Tsuyoshi Goto

Dual action of isoprenols from herbal medicines on both PPARγ and PPARα in 3T3‐L1 adipocytes and HepG2 hepatocytes

Several herbal medicines improve hyperlipidemia, diabetes and cardiovascular diseases. However, the molecular mechanism underlying this improvement has not yet been clarified. In this study, we found that several isoprenols, common components of herbal plants, activate human peroxisome proliferator‐activated receptors (PPARs) as determined using the novel GAL4 ligand‐binding domain chimera assay system with coactivator coexpression. Farnesol and geranylgeraniol that are typical isoprenols in herbs and fruits activated not only PPARγ but also PPARα as determined using the chimera assay system. These compounds also activated full‐length human PPARγ and PPARα in CV1 cells. Moreover, these isoprenols upregulated the expression of some lipid metabolic target genes of PPARγ and PPARα in 3T3‐L1 adipocytes and HepG2 hepatocytes, respectively. These results suggest that herbal medicines containing isoprenols with dual action on both PPARγ and PPARα can be of interest for the amelioration of lipid metabolic disorders associated with diabetes.

Overexpression and Ribozyme-mediated Targeting of Transcriptional Coactivators CREB-binding Protein and p300 Revealed Their Indispensable Roles in Adipocyte Differentiation through the Regulation of Peroxisome Proliferator-activated Receptor γ

The cAMP-response element-binding protein-binding protein (CBP) and p300 are common coactivators for several transcriptional factors. It has been reported that both CBP and p300 are significant for the activation of peroxisome proliferator-activated receptor γ (PPARγ), which is a crucial nuclear receptor in adipogenesis. However, it remains unclear whether CBP and/or p300 is physiologically essential to the activation of PPARγ in adipocytes and adipocyte differentiation. In this study, we investigated the physiological significance of CBP/p300 in NIH3T3 cells transiently expressing PPARγ and CBP and in 3T3-L1 preadipocytes stably expressing CBP- or p300-specific ribozymes. In PPARγ-transfected NIH3T3 cells, induction of expression of PPARγ target genes such as adipocyte fatty acid-binding protein (aP2) and lipoprotein lipase (LPL) by adding thiazolidinedione was enhanced, depending on the amount of a CBP expression plasmid transfected. Expression of aP2 and LPL genes, as well as glycerol-3-phosphate dehydrogenase activity and triacylglyceride accumulation after adipogenic induction, was largely suppressed in 3T3-L1 adipocytes expressing either the CBP- or p300-specific active ribozyme, but not in inactive ribozyme-expressing cells. These data suggest that both CBP and p300 are indispensable for the full activation of PPARγ and adipocyte differentiation and that CBP and p300 do not mutually complement in the process.

Activation of peroxisome proliferator-activated receptor-alpha stimulates both differentiation and fatty acid oxidation in adipocytes.

Peroxisome proliferator-activated receptor-α (PPARα) is a dietary lipid sensor, whose activation results in hypolipidemic effects. In this study, we investigated whether PPARα activation affects energy metabolism in white adipose tissue (WAT). Activation of PPARα by its agonist (bezafibrate) markedly reduced adiposity in KK mice fed a high-fat diet. In 3T3-L1 adipocytes, addition of GW7647, a highly specific PPARα agonist, during adipocyte differentiation enhanced glycerol-3-phosphate dehydrogenase activity, insulin-stimulated glucose uptake, and adipogenic gene expression. However, triglyceride accumulation was not increased by PPARα activation. PPARα activation induced expression of target genes involved in FA oxidation and stimulated FA oxidation. In WAT of KK mice treated with bezafibrate, both adipogenic and FA oxidation-related genes were significantly upregulated. These changes in mRNA expression were not observed in PPARα-deficient mice. Bezafibrate treatment enhanced FA oxidation in isolated adipocytes, suppressing adipocyte hypertrophy. Chromatin immunoprecipitation (ChIP) assay revealed that PPARα was recruited to promoter regions of both adipogenic and FA oxidation-related genes in the presence of GW7647 in 3T3-L1 adipocytes. These findings indicate that the activation of PPARα affects energy metabolism in adipocytes, and PPARα activation in WAT may contribute to the clinical effects of fibrate drugs.

