PPARδ modulates oxLDL-induced apoptosis of vascular smooth muscle cells through a TGF-β/FAK signaling axis

The peroxisome proliferator-activated receptor delta (PPARδ) has been implicated in the modulation of vascular homeostasis. However, its roles in the apoptotic cell death of vascular smooth muscle cells (VSMCs) are poorly understood. Here, we demonstrate that PPARδ modulates oxidized low-density lipoprotein (oxLDL)-induced apoptosis of VSMCs through the transforming growth factor-β (TGF-β) and focal adhesion kinase (FAK) signaling pathways. Activation of PPARδ by GW501516, which is a specific ligand, significantly inhibited oxLDL-induced cell death and generation of reactive oxygen species in VSMCs. These inhibitory effects were significantly reversed in the presence of small interfering (si)RNA against PPARδ, or by blockade of the TGF-β or FAK signaling pathways. Furthermore, PPARδ-mediated recovery of FAK phosphorylation suppressed by oxLDL was reversed by SB431542, a specific ALK5 receptor inhibitor, indicating that a TGF-β/FAK signaling axis is involved in the action of PPARδ. Among the protein kinases activated by oxLDL, p38 mitogen-activated protein kinase was suppressed by ligand-activated PPARδ. In addition, oxLDL-induced expression and translocation of pro-apoptotic or anti-apoptotic factors were markedly affected in the presence of GW501516. Those effects were reversed by PPARδ siRNA, or inhibitors of TGF-β or FAK, which also suggests that PPARδ exerts its anti-apoptotic effect via a TGF-β/FAK signaling axis. Taken together, these findings indicate that PPARδ plays an important role in the pathophysiology of disease associated with apoptosis of VSMC, such as atherosclerosis and restanosis.     

Peroxisome proliferator-activated receptor β/δ agonism protects the kidney against ischemia/reperfusion injury in diabetic rats

Diabetes is an important risk factor for ischemic acute kidney injury, whose pharmacological treatment remains an unmet medical need. The peroxisome proliferator-activated receptor (PPAR) β/δ is highly expressed in the kidney, although its role has not yet been elucidated. Here, we used an in vivo model of renal ischemia/reperfusion (I/R) in streptozotocin-induced diabetic rats (i) to evaluate whether diabetes increases kidney susceptibility to I/R injury and (ii) to investigate the effects of PPARβ/δ activation. The degree of renal injury (1h ischemia/6h reperfusion) was significantly increased in diabetic rats compared with nondiabetic littermates. PPARβ/δ expression was increased after I/R, with the highest levels in diabetic rats. Administration of the selective PPARβ/δ agonist GW0742 attenuated the renal dysfunction, leukocyte infiltration, and formation of interleukin-6 and tumor necrosis factor-α. These effects were accompanied by an increased expression of the suppressor of cytokine signaling (SOCS)-3, which plays a critical role in the cytokine-activated signaling pathway. The beneficial effects of GW0742 were attenuated by the selective PPARβ/δ antagonist GSK0660. Thus, we report herein that PPARβ/δ activation protects the diabetic kidney against I/R injury by a mechanism that may involve changes in renal expression of SOCS-3 resulting in a reduced local inflammatory response.     

Wnt and Cyclooxygenase-2 Cross-talk Accelerates Adenoma Growth

Both Wnt and cyclooxygenase (COX-2) pathways are activated in most sporadic and familial colorectal cancers, especially in those with chromosomal instability. We have recently shown that a common target of both signaling pathways, the peroxisome proliferator-activated receptor (PPAR)-?, is involved in intestinal adenoma growth. Activation of this receptor by synthetic agonist (GW501516) or COX-2-derived prostaglandin E2 (PGE2) accelerates intestinal adenoma growth in ApcMin mice. Moreover, these effects are lost in ApcMin mice lacking PPAR?. These findings implicate PPAR? as a focal point of cross-talk between the Wnt and prostaglandin signaling pathways. Based on this work it looks as if PPAR? agonists currently in development for treatment of dyslipidemias and obesity may increase the risk of tumor formation in humans. By contrast, antagonists of PPAR? may provide a novel approach for prevention and treatment of colorectal cancer.     

