15-deoxyprostaglandin-j2 and ciglitazone

Monocytes/macrophages link the innate and adaptive immune systems, and in inflammatory disorders their activation leads to tissue damage. 15-Deoxy-Δ(12,14)-prostaglandin J(2) (15d-PGJ(2)), a natural peroxisome proliferator-activated receptor gamma (PPARγ) ligand, has garnered much interest because it possesses anti-inflammatory properties in a number of experimental models. However, whether it regulates monocytes/macrophage pathophysiology is still unknown. This study was designed to examine the effects of 15d-PGJ(2) on the phagocytosis, proliferation and inflammatory cytokines generation in mouse monocyte/macrophage cell line RAW264.7 and J774A.1 cells upon lipopolysaccharide challenge. Our results showed that 15d-PGJ(2) inhibited the phagocytic activity and cell proliferation in a dose-dependent manner, and suppressed proinflammatory cytokines expression, such as tumor necrosis factor-α, transforming growth factor-β1, interleukin-6, and monocyte chemotactic protein-1. These effects were independent of PPARγ, because PPARγ agonist (troglitazone or ciglitazone) and PPARγ antagonist (GW9662) did not affect these activities mentioned above in cells. Treatment of 15d-PGJ(2) also did not modulate expression and distribution of PPARγ. However, these effects of 15d-PGJ(2) were abrogated by antioxidant N-acetylcysteine. Moreover, treatment of 15d-PGJ(2) induced a significant increase in reactive oxygen species production in RAW264.7 and J774A.1 cells. In conclusion, 15d-PGJ(2) attenuates the biological activities of mouse monocyte/macrophage cell line cells involving oxidative stress, independently of PPARγ. These data further underline the anti-inflammation potential of 15d-PGJ(2).

Topics: Acetylcysteine; Anilides; Animals; Cell Line; Cell Proliferation; Chemokine CCL2; Chromans; Interleukin-6; Lipopolysaccharides; Macrophages; Mice; Monocytes; Oxidative Stress; Phagocytosis; PPAR gamma; Prostaglandin D2; Reactive Oxygen Species; Thiazolidinediones; Troglitazone; Tumor Necrosis Factor-alpha

Peroxisome proliferator-activated receptors (PPARs) have been originally thought to be restricted to lipid metabolism or glucose homeostasis. Recently, evidence is growing that PPARγ ligands have inhibitory effects on tumor growth. To shed light on the potential therapeutic effects on melanoma we tested a panel of PPAR agonists on their ability to block tumor proliferation in vitro. Whereas ciglitazone, troglitazone and WY14643 showed moderate effects on proliferation, 15d-PGJ2 displayed profound anti-tumor activity on four different melanoma cell lines tested. Additionally, 15d-PGJ2 inhibited proliferation of tumor-associated fibroblasts and tube formation of endothelial cells. 15d-PGJ2 induced the tumor suppressor gene p21, a G(2)/M arrest and inhibited tumor cell migration. Shot gun proteome analysis in addition to 2D-gel electrophoresis and immunoprecipitation of A375 melanoma cells suggested that 15d-PGJ2 might exert its effects via modification and/or downregulation of Hsp-90 (heat shock protein 90) and several chaperones. Applying the recently established CPL/MUW database with a panel of defined classification signatures, we demonstrated a regulation of proteins involved in metastasis, transport or protein synthesis including paxillin, angio-associated migratory cell protein or matrix metalloproteinase-2 as confirmed by zymography. Our data revealed for the first time a profound effect of the single compound 15d-PGJ2 on melanoma cells in addition to the tumor-associated microenvironment suggesting synergistic therapeutic efficiency.

Topics: Antineoplastic Agents; Cell Cycle; Cell Line, Tumor; Cell Movement; Cell Proliferation; Chromans; Humans; Melanoma; PPAR gamma; Prostaglandin D2; Proteome; Pyrimidines; Thiazolidinediones; Troglitazone

Neural stem cells (NSCs) are a small population of resident cells that can grow, migrate and differentiate into neuro-glial cells in the central nervous system (CNS). Peroxisome proliferator-activated receptor gamma (PPARγ) is a nuclear receptor transcription factor that regulates cell growth and differentiation. In this study we analyzed the influence of PPARγ agonists on neural stem cell growth and differentiation in culture. We found that in vitro culture of mouse NSCs in neurobasal medium with B27 in the presence of epidermal growth factor (EGF) and basic fibroblast growth factor (bFGF) induced their growth and expansion as neurospheres. Addition of all-trans retinoic acid (ATRA) and PPARγ agonist ciglitazone or 15-Deoxy-Δ(12,14)-Prostaglandin J(2) (15d-PGJ2) resulted in a dose-dependent inhibition of cell viability and proliferation of NSCs in culture. Interestingly, NSCs cultured with PPARγ agonists, but not ATRA, showed significant increase in oligodendrocyte precursor-specific O4 and NG2 reactivity with a reduction in NSC marker nestin, in 3-7 days. In vitro treatment with PPARγ agonists and ATRA also induced modest increase in the expression of neuronal β-III tubulin and astrocyte-specific GFAP in NSCs in 3-7 days. Further analyses showed that PPARγ agonists and ATRA induced significant alterations in the expression of many stemness and differentiation genes associated with neuro-glial differentiation in NSCs. These findings highlight the influence of PPARγ agonists in promoting neuro-glial differentiation of NSCs and its significance in the treatment of neurodegenerative diseases.

Topics: Animals; Antigens; Cell Differentiation; Cell Proliferation; Cell Survival; Cells, Cultured; Dose-Response Relationship, Drug; Epidermal Growth Factor; Fibroblast Growth Factor 2; Gene Expression; Glial Fibrillary Acidic Protein; Intermediate Filament Proteins; Mice; Nerve Tissue Proteins; Nestin; Neural Stem Cells; Oligodendroglia; PPAR gamma; Prostaglandin D2; Proteoglycans; Thiazolidinediones; Tretinoin; Tubulin

Ovarian cancer accounts for the highest mortality among all gynecological cancers, mainly due to the fast developing chemoresistance. The death ligand TRAIL induces apoptosis and is able to sensitize tumor cells to cytostatic drugs without affecting physiological tissue. Combined treatment of TRAIL and the antidiabetic acting PPARγ ligands was shown to induce apoptosis synergistically in different ovarian cancer cell lines.. To investigate feasible TRAIL-dependent inhibition of proliferation and induction of apoptosis in chemoresistant ovarian cancer cell lines, the drug- and TRAIL-sensitive HEY cell line was utilized to develop subclones with selective resistance against cisplatin, etoposide, docetaxel, paclitaxel, gemcitabine and pemetrexed, as well as against TRAIL as control cell line. Expression of the key factors of the TRAIL signaling pathway, TRAIL receptors 1-4, caspase-8, FLIP and XIAP, was analyzed before and after TRAIL treatment by immunoblotting.. Cell proliferation experiments showed TRAIL-dependent inhibition that was further increased by combination treatment with the PPARγ ligands. Simultaneous exposure of TRAIL and the PPARγ ligands also resulted in enhanced induction of apoptosis even in partial TRAIL-resistant HEY cell lines. In the parental HEY cell line, additional treatment with the PPARγ ligands led to an increased protein expression of DR5 and a further decline of XIAP expression.. Therefore, the combinational treatment with TRAIL and PPARγ ligands might be a promising experimental therapy because the PPARγ ligands, especially d15-PGJ(2), sensitize drug-resistant ovarian cancer cells to TRAIL-induced apoptosis.

