Nonapoptotic cell death associated with S-phase arrest of prostate cancer cells via the peroxisome proliferator-activated receptor gamma ligand, 15-deoxy-delta12,14-prostaglandin J2.

15-Deoxy-delta12,14-prostaglandin J2 (15d-PGJ2) is a highly specific activator of the peroxisome proliferator-activated receptor gamma (PPAR-gamma). We investigated the effect of 15d-PGJ2 on three human prostate cancer cell lines, LNCaP, DU145, and PC-3. Western blotting demonstrated that PPAR-gamma1 is expressed predominantly in untreated prostate cancer cells. Treatment with 15d-PGJ2 caused an increase in the expression of PPAR-gamma2, whereas PPAR-gamma1 remained at basal levels. PPARs alpha and beta were not detected in these cells. Lack of lipid accumulation, increase in CCAAT/enhancer binding proteins (C/EBPs), or expression of aP2 mRNA indicated that adipocytic differentiation is not induced in these cells by 15d-PGJ2. 15d-PGJ2 and other PPAR-gamma activators induced cell death in all three cell lines at concentrations as low as 2.5 microM (similar to the Kd of PPAR-gamma for this ligand), coinciding with an accumulation of cells in the S-phase of the cell cycle. Activators for PPAR-alpha and beta did not induce cell death. Staining with trypan blue and propidium iodide suggested that, although the plasma membrane appears intact by electron microscopy, disturbances are evident as early as 2 h after treatment. Mitochondrial transmembrane potentials are significantly reduced by 15d-PGJ2 treatment. In addition, treatment with 15d-PGJ2 resulted in cytoplasmic changes, which are indicative of type 2 (autophagic), nonapoptotic programmed cell death.     

Peroxisome proliferator-activated receptors and cardiovascular remodeling

Peroxisome proliferator-activated receptors (PPARs) are nuclear receptors that heterodimerize with the retinoid X receptor and then modulate the function of many target genes. Three PPARs are known: alpha, beta/delta, and gamma. The better known are PPAR-alpha and PPAR-gamma, which may be activated by different synthetic agonists, although the endogenous ligands are unknown. PPAR-alpha is involved in fatty acid oxidation and expressed in the liver, kidney, and skeletal muscle, whereas PPAR-gamma is involved in fat cell differentiation, lipid storage, and insulin sensitivity. However, both have been shown to be present in variable amounts in cardiovascular tissues, including endothelium, smooth muscle cells, macrophages, and the heart. The activators of PPAR-alpha (fibrates) and PPAR-gamma (thiazolidinediones or glitazones) antagonized the actions of angiotensin II in vivo and in vitro and exerted cardiovascular antioxidant and anti-inflammatory effects. PPAR activators lowered blood pressure, induced favorable effects on the heart, and corrected vascular structure and endothelial dysfunction in several rodent models of hypertension. Activators of PPARs may become therapeutic agents useful in the prevention of cardiovascular disease beyond their effects on carbohydrate and lipid metabolism. Some side effects, such as weight gain, as well as documented aggravation of advanced heart failure through fluid retention by glitazones, may, however, limit their therapeutic application in prevention of cardiovascular disease.     

Effects of dietary mulberry, Korean red ginseng, and banaba on glucose homeostasis in relation to PPAR-alpha, PPAR-gamma, and LPL mRNA expressions

Despite lack of scientific evidences to support its therapeutic efficacy, the use of herbal supplements has significantly increased. The purpose of this study was to evaluate the effects of traditional anti-diabetic herbs on the progress of diabetes in db/db mice, a typical non-insulin-dependent model. Five different experimental diets were as follows: control diet, 0.5% mulberry leaf water extract diet, 0.5% Korean red ginseng diet, 0.5% banaba leaf water extract diet, and 0.5% combination diet (mulberry leaf water extract/Korean red ginseng/banaba leaf water extract, 1:1:1). Blood levels of glucose, insulin, HbA1c, and triglyceride were measured every 2 weeks. At 12 weeks of age, animals were sacrificed, and tissue mRNA levels of PPAR-alpha, PPAR-gamma, and LPL were determined. Results indicated that mulberry leaf water extract, Korean red ginseng, banaba leaf water extract, and the combination of above herbs effectively reduced blood glucose, insulin, TG, and percent HbA1c in study animals (p<0.05). We also observed that the increased expressions of liver PPAR-alpha mRNA and adipose tissue PPAR-gamma mRNA in animals fed diets supplemented with test herbs. The expression of liver LPL mRNA was also increased with experimental diets containing herbs. The efficacy was highest in animals fed the combination diet for all of the markers used. These results suggest that mulberry leaf water extract, Korean red ginseng, banaba leaf water extract, and the combination of these herbs fed at the level of 0.5% of the diet significantly increase insulin sensitivity, and improve hyperglycemia possibly through regulating PPAR-mediated lipid metabolism.     