Triiodothyronine induces UCP-1 expression and mitochondrial biogenesis in human adipocytes

Uncoupling protein (UCP)-1 expressed in brown adipose tissue plays an important role in thermogenesis. Recent data suggest that brown-like adipocytes in white adipose tissue (WAT) and skeletal muscle play a crucial role in the regulation of body weight. Understanding of the mechanism underlying the increase in UCP-1 expression level in these organs should, therefore, provide an approach to managing obesity. The thyroid hormone (TH) has profound effects on mitochondrial biogenesis and promotes the mRNA expression of UCP in skeletal muscle and brown adipose tissue. However, the action of TH on the induction of brown-like adipocytes in WAT has not been elucidated. Thus we investigate whether TH could regulate UCP-1 expression in WAT using multipotent cells isolated from human adipose tissue. In this study, triiodothyronine (T(3)) treatment induced UCP-1 expression and mitochondrial biogenesis, accompanied by the induction of the CCAAT/enhancer binding protein, peroxisome proliferator-activated receptor-γ coactivator-1α, and nuclear respiratory factor-1 in differentiated human multipotent adipose-derived stem cells. The effects of T(3) on UCP-1 induction were dependent on TH receptor-β. Moreover, T(3) treatment increased oxygen consumption rate. These findings indicate that T(3) is an active modulator, which induces energy utilization in white adipocytes through the regulation of UCP-1 expression and mitochondrial biogenesis. Our findings provide evidence that T(3) serves as a bipotential mediator of mitochondrial biogenesis.

6-Shogaol and 6-gingerol, the pungent of ginger, inhibit TNF-α mediated downregulation of adiponectin expression via different mechanisms in 3T3-L1 adipocytes

In this study, we demonstrated that the two ginger-derived components have a potent and unique pharmacological function in 3T3-L1 adipocytes via different mechanisms. Both pretreatment of 6-shogaol (6S) and 6-gingerol (6G) significantly inhibited the tumor necrosis factor-alpha (TNF-alpha) mediated downregulation of the adiponectin expression in 3T3-L1 adipocytes. Our study demonstrate that (1) 6S functions as a PPARgamma agonist with its inhibitory mechanism due to the PPARgamma transactivation, and (2) 6G is not a PPARgamma agonist, but it is an effective inhibitor of TNF-alpha induced c-Jun-NH(2)-terminal kinase signaling activation and thus, its inhibitory mechanism is due to this inhibitory effect.

Functional Food Targeting the Regulation of Obesity-Induced Inflammatory Responses and Pathologies

Obesity is associated with a low-grade systemic chronic inflammatory state, characterized by the abnormal production of pro- and anti-inflammatory adipocytokines. It has been found that immune cells such as macrophages can infiltrate adipose tissue and are responsible for the majority of inflammatory cytokine production. Obesity-induced inflammation is considered a potential mechanism linking obesity to its related pathologies, such as insulin resistance, cardiovascular diseases, type-2 diabetes, and some immune disorders. Therefore, targeting obesity-related inflammatory components may be a useful strategy to prevent or ameliorate the development of such obesity-related diseases. It has been shown that several food components can modulate inflammatory responses in adipose tissue via various mechanisms, some of which are dependent on peroxisome proliferator-activated receptor γ (PPARγ), whereas others are independent on PPARγ, by attenuating signals of nuclear factor-κB (NF-κB) and/or c-Jun amino-terminal kinase (JNK). In this review, we introduce the beneficial effects of anti-inflammatory phytochemicals that can help prevent obesity-induced inflammatory responses and pathologies.

Capsaicin inhibits the production of tumor necrosis factor α by LPS‐stimulated murine macrophages, RAW 264.7: a PPARγ ligand‐like action as a novel mechanism

Capsaicin, a major ingredient of hot pepper, is considered to exhibit anti‐inflammatory properties. Our previous study demonstrated that capsaicin inhibited the production of pro‐inflammatory mediators through NF‐κB inactivation in LPS‐stimulated macrophages. In order to further clarify the mechanism underlying the anti‐inflammatory action of capsaicin, we investigated whether capsaicin alters PPARγ activity, which regulates the production of the pro‐inflammatory cytokine TNFα. Capsaicin significantly inhibited the production of TNFα by macrophages in a dose‐dependent manner. Simultaneous exposure of the cells to capsaicin and PPARγ agonist troglitazone or RXR agonist LG100268 resulted in stronger inhibition of TNFα production compared to the cells treated with either capsaicin, troglitazone, or LG100268 alone. Luciferase reporter assay revealed that capsaicin induced GAL4/PPARγ chimera and full length PPARγ (PPRE) transactivations in a dose‐dependent manner. Furthermore, a specific PPARγ antagonist T0070907 abrogated the inhibitory action of capsaicin on LPS‐induced TNFα production by RAW 264.7 cells, indicating that capsaicin acts like a ligand for PPARγ. Our data demonstrate for the first time that the anti‐inflammatory action of capsaicin may be mediated by PPARγ activation in LPS‐stimulated RAW 264.7 cells.