Topics: Apoptosis; CASP8 and FADD-Like Apoptosis Regulating Protein; Caspase 8; Cell Line, Tumor; Chromans; Drug Resistance, Neoplasm; Drug Synergism; Female; Humans; Ovarian Neoplasms; PPAR gamma; Prostaglandin D2; Receptors, TNF-Related Apoptosis-Inducing Ligand; Thiazolidinediones; TNF-Related Apoptosis-Inducing Ligand; Troglitazone

Cyclopentenone prostaglandin 15-deoxy-Δ(12,14)-prostaglandin J(2) (15d-PGJ(2)), which is generated from the dehydration of PGD(2), is a natural ligand of peroxisome proliferator-activated receptor gamma (PPARγ) and a potential apoptotic mediator. The synthetic PPARγ ligands, troglitazone and ciglitazone, inhibit tumor progression in many cells by PPARγ activation, but the mechanism of 15d-PGJ(2) is still unclear. In this study, GW9662, an antagonist of PPARγ, and quercetin, a natural antioxidant, were used to study the apoptotic mechanism of 15d-PGJ(2) in A549 cells. Results showed that 15d-PGJ(2) induced apoptosis, which was associated with the production of reactive oxygen species (ROS) and the decrease of GSH levels. Furthermore, quercetin reduced the activity of caspases in 15d-PGJ(2)-induced apoptotic processes. These results suggest that 15d-PGJ(2) induces apoptosis in A549 cells mainly through the formation of ROS; it does not depend on PPARγ activation. Moreover, these findings support the use of quercetin and PPARγ agonists in non-small cell lung carcinoma.

Topics: Apoptosis; Carcinoma, Non-Small-Cell Lung; Caspases; Cell Line, Tumor; Glutathione; Humans; Lung Neoplasms; PPAR gamma; Prostaglandin D2; Quercetin; Reactive Oxygen Species; Thiazolidinediones

Ligands activating the transcription factor peroxisome proliferator-activated receptor-γ (PPARγ) have antiinflammatory effects. Vascular rejection induced by allogeneic T cells can be responsible for acute and chronic graft loss. Studies in rodents suggest that PPARγ agonists may inhibit graft vascular rejection, but human T-cell responses to allogeneic vascular cells differ from those in rodents, and the effects of PPARγ in human transplantation are unknown.. We tested the effects of PPARγ agonists on human vascular graft rejection using a model in which human artery is interposed into the abdominal aorta of immunodeficient mice, followed by adoptive transfer of allogeneic (to the artery donor) human peripheral blood mononuclear cells. Interferon-γ-dependent rejection ensues within 4 weeks, characterized by intimal thickening, T-cell infiltrates, and vascular cell activation, a response resembling clinical intimal arteritis. The PPARγ agonists 15-deoxy-prostaglandin-J(2), ciglitazone, and pioglitazone reduced intimal expansion, intimal infiltration of CD45RO(+) memory T cells, and plasma levels of inflammatory cytokines. The PPARγ antagonist GW9662 reversed the protective effects of PPARγ agonists, confirming the involvement of PPARγ-mediated pathways. In vitro, pioglitazone inhibited both alloantigen-induced proliferation and superantigen-induced transendothelial migration of memory T cells, indicating the potential mechanisms of PPARγ effects.. Our results suggest that PPARγ agonists inhibit allogeneic human memory T cell responses and may be useful for the treatment of vascular graft rejection.

Topics: Adoptive Transfer; Anilides; Animals; Arteries; Cell Movement; Cell Proliferation; Cytokines; Graft Rejection; Humans; Hypoglycemic Agents; Immunologic Memory; Isoantigens; Mice; Mice, SCID; Pioglitazone; PPAR gamma; Prostaglandin D2; Superantigens; T-Lymphocytes; Thiazolidinediones; Transplantation, Heterologous; Transplantation, Homologous

Side effects of peroxisome proliferator activated receptor gamma (PPARgamma) agonists such as ciglitazone include anemia, which in theory could be due to decreased formation or premature death of erythrocytes. A form of suicidal erythrocyte death is eryptosis, which is characterized by cell shrinkage and by breakdown of phosphatidylserine asymmetry leading to phosphatidylserine exposure at the cell surface. Phosphatidylserine-exposing erythrocytes are recognized by macrophages, engulfed, degraded and thus cleared from circulating blood. Triggers of eryptosis include increase in intracellular Ca(2+) concentration. The present study thus explored, whether the PPARgamma agonist ciglitazone or the natural PPARgamma ligand 15deoxy-delta12,14-prostaglandin J2 (15d-PGJ2) are capable to trigger eryptosis. Phosphatidylserine exposure was determined from annexin V binding and cell shrinkage from decrease of forward scatter of human erythrocytes in FACS analysis. Both, ciglitazone (>or= 5 microM) and 15d-PGJ2 (>or= 3 microM), within 24 hours increased phosphatidylserine exposure and at concentrations of 10 microM led to a significant loss of the cell volume. Ciglitazone further stimulated hemolysis, which, however, affected only a fraction of erythrocytes undergoing eryptosis. According to Fluo3 fluorescence of human erythrocytes, 10 microM ciglitazone or 15d-PGJ2 increased intracellular Ca(2+) activity. In conclusion, ciglitazone and 15d-PGJ2 trigger eryptosis at least in part by an increase in the cytosolic Ca(2+) concentration. The effect most likely contributes to the anemia observed following treatment with PPARgamma agonists.

Topics: Aniline Compounds; Annexin A5; Calcium; Cell Death; Erythrocytes; Hemolysis; Humans; Prostaglandin D2; Thiazolidinediones; Xanthenes

Tumor necrosis factor-alpha (TNF-alpha) plays critical roles in bone resorption at the site of inflammatory joints. The aim of this study is to evaluate the effect of peroxisome proliferator-activated receptor gamma (PPAR-gamma) agonists, a new class of anti-inflammatory compounds, on TNF-alpha-mediated osteoclastogenesis in human monocytes. Human monocytes were differentiated into osteoclasts in the presence of TNF-alpha and macrophage colony-stimulating factor. Tartrate-resistant acid phosphatase (TRAP) staining and a pit formation assay using dentin were used for the identification of activated osteoclasts. The protein and gene expressions of transcription factors were determined by immunofluorescence and real-time RT-PCR analysis, respectively. TNF-alpha-induced osteoclast generation from human peripheral monocytes in a dose-dependent manner, and the induction was not inhibited by osteoprotegerin, a decoy receptor for receptor activator of NF-kappaB ligand. The addition of PPAR-gamma agonists, 15-deoxy-Delta12,14-prostaglandin J2 (15d-PGJ2) or ciglitazone, to the culture resulted in a remarkably reduced number of generated osteoclasts. In addition, both agonists inhibited the protein and gene expressions of nuclear factor of activated T-cell isoform c1 (NFATc1), c-Fos, c-Jun and NF-kappaB p65, which are known to be associated with osteoclastogenesis. GW9662, an antagonist of PPAR-gamma, fully rescued ciglitazone-induced inhibition, but did not affect 15d-PGJ2-induced inhibition. Monocyte chemoattractant protein-1 (MCP-1), a CC chemokine related to osteoclastogenesis, was induced during TNF-alpha-mediated osteoclast differentiation, and the neutralizing antibody to MCP-1 reduced osteoclast formation by about 40%. 15d-PGJ2 and ciglitazone blocked the induction of MCP-1 by TNF-alpha. Moreover, the addition of MCP-1 rescued the inhibition of TRAP-positive multinucleated cell (TRAP-MNCs) formation by 15d-PGJ2 and ciglitazone, although generated TRAP-MNCs had no capacity to resorb dentin slices. Our data demonstrate that 15d-PGJ2 and ciglitazone down-regulate TNF-alpha-mediated osteoclast differentiation in human cells, in part via suppression of the action of MCP-1. These PPAR-gamma agonists may be a promising therapeutic application for rheumatoid arthritis and inflammatory bone-resorbing diseases.