The control of UCP1 is dissociated from that of PGC-1alpha or of mitochondriogenesis as revealed by a study using beta-less mouse brown adipocytes in culture

In rodent brown adipose tissue, the beta-adrenergic signaling is believed, by an action on PGC-1alpha, to control UCP1 expression and mitochondriogenesis. We addressed this hypothesis using beta(1)/beta(2)/beta(3)-adrenoceptor knockout (beta-less) brown adipocytes in primary culture. In these cells: (a) proliferation and differentiation into multilocular cells were normal; (b) UCP1 mRNA expression was dramatically decreased (by 93%), whereas PGC-1alpha and mtTFA mRNA expressions were not; (c) UCP1, PGC-1alpha and COX IV protein expressions were decreased by 97%, 62% and 22%, respectively. Altogether the data show a dissociation between the control of UCP1, which is mostly beta-adrenoceptor-dependent and that of PGC-1alpha and of mitochondriogenesis which are not.     

Rapid exercise-induced changes in PGC-1alpha mRNA and protein in human skeletal muscle

The mRNA of the nuclear coactivator peroxisome proliferator-activated receptor-gamma coactivator-1alpha (PGC-1alpha) increases during prolonged exercise and is influenced by carbohydrate availability. It is unknown if the increases in mRNA reflect the PGC-1alpha protein or if glycogen stores are an important regulator. Seven male subjects [23 +/- 1.3 yr old, maximum oxygen uptake (Vo(2 max)) 48.4 +/- 0.8 ml.kg(-1).min(-1)] exercised to exhaustion ( approximately 2 h) at 65% Vo(2 max) followed by ingestion of either a high-carbohydrate (HC) or low-carbohydrate (LC) diet (7 or 2.9 g.kg(-1).day(-1), respectively) for 52 h of recovery. Glycogen remained depressed in LC (P < 0.05) while returning to resting levels by 24 h in HC. PGC-1alpha mRNA increased both at exhaustion (3-fold) and 2 h later (6.2-fold) (P < 0.05) but returned to rest levels by 24 h. PGC-1alpha protein increased (P < 0.05) 23% at exhaustion and remained elevated for at least 24 h (P < 0.05). While there was no direct treatment effect (HC vs. LC) for PGC-1alpha mRNA or protein, there was a linear relationship between the changes in glycogen and those in PGC-1alpha protein during exercise and recovery (r = -0.68, P < 0.05). In contrast, PGC-1beta did not increase with exercise but rather decreased (P < 0.05) below rest level at 24 and 52 h, and the decrease was greater (P < 0.05) in LC. PGC-1alpha protein content increased in prolonged exercise and remained upregulated for 24 h, but this could not have been predicted by the changes in mRNA. The beta-isoform declined rather than increasing, and this was greater when glycogen was not resynthesized to rest levels.     

Changes in concentrations of follicular inhibin alpha and beta A subunit messenger ribonucleic acids and inhibin immunoactivity during preovulatory maturation in the pig.

Ovarian tissues were collected from 5 pigs on each of days 1, 3, 5, and 7 after withdrawal of an orally active progestin to determine changes in follicular inhibin subunit mRNAs during preovulatory maturation. Follicles (N = 146) were aspirated for fluid and homogenized in guanidinium isothiocyanate for RNA isolation. Follicular RNA and inhibin alpha and beta A subunit mRNA standards were dot-blotted, hybridized with [32P]-cDNA probes, and quantified by densitometry. Mean concentrations of alpha mRNA (pg/micrograms of RNA) increased (p < or = 0.05) by 140% as healthy follicles grew from medium (3-5 mm) to large (> 5 mm). Inhibin immunoactivity was greater (p < or = 0.05) in large than medium follicles. In contrast, mean concentrations of inhibin beta A subunit mRNA did not differ between healthy medium and large follicles. However, both alpha mRNA and beta A mRNA increased (p < or = 0.05) linearly as follicular diameter increased from 3 to 5 mm on Day 1 and from 3 to 9 mm on Day 3. On Day 5, alpha mRNA remained elevated, but was not significantly correlated with diameter. In contrast, beta A mRNA decreased linearly (p < 0.05) as diameter increased from 6 to 11 mm on Day 5. The molar ratio alpha mRNA to beta A mRNA was 20:1 in healthy, large follicles on Days 3 and 5. Mean concentration of alpha mRNA in large follicles decreased (p < 0.05) by 72% between Days 5 and 7, while beta A mRNA decreased to non-detectable levels on Day 7.(ABSTRACT TRUNCATED AT 250 WORDS)     

