Troglitazone inhibits cell growth and induces apoptosis of B-cell acute lymphoblastic leukemia cells with t(14;18)

Peroxisome proliferator-activated receptor-gamma (PPARgamma), a member of the nuclear receptor superfamily, has been detected in several human leukemia cells. Recent studies reported that PPARgamma ligands inhibit cell proliferation and induce apoptosis in both normal and malignant B-lineage cells. We investigated the expression of PPARgamma and the effects of PPARgamma ligands on UTree-O2, Bay91 and 380, three B-cell acute lymphoblastic leukemia (B-ALL) cell lines with t(14;18), which show a poor prognosis, accompanying c-myc abnormality. Western blot analysis identified expression of PPARgamma protein and real-time PCR that of PPARgamma mRNA on the three cell lines. Troglitazone (TGZ), a synthetic PPARgamma ligand, inhibited cell growth in these cell lines in a dose-dependent manner, which was associated with G(1) cell cycle arrest and apoptosis. We also found this effect PPARgamma independent since PPARgamma antagonists failed to reverse this effect. We assessed the expression of c-myc, an apoptosis-regulatory gene, since c-myc abnormality was detected in most B-ALL cells with t(14;18). TGZ was found to dose-dependently downregulate the expression of c-myc mRNA and c-myc protein in the three cell lines. These results suggest that TGZ inhibits cell growth via induction of G(1) cell cycle arrest and apoptosis in these cell lines and that TGZ-induced apoptosis, at least in part, may be related to the downregulation of c-myc expression. Moreover, the downregulation of c-myc expression by TGZ may depend on a PPARgamma-independent mechanism. Further studies indicate that PPARgamma ligands may serve as a therapeutic agent in B-ALL with t(14;18).     

Peroxisome proliferator-activated receptor (PPAR) alpha and PPARbeta/delta,but not PPARgamma,modulate the expression of genes involved in cardiac lipid metabolism

Long-chain fatty acids (FA) coordinately induce the expression of a panel of genes involved in cellular FA metabolism in cardiac muscle cells, thereby promoting their own metabolism. These effects are likely to be mediated by peroxisome proliferator-activated receptors (PPARs). Whereas the significance of PPARalpha in FA-mediated expression has been demonstrated, the role of the PPARbeta/delta and PPARgamma isoforms in cardiac lipid metabolism is unknown. To explore the involvement of each of the PPAR isoforms, neonatal rat cardiomyocytes were exposed to FA or to ligands specific for either PPARalpha (Wy-14,643), PPARbeta/delta (L-165041, GW501516), or PPARgamma (ciglitazone and rosiglitazone). Their effect on FA oxidation rate, expression of metabolic genes, and muscle-type carnitine palmitoyltransferase-1 (MCPT-1) promoter activity was determined. Consistent with the PPAR isoform expression pattern, the FA oxidation rate increased in cardiomyocytes exposed to PPARalpha and PPARbeta/delta ligands, but not to PPARgamma ligands. Likewise, the FA-mediated expression of FA-handling proteins was mimicked by PPARalpha and PPARbeta/delta, but not by PPARgamma ligands. As expected, in embryonic rat heart-derived H9c2 cells, which only express PPARbeta/delta, the FA-induced expression of genes was mimicked by the PPARbeta/delta ligand only, indicating that FA also act as ligands for the PPARbeta/delta isoform. In cardiomyocytes, MCPT-1 promoter activity was unresponsive to PPARgamma ligands. However, addition of PPARalpha and PPARbeta/delta ligands dose-dependently induced promoter activity. Collectively, the present findings demonstrate that, next to PPARalpha, PPARbeta/delta, but not PPARgamma, plays a prominent role in the regulation of cardiac lipid metabolism, thereby warranting further research into the role of PPARbeta/delta in cardiac disease.     