Topics: Cell Differentiation; Cells, Cultured; Chemokine CCL2; Humans; Monocytes; Osteoclasts; PPAR gamma; Prostaglandin D2; RANK Ligand; Thiazolidinediones; Transcription Factors; Tumor Necrosis Factor-alpha

Respiratory syncytial virus (RSV) is the major causative agent of severe lower respiratory tract disease and death in infants worldwide. The epithelial cells of the airways are the target cells for RSV infection and the site of the majority of the inflammation associated with the disease. However, despite five decades of intensive RSV research there exist neither an effective active vaccine nor a promising antiviral and anti-inflammatory therapy. Recently, peroxisome proliferator-activated receptor-gamma (PPAR-gamma), a member of the nuclear hormone receptor superfamily, has been shown to possess anti-inflammatory properties. Therefore, we hypothesized whether the detrimental increase of intercellular adhesion molecule-1 (ICAM-1) on RSV-infected lung epithelial cells (A549 and primary normal human bronchial epithelial cells (NHBE)) might be modulated by natural and synthetic PPAR-gamma agonists (15d-PGJ2, ciglitazone, troglitazone, Fmoc-Leu). Our data show that all PPAR-gamma agonists under study significantly down-regulated the RSV-induced expression of ICAM-1 on A549- and NHBE cells in a dose-dependent manner resulting in a reduced beta2 integrin-mediated adhesion of monocytic effector cells (U937) to RSV-infected A549 cell monolayers. In contrast, the PPAR-alpha agonist bezafibrate had no impact on the RSV-induced ICAM-1 expression. The reduced ICAM-1 expression was associated with a diminished ICAM-1 mRNA level and binding activity of nuclear factor-kappaB (p65/p50) in A549 cells. These findings suggest that PPARgamma agonists have beneficial effects in the suppression of the inflammatory response during RSV infection and therefore might have clinical efficacy in the course of severe RSV-infection.

Topics: Amino Acids; Cell Adhesion; Cell Line; Cells, Cultured; Chromans; Dose-Response Relationship, Drug; Down-Regulation; Epithelial Cells; Fluorenes; Humans; Intercellular Adhesion Molecule-1; Lung; NF-kappa B; PPAR gamma; Prostaglandin D2; Respiratory Mucosa; Respiratory Syncytial Virus Infections; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Thiazolidinediones; Troglitazone; U937 Cells

Peroxisome proliferator-activated receptor gamma (PPARgamma), a ligand-activated transcription factor, is considered as an anti-osteoblastic factor associated with adiposity and the elderly osteoporosis due to a defect in osteoblastogenesis. We have found that oral administration of PPARgamma activator rosiglitazone decreased tibia BMD and serum ALP but left serum calcium and osteoclast marker C-terminal telopeptide unaffected. In addition, we examined the inhibitory mechanisms of PPARgamma on the bone formation by using PPARgamma activators ciglitazone and 15-deoxy-Delta(12,14)-prostaglandin-J2 (15d-PGJ2). Our data indicated that PPARgamma ligands decreased both mineralized bone nodules and alkaline phosphatase (ALP) activities in cultured primary osteoblasts. Reverse transcription polymerase chain reaction (RT-PCR) showed that the expression of bone morphogenetic protein-2 (BMP-2) and osteocalcin (OCN) was inhibited by ciglitizone and 15d-PGJ2. Furthermore, PPARgamma ligands inhibited NF-kappaB associated downstream COX-2 and iNOS osteogenic signaling. The ultrasound (US)-induced elevation of COX-2 and iNOS expression and nitric oxide (NO) production were attenuated in the presence of PPARgamma ligands. Furthermore, local administration of PPARgamma ligands into the metaphysis of rat tibia decreased the bone volume in secondary spongiosa. These results suggest that the activation of PPARgamma inhibits osteoblastic differentiation and the expression of several anabolic mediators involved in bone formation. These data may reflect osteoporosis and less bone formation in the aging people and patients treated with thiazolidinediones.

Topics: Aging; Animals; Bone Morphogenetic Protein 2; Bone Morphogenetic Proteins; Cell Differentiation; Cell Proliferation; Cells, Cultured; Cyclooxygenase 2; Down-Regulation; Gene Expression Regulation, Enzymologic; Ligands; Nitric Oxide; Nitric Oxide Synthase Type II; Osteoblasts; Osteogenesis; PPAR gamma; Prostaglandin D2; Rats; RNA, Messenger; Rosiglitazone; Thiazolidinediones; Time Factors; Transforming Growth Factor beta

The signaling lipid molecule 15-deoxy-delta 12,14-prostaglandin J2 (15d-PGJ2) has multiple cellular functions, including anti-inflammatory and antineoplastic activities. Here, we report that 15d-PGJ2 blocks translation through inactivation of translational initiation factor eIF4A. Binding of 15d-PGJ2 to eIF4A blocks the interaction between eIF4A and eIF4G that is essential for translation of many mRNAs. Cysteine 264 in eIF4A is the target site of 15d-PGJ2. The antineoplastic activity of 15d-PGJ2 is likely attributed to inhibition of translation. Moreover, inhibition of translation by 15d-PGJ2 results in stress granule (SG) formation, into which TRAF2 is sequestered. The sequestration of TRAF2 contributes to the anti-inflammatory activity of 15d-PGJ2. These findings reveal a novel cross-talk between translation and inflammatory response, and offer new approaches to develop anticancer and anti-inflammatory drugs that target translation factors including eIF4A.

Topics: Anti-Inflammatory Agents; Arachidonic Acid; Arsenites; Chromans; Cyclopentanes; Cytoplasmic Granules; Dinoprostone; Emetine; Enzyme Inhibitors; Eukaryotic Initiation Factor-2; Eukaryotic Initiation Factor-4A; Gene Expression Regulation; HeLa Cells; Humans; Hypoglycemic Agents; Inflammation; Poly(A)-Binding Proteins; PPAR gamma; Prostaglandin D2; Prostaglandins A; Protein Biosynthesis; Protein Synthesis Inhibitors; Rosiglitazone; Signal Transduction; Sodium Compounds; T-Cell Intracellular Antigen-1; Thiazolidinediones; TNF Receptor-Associated Factor 2; Troglitazone; Tumor Necrosis Factor-alpha

The epithelial cells of the airways are the target cells for respiratory syncytial virus (RSV) infection and the site of the majority of the inflammation associated with the disease. Recently, peroxisome-proliferator-activated receptor gamma (PPARgamma), a member of the nuclear hormone receptor superfamily, has been shown to possess anti-inflammatory properties. Therefore, we investigated the role of PPARgamma agonists (15d-PGJ(2), ciglitazone and troglitazone) on the synthesis of RSV-induced cytokine release from RSV-infected human lung epithelial cells (A549). We observed that all PPARgamma ligands inhibited dose-dependently the release of TNF-alpha, GM-CSF, IL-1alpha, IL-6 and the chemokines CXCL8 (IL-8) and CCL5 (RANTES) from RSV-infected A549 cells. Concomitantly, the PPARgamma ligands diminished the cellular amount of mRNA encoding for IL-6, CXCL8 and CCL5 and the RSV-induced binding activity of the transcription factors NF-kappaB (p65/p50) and AP-1 (c-fos), respectively. Our data presented herein suggest a potential application of PPARgamma ligands in the anti-inflammatory treatment of RSV infection.

