Conjugated linoleic acid isomers and their precursor fatty acids regulate peroxisome proliferator-activated receptor subtypes and major peroxisome proliferator responsive element-bearing target genes in HepG2 cell model

The purpose of this study was to examine the induction profiles(as judged by quantitative reverse transcription polymerase chain reaction(qRT-PCR)) of peroxisome proliferator-activated receptor(PPAR) α,β,γ subtypes and major PPAR-target genes bearing a functional peroxisome proliferator responsive element(PPRE) in HepG2 cell model upon feeding with cis-9,trans-11-octadecadienoic acid(9-CLA) or trans-10,cis-12-octadecadienoic acid(10-CLA) or their precursor fatty acids(FAs).HepG2 cells were treated with 100 μmol/L 9-CLA or 10-CLA or their precursor FAs,viz.,oleic,linoleic,and trans-11-vaccenic acids against bezafibrate control to evaluate the induction/expression profiles of PPAR α,β,γ subtypes and major PPAR-target genes bearing a functional PPRE,i.e.,fatty acid transporter(FAT),glucose transporter-2(GLUT-2),liver-type FA binding protein(L-FABP),acyl CoA oxidase-1(ACOX-1),and peroxisomal bifunctional enzyme(PBE) with reference to β-actin as house keeping gene.Of the three housekeeping genes(glyceraldehyde 3-phosphate dehydrogenase(GAPDH),β-actin,and ubiquitin),β-actin was found to be stable.Dimethyl sulfoxide(DMSO),the common solubilizer of agonists,showed a significantly higher induction of genes analyzed.qRT-PCR profiles of CLAs and their precursor FAs clearly showed upregulation of FAT,GLUT-2,and L-FABP(～0.5-2.0-fold).Compared to 10-CLA,9-CLA decreased the induction of the FA metabolizing gene ACOX-1 less than did PBE,while 10-CLA decreased the induction of PBE less than did ACOX-1.Both CLAs and precursor FAs upregulated PPRE-bearing genes,but with comparatively less or marginal activation of PPAR subtypes.This indicates that the binding of CLAs and their precursor FAs to PPAR subtypes results in PPAR activation,thereby induction of the target transporter genes coupled with downstream lipid metabolising genes such as ACOX-1 and PBE.To sum up,the expression profiles of these candidate genes showed that CLAs and their precursor FAs are involved in lipid signalling by modulating the PPAR α,β,or γ subtype for the indirect activation of the PPAR-target genes,which may in turn be responsible for the supposed health effects of CLA,and that care should be taken while calculating the actual fold induction values of candidate genes with reference to housekeeping gene and DMSO as they may impart false positive results.