Microsomal Fatty Acid Oxidation Research Articles

Hepatic Peroxisomal Fatty Acid β-Oxidation Is Regulated by Liver X Receptor α

Peroxisomes are the exclusive site for the beta-oxidation of very-long-chain fatty acids of more than 20 carbons in length (VLCFAs). Although the bulk of dietary long-chain fatty acids are oxidized in the mitochondria, VLCFAs cannot be catabolized in mitochondria and must be shortened first by peroxisomal beta-oxidation. The regulation of peroxisomal, mitochondrial, and microsomal fatty acid oxidation systems in liver is mediated principally by peroxisome proliferator-activated receptor alpha (PPARalpha). In this study we provide evidence that the liver X receptor (LXR) regulates the expression of the genetic program for peroxisomal beta-oxidation in liver. The genes encoding the three enzymes of the classic peroxisomal beta-oxidation cycle, acyl-coenzyme A (acyl-CoA) oxidase, enoyl-CoA hydratase/L-3-hydroxyacyl-CoA dehydrogenase, and 3-ketoacyl-CoA thiolase, are activated by the LXR ligand, T0901317. Accordingly, administration of T0901317 in mice promoted a dose-dependent and greater than 2-fold increase in the rate of peroxisomal beta-oxidation in the liver. The LXR effect is independent of PPARalpha, because T0901317-induced peroxisomal beta-oxidation in the liver of PPARalpha-null mice. Interestingly, T0901317-induced peroxisomal beta-oxidation is dependent on the LXRalpha isoform, but not the LXRbeta isoform. We propose that induction of peroxisomal beta-oxidation by LXR agonists may serve as a counterregulatory mechanism for responding to the hypertriglyceridemia and liver steatosis that is promoted by potent LXR agonists in vivo; however, additional studies are warranted.

Induction of overlapping genes by fasting and a peroxisome proliferator in pigs: evidence of functional PPARα in nonproliferating species

Peroxisome proliferator-activated receptor alpha (PPARalpha), a key regulator of fatty acid oxidation, is essential for adaptation to fasting in rats and mice. However, physiological functions of PPARalpha in other species, including humans, are controversial. A group of PPARalpha ligands called peroxisome proliferators (PPs) causes peroxisome proliferation and hepatocarcinogenesis only in rats and mice. To elucidate the role of PPARalpha in adaptation to fasting in nonproliferating species, we compared gene expressions in pig liver from fasted and clofibric acid (a PP)-fed groups against a control diet-fed group. As in rats and mice, fasting induced genes involved with mitochondrial fatty acid oxidation and ketogenesis in pigs. Those genes were also induced by clofibric acid feeding, indicating that PPARalpha mediates the induction of these genes. In contrast to rats and mice, little or no induction of genes for peroxisomal or microsomal fatty acid oxidation was observed in clofibric acid-fed pigs. Histology showed no significant hyperplasia or hepatomegaly in the clofibric acid-fed pigs, whereas it showed a reduction of glycogen by clofibric acid, an effect of PPs also observed in rats. Copy number of PPARalpha mRNA was higher in pigs than in mice and rats, suggesting that peroxisomal proliferation and hyperresponse of several genes to PPs seen only in rats and mice are unrelated to the abundance of PPARalpha. In conclusion, PPARalpha is likely to play a central role in adaptation to fasting in pig liver as in rats and mice.

Weight-of-evidence versus strength-of-evidence in toxicologic hazard identification: Di(2-ethylhexyl)phthalate (DEHP)

Toxicokinetic and mode of action data for DEHP reduce the concern for its potential carcinogenic hazard to human health. Chronic, high dose ingestion of DEHP and related peroxisome proliferators (PP) by mice and rats precipitate the following: activation of peroxisome proliferator activated receptor (PPARα) and its binding to peroxisome proliferator response elements (PPREs) within promoters of PP-responsive genes, peroxisome proliferation, increased microsomal fatty acid oxidation, increased hepatic hydrogen peroxide, hepatomegaly, hyperplasia and subsequent neoplasia. Neither peroxisome proliferation nor increased liver cancer occur in patients treated with pharmacologic doses of PP. Species differences in endogenous PPARα expression and differential activity of the peroxisome proliferator response element (PPRE) contribute to the failure of humans to respond in a manner qualitatively similar to that of rats or mice. Where it can be demonstrated that a mechanism for rodent tumor formation has no relevance for humans, then a substance which elicits a carcinogenic response in the test species via that mechanism should not be classified as anything other than an animal carcinogen. Systemic noncarcinogenic endpoints are available for definition of a DEHP reference dose. Considerable difficulty is encountered in the revision of promulgated regulations and in public risk communication when a material is no longer considered a carcinogenic hazard to humans.

