Role Of PPAR-? Modulating Lipid Induced Kupffer Cell Polarization Shift In Non-alcoholic Fatty Liver Disease

Background and Aims: Abnormal lipid mediated hepatic inflammatory-immune dysfunction related systematic low-grade inflammation plays an important role in the development of non-alcoholic fatty liver disease(NAFLD).As one of the prominent cells of metabolic-immune-inflammatory responses in the liver,Kupffer cells critically contribute to the progression of NAFLD.Macrophages(including the resident macrophages in the liver,Kupffer cells)are key cellular components of the immune system,characterized by their plasticity and heterogeneity,responding to different micro-environmental stimuli by displaying diverse functional phenotypes,which determined as polarization.Peroxisome proliferator-activated receptor-?(PPAR-?)and nuclear factor-?B(NF-?B)pathway are involved in the regulation of energy metabolism and inflammatory-immune responses as well as macrophage polarization.In the present study,we established high-fat diet mice model and in vitro dietary fatty acids treatment to characterize their effect on Kupffer cell polarization and the role of PPAR-? and NF-?B pathway.We also determined the effect of PPAR-? activation on lipid-induced macrophage M1-M2 polarization shift and the underlying mechanism.In addition,we investigated the potential of PPAR-? agonist in vivo on high-fat diet induced Kupffer cell polarization and hepatic steatosis.Methods: C57BL/6 mice were fed high-fat diet to establish NAFLD model.Hepatic steatosis was determined by histochemistry and the count,distribution as well as phenotypes of Kupffer cells were determined by immunohistochemistry.Kupffer cells were isolated by in situ perfusion of the liver with collagenase.Kupffer cells/macrophages were treated with either saturated fatty acids(SFAs)or polyunsaturated fatty acids(PUFAs)in vitro as well as pre-treated with PPAR-? agonist or antagonist and then combined with SFAs or PUFAs.M1/M2 phenotype markers were detected by real-time PCR.Secretion level of pro-inflammatory cytokines was measured by ELISA.The expression of NF-?B pathway and PPAR-? proteins was determined by Western Blot.The protein interaction between PPAR-? and NF-?Bp65 in macrophage nuclei was detected by co-immunoprecipitation.To explore the effect of PPAR-? agonist in vivo on high-fat diet induced Kupffer cell polarization and hepatic steatosis,the NAFLD mice were received PPAR-? agonist rosiglitazone administration for consecutive 2-4 weeks.Results:(1)The dynamic change of Kupffer cell count,distribution and predominant phenotype was consistent with the extent of high-fat diet induced hepatic steatosis and local pro-inflammatory response.Long-term(16-week)high-fat diet induced increasing infiltrating Kupffer cells mainly around hepatic fat vacuoles with predominant M1 polarization and increasing production of pro-inflammatory cytokines.(2)SFA-palmitic acid(PA)polarized Kupffer cells/macrophages to an M1 predominant M1/M2 mixed phenotype with increase of pro-inflammatory cytokines and significantly up-regulated protein phosphorylating activation of NF-?B pathway as well as protein expression of PPAR-?,while PUFA-docosahexaenoic acid(DHA)polarized Kupffer cells/macrophages to an M2 phenotype and remarkably increased PPAR-? protein expression but had no obvious effect on NF-?B pathway.(3)PPAR-? agonist shifted SFA induced predominant M1 polarization of macrophages to M2 polarization and enhanced PUFA induced M2 polarization of macrophages,while PPAR-? antagonist decreased both SFA and PUFA induced M2 polarization of macrophages but increased SFA induced M1 polarization of macrophages.PPAR-? agonist inhibited NF-?B pathway but enhanced protein interaction between PPAR-? and NF-?Bp65 treated with fatty acids,while PPAR-? antagonist had an opposite effect.(4)PPAR-? agonist rosiglitazone administration inhibited high-fat diet induced M1 polarization of Kupffer cells and alleviated hepatic steatosis as well as local pro-inflammatory response.Conclusions:(1)The dynamic change of Kupffer cell count,distribution and predominant phenotype was consistent with the extent of high-fat diet induced hepatic steatosis and local pro-inflammatory response.(2)Different fatty acids had different effect on Kupffer cell/macrophages polarization.SFA induced M1 polarization of macrophages was associated with activation of NF-?B pathway,while PPAR-? played an important role in different fatty acids induced M2 polarization of macrophages.(3)Enhanced PPAR-? expression caused M2 shift of fatty acids induced macrophage polarization,probably associated with the increasing protein interaction between PPAR-? and NF-?Bp65 leading to decreasing activity of NF-?B pathway.On the contrary,reductive PPAR-? expression had an opposite effect.(4)In vivo PPAR-? agonist administration inhibited high-fat diet induced M1 polarization of Kupffer cells and alleviated hepatic steatosis.