gw-501516 and Carcinoma--Hepatocellular

The peroxisome proliferator-activated receptor delta (PPARdelta) is a transcription factor that regulates genes of importance in lipid and glucose metabolism. ApoA-II is one of the major proteins of the HDL-particle. The aim of this study was to investigate the regulation of apoA-II gene expression by PPARdelta. Treatment of HepG2 cells with the PPARdelta specific agonist GW501516 increased apoA-II mRNA expression. Likewise, reporter gene assays using a construct containing 2.7 kb of the proximal apoA-II promoter showed increased activity after treatment with GW501516, both in HepG2 and in HuH-7 cells. Mutation of two putative PPAR response elements (PPREs) in this region showed that the PPRE at position -737/-717 is the functional site. Binding of PPARdelta to this site was confirmed by chromatin immunoprecipitation and gel retardation analyses. In conclusion, PPARdelta increases the expression of the human apoA-II gene in liver cells via a PPRE in the proximal promoter.

Topics: Apolipoprotein A-II; Base Sequence; Binding Sites; Carbohydrate Metabolism; Carcinoma, Hepatocellular; Cell Line, Tumor; Chromatin Immunoprecipitation; Electrophoretic Mobility Shift Assay; Gene Expression; Humans; Lipid Metabolism; Mutation; PPAR delta; Promoter Regions, Genetic; Protein Binding; Response Elements; Sequence Homology, Nucleic Acid; Thiazoles

Interleukin-6 and downstream liver effectors acute phase reactants are implicated in the systemic inflammatory reaction. Peroxisome proliferator-activated receptor delta (PPARdelta), which binds to and is activated by a variety of fatty acids, was recently shown to have anti-inflammatory actions.. We examined the ability of the synthetic PPARdelta agonist GW501516 to suppress interleukin-6-induced expression of acute phase proteins in human hepatoma HepG2 cells and rat primary hepatocytes. Results GW501516 dose-dependently suppressed interleukin-6-induced mRNA expression of the acute phase protein alpha1-antichymotrypsin in HepG2 cells. The compound also suppressed interleukin-6-induced mRNA expression of alpha2-acid glycoprotein, beta-fibrinogen and alpha2-macroglobulin in and the secretion of C-reactive protein by rat primary hepatocytes. Depletion of the PPARdelta receptor, but not of PPARalpha or gamma, attenuated the suppressive effect of GW501516 on interleukin-6-induced alpha1-antichymotrypsin mRNA expression, indicating that PPARdelta specifically mediated this effect. Since interleukin-6 stimulates the transcriptional activity of the alpha1-antichymotrypsin promoter by activating the signal transducer and activator of transcription (STAT) 3, we examined functional interaction of this transcription factor and PPARdelta on this promoter. Overexpression of PPARdelta enhanced the suppressive effect of GW501516 on STAT3-activated transcriptional activity of the alpha1-antichymotrypsin promoter, while GW501516 suppressed interleukin-6-induced binding of this transcription factor to this promoter.. These findings indicate that agonist-activated PPARdelta interferes with interleukin-6-induced acute phase reaction in the liver by inhibiting the transcriptional activity of STAT3. PPARdelta agonists might be useful for the suppression of systemic inflammatory reactions in which IL-6 plays a central role.

Topics: Acute-Phase Proteins; Acute-Phase Reaction; Animals; Carcinoma, Hepatocellular; Humans; Interleukin-6; Liver Neoplasms, Experimental; PPAR delta; Rats; STAT3 Transcription Factor; Thiazoles

Cyclooxygenase-2 (COX-2) has been suggested to be associated with carcinogenesis. Recently, many studies have shown increased expression of COX-2 in a variety of human malignancies, including hepatocellular carcinoma (HCC). Therefore, it becomes important to know more about what determines COX-2 expression. In this work, we have studied the effect of PPARdelta activation on COX-2 expression using a selective agonist (GW501516) in human hepatocellular carcinoma (HepG2) cells. Activation of PPARdelta resulted in increased COX-2 mRNA and protein expression. The mechanism behind the induction seems to be increased activity of the proximal promoter of the COX-2 gene, spanning nucleotides -327 to +59. The increased COX-2 protein expression and promoter activity induced by the GW501516 was also confirmed in the monocytic cell line THP-1. Induced levels of COX-2 have previously been associated with resistance to apoptosis and increased cell proliferation in many cell types. In HepG2 cells, we observed a dose-dependent increase in cell number by GW501516 treatment for 72h. The levels of PCNA, used as an indicator of cell division were induced, and the cell survival promoting complex p65 (NF-kappaB) was phosphorylated under GW501516 treatment. We conclude that PPARdelta activation in HepG2 cells results in induced COX-2 expression and increased cellular proliferation. These results may suggest that PPARdelta plays an important role in the development of HCC by modulating expression of COX-2.

Topics: Carcinoma, Hepatocellular; Cell Division; Cyclooxygenase 2; Gene Expression Regulation, Enzymologic; Gene Expression Regulation, Neoplastic; Humans; Isoenzymes; Liver Neoplasms; Membrane Proteins; Promoter Regions, Genetic; Prostaglandin-Endoperoxide Synthases; Receptors, Cytoplasmic and Nuclear; Thiazoles; Transcription Factors; Tumor Cells, Cultured