Fish Oil Feeding Decreases Mature Sterol Regulatory Element-binding Protein 1 (SREBP-1) by Down-regulation of SREBP-1c mRNA in Mouse Liver: A POSSIBLE MECHANISM FOR DOWN-REGULATION OF LIPOGENIC ENZYME mRNAs

Dietary fish oil induces hepatic peroxisomal and microsomal fatty acid oxidation by peroxisome proliferator-activator receptor alpha activation, whereas it down-regulates lipogenic gene expression by unknown mechanism(s). Because sterol regulatory element-binding proteins (SREBPs) up-regulated lipogenic genes, investigation was made on the effects of fish oil feeding on SREBPs and sterol regulatory element (SRE)-dependent gene expression in C57BL/6J mice. Three forms of SREBPs, SREBP-1a, -1c, and -2, are expressed in liver, and their truncated mature forms activate transcription of sterol-regulated genes. C57BL/6J mice were divided into three groups; the first group was given a high carbohydrate diet, and the other two groups were given a high fat diet (60% of total energy), with the fat in the form of safflower oil or fish oil, for 5 months. Compared with safflower oil feeding, fish oil feeding decreased triglyceride and cholesterol concentrations in liver. There were no differences in amount of SREBP-1 and -2 in both precursor and mature forms between carbohydrate- and safflower oil-fed mice. However, compared with safflower oil feeding, fish oil feeding reduced the amounts of precursor SREBP-1 in membrane fraction by 90% and of mature SREBP-1 in liver nuclei by 57%. Fish oil feeding also reduced precursor SREBP-2 by 65% but did not alter the amount of mature SREBP-2. Compared with safflower oil feeding, fish oil feeding decreased liver SREBP-1c mRNA level by 86% but did not alter SERBP-1a mRNA. Consistent with decrease of mature SREBP-1, compared with safflower oil feeding, fish oil feeding down-regulated the expression of liver SRE-dependent genes, such as low density lipoprotein receptor, 3-hydroxy-3-methylglutaryl-CoA reductase, 3-hydroxy-3-methylglutaryl-CoA synthase, fatty acid synthase, acetyl-CoA carboxylase, and stearoyl-CoA desaturase-1. These data suggested that in liver, fish oil feeding down-regulates the mature form of SREBP-1 by decreasing SREBP-1c mRNA expression, with corresponding decreases of mRNAs of cholesterologenic and lipogenic enzymes.

Multiple mechanisms for polyunsaturated fatty acid regulation of hepatic gene transcription

Dietary polyunsaturated fatty acids (PUFA) have profound effects on hepatic gene transcription leading to significant changes in lipid metabolism. Highly unsaturated n-3 PUFA suppress the transcription of genes encoding specific lipogenic enzymes and induce the expression of genes encoding specific enzymes involved in peroxisomal and microsomal fatty acid oxidation. Our studies have shown that fatty acid effects on hepatic gene expression may involve at least three distinct pathways. One pathway involves peroxisome proliferator-activated receptor (PPARα), a fatty acid activated nuclear receptor. PPARα is required for the PUFA induction of mRNAs encoding enzymes involved in fatty acid oxidation. However, PPARα is not required for PUFA suppression of mRNAs encoding proteins involved in lipogenesis. A second pathway involves prostanoids. In cultured 3T3-L1 adipocytes, cyclooxygenase derived 20:4 n-6 metabolites, like PGE 2, suppress mRNAs encoding proteins involved in lipogenesis. However, in hepatic parenchymal cells, 20:4 n-6 suppression of lipogenic gene expression does not require a cyclooxygenase. Nevertheless, PGE 2 and PGF 2α suppress hepatic lipogenic gene expression. 20:4 n-6 cyclooxygenase products can arise from non-parenchymal cells and through a paracrine control process act on a G-protein linked receptor signaling cascade to suppress lipogenic gene expression. The fact that n-3 and n-6 PUFA suppression of lipogenic gene expression does not require PPARα or cyclooxygenase activity indicates the presence of a third pathway for the control of hepatic gene transcription. These studies indicate that the pleiotropic effects of PUFA on hepatic lipid metabolism cannot be attributed to a single regulatory mechanism.

