Involvement of PPARα and PPARγ in apoptosis and proliferation of human hepatocarcinoma HepG2 cells

Peroxisome proliferator‐activated receptors (PPARs) mediate the effects of various ligands, known as peroxisome proliferators, a heterogeneous class of compounds including industrial chemicals, pharmaceuticals, and biomolecules such as fatty acids and eicosanoids. Among peroxisome proliferators, fibrate derivatives are considered specific ligands for PPARα, whereas eicosanoids, such as PGJ2, for PPARγ. The study aimed to clarify the relation between PPARs and apoptosis or proliferation on the same type of cells, using clofibrate as specific ligand of PPARα and PGJ2 as specific ligand of PPARγ. The cells used were human hepatocarcinoma HepG2 cells. The results showed that PPARα protein content increased in HepG2 cells treated with clofibrate, causing apoptosis in a time‐ and concentration‐dependent way, as evidenced by the citofluorimetric assay and determination of BAD, myc and protein phosphatase 2A protein content. It also emerged that PPARγ increased in the same cells when treated with a specific ligand of this PPAR; in this case the increase of PPARγ did not cause an increase of apoptosis, but a time‐ and concentration‐dependent inhibition of cell proliferation, evidenced by decreased cell numbers and increased number of cells in the G0/G1 phase of the cycle. It may be concluded that PPARα is chiefly related to apoptosis and PPARγ to cell proliferation. Copyright © 2010 John Wiley & Sons, Ltd.     

PPARγ activation induces autophagy in breast cancer cells

It has been previously shown that PPARγ ligands induce apoptotic cell death in a variety of cancer cells. Given the evidence that these ligands have a receptor-independent function, we further examined the specific role of PPARγ activation in this biological process. Surprisingly, we failed to demonstrate that MDA-MB-231 breast cancer cells undergo apoptosis when treated with sub-saturation doses of troglitazone and rosiglitazone, which are synthetic PPARγ ligands. Acridine orange (AO) staining showed acidic vesicular formation within ligand-treated cells, indicative of autophagic activity. This was confirmed by autophagosome formation as indicated by redistribution of LC3, an autophagy-specific protein, and the appearance of double-membrane autophagic vacuoles by electron microscopy following exposure to ligand. To determine the mechanism by which PPARγ induces autophagy, we transduced primary mammary epithelial cells with a constitutively active mutant of PPARγ and screened gene expression associated with PPARγ activation by genome-wide array analysis. HIF1α and BNIP3 were among 42 genes up-regulated by active PPARγ. Activation of PPARγ induced HIF1α and BNIP3 protein and mRNA abundance. HIF1α knockdown by shRNA abolished the autophagosome formation induced by PPARγ activation. In summary, our data shows a specific induction of autophagy by PPARγ activation in breast cancer cells providing an understanding of distinct roles of PPARγ in tumorigenesis.     

The Pro12Ala Polymorphism of the PPARγ2 Gene Regulates Weight from Birth to Adulthood

Objective: The Pro12Ala polymorphism in exon B of peroxisome proliferator‐activated receptor γ 2 (PPARγ2) gene has been related to obesity, insulin resistance, and risk of type 2 diabetes. In this study, the effect of the Pro12Ala polymorphism on long‐term changes in weight and body composition was investigated. Research Methods and Procedures: The Pro12Ala polymorphism was genotyped in 311 subjects who participated in our previous population‐based study. In that study, weight at birth, 7 years, 20 years, and 41 years, and ponderal index at birth and BMI and waist circumference at 41 years were recorded. Results: The Ala12 allele of the PPARγ2 gene was associated with high ponderal index at birth (2.77 ± 0.27 kg/m³ in subjects with the Ala12Ala genotype, 2.79 ± 0.29 kg/m³ in subjects with the Pro12Ala genotype, and 2.63 ± 0.25 kg/m³ in subjects with the Pro12Pro genotype, p = 0.007, adjusted for gender) and weight at 7 years (p = 0.045) and tended to be associated with high birth weight (p = 0.094). Subjects with this allele gained less weight between 7 and 20 years (p = 0.043) and more weight between 20 and 41 years (p = 0.001) and ended up having higher waist circumference (p = 0.040) in adulthood than did subjects with the Pro12Pro genotype. Discussion: We conclude that the Pro12Ala polymorphism of the PPARγ2 gene regulates weight and body composition from utero to adulthood.     

