Stable over-expression of PPARβ/δ and PPARγ to examine receptor signaling in human HaCaT keratinocytes

Peroxisome proliferator-activated receptor-beta/delta (PPAR beta/delta) function and receptor cross-talk with other nuclear receptors, including PPAR gamma and retinoic acid receptors (RARs), was examined using stable human HaCaT keratinocyte cell lines over-expressing PPAR beta/delta or PPAR gamma. Enhanced ligand-induced expression of two known PPAR target genes, adipocyte differentiation-related protein (ADRP) and angiopoietin-like protein 4 (ANGPTL4), was found in HaCaT keratinocytes over-expressing PPAR beta/delta or PPAR gamma. Over-expression of PPAR beta/delta did not modulate the effect of a PPAR gamma agonist on up-regulation of ADRP or ANGPTL4 mRNA in HaCaT keratinocytes. All-trans retinoic acid (atRA) increased expression of a known RAR target gene, yet despite a high ratio of fatty acid binding protein 5 (FABP5) to cellular retinoic acid binding protein II, did not increase expression of ANGPTL4 or 3-phosphoinositide-dependent-protein kinase 1 (PDPK1), even in HaCaT keratinocytes expressing markedly higher levels of PPAR beta/delta. While PPAR beta/delta-dependent attenuation of staurosporine- or UVB-induced poly (ADP-ribose) polymerase (PARP) cleavage was not observed, PPAR beta/delta- and PPAR gamma-dependent repression of UVB-induced expression and secretion of inflammatory cytokines was found in HaCaT keratinocytes over-expressing PPAR beta/delta or PPAR gamma. These studies suggest that FABP5 does not transport atRA or GW0742 to PPAR beta/delta and promote anti-apoptotic activity by increasing expression of PDPK1, or that PPAR beta/delta interferes with PPAR gamma transcriptional activity. However, these studies demonstrate that stable over-expression of PPAR beta/delta or PPAR gamma significantly increases the efficacy of ligand activation and represses UVB-induced expression of tumor necrosis factor et (TNF alpha), interleukin 6 (IL6), or IL8 in HaCaT keratinocytes, thereby establishing an excellent model to study the functional role of these receptors in human keratinocytes. (C) 2011 Elsevier Inc. All rights reserved.