PPARδ coordinates angiotensin II-induced senescence in vascular smooth muscle cells through PTEN-mediated inhibition of superoxide generation

Cellular senescence-associated changes in blood vessels have been implicated in aging and age-related cardiovascular disorders. Here, we demonstrate that peroxisome proliferator-activated receptor (PPAR) δ coordinates angiotensin (Ang) II-induced senescence of human vascular smooth muscle cells (VSMCs). Activation of PPARδ by GW501516, a specific ligand for PPARδ, significantly attenuated Ang II-induced generation of superoxides and suppressed senescence of VSMCs. A marked increase in the levels of p53 and p21 induced by Ang II was blunted by the treatment with GW501516. Ligand-activated PPARδ up-regulated expression of phosphatase and tensin homolog deleted on chromosome 10 (PTEN) and suppressed the phosphatidylinositol 3-kinase (PI3K)/Akt pathway. Knockdown of PTEN with siRNA abrogated the effects of PPARδ on cellular senescence, on PI3K/Akt signaling, and on generation of ROS in VSMCs treated with Ang II. Finally, administration of GW501516 to apoE-deficient mice treated with Ang II significantly reduced the number of senescent cells in the aorta, where up-regulation of PTEN with reduced levels of phosphorylated Akt and ROS was demonstrated. Thus, ligand-activated PPARδ confers resistance to Ang II-induced senescence by up-regulation of PTEN and ensuing modulation of the PI3K/Akt signaling to reduce ROS generation in vascular cells.     

The PPARδ ligand L-165041 inhibits VEGF-induced angiogenesis, but the antiangiogenic effect is not related to PPARδ

Peroxisome proliferator-activated receptor (PPAR)δ is known to be expressed ubiquitously and involved in lipid and glucose metabolism. Recent studies have demonstrated that PPARδ is expressed in endothelial cells (ECs) and plays a potential role in endothelial survival and proliferation. Although PPARα and PPARγ are well recognized to play anti-inflammatory, antiproliferative, and antiangiogenic roles in ECs, the general effect of PPARδ on angiogenesis in ECs remains unclear. Thus, we investigated the effect of the PPARδ ligand L-165041 on vascular EC proliferation and angiogenesis in vitro as well as in vivo. Our data show that L-165041 inhibited VEGF-induced cell proliferation and migration in human umbilical vein ECs (HUVECs). L-165041 also inhibited angiogenesis in the Matrigel plug assay and aortic ring assay. Flow cytometric analysis indicated that L-165041 reduced the number of ECs in the S phase and the expression levels of cell cycle regulatory proteins such as cyclin A, cyclin E, CDK2, and CDK4; phosphorylation of the retinoblastoma protein was suppressed by pretreatment with L-165041. We confirmed whether these antiangiogenic effects of L-165041 were PPARδ-dependent using GW501516 and PPARδ siRNA. GW501516 treatment did not inhibit VEGF-induced angiogenesis, and transfection of PPARδ siRNA did not reverse this antiangiogenic effect of L-165041, suggesting that the antiangiogenic effect of L-165041 on ECs is PPARδ-independent. Together, these data indicate that the PPARδ ligand L-165041 inhibits VEGF-stimulated angiogenesis by suppressing the cell cycle progression independently of PPARδ. This study highlights the therapeutic potential of L-165041 in the treatment of many disorders related to pathological angiogenesis.     

Sorting out the functional role(s) of peroxisome proliferator-activated receptor-β/δ (PPARβ/δ) in cell proliferation and cancer

Peroxisome proliferator-activated receptor-β/δ (PPARβ/δ) has many beneficial physiological functions ranging from enhancing fatty acid catabolism, improving insulin sensitivity, inhibiting inflammation and increasing oxidative myofibers allowing for improved athletic performance. Thus, given the potential for targeting PPARβ/δ for the prevention and/or treatment of diseases including diabetes, dyslipidemias, metabolic syndrome and cancer, it is critical to clarify the functional role of PPARβ/δ in cell proliferation and associated disorders such as cancer. However, there is considerable controversy whether PPARβ/δ stimulates or inhibits cell proliferation. This review summarizes the literature describing the influence of PPARβ/δ on cell proliferation, with an emphasis toward dissecting the data that give rise to opposing hypotheses. Suggestions are offered to standardize measurements associated with these studies so that interlaboratory comparisons can be accurately assessed.     

PPARδ activation inhibits angiotensin II induced cardiomyocyte hypertrophy by suppressing intracellular Ca2+ signaling pathway

