PPARγ-independent Pathway Research Articles

Cisplatin-induced changes in calcitonin gene-related peptide or TNF-α release in rat dorsal root ganglia in vitro model of neurotoxicity are not reverted by rosiglitazone

IntroductionChemotherapy-induced peripheral neuropathy (CIPN) is a common side effect of anticancer drugs which affect the peripheral nervous system, as occurs with cisplatin treatment. Nowadays, one strategy in development to prevent, minimize and/or revert CIPN is neuroprotection. Therefore, we have evaluated the signaling pathways involved in CIPN and the effect of rosiglitazone, a peroxisome proliferator-activated receptor γ (PPAR-γ) agonist. MethodsDorsal Root Ganglia (DRG) were harvested from Wistar rats (Rattus norvegicus), the cells were dissociated, plated, and maintained with nerve growth factor for 9 days. On day 8, the cells were treated with cisplatin, rosiglitazone and/or T0070907 (PPAR-γ antagonist) for 24 h. The cell viability was measured by trypan blue exclusion method, the mRNA was quantified by real-time RT-PCRq and the release of TNF-α and calcitonin gene-related peptide (CGRP) was evaluated by ELISA. ResultsCisplatin, rosiglitazone or T0070907 treatments did not decreased the cell viability on the primary DRG cultures cells. Cisplatin treatment induced a decrease of PPAR-γ and -β/δ mRNA, while the co-treatment with rosiglitazone inhibited this cisplatin-induced effect. Moreover, T0070907 did not change the observed results, indicating that the rosiglitazone’s effect could be due to mechanisms beyond PPAR-γ activation. Also, the rosiglitazone effect is not exclusively to DRG cells since there was an increase of PPAR-γ mRNA expression in 3T3-L1 cells. Furthermore, rosiglitazone did not modulate the cisplatin decrease neuronal function of DRG cells (TNF-α and CGRP release). ConclusionCisplatin decreased the gene expression of PPAR-γ and -β/δ, while the rosiglitazone treatment inhibited these effects via PPAR-γ independent pathway. Rosiglitazone did not show improvement in modulation of TNF-α or CGRP release impaired by cisplatin.

Anti-Diabetic Countermeasures Against Tobacco Smoke-Dependent Cerebrovascular Toxicity: Use and Effect of Rosiglitazone.

Tobacco smoking (TS) is one of the most addictive habit sand a main public health hazards, impacting the vascular endothelium through oxidative stress (OS) stimuli, exposure to nicotine, and smoking-induced inflammation in a dose-dependent manner. Increasing evidence also suggested that TS increases glucose intolerance and the risk factor of developing type-2 diabetes mellitus (2DM), which, along with TS, is connected to blood–brain barrier (BBB) injuries, and heightens the risk of cerebrovascular disorders. Although the exact mechanism of rosiglitazone (RSG) is unknown, our previous in vitro work showed how RSG, an oral anti-diabetic drug belonging to the family of thiazolidinedione class, can protect BBB integrity through enhancement of nuclear factor erythroid 2-related factor (Nrf2) activity. Herein, we have validated the protective role of rosiglitazone against TS-induced BBB impairment in vivo. Our results revealed that RSG as a peroxisome proliferator-activated receptor gamma (PPARγ), activates counteractive mechanisms primarily associated with the upregulation of Nrf2 and PPARγ pathways which reduce TS-dependent toxicity at the cerebrovascular level. In line with these findings, our results show that RSG reduces inflammation and protects BBB integrity. In conclusion, RSG offers a novel and promising therapeutic application to reduce TS-induced cerebrovascular dysfunction through activation of the PPARγ-dependent and/or PPARγ-independent Nrf2 pathway.

15-Deoxy-Δ12,14-prostaglandin J2 as a potential regulator of bone metabolism via PPARγ-dependent and independent pathways: a review.