Glucagon‐like peptide‐1 ameliorates cardiac lipotoxicity in diabetic cardiomyopathy via the PPARα pathway

Lipotoxicity cardiomyopathy is the result of excessive accumulation and oxidation of toxic lipids in the heart. It is a major threat to patients with diabetes. Glucagon‐like peptide‐1 (GLP‐1) has aroused considerable interest as a novel therapeutic target for diabetes mellitus because it stimulates insulin secretion. Here, we investigated the effects and mechanisms of the GLP‐1 analog exendin‐4 and the dipeptidyl peptidase‐4 inhibitor saxagliptin on cardiac lipid metabolism in diabetic mice (DM). The increased myocardial lipid accumulation, oxidative stress, apoptosis, and cardiac remodeling and dysfunction induced in DM by low streptozotocin doses and high‐fat diets were significantly reversed by exendin‐4 and saxagliptin treatments for 8 weeks. We found that exendin‐4 inhibited abnormal activation of the (PPARα)‐CD36 pathway by stimulating protein kinase A (PKA) but suppressing the Rho‐associated protein kinase (ROCK) pathway in DM hearts, palmitic acid (PA)‐treated rat h9c2 cardiomyocytes (CMs), and isolated adult mouse CMs. Cardioprotection in DM mediated by exendin‐4 was abolished by combination therapy with the PPARα agonist wy‐14643 but mimicked by PPARα gene deficiency. Therefore, the PPARα pathway accounted for the effects of exendin‐4. This conclusion was confirmed in cardiac‐restricted overexpression of PPARα mediated by adeno‐associated virus serotype‐9 containing a cardiac troponin T promoter. Our results provide the first direct evidence that GLP‐1 protects cardiac function by inhibiting the ROCK/PPARα pathway, thereby ameliorating lipotoxicity in diabetic cardiomyopathy.     

Chondroprotection of PPARα activation by WY14643 via autophagy involving Akt and ERK in LPS‐treated mouse chondrocytes and osteoarthritis model

Autophagy maintains cellular homoeostasis. The enhancement of autophagy in chondrocytes could prevent osteoarthritis (OA) progression in articular cartilage. Peroxisome proliferator‐activated receptor α (PPARα) activation may also protect articular chondrocytes against cartilage degradation in OA. However, whether the protective effect of activated PPARα is associated with autophagy induction in chondrocytes is not determined. In this study, we investigated the effect of PPARα activation by its agonist, WY14643, on the protein expression level of Aggrecan and ADAMTS5, and the protein expression level of autophagy biomarkers, including LC3B and P62, using Western blotting analysis in isolated mouse chondrocytes pre‐treated with lipopolysaccharides (LPS, mimicking OA chondrocytes) with or without the autophagy inhibitor chloroquine diphosphate salt. Furthermore, Akt and ERK phosphorylation was detected in LPS‐treated chondrocytes in response to WY14643. In addition, the effect of intra‐articularly injected WY14643 on articular cartilage in a mouse OA model established by the destabilization of the medial meniscus was assessed using the Osteoarthritis Research Society International (OARSI) histopathology assessment system, along with the detection of Aggrecan, ADAMTS5, LC3B and P62 protein levels using immunohistochemistry assay. The results indicated that PPARα activation by WY14643 promoted proteoglycan synthesis by autophagy enhancement in OA chondrocytes in vivo and in vitro concomitant with the elevation of Akt and ERK phosphorylation. Therefore, autophagy could contribute to the chondroprotection of PPARα activation by WY14643, with the implication that PPARα activation by WY14643 may be a potential approach for OA therapy.