Abietic acid activates peroxisome proliferator-activated receptor-γ (PPARγ) in RAW264.7 macrophages and 3T3-L1 adipocytes to regulate gene expression involved in inflammation and lipid metabolism

Abietic acid is one of the terpenoids, which are multifunctional natural compounds. It has been reported that abietic acid suppresses effects on inflammation. However, the mechanism underlying the anti‐inflammatory effects remains unclear. The present work indicates that abietic acid suppresses the protein expression of tumor necrosis factor‐α and cyclooxygenase 2, which are involved in inflammation, in lipopolysaccharide‐stimulated macrophages. Moreover, this effect resembles that of thiazolidinedione, a synthetic peroxisome proliferator‐activated receptor‐γ (PPARγ) ligand. Indeed, abietic acid activates PPARγ in luciferase reporter assays. The activity of abietic acid induces PPARγ target gene expression in RAW264.7 macrophages and 3T3‐L1 adipocytes. These data indicate that abietic acid is a PPARγ ligand and that its anti‐inflammatory effect is partly due to the activation of PPARγ in stimulated macrophages. The present work suggests a novel possibility that abietic acid, a naturally occurring compound, can be used not only for anti‐inflammation but also for regulating lipid metabolism and atherosclerosis.

Bixin regulates mRNA expression involved in adipogenesis and enhances insulin sensitivity in 3T3-L1 adipocytes through PPARγ activation

Insulin resistance is partly due to suppression of insulin-induced glucose uptake into adipocytes. The uptake is dependent on adipocyte differentiation, which is controlled at mRNA transcription level. The peroxisome proliferator-activated receptor (PPAR), a ligand-regulated nuclear receptor, is involved in the differentiation. Many food-derived compounds serve as ligands to activate or inactivate PPAR. In this study, we demonstrated that bixin and norbixin (annatto extracts) activate PPARgamma by luciferase reporter assay using GAL4-PPAR chimera proteins. To examine the effects of bixin on adipocytes, 3T3-L1 adipocytes were treated with bixin or norbixin. The treatment induced mRNA expression of PPARgamma target genes such as adipocyte-specific fatty acid-binding protein (aP2), lipoprotein lipase (LPL), and adiponectin in differentiated 3T3-L1 adipocytes and enhanced insulin-dependent glucose uptake. The observations indicate that bixin acts as an agonist of PPARgamma and enhances insulin sensitivity in 3T3-L1 adipocytes, suggesting that bixin is a valuable food-derived compound as a PPAR ligand to regulate lipid metabolism and to ameliorate metabolic syndrome.

Tiliroside, a glycosidic flavonoid, ameliorates obesity-induced metabolic disorders via activation of adiponectin signaling followed by enhancement of fatty acid oxidation in liver and skeletal muscle in obese–diabetic mice

Tiliroside contained in several dietary plants, such as rose hips, strawberry and raspberry, is a glycosidic flavonoid and possesses anti-inflammatory, antioxidant, anticarcinogenic and hepatoprotective activities. Recently, it has been reported that the administration of tiliroside significantly inhibited body weight gain and visceral fat accumulation in normal mice. In this study, we evaluated the effects of tiliroside on obesity-induced metabolic disorders in obese-diabetic KK-A(y) mice. In KK-A(y) mice, the administration of tiliroside (100 mg/kg body weight/day) for 21 days failed to suppress body weight gain and visceral fat accumulation. Although tiliroside did not affect oxygen consumption, respiratory exchange ratio was significantly decreased in mice treated with tiliroside. In the analysis of metabolic characteristics, it was shown that plasma insulin, free fatty acid and triglyceride levels were decreased, and plasma adiponectin levels were increased in mice administered tiliroside. The messenger RNA expression levels of hepatic adiponectin receptor (AdipoR)-1 and AdipoR2 and skeletal muscular AdipoR1 were up-regulated by tiliroside treatment. Furthermore, it was indicated that tiliroside treatment activated AMP-activated protein kinase in both the liver and skeletal muscle and peroxisome proliferator-activated receptor α in the liver. Finally, tiliroside inhibited obesity-induced hepatic and muscular triglyceride accumulation. These findings suggest that tiliroside enhances fatty acid oxidation via the enhancement adiponectin signaling associated with the activation of both AMP-activated protein kinase and peroxisome proliferator-activated receptor α and ameliorates obesity-induced metabolic disorders, such as hyperinsulinemia and hyperlipidemia, although it does not suppress body weight gain and visceral fat accumulation in obese-diabetic model mice.