Bone metabolism is a complex physiological process that primarily involves osteoblast-mediated bone formation and osteoclast-mediated bone resorption, both of which are regulated by a variety of biological factors. There is increasing evidence that peroxisome proliferator-activated receptor γ (PPARγ) is a member of the nuclear receptor superfamily and plays an important role in lipid metabolism and bone metabolism. Through the PPARγ-dependent pathway, 15-deoxy-Δ12,14-prostaglandin J2 (15d-PGJ2) promotes the formation of marrow adipocytes and inhibits the formation of osteoblasts, resulting in bone loss and increasing the risk of fracture and osteoporosis. Recent studies have found that through the PPARγ-independent pathway, 15d-PGJ2 plays a regulatory role in bone metastasis of breast cancer, which can inhibit osteoclastogenesis and reduce bone destruction. The purpose of our review is to summarize the recent progress in elucidating the mechanisms and effects of 15d-PGJ2 in bone metabolism, which can serve as a novel therapeutic target for bone tumors, osteoporosis, rheumatoid arthritis (RA), and other bone diseases.

Pioglitazone inhibits cancer cell growth through STAT3 inhibition and enhanced AIF expression via a PPARγ-independent pathway.

Pioglitazone is an anti-diabetic agent that belongs to the thiazolidinedione class, which target peroxisome proliferator-activated receptor γ (PPARγ), a transcription factor in the nuclear receptor family. Different cancer cells expressing high levels of PPARγ and PPARγ ligands induce cell cycle arrest, cell differentiation, and apoptosis. However, the mechanisms underlying these processes remain unknown. Here, we investigated the mechanism underlying pioglitazone-induced apoptosis in human cancer cells. We showed that at similar concentrations, pioglitazone induced death in cancer cells expressing high or low levels of PPARγ. Combined treatment of pioglitazone and GW9662, a PPARγ antagonist, did not rescue this cell death phenotype. Z-VAD-fmk, a pan-caspase inhibitor, did not reverse pioglitazone-induced apoptosis in cancer cells expressing PPARγ at high or low levels. Pioglitazone suppressed the activation of signal transducers and activator of transcription 3 (STAT3) and Survivin expression, and enhanced the apoptosis-inducing factor (AIF) levels in these cells. Furthermore, pioglitazone enhanced the cytotoxic effect of cisplatin and oxaliplatin by suppressing Survivin and increasing AIF expression. These results indicated that pioglitazone induced apoptosis via a PPARγ-independent pathway, thus describing pioglitazone as a potential therapeutic agent for controlling the progression of different cancers.

Trans-10, cis-12 Conjugated Linoleic Acid Modulates Tumor Necrosis Factor-${alpha}$ Production and Nuclear Factor-${kappa}B$ Activation in RAW 264.7 Macrophages Through Formation of Reactive Oxygen Species

The aims of this study were to explore the effects of conjugated linoleic acid (CLA) on reactive oxygen species (ROS) production in lipopolysaccharide (LPS)-naive and LPS-stimulated RAW 264.7 macrophages and to examine whether these effects affect the regulation of tumor necrosis factor-alpha (TNF-α) production, and nuclear factor-kappa B (NF-κB) and peroxisome proliferator-activated receptor gamma (PPARγ) activation. Trans-10, cis-12 (t10c12)-CLA increased the production of ROS, as well as TNF-α in LPS-naive RAW 264.7 cells. The CLA-induced TNF-α production was suppressed by treatment of diphenyleneiodonium chloride (DPI), a NADPH oxidase inhibitor. In addition, CLA enhanced the activities of NF-κB and PPARγ in LPS-naive RAW 264.7 cells, and this effect was abolished with DPI treatment. LPS treatment increased ROS production, whereas CLA reduced LPS-induced ROS production. LPS increased both TNF-α production and NF-κB activity, whereas t10c12-CLA reduced TNF-α production and NF-κB activity in LPS-stimulated RAW 264.7 cells. DPI treatment suppressed LPS-induced ROS production and NF-κB activity. Moreover, DPI enhanced the inhibitory effects of t10c12-CLA on TNF-α production and NF-κB activation in LPS-stimulated RAW 264.7 cells. However, neither t10c12-CLA nor DPI affected PPARγ activity in LPSstimulated RAW 264.7 cells. Taken together, these data indicate that t10c12-CLA induces TNF-α production by increasing ROS production in LPS-naive RAW 264.7 cells, which is mediated by the enhancement of NF-κB activity via PPARγ activation. By contrast, t10c12-CLA suppresses TNF-α production by inhibiting ROS production and NF- κB activation via a PPARγ-independent pathway in LPS-stimulated RAW 264.7 cells. These results suggest that t10c12- CLA can modulate TNF-α production and NF-κB activation through formation of ROS in RAW 264.7 macrophages.