Modulation of peroxisomal and microsomal fatty acid oxidation by acetone. A comparative study between liver and kidney

The effect of acetone consumption on some microsomal and peroxisomal activities was studied in rat kidney and these results were compared with data from former investigations in liver. Acetone increased the microsomal lauric acid hydroxylation, the aminopyrine N-demethylation catalyzed by cytochrome P450 and the microsomal UDP-glucuronyltransferase activity. Also, acetone increased the peroxisomal β-oxidation of palmitoyl CoA and catalase activities in kidney. These studies suggest that acetone is a common inducer of the microsomal and peroxisomal fatty acid oxidation, as previously shown in both starved and ethanol treated rats. Our results support the hypothesis that microsomal fatty acid ω-hydroxylation results in the generation of substrates being supplied for peroxisomal β-oxidation. We propose that the final purpose of these linked fatty acid oxidations could be the catabolism of fatty acids or the generation of a substrate for the synthesis of glucose from fatty acids. This pathway would be triggered by acetone treatment in a similar way in liver and kidney.

Peroxisomal and Microsomal Fatty Acid Oxidation in Liver of Rats after Chronic Ethanol Consumption

1. Microsomal P450 and peroxisomal fatty acid oxidation activities were studied in liver of rats after long-term ethanol consumption. 2. Ethanol increased the microsomal lauric acid ω-hydroxylation and the aminopyrine N-demethylation catalyzed by cytochrome P450. 3. Ethanol increased peroxisomal β-oxidation of palmitoyl CoA and catalase activity in liver. 4. Both microsomal and peroxisomal activities behaved in a coordinate way in the liver of rats with long-term ethanol consumption. 5. These results would support a role of microsomal ω-hydroxylation and peroxisomal β-oxidation of fatty acids in an extramitochondrial pathway of lipid oxidation in the liver.

Do Peroxisome Proliferating Compounds Pose a Hepatocarcinogenic Hazard to Humans?

The purpose of the workshop “Do Peroxisome Proliferating Compounds Pose a Hepatocarcinogenic Hazard to Humans?” was to provide a review of the current state of the science on the relationship between peroxisome proliferation and hepatocarcinogenesis. There has been much debate regarding the mechanism by which peroxisome proliferators may induce liver tumors in rats and mice and whether these events occur in humans. A primary goal of the workshop was to determine where consensus might be reached regarding the interpretation of these data relative to the assessment of potential human risks. A core set of biochemical and cellular events has been identified in the rodent strains that are susceptible to the hepatocarcinogenic effects of peroxisome proliferators, including peroxisome proliferation, increases in fatty acyl-CoA oxidase levels, microsomal fatty acid oxidation, excess production of hydrogen peroxide, increases in rates of cell proliferation, and expression and activation of the α subtype of the peroxisome proliferator-activated receptor (PPAR-α). Such effects have not been identified clinically in liver biopsies from humans exposed to peroxisome proliferators or inin vitrostudies with human hepatocytes, although PPAR-α is expressed at a very low level in human liver. Consensus was reached regarding the significant intermediary roles of cell proliferation and PPAR-α receptor expression and activation in tumor formation. Information considered necessary for characterizing a compound as a peroxisome proliferating hepatocarcinogen include hepatomegaly, enhanced cell proliferation, and an increase in hepatic acyl-CoA oxidase and/or palmitoyl-CoA oxidation levels. Given the lack of genotoxic potential of most peroxisome proliferating agents, and since humans appear likely to be refractive or insensitive to the tumorigenic response, risk assessments based on tumor data may not be appropriate. However, nontumor data on intermediate endpoints would provide appropriate toxicological endpoints to determine a point of departure such as the LED10or NOAEL which would be the basis for a margin-of-exposure (MOE) risk assessment approach. Pertinent factors to be considered in the MOE evaluation would include the slope of the dose–response curve at the point of departure, the background exposure levels, and variability in the human response.

