PPARalpha inhibits TGF-beta-induced beta5 integrin transcription in vascular smooth muscle cells by interacting with Smad4

• Integrins play an important role in vascular smooth muscle cell (VSMC) migration, a crucial event in the development of restenosis and atherosclerosis. Transforming growth factor-beta (TGF-beta) is highly expressed in restenotic and atherosclerotic lesions, and known to induce integrin expression. Peroxisome proliferator-activated receptor alpha (PPARalpha), a member of the nuclear receptor superfamily, regulates gene expression in a variety of vascular cells. We investigated the effects of PPARalpha ligands on TGF-beta-induced beta3 and beta5 integrin expression and potential interaction between PPARalpha and TGF-beta signaling. PPARalpha ligands WY-14643 (100 micromol/L) and 5,8,11,14-eicosatetranoic acid (ETYA, 50 micromol/L) inhibited TGF-beta-induced beta5 integrin protein expression by 72+/-6.8% and 73+/-7.1%, respectively (both P<0.05). TGF-beta-stimulated beta3 integrin expression was not affected by PPARalpha ligands. Both PPARalpha ligands also suppressed TGF-beta-induced beta5 integrin mRNA levels. PPARalpha ligands inhibited TGF-beta-inducible transcription of beta5 integrin by an interaction with a TGF-beta response element between nucleotides -63 and -44, which contains a Sp1/Sp3 transcription factor binding site. Nuclear complexes binding to the TGF-beta response region contained Sp1/Sp3 and TGF-beta-regulated Smad 2, 3, and 4 transcription factors. TGF-beta-stimulated Sp1/Smad4 nuclear complex formation was inhibited by WY-14643 and ETYA with a parallel induction of PPARalpha/Smad4 interactions. However, in vitro pull-down experiments failed to demonstrate direct binding between PPARalpha/Smad4. Both PPARalpha ligands blocked PDGF-directed migration of TGF-beta-pretreated VSMCs, a process mediated, in part, by beta5 integrins. The present study demonstrates that PPARalpha activators inhibit TGF-beta-induced beta5 integrin transcription in VSMCs through a novel indirect interaction between ligand-activated PPARalpha and the TGF-beta-regulated Smad4 transcription factors.

Nyckelord

5;8;11;14-Eicosatetraynoic Acid/pharmacology

Animals

Cell Movement/drug effects

Cells; Cultured

DNA-Binding Proteins/*metabolism

Gene Expression/drug effects

Genes; Reporter

Integrin beta Chains/*biosynthesis/genetics

Integrin beta3/biosynthesis/genetics

Ligands

Macromolecular Substances

Muscle; Smooth; Vascular/cytology/drug effects/*metabolism

Nuclear Proteins/metabolism

Platelet-Derived Growth Factor/pharmacology

Promoter Regions (Genetics)/genetics

Protein Binding/drug effects/physiology

Pyrimidines/pharmacology

RNA; Messenger/metabolism

Rats

Rats; Sprague-Dawley

Receptors; Cytoplasmic and Nuclear

Signal Transduction/drug effects/physiology

Smad4 Protein

Trans-Activators/*metabolism

Transcription Factors/*pharmacology

Transcription; Genetic/*drug effects

Transforming Growth Factor beta/*pharmacology

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