Troglitazone and Δ2Troglitazone enhance adiponectin expression in monocytes/macrophages through the AMP-activated protein kinase pathway.

Accumulating evidence indicates that the regimen to increase adiponectin will provide a novel therapeutic strategy for inflammation and cardiovascular disorders. Here, we tested the effect of troglitazone (TG) and its newly synthesized derivative, 5-[4-(6-hydroxy-2,5,7,8-tetramethyl-chroman-2-yl-methoxy)-benzylidene]-2,4-thiazolidinedione (Δ2troglitazone, (Δ2TG)), on the adiponectin expression in monocytes/macrophages and the relative mechanisms. The expression of adiponectin was located in macrophages of atherosclerotic lesions from patients and cholesterol-fed rabbits. TG and Δ2TG enhanced adiponectin mRNA and protein expression in THP-1 cells by quantitative real-time PCR, Western blot, and immunocytochemistry. TG induced adiponectin mRNA expression through a PPARγ-dependent pathway whereas Δ2TG enhanced adiponectin mRNA expression through a PPARγ-independent pathway in THP-1 cells. Both TG and Δ2TG enhanced adiponectin mRNA expression through AMP-activated protein kinase (AMPK) activation. TG and Δ2TG decreased the adhesion of THP-1 cells to TNF-α-treated HUVECs and the inhibitory effect was abolished by specific antiadiponectin antibodies. TG- and Δ2TG-induced suppression on monocyte adhesion were inhibited by a selective AMPK inhibitor compound C. Our data suggest that the inhibitory effect of TG and Δ2TG on monocyte adhesion might be at least in part through de novo adiponectin expression and activation of an AMPK-dependent pathway, which might play an important role in anti-inflammation and antiatherosclerosis.

188 DRUG-ELUTING STENTS CONTAINING IBUPROFEN: A NOVEL STRATEGY TO REDUCE RESTENOSIS AND PREVENT LATE IN-STENT THROMBOSIS

<h3>Introduction</h3> Drug-eluting stents (DES) have reduced the rates of restenosis to less than 10%; however, increased incidence of late in-stent thrombosis, is now a major clinical problem of using DES for its consequences in terms of morbidity and mortality. Interference with the process of re-endothelialization appears to play a major role in this latent complication. Thus, the search for molecular compounds capable of preventing restenosis that also can reduce the risk of late in-stent thrombosis is of utmost importance. Previously, we have shown that certain non-steroidal anti-inflammatory drugs (NSAIDs) are capable of inhibiting rat vascular smooth muscle cell proliferation, a key element in the process of restenosis post stent deployment. Here, we evaluated whether proliferation and migration of human coronary artery smooth muscle cells (HCASMCs) is reduced by the NSAID, ibuprofen, whether this correlated with changes in the phenotype of HCASMCs and the potential molecular mechanisms involved. Also, we investigated whether human coronary artery endothelial cell (HCAEC) migration was affected by ibuprofen. <h3>Methods</h3> Cell proliferation was evaluated by trypan blue exclusion. Cell migration was assessed by wound healing assay and by time lapse videomicroscopy. Protein expression was assessed by immunoblotting and morphology by immunocytochemistry. The involvement of the PPARγ pathway was studied with the selective agonist, troglitazone, and with the PPARγ antagonists, PGF2α and GW9662. <h3>Results</h3> We show that ibuprofen inhibited proliferation as well as migration of HCASMCs in a dose-dependent manner (IC₅₀ of 680 and 410 µM, respectively). Interestingly, we found that ibuprofen induced a switch in HCASMCs towards a differentiated phenotype, with a characteristic spindle shape, and a significant increase in the expression of contractile protein markers, e.g. SMα-actin, SM22α and F-actin. PPARγ antagonists, PGF2α and GW9662, almost completely abrogated the proliferative and migratory responses of HCASMCs, as well the changes in morphology. However, PPARγ antagonists did not affect the expression pattern of contractile proteins, suggesting that these effects are mediated by a PPARγ-independent pathway. Importantly, ibuprofen did not affect migration of HCAECs even at high doses (up to 1000 µM), suggesting that the effects of ibuprofen on HCASMCs are selective. Interestingly, the protein levels of PPARγ are higher in HCAECs compared to those in HCASMCs. <h3>Conclusions</h3> Taken together, our results suggest that ibuprofen could be an effective treatment for the development of novel DES. Its effects on migration and proliferation of HCASMCs, and induction of a differentiated phenotype, could translate into reduced rates of restenosis, whilst the lack of an effect on HCAEC migration could result in a reduced risk of late in-stent thrombosis. Further studies in animal models will be required to evaluate the performance of an ibuprofen DES <i>in vivo</i>.