Topics: Cell Line, Tumor; Chemokine CCL5; Chemokines, CC; Chromans; Cytokines; Epithelial Cells; Gene Expression; Granulocyte-Macrophage Colony-Stimulating Factor; Humans; Interleukin-1; Interleukin-6; Interleukin-8; NF-kappa B; PPAR gamma; Prostaglandin D2; Respiratory Syncytial Viruses; RNA, Messenger; Thiazolidinediones; Transcription Factor AP-1; Troglitazone; Tumor Necrosis Factor-alpha

2-Arachidonyl glycerol (2-AG) is an endogenous arachidonic acid derivative that binds cannabinoid receptors CB1 and CB2 and is hence termed an endocannabinoid. 2-AG also modulates a variety of immunological responses, including expression of the autocrine/paracrine T cell growth factor interleukin (IL)-2. The objective of the present studies was to determine the mechanism responsible for IL-2 suppression by 2-AG. Because of the labile properties of 2-AG, 2-AG ether, a nonhydrolyzable analog of 2-AG, was also used. Both 2-AG and 2-AG ether suppressed IL-2 expression independently of CB1 and CB2, as demonstrated in leukocytes derived from CB1/CB2-null mice. Moreover, we demonstrated that both 2-AG and 2-AG ether treatment activated peroxisome proliferator-activated receptor gamma (PPARgamma), as evidenced by forced differentiation of 3T3-L1 cells into adipocytes, induction of aP2 mRNA levels, and activation of a PPARgamma-specific luciferase reporter in transiently transfected 3T3-L1 cells. Consequently, the putative role of PPARgamma in IL-2 suppression by 2-AG and 2-AG ether was examined in Jurkat T cells. Concordant with PPARgamma involvement, the PPARgamma-specific antagonist 2-chloro-5-nitro-N-(4-pyridyl)-benzamide (T0070907) blocked 2-AG- and 2-AG ether-mediated IL-2 suppression. Likewise, 2-AG suppressed the transcriptional activity of two transcription factors crucial for IL-2 expression, nuclear factor of activated T cells and nuclear factor kappaB, in the absence but not in the presence of T0070907. 2-AG treatment also induced PPARgamma binding to a PPAR response element in activated Jurkat T cells. Together, the aforementioned studies identify PPARgamma as a novel intracellular target of 2-AG, which mediates the suppression of IL-2 by 2-AG in a manner that is independent of CB1 and/or CB2.

Topics: 3T3-L1 Cells; Adipogenesis; Animals; Arachidonic Acids; Benzamides; Cells, Cultured; Endocannabinoids; Fatty Acid-Binding Proteins; Female; Gene Expression; Glycerides; Humans; Interferon-gamma; Interleukin-2; Interleukin-4; Jurkat Cells; Mice; Mice, Inbred C57BL; Mice, Knockout; NF-kappa B; NFATC Transcription Factors; Peroxisome Proliferator-Activated Receptors; PPAR gamma; Prostaglandin D2; Protein Binding; Pyridines; Receptor, Cannabinoid, CB1; Receptor, Cannabinoid, CB2; Response Elements; T-Lymphocytes; Thiazolidinediones

We have previously shown that peroxisome proliferator-activated receptor-gamma (PPARgamma) agonists inhibited the inflammatory response of RSV-infected human lung epithelial cells. In this study, we supply evidence that specific PPARgamma agonists (15d-PGJ2, ciglitazone, troglitazone, Fmoc-Leu) efficiently blocked the RSV-induced cytotoxicity and development of syncytia in tissue culture (A549, HEp-2). All PPARgamma agonists under study markedly inhibited the cell surface expression of the viral G and F protein on RSV-infected A549 cells. This was paralleled by a reduced cellular amount of N protein-encoding mRNA determined by real-time RT-PCR. Concomitantly, a reduced release of infectious progeny virus into the cell supernatants of human lung epithelial cells (A549, normal human bronchial epithelial cells (NHBE)) was observed. Similar results were obtained regardless whether PPARgamma agonists were added prior to RSV infection or thereafter, suggesting that the agonists inhibited viral gene expression and not the primary adhesion or fusion process.

Topics: Chromans; Humans; Lung; PPAR gamma; Prostaglandin D2; Respiratory Mucosa; Respiratory Syncytial Viruses; Thiazolidinediones; Troglitazone; Virus Replication

Peroxisome proliferator-activated receptor gamma (PPARgamma) agonists cause cell death in several types of cancer cells. The aim of this study was to examine the effects of two PPARgamma agonists, ciglitazone and 15-deoxy-delta(12,14)-prostaglandin J2 (15dPGJ2), on the survival of thyroid carcinoma CGTH W-2 cells. Both ciglitazone and 15dPGJ2 decreased cell viability in a time- and dose-dependent manner. Cell death was mainly due to apoptosis, with a minor contribution from necrosis. Increased levels of active caspase 3, cleaved poly (ADP-ribose) polymerase (PARP), and cytosolic cytochrome-c were noted. In addition, ciglitazone and 15dPGJ2 induced detachment of CGTH W-2 cells from the culture substratum. Both the protein levels and immunostaining signals of focal adhesion (FA) proteins, including vinculin, integrin beta1, focal adhesion kinase (FAK), and paxillin were decreased after PPARgamma agonist treatment. Meanwhile, reduced phosphorylation of FAK and paxillin was noted. Furthermore, PPARgamma agonists induced expression of protein tyrosine phosphatase-PEST (PTP-PEST), and of phosphatase and tensin homologue deleted on chromosome ten (PTEN). The upregulation of these phosphatases might contribute to the dephosphorylation of FAK and paxillin, since pre-treatment with orthovanadate prevented PPARgamma agonist-induced dephosphorylation of FAK and paxillin. Perturbation of CGTH W-2 cells with anti-integrin beta1 antibodies induced FA disruption and apoptosis in the same cells, thus the downregulation of integrin beta1 by PPARgamma agonists resulted in FA disassembly and might induce apoptosis via anoikis. Our results suggested the presence of crosstalk between apoptosis and integrin-FA signaling. Moreover, upregulation and activation of PTEN was correlated with reduced phosphorylation of Akt, and this consequence disfavored cell survival. In conclusion, PPARgamma agonists induced apoptosis of thyroid carcinoma cells via the cytochrome-c caspase 3 and PTEN-Akt pathways, and induced necrosis via the PARP pathway.

