PPAR-γ Ligands Repress TGFβ-Induced Myofibroblast Differentiation by Targeting the PI3K/Akt Pathway: Implications for Therapy of Fibrosis

Transforming growth factor beta (TGFβ) induced differentiation of human lung fibroblasts to myofibroblasts is a key event in the pathogenesis of pulmonary fibrosis. Although the typical TGFβ signaling pathway involves the Smad family of transcription factors, we have previously reported that peroxisome proliferator-activated receptor-γ (PPAR-γ) ligands inhibit TGFβ-mediated differentiation of human lung fibroblasts to myofibroblasts via a Smad-independent pathway. TGFβ also activates the phosphatidylinositol 3 kinase/protein kinase B (PI3K/Akt) pathway leading to phosphorylation of AktS473. Here, we report that PPAR-γ ligands, 2-cyano-3,12-dioxooleana-1,9-dien-28-oic acid (CDDO) and 15-deoxy-(12,14)-15d-prostaglandin J2 (15d-PGJ2), inhibit human myofibroblast differentiation of normal and idiopathic pulmonary fibrotic (IPF) fibroblasts, by blocking Akt phosphorylation at Ser473 by a PPAR-γ-independent mechanism. The PI3K inhibitor LY294002 and a dominant-negative inactive kinase-domain mutant of Akt both inhibited TGFβ-stimulated myofibroblast differentiation, as determined by Western blotting for α-smooth muscle actin and calponin. Prostaglandin A1 (PGA1), a structural analogue of 15d-PGJ2 with an electrophilic center, also reduced TGFβ-driven phosphorylation of Akt, while CAY10410, another analogue that lacks an electrophilic center, did not; implying that the activity of 15d-PGJ2 and CDDO is dependent on their electrophilic properties. PPAR-γ ligands inhibited TGFβ-induced Akt phosphorylation via both post-translational and post-transcriptional mechanisms. This inhibition is independent of MAPK-p38 and PTEN but is dependent on TGFβ-induced phosphorylation of FAK, a kinase that acts upstream of Akt. Thus, PPAR-γ ligands inhibit TGFβ signaling by affecting two pro-survival pathways that culminate in myofibroblast differentiation. Further studies of PPAR-γ ligands and small electrophilic molecules may lead to a new generation of anti-fibrotic therapeutics

Stabilization of β-catenin by a Wnt-independent mechanism regulates cardiomyocyte growth

β-Catenin is a transcriptional activator that regulates embryonic development as part of the Wnt pathway and also plays a role in tumorigenesis. The mechanisms leading to Wnt-induced stabilization of β-catenin, which results in its translocation to the nucleus and activation of transcription, have been an area of intense interest. However, it is not clear whether stimuli other than Wnts can lead to important stabilization of β-catenin and, if so, what factors mediate that stabilization and what biologic processes might be regulated. Herein we report that β-catenin is stabilized in cardiomyocytes after these cells have been exposed to hypertrophic stimuli in culture or in vivo. The mechanism by which β-catenin is stabilized is distinctly different from that used by Wnt signaling. Although, as with Wnt signaling, inhibition of glycogen synthase kinase-3 remains central to hypertrophic stimulus-induced stabilization of β-catenin, the mechanism by which this occurs involves the recruitment of activated PKB to the β-catenin-degradation complex. PKB stabilizes the complex and phosphorylates glycogen synthase kinase-3 within the complex, inhibiting its activity directed at β-catenin. Finally, we demonstrate via adenoviral gene transfer that β-catenin is both sufficient to induce growth in cardiomyocytes in culture and in vivo and necessary for hypertrophic stimulus-induced growth. Thus, in these terminally differentiated cells, β-catenin is stabilized by hypertrophic stimuli acting via heterotrimeric G protein-coupled receptors. The stabilization occurs via a unique Wnt-independent mechanism and results in cellular growth