Peroxisome proliferator-activated receptors in reproductive tissues: from gametogenesis to parturition

Peroxisome proliferator-activated receptors (PPARalpha, PPARbeta/delta and PPARgamma) are a family of nuclear receptors that are activated by binding of natural ligands, such as polyunsaturated fatty acids or by synthetic ligands. Synthetic molecules of the glitazone family, which bind to PPARgamma, are currently used to treat type II diabetes and also to attenuate the secondary clinical symptoms frequently associated with insulin resistance, including polycystic ovary syndrome (PCOS). PPARs are expressed in different compartments of the reproductive system (hypothalamus, pituitary, ovary, uterus and testis). Conservative functions of PPARs in mammalian species could be suggested through several in vivo and in vitro studies, especially in the ovary and during placental development. Several groups have described a strong expression of PPARgamma in ovarian granulosa cells, and glitazones modulate granulosa cell proliferation and steroidogenesis in vitro. All these recent data raise new questions about the biologic actions of PPARs in reproduction and their use in therapeutic treatments of fertility troubles such as PCOS or endometriosis. In this review, we first describe the roles of PPARs in different compartments of the reproductive axis (from male and female gametogenesis to parturition), with a focus on PPARgamma. Secondly, we discuss the possible molecular mechanisms underlying the effect of glitazones on PCOS. Like other 'insulin sensitizer' molecules, such as metformin, glitazones may in fact act directly on ovarian cells. Finally, we discuss the eventual actions of PPARs as mediators of environmental toxic substances for reproductive function.     

Platelet-derived growth factor receptor independent proliferation of human glioblastoma cells: selective tyrosine kinase inhibitors lack antiproliferative activity

Purpose: The aim of this study was to investigate the role of platelet-derived growth factor (PDGF) and PDGF receptors (PDGFRs) in the proliferation of human glioblastoma cells as a prerequisite for a new therapeutic approach to the treatment of malignant brain tumors with selective tyrosine kinase inhibitors such as imatinib. Methods and results: In the human glioblastoma cell lines U-87 MG, U-118 MG and U-373 MG different PDGF and PDGFR mRNAs were detected by RT-PCR, and the expression of the receptor proteins was demonstrated by immunostaining and flow cytometry. Moreover, functional activity of PDGFRs was demonstrated in PDGFRβ expressing glioblastoma cell variants by measuring the mobilization of intracellular Ca²⁺ upon PDGF-BB stimulation. However, addition of PDGF-BB to the serum-free culture medium had no stimulatory effect on cell proliferation. Furthermore, cell growth in serum-supplemented and serum-free medium was not affected by imatinib, leflunomide and AG-1296 at therapeutically relevant concentrations. Conclusion: Our results suggest that clinical antitumor effects of imatinib on glioblastoma, if any, are not mediated by the PDGFR.     

过氧化物酶体增生因子激活受体α和γ配体对游离脂肪酸介导的β细胞损害的干预作用

目的：研究过氧化物酶体增生因子激活受体α和γ(PPARα和PPARγ)配体对游离脂肪酸(FFA)介导的胰岛β细胞损害的干预作用。方法：应用PPARα配体氯贝丁酯及PPARγ配体曲格列酮(troglitazone,TGZ)和噻唑烷二酮(thiazolidinedione,TZD)处理大鼠胰岛素瘤细胞系β细胞(INS-1细胞),采用细胞活力和DNA片段梯度分析评价PPARα和PPARγ配体对FFA诱导的INS-1细胞损害的影响。结果：INS-1细胞与0．25～1mmol／L的FFA孵育24h后细胞活力下降,1mmol／L的FFA可诱导INS-1细胞发生凋亡。比较是否使用氯贝丁酯(100μmol／L)、TGZ(10μmol／L)和TZD(100μmol/L)处理β细胞的结果,发现这些配体可保护INS-1细胞免于FFA的细胞毒性(包括脂性凋亡)作用。结论：FFA可介导β细胞发生明显的脂毒性和脂性凋亡,应用PPARα和PPARγ配体可能具有保护β细胞免于FFA的细胞毒性的作用。     

PPARdelta modulates lipopolysaccharide-induced TNFalpha inflammation signaling in cultured cardiomyocytes

Peroxisome proliferator activated receptors (PPARs: PPARalpha, gamma and delta) regulate fatty acid metabolism, glucose homeostasis, cell proliferation, differentiation and inflammation. Tumor necrosis factor alpha (TNFalpha) is one of the important pathological factors in inflammatory responses during the pathological progression of myocardial ischemic/reperfusion and hypertrophy. Accumulating evidence shows that synthetic ligands of PPARalpha and PPARgamma suppress myocardial inflammatory responses, such as the production of TNFalpha, thus exerting beneficial effects in animals who had undergone ischemia/reperfusion injury or cardiac hypertrophy. However, it remains obscured if PPARdelta and its ligands exert any effect on the production of TNFalpha, thus influencing cardiac inflammatory responses. In this study, we investigated the effects of PPARdelta and its synthetic ligand GW0742 on TNFalpha production in cultured cardiomyocytes. Our studies indicate that a PPARdelta-selective ligand inhibited lipopolysaccharide (LPS)-induced TNFalpha production from cardiomyocytes. Adenoviral-mediated overexpression of PPARdelta substantially inhibited TNFalpha expression in cultured cardiomyocytes compared to controls, whereas overexpression of a PPARdelta mutant with ablated ligand binding domain did not show similar effect. Conversely, absence of PPARdelta in cardiomyocytes further exaggerated LPS-induced TNFalpha production. Moreover, activation of PPARdelta abrogated LPS-induced degradation of IkappaBs, thus suppressing LPS-induced nuclear factor kappaB (NF-kappaB) activities. Therefore, PPARdelta is an important determinant of TNFalpha expression via the NF-kappaB signaling pathway, thus serving as therapeutic targets to attenuate inflammation that are involved in cardiac pathological progression.     