Topics: Apoptosis; Asian People; Carcinoma; Cell Line, Tumor; Cell Survival; Dose-Response Relationship, Drug; Focal Adhesions; Humans; Hypoglycemic Agents; Necrosis; Neoplasm Proteins; PPAR gamma; Prostaglandin D2; Signal Transduction; Thiazolidinediones; Thyroid Neoplasms

Peroxisome proliferator-activated receptor gamma (PPARgamma) is a transcription factor important for adipogenesis and more recently has been shown to be an anticancer target. PPARgamma ligands, including the endogenous ligand 15-deoxy-Delta12,14-PGJ2 (15d-PGJ2) and synthetic ligands like ciglitazone and troglitazone, all induce apoptosis in normal and malignant human B lymphocytes, but the dependency of PPARgamma for apoptosis induction is unknown. In this study, we used a PPARgamma dominant-negative approach and a small molecule irreversible PPARgamma antagonist and found that these inhibitors prevented PPARgamma activation but did not prevent B cell apoptosis induced by 15d-PGJ2 or ciglitazone. In addition, a PPARgamma agonist that is a structural analog of 15d-PGJ2, and lacks the electrophilic carbon of the 15d-PGJ2 cyclopentenone ring, activated PPARgamma but did not kill B lymphocytes, further supporting a non-PPARgamma-mediated mechanism. To further investigate the apoptotic mechanism, the effects of 15d-PGJ2 and ciglitazone on reactive oxygen species were investigated. 15d-PGJ2, but not ciglitazone, potently induced reactive oxygen species in B lymphocytes, implicating the reactive nature of the 15d-PGJ2 structure in the apoptosis mechanism. In addition, 15d-PGJ2 caused an almost complete depletion of intracellular glutathione. Moreover, incubation with glutathione reduced ethyl ester, an antioxidant, prevented apoptosis induced by 15d-PGJ2, but not by ciglitazone. These findings indicate that the expression of PPARgamma may not be predictive of whether a normal or malignant B lineage cell is sensitive to PPARgamma agonists. Furthermore, these new findings support continued investigation into the use of PPARgamma agonists as agents to attenuate normal B cell responses and as anti-B cell lymphoma agents.

Topics: Apoptosis; B-Lymphocytes; Humans; Ligands; Lymphoma, B-Cell; PPAR gamma; Prostaglandin D2; Reactive Oxygen Species; Thiazolidinediones

Brain abscesses arise from a focal parenchymal infection by various pathogens, particularly Staphylococcus aureus. We have shown that astrocytes are activated upon exposure to S. aureus and may contribute to the excessive tissue damage characteristic of brain abscess. Therefore, modulating astrocyte activation may facilitate a reduction in brain abscess severity. Peroxisome proliferator activated receptor-gamma (PPAR-gamma) agonists are potent inhibitors of microglial activation; however, the effects of these compounds on S. aureus-dependent astrocyte activation have not yet been examined. Here, we demonstrate that two chemically distinct PPAR-gamma agonists, 15-deoxy-delta12,14-prostaglandin J2 (15d-PGJ2) and ciglitazone, suppress the production of several pro-inflammatory molecules in S. aureus-stimulated astrocytes including interleukin-1beta and nitric oxide (NO). Interestingly, 15d-PGJ2 attenuated Toll-like receptor 2 (TLR2) and inducible nitric oxide synthase expression, but failed to modulate macrophage inflammatory protein-2 (MIP-2/CXCL2) production, suggesting that 15d-PGJ2 is not a global inhibitor of astrocyte activation. Another novel finding of this study was the fact that both 15d-PGJ2 and ciglitazone were capable of attenuating pre-existing astrocyte activation, indicating their potential benefit in a therapeutic setting. Importantly, 15d-PGJ2 and ciglitazone were still capable of inhibiting S. aureus-induced pro-inflammatory mediator release in PPAR-gamma-deficient astrocytes, supporting PPAR-gamma-independent effects of these compounds. Collectively, these results suggest that 15d-PGJ2 and ciglitazone exert their anti-inflammatory actions on astrocytes primarily independent of the PPAR-gamma pathway.

Topics: Animals; Animals, Newborn; Astrocytes; Cells, Cultured; Hypoglycemic Agents; Inflammation Mediators; Mice; Mice, Inbred C57BL; Mice, Knockout; Nitric Oxide Synthase Type II; PPAR gamma; Prostaglandin D2; Staphylococcus aureus; Thiazolidinediones; Toll-Like Receptor 2

Recently, we demonstrated that the peroxisome proliferator-activated receptor-gamma (PPAR-gamma) ligands, either 15-deoxy-Delta12,14-prostaglandin J2 (15d-PGJ2) or ciglitazone, increased endothelial nitric oxide (.NO) release without altering endothelial nitric oxide synthase (eNOS) expression (4). However, the precise molecular mechanisms of PPAR-gamma-stimulated endothelial.NO release remain to be defined. Superoxide anion radical (O2-.) combines with .NO to decrease.NO bioavailability. NADPH oxidase, which produces O2-., and Cu/Zn-superoxide dismutase (Cu/Zn-SOD), which degrades O2-., thereby contribute to regulation of endothelial cell.NO metabolism. Therefore, we examined the ability of PPAR-gamma ligands to modulate endothelial O2-. metabolism through alterations in the expression and activity of NADPH oxidase or Cu/Zn-SOD. Treatment with 10 microM 15d-PGJ2 or ciglitazone for 24 h decreased human umbilical vein endothelial cell (HUVEC) membrane NADPH-dependent O2-. production detected with electron spin resonance spectroscopy. Treatment with 15d-PGJ2 or ciglitazone also reduced relative mRNA levels of the NADPH oxidase subunits, nox-1, gp91phox (nox-2), and nox-4, as measured using real-time PCR analysis. Concordantly, Western blot analysis demonstrated that 15d-PGJ2 or ciglitazone decreased nox-2 and nox-4 protein expression. PPAR-gamma ligands also stimulated both activity and expression of Cu/Zn-SOD in HUVEC. These data suggest that in addition to any direct effects on endothelial.NO production, PPAR-gamma ligands enhance endothelial.NO bioavailability, in part by altering endothelial O2-. metabolism through suppression of NADPH oxidase and induction of Cu/Zn-SOD. These findings further elucidate the molecular mechanisms by which PPAR-gamma ligands directly alter vascular endothelial function.

Topics: Blotting, Western; Cell Membrane; Cells, Cultured; Electron Spin Resonance Spectroscopy; Endothelial Cells; Humans; Hypoglycemic Agents; Ligands; NADPH Oxidases; Nitric Oxide; PPAR gamma; Prostaglandin D2; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Superoxide Dismutase; Superoxides; Thiazolidinediones; Umbilical Veins

Pulmonary fibrosis is a progressive life-threatening disease for which no effective therapy exists. Myofibroblasts are one of the key effector cells in pulmonary fibrosis and are the primary source of extracellular matrix production. Drugs that inhibit the differentiation of fibroblasts to myofibroblasts have potential as antifibrotic therapies. Peroxisome proliferator-activated receptor (PPAR)-gamma is a transcription factor that upon ligation with PPARgamma agonists activates target genes containing PPAR response elements. PPARgamma agonists have anti-inflammatory activities and may have potential as antifibrotic agents. In this study, we examined the abilities of PPARgamma agonists to block two of the most important profibrotic activities of TGF-beta on pulmonary fibroblasts: myofibroblast differentiation and production of excess collagen. Both natural (15d-PGJ2) and synthetic (ciglitazone and rosiglitazone) PPARgamma agonists inhibited TGF-beta-driven myofibroblast differentiation, as determined by alpha-smooth muscle actin-specific immunocytochemistry and Western blot analysis. PPARgamma agonists also potently attenuated TGF-beta-driven type I collagen protein production. A dominant-negative PPARgamma partially reversed the inhibition of myofibroblast differentiation by 15d-PGJ2 and rosiglitazone, but the irreversible PPARgamma antagonist GW-9662 did not, suggesting that the antifibrotic effects of the PPARgamma agonists are mediated through both PPARgamma-dependent and independent mechanisms. Thus PPARgamma agonists have novel and potent antifibrotic effects in human lung fibroblasts and may have potential for therapy of fibrotic diseases in the lung and other tissues.

