and monkey liver in the absence or presence of clofibric acid or beclobric acid. It was demonstrated that it is possible to differentiate between peroxisomal and mitochondrial beta-oxidation activities in intact cells. Overall beta-oxidation of palmitate was ca. 30% higher in rat hepatocytes than in monkey liver cells. In both monkey and rat cell cultures the mitochondrial component was over 90% of the total palmitate beta-oxidation. In rat hepatocyte culture clofibric acid and beclobric acid caused a 5- to 8-fold stimulation of peroxisomal beta-oxidation, while in monkey cells this activity was not significantly increased. However, in cells derived from both species mitochondrial palmitate beta-oxidation was increased (rat 2.5-fold; monkey 1.5-fold). These results indicate that the species differences in the increase in peroxisomal fatty acid oxidation are not a result of an inability to metabolize fatty acids in rat liver cell mitochondria. A comparison of the activity of enzymes involved in the detoxification of hydrogen peroxide showed that catalase and glutathione-S-transferase activity is 2.9-fold higher in monkey hepatocytes than in rat liver cells, while glutathione peroxidase activity was 1.6-fold higher in rat cells.(ABSTRACT TRUNCATED AT 250 WORDS)

Dietary gamma-linolenic acid-enriched oil reduces body fat content and induces liver enzyme activities relating to fatty acid beta-oxidation in rats.

The objectives of this study were to examine the effects of dietary gamma-linolenic acid-enriched oil extracted from fungi on rat body composition and on the various enzyme activities relating to fat metabolism in the liver. The oil contained 25.3 g gamma-linolenic acid/100 g fatty acids. The levels of gamma-linolenic acid-enriched oil in the diets were 0, 1.5 and 4%, to give 0, 2.88 and 7.68 g gamma-linolenic acid/kg diet. The control diet contained 8% soybean oil. The rats were given free access to these diets for 4 wk. Body weight gain was less in the gamma-linolenic acid oil-fed groups than in the control group, although food intake was similar among the three groups. Absolute and relative carcass fat weights were significantly lower in the gamma-linolenic acid oil-fed groups than in the control group. Carcass protein and water contents were not different among the three groups, although values were slightly greater than controls in gamma-linolenic acid-fed groups when expressed relative to body weight. Plasma total cholesterol and free fatty acid concentrations generally were lower in the gamma-linolenic acid oil-fed groups than in the control group. In the liver, there were no significant differences in activities of malic enzyme and citrate cleavage enzyme among the three groups. However, the activities of carnitine palmitoyl-transferase and peroxisomal beta-oxidation were significantly higher in the gamma-linolenic acid oil-fed groups than in the control group. These results clearly demonstrate that dietary gamma-linolenic acid oil reduces body fat content and facilitates fatty acid beta-oxidation in the liver.