Peroxisome proliferator‐activated receptors δ (PPARδ) is known to be expressed ubiquitously, and the predominant PPAR subtype of cardiac cells. However, relatively less is known regarding the role of PPARδ in cardiac cells except that PPARδ ligand treatment protects cardiac hypertrophy by inhibiting NF‐κB activation. Thus, in the present study, we examined the effect of selective PPARδ ligand L‐165041 on angiotensin II (AngII) induced cardiac hypertrophy and its underlying mechanism using cardiomyocyte. According to our data, L‐165041 (10 µM) inhibited AngII‐induced [³H] leucine incorporation, induction of the fetal gene atrial natriuretic factor (ANF) and increase of cardiomyocyte size. Previous studies have implicated the activation of focal adhesion kinase (FAK) in the progress of cardiomyocyte hypertrophy. L‐165041 pretreatment significantly inhibited AngII‐induced intracellular Ca²⁺ increase and subsequent phosphorylation of FAK. Further experiment using Ca²⁺ ionophore A23187 confirmed that Ca²⁺ induced FAK phosphorylation, and this was also blocked by L‐165041 pretreatment. In addition, overexpression of PPARδ using adenovirus significantly inhibited AngII‐induced intracellular Ca²⁺ increase and FAK expression, while PPARδ siRNA treatment abolished the effect of L‐165041. These data indicate that PPARδ ligand L‐165041 inhibits AngII induced cardiac hypertrophy by suppressing intracellular Ca²⁺/FAK/ERK signaling pathway in a PPARδ dependent mechanism. J. Cell. Biochem. 106: 823–834, 2009. © 2009 Wiley‐Liss, Inc.     

Agonist-induced activation releases peroxisome proliferator-activated receptor β/δ from its inhibition by palmitate-induced nuclear factor-κB in skeletal muscle cells

The mechanisms by which elevated levels of free fatty acids cause insulin resistance are not well understood, but there is a strong correlation between insulin resistance and intramyocellular lipid accumulation in skeletal muscle. In addition, accumulating evidence suggests a link between inflammation and type 2 diabetes. The aim of this work was to study whether the exposure of skeletal muscle cells to palmitate affected peroxisome proliferator-activated receptor (PPAR) β/δ activity. Here, we report that exposure of C2C12 skeletal muscle cells to 0.75 mM palmitate reduced (74%, P<0.01) the mRNA levels of the PPARβ/δ-target gene pyruvatedehydrogenase kinase 4 (PDK-4), which is involved in fatty acid utilization. This reduction was not observed in the presence of the PPARβ/δ agonist L-165041. This drug prevented palmitate-induced nuclear factor (NF)-κB activation. Increased NF-κB activity after palmitate exposure was associated with enhanced protein–protein interaction between PPARβ/δ and p65. Interestingly, treatment with the PPARβ/δ agonist L-165041 completely abolished this interaction. These results indicate that palmitate may reduce fatty acid utilization in skeletal muscle cells by reducing PPARβ/δ signaling through increased NF-κB activity.     

The role of peroxisome proliferator-activated receptor-beta/delta in epidermal growth factor-induced HaCaT cell proliferation

Epidermal growth factor (EGF) has been shown to be a potent mitogen for epidermal cells both in vitro and in vivo, thus contributing to the development of an organism. It has recently become clear that peroxisome proliferator-activated receptor-β/δ (PPARβ/δ) expression and activation is involved in the cell proliferation. However, little is known about the role of PPARβ/δ in EGF-induced proliferation of HaCaT keratinocytes. In this study, HaCaT cells were cultured in the presence and absence of EGF and we identified that EGF induced an increase of PPARβ/δ mRNA and protein level expression in time-dependent and dose-dependent manner, and AG1487, an EGF receptor (EGFR) special inhibitor, caused attenuation of PPARβ/δ protein expression. Electrophoretic mobility shift assay (EMSA) revealed that EGF significantly increased PPARβ/δ binding activity in HaCaT keratinocytes. Antisense phosphorothioate oligonucleotides (asODNs) against PPARβ/δ caused selectively inhibition of PPARβ/δ protein content induced by EGF and significantly attenuated EGF-mediated cell proliferation. Treatment of the cells with L165041, a specific synthetic ligand for PPARβ/δ, significantly enhanced EGF-mediated cell proliferation. Finally, c-Jun ablation inhibited PPARβ/δ up-regulation induced by EGF, and chromatin immunoprecipitation (ChIP) showed that c-Jun bound to the PPARβ/δ promoter and the binding increased in EGF-stimulated cells. These results demonstrate that EGF induces PPARβ/δ expression in a c-Jun-dependent manner and PPARβ/δ plays a vital role in EGF-stimulated proliferation of HaCaT cells.     

Role of the transforming growth factor-β signaling pathway in the pathogenesis of colorectal cancer

The transforming growth factor-β (TGF-β) signaling pathway plays an important role in cancer cell proliferation, growth, metastasis, and apoptosis. It has been shown that TGF-β acts as a tumor suppressor in the early stages of the disease, and as a tumor promoter in its late stages. Mutations in the TGF-β signaling components, the TGF-β receptors and cytoplasmic signaling transducers, are frequently observed in colorectal carcinomas. Exploiting specific TGF-β receptor agonist and antagonist with antitumor properties may be a way of controlling cancer progression. This review summarizes the regulatory role of TGF-β signaling in the pathogenesis of colorectal cancer.