Topics: Actins; Anilides; Cell Differentiation; Collagen Type I; Fibroblasts; Humans; Hypoglycemic Agents; Lung; Muscle, Smooth; PPAR gamma; Prostaglandin D2; Pulmonary Fibrosis; Rosiglitazone; Thiazolidinediones; Transforming Growth Factor beta

We recently reported that the peroxisome proliferator-activated receptor gamma (PPARgamma) ligands 15-deoxy-Delta(12,14)-prostaglandin J2 (15d-PGJ2) and ciglitazone increased cultured endothelial cell nitric oxide (NO) release without increasing the expression of endothelial nitric oxide synthase (eNOS). The current study was designed to characterize further the molecular mechanisms underlying PPARgamma-ligand-stimulated increases in endothelial cell NO production.. Treating human umbilical vein endothelial cells (HUVEC) with PPARgamma ligands (10 micromol/L 15d-PGJ2, ciglitazone, or rosiglitazone) for 24 hours increased NOS activity and NO release. In selected studies, HUVEC were treated with PPARgamma ligands and with the PPARgamma antagonist GW9662 (2 micromol/L), which fully inhibited stimulation of a luciferase reporter gene, or with small interfering RNA to PPARgamma, which reduced HUVEC PPARgamma expression. Treatment with either small interfering RNA to PPARgamma or GW9662 inhibited 15d-PGJ2-, ciglitazone-, and rosiglitazone-induced increases in endothelial cell NO release. Rosiglitazone and 15d-PGJ2, but not ciglitazone, increased heat shock protein 90-eNOS interaction and eNOS ser1177 phosphorylation. The heat shock protein 90 inhibitor geldanamycin attenuated 15d-PGJ2- and rosiglitazone-stimulated NOS activity and NO production.. These findings further clarify mechanisms involved in PPARgamma-stimulated endothelial cell NO release and emphasize that individual ligands exert their effects through distinct PPARgamma-dependent mechanisms.

Topics: Anilides; Cells, Cultured; Endothelium, Vascular; Genes, Reporter; Humans; Hypoglycemic Agents; Ligands; Nitric Oxide; PPAR gamma; Prostaglandin D2; RNA, Small Interfering; Rosiglitazone; Signal Transduction; Thiazolidinediones; Umbilical Veins

Subject- Peroxisome proliferator-activated receptor (PPAR)-gamma agonists are emerging as potential protectors against inflammatory cardiovascular diseases including atherosclerosis and diabetic complications. However, their molecular mechanism of action within vasculature remains unclear. We report here that PPARgamma agonists, thiazolidinedione class drugs (TZDs), or 15-deoxy-Delta12,14-prostaglandin J2 (15d-PGJ2) were capable of activating diacylglycerol (DAG) kinase (DGK), resulting in attenuation of DAG levels and inhibition of protein kinase C (PKC) activation. The PPARgamma agonist-induced DGK was completely blocked by a dominant-negative mutant of PPARgamma, indicating an essential receptor-dependent action. Importantly, the suppression of DAG-PKC signaling pathway was functional linkage to the anti-inflammatory properties of PPARgamma agonists in endothelial cells (EC), characterized by the inhibition of proinflammatory adhesion molecule expression and adherence of monocytes to the activated EC induced by high glucose. These findings thus demonstrate a novel molecular action of PPARgamma agonists to suppress the DAG-PKC signaling pathway via upregulation of an endogenous attenuator, DGK.

Topics: Animals; Aorta; Cattle; Cell Adhesion; Cell Adhesion Molecules; Cells, Cultured; Chromans; Diabetic Angiopathies; Diacylglycerol Kinase; Diglycerides; Endothelial Cells; Endothelium, Vascular; Enzyme Induction; Glucose; Humans; PPAR gamma; Prostaglandin D2; Protein Kinase C; Protein Kinase C beta; Protein Transport; Recombinant Fusion Proteins; Signal Transduction; Thiazolidinediones; Transfection; Troglitazone; U937 Cells; Umbilical Veins; Vasculitis

Topics: Antineoplastic Agents; Apoptosis; Chromans; Cyclin D1; Humans; Lung Neoplasms; Prostaglandin D2; Proto-Oncogene Proteins p21(ras); Receptors, Cytoplasmic and Nuclear; RNA, Messenger; Thiazolidinediones; Transcription Factors; Troglitazone

Background. Peroxisome proliferator activated receptors (PPARs) are ligand-activated transcription factors with multiple effects on target cell function. PPAR gamma activity is regulated by extracellular signal-regulated protein kinase (ERK), mitogen-activated protein (MAP) kinase, and PPAR gamma ligands have varying effects on activity of ERK. Different PPAR gamma ligands have been shown to have both protective and detrimental effects in the kidney. Since transcriptional activation by different PPAR agonists is ligand- and depot-specific PPAR gamma, we have examined the effects of different agonists on PPAR activity in the proximal tubule.. Opossum kidney cells were used in all experiments, transiently transfected with a PPAR response element luciferase reporter and subject to stimulation with various PPAR ligands. The role of ERK and phosphorylation in PPAR gamma activation were studied, as were the effects of PPAR agonists on ERK activation and cell proliferation.. Transcriptional activity of PPAR was not stimulated by PPAR alpha agonists, and only very modestly stimulated by a PPAR beta agonist. The PPAR gamma agonists 15-deoxy-Delta(12,14)-prostaglandin J(2) (15d-PGJ(2)), ciglitazone, and troglitazone stimulated significant transcriptional activation and phosphorylation of PPAR gamma. These effects were more marked with 15d-PGJ(2). Thiazolidinediones attenuated 15d-PGJ(2) evoked PPAR gamma activation and phosphorylation. ERK activity positively regulated PPAR activation. Only 15d-PGJ(2) stimulated ERK activity and cell proliferation, and these effects were also inhibited by thiazolidinediones.. PPAR gamma agonists exert differential effects in proximal tubule cells with thiazolidinediones behaving as partial agonists.