Telmisartan ameliorates insulin sensitivity by activating the AMPK/SIRT1 pathway in skeletal muscle of obese db/db mice

BackgroundTelmisartan is a well-established angiotensin II type 1 receptor blocker that improves insulin sensitivity in animal models of obesity and insulin resistance, as well as in humans. Telmisartan has been reported to function as a partial agonist of the peroxisome proliferator-activated receptor (PPAR) γ, which is also targeted by the nicotinamide adenine dinucleotide (NAD)-dependent deacetylase (SIRT1). Here, we investigated the pathways through which telmisartan acts on skeletal muscle, in vitro as well as in vivo.MethodsNine-week-old male db/db mice were fed a 60% high-fat diet, with orally administrated either vehicle (carboxymethyl-cellulose, CMC), 5 mg/kg telmisartan, or 5 mg/kg telmisartan and 1 mg/kg GW9662, a selective irreversible antagonist of PPARγ, for 5 weeks. Effects of telmisartan on Sirt1 mRNA, AMPK phosphorylation, and NAD+/NADH ratio were determined in C2C12 cultured myocytes.Results and discussionTelmisartan treatment improved insulin sensitivity in obese db/db mice fed a high-fat diet and led to reduction in the size of hypertrophic pancreatic islets in these mice. Moreover, in vitro treatment with telmisartan led to increased expression of Sirt1 mRNA in C2C12 skeletal muscle cells; the increase in Sirt1 mRNA in telmisartan-treated C2C12 myoblasts occurred concomitantly with an increase in AMPK phosphorylation, an increase in NAD+/NADH ratio, and increases in the mRNA levels of PGC1α, FATP1, ACO, and GLUT4.ConclusionsOur results indicate that telmisartan acts through a PPARγ-independent pathway, but at least partially exerts its effects by acting directly on skeletal muscle AMPK/SIRT1 pathways.

Cytotoxicity of 15-Deoxy-Δ12,14-prostaglandin J2 through PPARγ-independent Pathway and the Involvement of the JNK and Akt Pathway in Renal Cell Carcinoma

Introduction: Agonists of peroxisome proliferator-activated receptor gamma (PPARγ) have been examined as chemopreventive and chemotherapeutic agents. The aim was to investigate the cytotoxicity and action mechanisms of 15-deoxy-Δ12,14-prostaglandin J2 (15d-PGJ2), one of endogenous ligands for PPARγ, in terms of PPARγ-dependency and the mitogen-activated protein kinase (MAPK) and Akt pathway in three human renal cell carcinoma (RCC)-derived cell lines.Methods: 786-O, Caki-2 and ACHN cells were used as human RCC-derived cell lines. Cell viability and caspase-3 activity was detected by fluorescent reagents, and chromatin-condensation was observed with a brightfield fluorescent microscope after staining cells with Hoechst33342. The expression levels of proteins were detected by Western blot analysis.Results: 15d-PGJ2 showed cytotoxicity in dose-dependent manner. 15d-PGJ2 induced chromatin-condensation and elevated caspase-3 activity, and the cell viability was restored by co-treatment with a pan-caspase inhibitor, Z-VAD-FMK, indicating the involvement of caspase-dependent apoptosis. The cytotoxicity was not impaired by a PPARγ inhibitor, GW9662, suggesting that 15d-PGJ2 exerted the cytotoxicity in a PPARγ-independent manner. Some antioxidants rescued cells from cell death induced by 15d-PGJ2, but some did not, suggesting that reactive oxygen species (ROS) did not contribute to the apoptosis. 15d-PGJ2 also increased the expression levels of phospho-c-Jun N terminal kinase (JNK) in Caki-2 cells, and decreased those of phospho-Akt in 786-O cells, indicating that the JNK MAPK and the Akt pathways participated in the anticancer effects of 15d-PGJ2 in some cell lines.Conclusion: 15d-PGJ2 exerted cytotoxic effects accompanying caspase-dependent apoptosis, and this effect was elicited in a PPARγ-independent manner in three cell lines. In addition, the JNK MAPK and Akt pathway was involved in the cytotoxicity of 15d-PGJ2 to some extent in some cell line. Therefore, our study showed the 15d-PGJ2 to potentially be an interesting approach for RCC treatment.