Functional expression of bombesin receptor in most adult and pediatric human glioblastoma cell lines; role in mitogenesis and in stimulating the mitogen-activated protein kinase pathway

Functional bombesin receptors were identified in most human glioblastoma cell lines examined (85% of lines). Bombesin stimulated the release of intracellular Ca²⁺ in human adult (U-373MG, D-247MG, U-118MG, U-251MG, D-245MG, U-105MG, D-54MG, A-172MG, and D-270MG lines) and pediatric (SJ-S6 and SJ-G2 lines) glioblastoma cell lines. Stimulation of the glioblastoma cell line U-373MG with bombesin or gastrin-releasing peptide (GRP) induced mitogenesis, measured by [³H]thymidine incorporation into DNA, and stimulated the tyrosine phosphorylation of the mitogen-activated protein (MAP) kinases (Erk1 and Erk2). The stimulation of the MAP kinase phosphorylation in U-373MG cells was time- and peptide concentration-dependent. Both bombesin and GRP showed similar potencies in stimulation of intracellular Ca²⁺ release and activation of the MAP kinase pathway in U-373MG cells, whereas neuromedin B (NMB) peptide was less potent. Bombesin and GRP induced the release of cytosolic Ca²⁺ in a concentration-dependent manner. Because bombesin and GRP were more potent than NMB peptide in increasing the cytosolic Ca²⁺ levels in U-373MG cells, we concluded that the BB₂ subtype (also known as GRP-preferring receptor subtype) of the bombesin receptor is expressed in this cell line. The bombesin receptor antagonist ([Leu¹³-ψ(CH₂NH)Leu¹⁴]bombesin) blocked bombesin induced Ca²⁺ release and attenuated MAP kinase activation in U-373MG cells demonstrating that bombesin is acting through a receptor-dependent mechanism. This study indicates that functional bombesin receptors are widely expressed in human glioblastoma cell lines.     

Dual activation of PPARalpha and PPARgamma by mono-(2-ethylhexyl) phthalate in rat ovarian granulosa cells

Peroxisome proliferator-activated receptors (PPARs) are key regulators of lipid metabolism and cell differentiation. The plasticizer di-(2-ethylhexyl) phthalate is a peroxisome proliferator, and its active metabolite mono-(2-ethylhexyl) phthalate (MEHP) activates PPARalpha and PPARgamma in cell transactivation assays. MEHP is a female reproductive toxicant and decreases activity, mRNA, and protein levels of aromatase, the rate-limiting enzyme that converts testosterone to estradiol in ovarian granulosa cells. To test the hypothesis that MEHP suppresses aromatase through PPAR pathways, granulosa cells were cultured with MEHP (50 microM) or selective activators of PPARgamma or PPARalpha for 48 h and gene expression was analyzed by real time RT-PCR. Both PPARalpha and PPARgamma activators significantly decreased aromatase mRNA and estradiol production like MEHP. The PPARgamma-selective antagonist GR 259662 partially blocked the suppression of aromatase by MEHP, suggesting that MEHP acts through PPARgamma, but not exclusively. MEHP and the PPARalpha-selective agonist GW 327647 induced expression of 17beta-hydroxysteroid dehydrogenase IV, a known PPARalpha-regulated gene, and induction was maintained with addition of the PPARgamma-selective antagonist. PPARalpha-selective activation also induced expression of aryl hydrocarbon receptor (AhR), CYP1B1, and epoxide hydrolase in the granulosa cell. These data support a model in which MEHP activates both PPARalpha and PPARgamma to suppress aromatase and alter other genes related to metabolism and differentiation in the granulosa cell.