Polyunsaturated Fatty Acid Suppression of Hepatic Fatty Acid Synthase and S14 Gene Expression Does Not Require Peroxisome Proliferator-activated Receptor α

Dietary polyunsaturated fatty acids (PUFA) induce hepatic peroxisomal and microsomal fatty acid oxidation and suppress lipogenic gene expression. The peroxisome proliferator-activated receptor alpha (PPARalpha) has been implicated as a mediator of fatty acid effects on gene transcription. This report uses the PPARalpha-deficient mouse to examine the role of PPARalpha in the PUFA regulation of mRNAs encoding hepatic lipogenic (fatty acid synthase (FAS) and the S14 protein (S14)), microsomal (cytochrome P450 4A2 (CYP4A2)), and peroxisomal (acyl-CoA oxidase (AOX)) enzymes. PUFA ingestion induced mRNAAOX (2.3-fold) and mRNACYP4A2 (8-fold) and suppressed mRNAFAS and mRNAS14 by >/=80% in wild type mice. In PPARalpha-deficient mice, PUFA did not induce mRNAAOX or mRNACYP4A2, indicating a requirement for PPARalpha in the PUFA-mediated induction of these enzymes. However, PUFA still suppressed mRNAFAS and mRNAS14 in the PPARalpha-deficient mice. Studies in rats provided additional support for the differential regulation of lipogenic and peroxisomal enzymes by PUFA. These studies provide evidence for two distinct pathways for PUFA control of hepatic lipid metabolism. One requires PPARalpha and is involved in regulating peroxisomal and microsomal enzymes. The other pathway does not require PPARalpha and is involved in the PUFA-mediated suppression of lipogenic gene expression.

Starvation effect on rat kidney peroxisomal and microsomal fatty acid oxidation: A comparative study between liver and kidney

Microsomal lauric acid 12-hydroxy lauric acid (ω)-hydroxylation and fatty acid peroxisomal β-oxidation were studied in kidney tissue from starved rats. Starvation increased the microsomal ω-hydroxylation and peroxisomal β-oxidation of fatty acids with a high correlation between both processes. Earlier, we reported similar results in liver. Our results support the hypothesis that the role of microsomal fatty acids ω-hydroxylation is the generation of substrate for peroxisomal β-oxidation, with the final purpose of contributing to a catabolic or gluconeogenic pathway from fatty acids.

Modulation of rat liver peroxisomal and microsomal fatty acid oxidation by starvation

In this work the microsomal lauric acid ω-hydroxylation, fatty acid peroxisomal β-oxidation, and the levels of cytochrome P-450 IVAI were studied in liver tissue from starved rats. Starvation increased the peroxisomal β-oxidation and the microsomal hydroxylation of fatty acids. The correlation between these activities would support the proposal that both processes are linked, contributing in part to catabolism of fatty acids in liver of starved rats.

Effects of Octanoate and Oleate on Energy Metabolism in the Perfused Rat Liver

The effects of octanoate and oleate were studied in the isolated fasted rat liver perfused without substrate and were compared with the effects of lactate. Measurements were made of hepatic adenine nucleotide content, formation of ketone bodies, urea and glucose and uptake of fatty acids, lactate and oxygen. Calculations of the Krebs cycle activity and the flux through the oxidation at C‐3 were carried out. Comparing the extra oxygen consumption with extra ATP needs after addition of the fatty acids or lactate, ADP: O ratios were estimated for fatty acid oxidation and lactate gluconeogenesis.1. Octanoate as well as oleate induced a net decrease in ATP and an increase in AMP content of the liver. Total adenine nucleotides were unaltered. ATP:ADP ratios were lowered while the adenylate kinase mass‐action ratio increased.2. The Krebs cycle activity was suppressed by both fatty acids and enhanced by lactate. Oxidation at C‐3 was strongly stimulated by the fatty acids and was unaffected by lactate.3. An apparent ADP:O ratio of 2.7 was obtained after lactate addition, indicating a tightly coupled oxidative phosphorylation for the liver preparations used. Octanoate and oleate gave extremely low ratios of near one and near zero respectively.4. Perfusion with oligomycin caused a severe drop in oxygen uptake by the liver, which was unaltered after addition of fatty acids. Oligomycin added after the fatty acids caused an immediate fall in oxygen uptake to the level observed with oligomycin alone. 2,4‐Dinitrophenol was able to stimulate the oligomycin‐depressed respiration, both in the absence and in the presence of fatty acids. These results indicate that under our experimental conditions the fatty acids had no uncoupling effect and that microsomal fatty acid oxidation had to be minimal.5. Incorporation of [3H]leucine into proteins in liver and medium remained unchanged after addition of fatty acids, indicating that the low apparent ADP:O ratios are not the result of an enhanced synthesis of proteins.6. The possible relationship between fatty acid transport, energy‐wasting futile cycles and the low apparent ADP:O ratios observed during fatty acid oxidation is discussed.