Topics: Animals; Cells, Cultured; Chromans; Extracellular Signal-Regulated MAP Kinases; Kidney Tubules, Proximal; Ligands; Opossums; PPAR gamma; Prostaglandin D2; Thiazolidinediones; Transcriptional Activation; Transfection; Troglitazone

Several studies have shown that PPARgamma agonists play a role in the regulation of lymphocytes function and apoptosis. However, the molecular mechanism(s) underlying the immunomodulatory effects of PPARgamma agonists are not defined yet. In this study, the effects of PPARgamma (15d PGJ2 and ciglitizone) ligands on proliferation, cytokine production and apoptosis of Jurkat and Raji cells (human T and B lymphocytes, respectively) were examined. Ciglitizone and 15d PGJ2 presented antiproliferative and cytotoxic effects on Jurkat and Raji cells as shown by [14C]-thymidine incorporation and cell viability assay. In addition, 15d PGJ2 inhibited cytokine production (IL-2 in Jurkat cells and IL-10 in Raji cells). The mechanism whereby PPARgamma agonists induced cytotoxicity is via apoptosis as shown by DNA fragmentation, nuclear condensation and phosphatidylserine externalization. The induction of apoptosis by ciglitizone and 15d PGJ2 on Jurkat and Raji cells may explain the suppression of cytokine production and the decrease in proliferation observed in both cell types. The apoptotic process was associated with a decrease in mitochondrial membrane potential and a marked down-regulation of the c-myc expression. These findings might play a key role in the apoptosis of T and B lymphocytes induced by PPARgamma agonists.

Topics: Annexin A5; Apoptosis; Benzimidazoles; Cell Membrane; Cell Proliferation; Chromatin; Cytokines; DNA Fragmentation; Fluorescent Dyes; Genes, myc; Humans; Immunohistochemistry; Jurkat Cells; Ligands; Membrane Potentials; Mitochondria; Phosphatidylserines; PPAR gamma; Prostaglandin D2; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Thiazolidinediones

Interferon-gamma (IFN-gamma) production and cytolytic activity are 2 major biologic functions of natural killer (NK) cells that are important for innate immunity. We demonstrate here that these functions are compromised in human NK cells treated with peroxisome proliferator-activated-gamma (PPAR-gamma) ligands via both PPAR-gamma-dependent and -independent pathways due to variation in PPAR-gamma expression. In PPAR-gamma-null NK cells, 15-deoxy-Delta(12,14) prostaglandin J(2) (15d-PGJ(2)), a natural PPAR-gamma ligand, reduces IFN-gamma production that can be reversed by MG132 and/or chloroquine, and it inhibits cytolytic activity of NK cells through reduction of both conjugate formation and CD69 expression. In PPARgamma-positive NK cells, PPAR-gamma activation by 15d-PGJ(2) and ciglitazone (a synthetic ligand) leads to reduction in both mRNA and protein levels of IFN-gamma. Overexpression of PPAR-gamma in PPAR-gamma-null NK cells reduces IFN-gamma gene expression. However, PPAR-gamma expression and activation has no effect on NK cell cytolytic activity. In addition, 15d-PGJ(2) but not ciglitazone reduces expression of CD69 in human NK cells, whereas CD44 expression is not affected. These results reveal novel pathways regulating NK cell biologic functions and provide a basis for the design of therapeutic agents that can regulate the function of NK cells within the innate immune response.

Topics: Humans; Hypoglycemic Agents; Interferon-gamma; Killer Cells, Natural; Ligands; PPAR gamma; Prostaglandin D2; Thiazolidinediones

Neuroblastoma is a childhood cancer, which spontaneously regresses. This has led to a search for agents that mimic this process. We show that both natural and synthetic ligands of PPARgamma (peroxisome-proliferator-activated receptor gamma) inhibit the growth of neuroblastoma cells in vitro. The degree of PPAR activation was attenuated however in the presence of the retinoblastoma protein. Addition of trichostatin A, a histone deacetylase inhibitor, abolished retinoblastoma protein repression of PPAR activity. Moreover, enhanced growth inhibition was observed when neuroblastoma cells were treated with a PPARgamma ligand and a histone deacetylase inhibitor, suggesting a combination therapy to treat neuroblastoma might prove more effective than using either agent alone.

Topics: Carboxy-Lyases; Cell Line, Tumor; Dose-Response Relationship, Drug; Enzyme Inhibitors; Genes, Reporter; Histones; Humans; Hydroxamic Acids; Ligands; Neuroblastoma; PPAR gamma; Prostaglandin D2; Retinoblastoma Protein; Thiazolidinediones; Time Factors; Transcription, Genetic; Transfection

During chronic inflammatory response, mono- cytes/macrophages produce 92-kDa matrix metalloproteinase-9 (MMP-9), which may contribute to their extravasation, migration and tissue remodeling. Activation of peroxisome proliferator- activated factor receptor-g (PPAR-g) has been shown to inhibit MMP-9 activity. To evaluate whether ox-LDL, a PPAR-g activator, inhibits PMA-induced MMP-9 expression and activity, and if so, whether CD36 and PPAR-g are involved in this process, we investigated the effect of ox-LDL on MMP-9 expression and activity in PMA-activated human monocytic cell line U937. PMA-induced MMP-9 expression and activity were suppressed by the treatment with ox-LDL (50 mg/ml) or PPAR-g activators such as troglitazone (5 mM), ciglitazone (5 mM), and 15d- PGJ2 (1 mM) for 24 h. This ox-LDL or PPAR-g activator-mediated inhibition of MMP-9 activity was diminished by the pre-treatment of cells with a blocking antibody to CD36, or PGF2a (0.3 mM), which is a PPAR-g inhibitor, as well as overexpression of a dominant-negative form of CD36. Taken together, these results suggest that ox-LDL suppresses PMA-induced MMP-9 expression and activity through CD36-mediated activation of PPAR-g.

Topics: Antibodies, Blocking; CD36 Antigens; Cells, Cultured; Chromans; Humans; Lipoproteins, LDL; Matrix Metalloproteinase 9; Matrix Metalloproteinase Inhibitors; Monocytes; NF-kappa B; PPAR gamma; Prostaglandin D2; RNA, Messenger; Tetradecanoylphorbol Acetate; Thiazolidinediones; Transcription, Genetic; Troglitazone

Peroxisome proliferator-activated receptor gamma (PPARgamma) is a transcription factor in the steroid nuclear receptor superfamily. Ligand activation of PPARgamma is associated with differentiation and an inhibition of proliferation in normal and malignant cells, including adipocytes, monocytes, and tumor cells in colon, prostate, and breast cancers. The current studies were undertaken to assess both the expression and functional activity of PPARgamma in cultured normal human mammary epithelial cells (HMECs) and tissue samples. Analyses by northern hybridization, immunoblotting, and immunohistochemistry demonstrate PPARgamma gene expression in HMECs and breast tissue specimens. DNA binding and transactivation assays indicate the presence of functionally active PPARgamma in HMECs. Treatment with PPARgamma selective ligands, 15-deoxy-delta-(12,14)-prostaglandin J2 (15dPGJ2) and ciglitazone, inhibits the growth of HMECs in a dose-dependent manner. This growth inhibition is associated with alterations in cell cycle progression and the induction of apoptosis.