Signaling of mitochondrial biogenesis following oxidant injury

Mitochondrial dysfunction is a common consequence of ischemia-reperfusion and drug injuries. For example, sublethal injury of renal proximal tubular cells (RPTCs) with the model oxidant tert-butylhydroperoxide (TBHP) causes mitochondrial injury that recovers over the course of six days. Although regeneration of mitochondrial function is integral to cell repair and function, the signaling pathway of mitochondrial biogenesis following oxidant injury has not been examined. A 10-fold overexpression of the mitochondrial biogenesis regulator PPAR-gamma cofactor-1alpha (PGC-1alpha) in control RPTCs resulted in a 52% increase in mitochondrial number, a 27% increase in respiratory capacity, and a 30% increase in mitochondrial protein markers, demonstrating that PGC-1alpha mediates mitochondrial biogenesis in RPTCs. RPTCs sublethally injured with TBHP exhibited a 50% decrease in mitochondrial function and increased mitochondrial autophagy. Compared with the controls, PGC-1alpha levels increased 12-fold on days 1, 2, and 3 post-injury and returned to base line on day 4 as mitochondrial function returned. Inhibition p38 MAPK blocked the up-regulation of PGC-1alpha following oxidant injury, whereas inhibition of calcium-calmodulin-dependent protein kinase, calcineurin A, nitric-oxide synthase, and phosphoinositol 3-kinase had no effect. The epidermal growth factor receptor (EGFR) was activated following TBHP exposure, and the EGFR inhibitor AG1478 blocked the up-regulation of PGC-1alpha. Additional inhibitor studies revealed that the sequential activation of Src, p38 MAPK, EGFR, and p38 MAPK regulate the expression of PGC-1alpha following oxidant injury. In contrast, although Akt was activated following oxidant injury, it did not play a role in PGC-1alpha expression. We suggest that mitochondrial biogenesis following oxidant injury is mediated by p38 and EGFR activation of PGC-1alpha.     

Effects of alpha/beta-androstenediol immune regulating hormones on bone remodeling and apoptosis in osteoblasts

A large body of evidence suggests that the immune system directly impacts bone physiology. We tested whether immune regulating hormones (IRH), 17beta-androstenediol (beta-AED), 7beta,17beta-androstenetriol (beta-AET) or the 17alpha-androstenediol (alpha-AED), and 7alpha,17beta-androstenetriol (alpha-AET) metabolites could directly influence bone remodeling in vitro using human fetal osteoblasts (FOB-9). The impact on bone remodeling was examined by comparing the ratio of RANKL/OPG gene expression in response to AED and AET compounds. The alpha-AED was found to significantly increase in the ratio of RANKL/OPG gene expression and altering the morphology of RANKL stained FOB-9 cells. Cell viability was assessed using a Live/Dead assay. Again alpha-AED was unique in its ability to reduce the proportion of viable cells, and to induce mild apoptosis of FOB-9 cells. Treatment of FOB-9 cells with WY14643, an activator of PPAR-alpha and -gamma, also significantly elevated the percentage of dead cells. This increase was abolished by co-treatment with GW9962, a specific inhibitor of PPAR-gamma. Analysis of PPAR-gamma mRNA by Quantitative RT-PCR and its activation by DNA binding demonstrated that alpha-AED increased PPAR-gamma activation by 19%, while beta-AED conferred a 37% decrease in PPAR-gamma activation. In conclusion, alpha-AED opposed beta-AED by elevating a bone resorption scenario in osteoblast cells. The increase in RANKL/OPG is modulated by an activation of PPAR-gamma that in turn caused mild apoptosis of FOB-9 cells.     

