Effect Of The Novel Ligand Of PPARγ On Lipid Metabolism In Macrophages

It has a positive impact that removed excess cholesterol from the intracellular to the liverin atherosclerosis.Reverse cholesterol transport (RCT) is a pathway by which to describe theProcess by which extrahepatic (peripheral) cholesterol is returned to the liver for excretion inthe bile and ultimately the feces.Cholesterol efflux,part of the RCT process, is a major processby which macrophages within the vessel wall secrete cholesterol outside cells.Overexpressionof ATP binding cassette transporter A1(ABCA1) stimulates efflux of cholesterol,ABCA1istherefore an important target for anti-atherogenic therapies. It has been reported that activationof Peroxisome proliferator-activated receptor γ(PPARγ) by natural or synthetic ligands resultsin thetransactivation of ABCA1:Initially, uptake of oxidized lowdensity lipoprotein(oxLDL)by macrophages results in induction of PPARγ,release of its ligands, on the one hand，increasedexpression of ABCA1，promoted thecholesterol efflux,on theother hand,up-regulated expression of scavenger receptor CD36and stimulate uptake of oxLDL,keepcholesterol balance.We previously purified a negatively charged lipid moiety from Cu2+induced oxLDL,7-ketocholesteryl-9-carboxynonanoate(oxLig-1),which have the similarstructure with PPARγ ligands.Theeffect of oxLig-1onABCA1expressionmediated by nuclearreceptors (NRs) has not been elucidated.In the present study, we investigated the signalprocess by which oxLig-1regulates ABCA1expression and promotes ApoA-I-mediatedcholesterol efflux from macrophages by the mediation of PPARγ.Methods: First,we docked oxLig-1and the ligand-binding domain (LBD) of PPARγ byELISA assay.Dual-Luciferase reporter assay detected the effect of oxLig-1on PPARγtranscriptional activity.Then analysed the influence of oxLig-1on PPARγ-med iated Lipidmetabolic signaling pathways by the methods of Western blotting,3-Dodecanoyl-NBD-chol-esterol(NBD-Cholesterol),Flow cytometry and shRNA.Results: It was found that oxLig-1was able to bind to PPARγ LBD,and oxLig-1was aneffective competitor for troglitazone binding.oxLig-1also improve the transcription activityof PPARγ.ABCA1expression was increased in the presence of oxLig-1at differentconcentrations (15~30μg/mL) within6h. The expression of ABCA1reached the peak levelwith20μg/mL oxLig-1for2hrs. A peak efflux rate was observed with20μg/mL oxLig-1for4h.As expected, PPARγ-shRNA resulted in a reduction of90%in PPARγ proteinexpression.Under PPARγ knockdown, oxLig-1-induced ABCA1expression and cholesterolefflux was blocked by62%and25%respectively. CD36expression was increased in thepresence of oxLig-1also mediated by PPARγ.Conclusions:These observations suggest that oxLig-1is possibly a novel ligand for PPARγ, play an important role in regulating lipid metabolism balance by increasingABCA1,CD36expression via induction of PPARγ.