PPAR alpha is necessary for the lipopenic action of hyperleptinemia on white adipose and liver tissue

Abstract

Adenovirus-induced hyperleptinemia causes rapid disappearance of body fat in normal rats, presumably by up-regulating fatty acid oxidation within white adipocytes. To determine the role of peroxisomal proliferation-activated receptor (PPAR)alpha expression, which was increased during the rapid loss of fat, we infused adenovirus-leptin into PPAR alpha(-/-) and PPAR alpha(+/+) mice. Despite similar degrees of hyperleptinemia and reduction in food intake, epididymal fat pad weight declined 55% in wild-type but only 6% in PPAR alpha(-/-) mice; liver triacylglycerol fell 39% in the wild-type group but was unchanged in PPAR(-/-) mice. Carnitine palmitoyl transferase-1 mRNA rose 52% in the wild-type mice but did not increase in PPAR alpha(-/-) mice. PPAR gamma coactivator-1 alpha rose 3-fold in the fat and 46% in the liver of wild-type mice but was unchanged in PPAR alpha(-/-) mice. Although AMP-activated protein kinase could not be implicated in the lipopenic actions of hyperleptinemia, acetyl CoA carboxylase protein was reduced in the liver of wild-type but not in PPAR alpha(-/-) mice. Thus, in PPAR alpha(-/-) mice, up-regulation of carnitine palmitoyl transferase-1 mRNA in fat, down-regulation of acetyl CoA carboxylase in liver, and up-regulation of PPAR gamma coactivator-1 alpha mRNA in both tissues are abolished, as is the reduction in their triacylglycerol content.

Figure 1

(A) Comparison by reverse transcription–PCR of PPARα mRNA in the liver of a wild-type (+/+) and a PPARα knockout (−/−) mouse. (B) The appearance of adipose tissue in a PPAR^+/+ and a PPAR^−/− mouse 7 days after the induction of hyperleptinemia by the i.v. administration of AdCMV-leptin.

Figure 2

(A) Comparison of mRNA, measured by real-time reverse transcription–PCR, of the oxidative enzymes CPT-1 and ACO in the epididymal fat (WAT) of PPAR^+/+ and PPAR^−/− mice 7 days after the i.v. administration of AdCMV-leptin or AdCMV-β-gal as a control. (B) Comparison of the mRNA of SREBP-1c and ACC in the liver of PPARα^+/+ and PPARα^−/− mice 7 days after the i.v. administration of AdCMV-leptin or AdCMV-β-gal. (C) Comparison of the mRNA ratios to β-actin of PGC-1α in WAT and liver of PPARα^+/+ and PPARα^−/− mice 7 days after the i.v. administration of AdCMV-leptin or AdCMV-β-gal. The mRNAs of interest are expressed as percent of that in AdCMV-β-gal-treated controls (mean ± SEM).

Figure 3

FAS and ACL content of liver and adipose tissue. Shown are immunoblots of FAS and ACL in replicate extracts prepared from PPARα^+/+ and PPARα^−/− mice 7 days after the i.v. administration of adCMV-leptin or AdCMV-β-gal as control.

Figure 4

ACC content in mouse adipose tissue. Shown are streptavidin–horseradish peroxidase blots of extracts of mouse adipose tissue (in duplicate or triplicate) prepared from PPAR^+/+ and PPAR^−/− mice 7 days after the i.v. administration of AdCMV-leptin or AdCMV-β-gal as a control. Visible are the two known ACC isoforms [ACCβ (280 kDa) and ACCα (265 kDa)].

Figure 5

ACC content in mouse liver. Shown are streptavidin–horseradish peroxidase blots (Top and Middle) of extracts of mouse adipose (in triplicate) prepared from PPAR^+/+ and PPAR^−/− mice 7 days after the i.v. administration of AdCMV-leptin or AdCMV-β-gal as a control. Visible are the two known ACC isoforms, ACCβ (280 kDa) and ACCα (265 kDa). (Bottom) The same samples have been blotted with an antibody that specifically recognizes Ser79p, the regulatory phosphorylation site of ACC.

Figure 6

AMPK activity and phosphorylation state in mouse adipose tissue and liver. (Upper) Results of triplicate determinations (mean ± SEM) of AMPK activity, measured as in Materials and Methods, in the presence of saturating concentrations of AMP (200 μM). The data are expressed as pmol P incorporated per minute per milligram total protein immunoprecipiates (in triplicate), blotted with an antibody specific for Thr-172, the activating phosphorylation site of AMPKα.