PPARs在长链脂肪酸调控能量代谢中的作用

过氧化物酶体增殖物激活受体α(PPARα)主要在肝脏中表达,饥饿时能诱导β-氧化与生酮作用相关基因和成纤维化生长因子21(FGF21)表达,这在肝脏的饥饿代谢适应中起重要作用。饥饿与耐力训练时,骨骼肌中,过氧化物酶体增殖物激活受体δ(PPARδ)能诱导长链脂肪酸(LCFAs)氧化基因、叉头转录因子(FOXO1)及PPARδ共激活物α1(PGC1α)表达,其中,FOXO1和PGC1α能调控糖代谢与线粒体生物发生。脂肪细胞中,PPARγ能介导LCFAs调控能量代谢,活化的PPARγ能诱导与LCFAs转化为甘油三酯形式储存相关的基因表达。脂联素,PPARγ的另一靶基因,能维持脂肪细胞的胰岛素敏感性。本文就PPARs在LCFAs调控能量代谢中的作用做一综述。     

Rosiglitazone prevents the impairment of human islet function induced by fatty acids: evidence for a role of PPARgamma2 in the modulation of insulin secretion

Peroxisome proliferator-activated receptors (PPARs) are a subgroup of the superfamily of nuclear receptors, with three distinct main types: alpha, beta and gamma (subdivided into gamma(1) and gamma(2)). Recently, the presence of PPARgamma has been reported in human islets. Whether other PPAR types can be found in human islets, how islet PPARgamma mRNA expression is regulated by the metabolic milieu, their role in insulin secretion, and the effects of a PPARgamma agonist are not known. In this study, human pancreatic islets were prepared by collagenase digestion and density gradient purification from nonobese adult donors. The presence of PPAR mRNAs was assessed by RT-PCR, and the effect was evaluated of exposure for up to 24 h to either 22.2 mmol/l glucose and/or 0.25, 0.5, or 1.0 mmol/l long-chain fatty acid mixture (oleate to palmitate, 2:1). PPARbeta and, to a greater extent, total PPARgamma and PPARgamma(2) mRNAs were expressed in human islets, whereas PPARalpha mRNA was not detected. Compared with human adipose tissue, PPARgamma mRNA was expressed at lower levels in the islets, and PPARbeta at similar levels. The expression of PPARgamma(2) mRNA was not affected by exposure to 22.2 mmol/l glucose, whereas it decreased markedly and time-dependently after exposure to progressively higher free fatty acids (FFA). This latter effect was not affected by the concomitant presence of high glucose. Exposure to FFA caused inhibition of insulin mRNA expression, glucose-stimulated insulin release, and reduction of islet insulin content. The PPARgamma agonists rosiglitazone and 15-deoxy-Delta-(12,14)prostaglandin J(2) prevented the cytostatic effect of FFA as well as the FFA-induced changes of PPAR and insulin mRNA expression. In conclusion, this study shows that PPARgamma mRNA is expressed in human pancreatic islets, with predominance of PPARgamma(2); exposure to FFA downregulates PPARgamma(2) and insulin mRNA expression and inhibits glucose-stimulated insulin secretion; exposure to PPARgamma agonists can prevent these effects.     

Free fatty acids increase PGC-1alpha expression in isolated rat islets

PGC-1alpha mRNA and protein are elevated in islets from multiple animal models of diabetes. Overexpression of PGC-1alpha impairs glucose-stimulated insulin secretion (GSIS). However, it is not well known which metabolic events lead to upregulation of PGC-1alpha in the beta-cells under pathophysiological condition. In present study, we have investigated effects of chronic hyperlipidemia and hyperglycemia on PGC-1alpha mRNA expression in isolated rat islets. Isolated rat islets are chronically incubated with 0, 0.2 and 0.4 mM oleic acid/palmitic acid (free fatty acids, FFA) or 5.5 and 25 mM glucose for 72 h. FFA dose-dependently increases PGC-1alpha mRNA expression level in isolated islets. FFA also increases PGC-1alpha expression in mouse beta-cell-derived beta TC3 cell line. In contrast, 25 mM glucose decreases expression level of PGC-1alpha. Inhibition of PGC-1alpha by siRNA improves FFA-induced impairment of GSIS in islets. These data suggest that hyperlipidemia and hyperglycemia regulate PGC-1alpha expression in islets differently, and elevated PGC-1alpha by FFA plays an important role in chronic hyperlipidemia-induced beta-cell dysfunction.