Fasting causes lipolysis in adipose tissue leading to the release of large quantities of free fatty acids into circulation that reach the liver where they are metabolized to generate ketone bodies to serve as fuels for other tissues. Since fatty acid-metabolizing enzymes in the liver are transcriptionally regulated by peroxisome proliferator-activated receptor ␣ (PPAR␣), we investigated the role of PPAR␣ in the induction of these enzymes in response to fasting and their relationship to the development of hepatic steatosis in mice deficient in PPAR␣ (PPAR␣ ؊/؊ ), peroxisomal fatty acyl-CoA oxidase (AOX ؊/؊ ), and in both PPAR␣ and AOX (double knockout (DKO)). Fasting for 48 -72 h caused profound impairment of fatty acid oxidation in both PPAR␣؊/؊ and DKO mice, and DKO mice revealed a greater degree of hepatic steatosis when compared with PPAR␣ ؊/؊ mice. The absence of PPAR␣ in both PPAR␣ ؊/؊ and DKO mice impairs the induction of mitochondrial ␤-oxidation in liver following fasting which contributes to hypoketonemia and hepatic steatosis. Pronounced steatosis in DKO mouse livers is due to the added deficiency of peroxisomal ␤-oxidation system in these animals due to the absence of AOX. In mice deficient in AOX alone, the sustained hyperactivation of PPAR␣ and up-regulation of mitochondrial ␤-oxidation and microsomal -oxidation systems as well as the regenerative nature of a majority of hepatocytes containing numerous spontaneously proliferated peroxisomes, which appear refractory to store triglycerides, blunt the steatotic response to fasting. Starvation for 72 h caused a decrease in PPAR␣ hepatic mRNA levels in wild type mice, with no perceptible compensatory increases in PPAR␥ and PPAR␦ mRNA levels. PPAR␥ and PPAR␦ hepatic mRNA levels were lower in fed PPAR␣ ؊/؊ and DKO mice when compared with wild type mice, and fasting caused a slight increase only in PPAR␥ levels and a decrease in PPAR␦ levels. Fasting did not change the PPAR isoform levels in AOX ؊/؊ mouse liver. These observations point to the critical importance of PPAR␣ in the transcriptional regulatory responses to fasting and in determining the severity of hepatic steatosis.

Fatty acid ␤-oxidation occurs in both mitochondria and peroxisomes. Long chain fatty acids are also metabolized by the cytochrome P450 CYP4A -oxidation enzymes to toxic dicarboxylic acids (DCAs) that serve as substrates for peroxisomal ␤-oxidation. Synthetic peroxisome proliferators interact with peroxisome proliferator activated receptor ␣ (PPAR␣) to transcriptionally activate genes that participate in peroxisomal, microsomal, and mitochondrial fatty acid oxidation. Mice lacking PPAR␣ (PPAR␣ ؊/؊ ) fail to respond to the inductive effects of peroxisome proliferators, whereas those lacking fatty acyl-CoA oxidase (AOX ؊/؊ ), the first enzyme of the peroxisomal ␤-oxidation system, exhibit extensive microvesicular steatohepatitis, leading to hepatocellular regeneration and massive peroxisome proliferation, implying sustained activation of PPAR␣ by natural ligands. We now report that mice nullizygous for both PPAR␣ and AOX (PPAR␣ ؊/؊ AOX ؊/؊ ) failed to exhibit spontaneous peroxisome proliferation and induction of PPAR␣-regulated genes by biological ligands unmetabolized in the absence of AOX. In AOX ؊/؊ mice, the hyperactivity of PPAR␣ enhances the severity of steatosis by inducing CYP4A family proteins that generate DCAs and since they are not metabolized in the absence of peroxisomal ␤-oxidation, they damage mitochondria leading to steatosis. Blunting of microvesicular steatosis, which is restricted to few liver cells in periportal regions in PPAR␣ ؊/؊ AOX ؊/؊ mice, suggests a role for PPAR␣-induced genes, especially members of CYP4A family, in determining the severity of steatosis in livers with defective peroxisomal ␤-oxidation. In agematched PPAR␣ ؊/؊ mice, a decrease in constitutive mitochondrial ␤-oxidation with intact constitutive peroxisomal ␤-oxidation system contributes to large droplet fatty change that is restricted to centrilobular hepatocytes. These data define a critical role for both PPAR␣ and AOX in hepatic lipid metabolism and in the pathogenesis of specific fatty liver phenotype.In animal cells, mitochondria as well as peroxisomes oxidize fatty acids via ␤-oxidation, with long chain and very long chain fatty acids (LCFAs and VLCFAs) 1 being preferentially oxidized by peroxisomes (1-3). Peroxisomal ␤-oxidation is carried out by two distinct groups of enzymes. The classical first group utilizes straight chain saturated fatty acyl-CoAs as substrates, whereas the second group acts on branched chain acyl-CoAs (3, 4). In the classical L-3-hydroxy-specific ␤-oxidation spiral, dehydrogenation of acyl-CoA esters to their corresponding trans-2-enoyl-CoAs is catalyzed by fatty acyl-CoA oxidase (AOX), whereas the second and third reactions, hydration and dehydrogenation of enoyl-CoA esters to 3-ketoacyl-CoA, are catalyzed by a single enzyme, enoyl-CoA hydratase/L-3-hydroxyacyl-CoA dehydrogenase (L-bifunctional enzyme (L-PBE)) (3). The third enzyme of this classical system, 3-ketoacyl-CoA thiolase (PTL), cleaves 3-ketoacyl-CoAs to acetyl-CoA and an acylCoA that is two carbon atoms shorter than the original mo...

Adenovirus-induced hyperleptinemia rapidly depletes body fat in normal rats without increasing free fatty acids and ketogenesis, implying that fat-storing adipocytes are oxidizing the fat. To analyze the ultrastructural changes of adipocytes accompanying this functional transformation, we examined the fat tissue by electron microscopy. After 14 days of hyperleptinemia, adipocytes had become shrunken, fatless, and encased in a thick basementmembrane-like matrix. They were crowded with mitochondria that were much smaller than those of brown adipocytes. Their gene expression profile revealed striking up-regulation of peroxisome proliferator-activated receptor ␥ coactivator 1␣ (an up-regulator of mitochondrial biogenesis not normally expressed in white fat), increased uncoupling proteins-1 and -2, and down-regulation of lipogenic enzymes. Phosphorylation of both acetyl CoA carboxylase and AMP-activated protein kinase was increased, thus explaining the increase in fatty acid oxidation. The ability to transform adipocytes into unique fat-burning cells may suggest novel therapeutic strategies for obesity.