Topics: Apoptosis; Breast; Cell Cycle; Cell Division; Cells, Cultured; Epithelial Cells; Female; Gene Expression; Humans; Hypoglycemic Agents; Ligands; Prostaglandin D2; Receptors, Cytoplasmic and Nuclear; Repressor Proteins; Thiazoles; Thiazolidinediones; Transcription Factors

To explore the effects of peroxisome proliferator-activated receptor-gamma (PPAR-gamma) on the growth of human lung cancer cell lines and its possible mechanism.. Human non-small cell lung cancer (NSCLC) cells of the A549 line and human small cell lung cancer (SCLC) of the LTEP-P line were cultured and were divided into 3 groups respectively: control group, 15d-PGJ(2) group (15d-PGJ(2), a PPAR-gamma activator, was added), and ciglitazone group (ciglitazone, am antidiabetic drug, was added). Twenty-four, forty-eight, and seventy-two hours later, nested RT-PCR was used to detect t the expression of PPAR-gamma mRNA, Western blotting technique was used to detect the expression of PPAR-gamma protein, MTT staining was used to observe the proliferation of cells induced by PPAR-gamma agonists, TUNEL method was used to observe the apoptosis of cells affected by the ligands of PPAR-gamma, the expressions of bax, and bcl-2 mRN and proteins were examined by in situ hybridization and immunohistochemistry, and the expression of caspase-3 was detected by immunohistochemistry.. PPAR-gamma was expressed in the two lung cancer cell lines. The cell proliferation was inhibited by 15d-PGJ(2) and ciglitazone, especially the former, in dose-dependent and time-dependent manners. The apoptosis rates were (1.9 +/- 0.5)%, (9.8 +/- 1.5)%, and (5.6 +/- 1.1)% respectively in the control, 15d-PGJ(2), and ciglitazone groups with a significant difference between ant 2 groups (all P < 0.05). The expression rate of bax were (9,2 +/- 1.5)%, (63 +/- 10)%, and (31 +/- 6)% respectively in the control, 15d-PGJ(2), and ciglitazone groups with a very significant difference between ant 2 groups (all P < 0.01). he expression rate of bcl-2 were (18 +/- 3)%, (36 +/- 9)%, and (33 +/- 7)% respectively in the control, 15d-PGJ(2), and ciglitazone groups with a very significant difference between the control group and any of the agonist-treated groups (all P < 0.01) and without significant difference between the two treated groups. The expression rates of caspase-3 were (6.5 +/- 1.0)%, (65 +/- 11)%, and (40 +/- 7)% respectively in the control, 15d-PGJ(2), and ciglitazone groups with a significant difference between any 2 group (all P < 0.01). The caspase-3 level was positively correlated with the level of apoptosis.. Activated by ligands, PPAR-gamma remarkably inhibits the growth of human lung cancer cells through inducing apoptosis. Caspase-3 and bax/bcl-2 play a role in this process, PPAR-gamma is so important in the pathogenesis and/or progression of lung cancer that it may be a novel therapeutical target against lung cancer.

Topics: Apoptosis; bcl-2-Associated X Protein; Blotting, Western; Caspase 3; Caspases; Cell Division; Cell Line, Tumor; Gene Expression Regulation, Neoplastic; Humans; Immunohistochemistry; In Situ Hybridization; In Situ Nick-End Labeling; Prostaglandin D2; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-bcl-2; Receptors, Cytoplasmic and Nuclear; RNA, Messenger; Thiazolidinediones; Time Factors; Transcription Factors

Peroxisome proliferator-activated receptor-gamma (PPAR-gamma) plays a critical role in adipocytes differentiation and insulin sensitivity and is also related to regulation of inflammation and cell proliferation. The aim of this study was to investigate the PPAR-gamma agonist-induced apoptosis and effects of PPAR-gamma agonist on Fas-mediated apoptosis in a human colon cancer cell line.. Cell survival and apoptosis of HT-29 cells were measured by trypan blue exclusion method and FACScan after treatment with 15d-PGJ2, ciglitazone and IgM anti-Fas antibody (CH11), respectively or simultaneously. Also, activation of caspase-3 and caspase-8 was analyzed to assess the effects of PPAR-gamma and Fas on apoptosis signaling pathways.. CH11 induced apoptosis of HT-29 cells. 15d-PGJ2 or ciglitazone alone did not induce apoptosis, but combined stimulation with CH11 synergistically induced apoptosis. Also, 15d-PGJ2 alone did not activate caspase-3, but CH11 and 15d-PGJ2 synergistically activated caspase-3. CH11 activated procaspase-8, but 15d-PGJ2 did not.. PPAR-gamma was not an enough condition to induce apoptosis of HT-29 cells. Apoptosis was induced by high dose Fas, and was enhanced with PPAR-gamma agonist. PPAR-gamma agonist seems to enhance Fas-mediated apoptosis by affecting the way between caspase-8 and caspase-3. Further research is needed to use PPAR-gamma agonists as chemopreventive and therapeutic agent for colon cancer and to find the pathways of PPAR-gamma on apoptotic cascade of colon cancer cells.

Topics: Apoptosis; Caspases; Cell Survival; Cells, Cultured; Chalcone; Chalcones; Enzyme Activation; fas Receptor; HT29 Cells; Humans; Prostaglandin D2; Receptors, Cytoplasmic and Nuclear; Thiazolidinediones; Transcription Factors

Peroxisome proliferator-activated receptor-gamma (PPAR-gamma) has been implicated in the regulation of multiple inflammatory processes. However, little is known of PPAR-gamma in the regulation of interleukin (IL)-4 expression in T cells. In this study, the effects of PPAR-gamma ligands on production of IL-4, a pro-inflammatory cytokine associated with the pathophysiology of allergic diseases, were investigated. 15-Deoxy-Delta12,14 prostaglandin J2 (15d-PGJ2) and ciglitazone, two representative PPAR-gamma ligands, significantly inhibited IL-4 production in both antigen-stimulated primary CD4+ T cells and the phorbol 12-myristate 13-acetate (PMA)/ionomycin-activated EL-4 T cell line. 15d-PGJ2 and ciglitazone inhibited the activation of IL-4 gene promoter in EL-4 T cells transiently transfected with IL-4 promoter/reporter constructs, and the repressive effect mapped to a region in the IL-4 promoter containing binding sites for nuclear factor of activated T cells (NF-AT). The activation of T cells by PMA/ionomycin resulted in a marked enhancement of the binding activities to the NF-AT site that was significantly inhibited by the addition of PPAR-gamma ligands. In cotransfected EL-4 T cells, PPAR-gamma also inhibited the activation of the IL-4 promoter at multiple NF-AT sites in a ligand-dependent manner. NF-ATc1 bound PPAR-gamma both in vivo and in vitro, and the interaction interfaces involved the Rel similarity domain of NF-ATc1. In cotransfections of HeLa cells, PPAR-gamma inhibited the NF-ATc1 transactivation in a ligand-dependent manner. Coexpression of p300 or AP-1 relieved the PPAR-gamma ligand-mediated inhibition of the NF-AT transactivation. From these results, we propose that PPAR-gamma ligand-mediated suppression of IL-4 production in CD4+ T cells may involve both inhibition of the NFAT-DNA interactions and competitive recruitment of transcription integrators between NF-AT and PPAR-gamma.

Topics: Animals; Benzhydryl Compounds; Binding Sites; Carcinogens; CD4-Positive T-Lymphocytes; Cell Line; DNA-Binding Proteins; Drug Interactions; E1A-Associated p300 Protein; Epoxy Compounds; Interleukin-4; Ionomycin; Ligands; Mice; Mice, Inbred BALB C; NFATC Transcription Factors; Nuclear Proteins; Promoter Regions, Genetic; Prostaglandin D2; Receptors, Cytoplasmic and Nuclear; Tetradecanoylphorbol Acetate; Thiazolidinediones; Trans-Activators; Transcription Factor AP-1; Transcription Factors; Tumor Cells, Cultured