Cytotoxicity of troglitazone through PPARγ-independent pathway and p38 MAPK pathway in renal cell carcinoma

Agonists of peroxisome proliferator-activated receptor gamma (PPARγ) have been examined as chemopreventive and chemotherapeutic agents. The aim was to investigate the cytotoxicity of troglitazone (TGZ) and its mechanisms in terms of PPARγ dependency and the p38 mitogen-activated protein kinase (MAPK) pathway in three human renal cell carcinoma (RCC) cell lines, 786-O, Caki-2 and ACHN cells. TGZ induced apoptosis and exerted cytotoxicity in a PPARγ-independent manner. We demonstrated that TGZ activated the p38 MAPK pathway and was involved in the cytotoxicity of TGZ. It was also revealed that TGZ induced G 2/M cell cycle arrest through activation of p38 MAPK.

New troglitazone derivatives devoid of PPARγ agonist activity display an increased antiproliferative effect in both hormone-dependent and hormone-independent breast cancer cell lines

Numerous recent studies indicate that most anticancer effects of PPARγ agonists like thiazolidinediones are the result of PPARγ-independent pathways. These conclusions were obtained by several approaches including the use of thiazolidinedione derivatives like Δ2-Troglitazone (Δ2-TGZ) that does not activate PPARγ. Since biotinylation has been proposed as a mechanism able to increase the specificity of drug delivery to cancer cells which could express a high level of vitamin receptor, a biotinylated derivative of Δ2-TGZ (bΔ2-TGZ) has been synthetized. In the present article, we have studied the in vitro effects of this molecule on both hormone-dependent (MCF-7) and hormone-independent (MDA-MB-231) breast cancer cells. In both cell lines, bΔ2-TGZ was more efficient than Δ2-TGZ to decrease cell viability. bΔ2-TGZ was also more potent than Δ2-TGZ to induce the proteasomal degradation of cyclin D1 in both cell lines and those of ERα in MCF-7 cells. However, in competition experiments, the presence of free biotin in the culture medium did not decrease the antiproliferative action of bΔ2-TGZ. Besides, other compounds that had no biotin but that were substituted at the same position of the phenolic group of the chromane moiety of Δ2-TGZ decreased cell viability similarly to bΔ2-TGZ. Hence, we concluded that the increased antiproliferative action of bΔ2-TGZ was not due to biotin itself but to the functionalization of the terminal hydroxyl group. This should be taken into account for the design of new thiazolidinedione derivatives able to affect not only hormone-dependent but also hormone-independent breast cancer cells in a PPARγ-independent pathway.

Troglitazone increases expression of E-cadherin and claudin 4 in human pancreatic cancer cells

We examined the effects of troglitazone on expression of E-cadherin and claudin 4 in human pancreatic cancer cells. Troglitazone dose-dependently increased expression of E-cadherin and claudin 4 mRNA and protein in PK-1 cells. Snail, Slug and ZEB1, mRNAs were not changed by troglitazone, indicating that these three transcriptional repressors would not play a role in the induction of E-cadherin by troglitazone. GW9662, a PPARγ antagonist, failed to block the increased expression of E-cadherin or claudin 4 mRNA, suggesting a PPARγ-independent pathway. A MEK inhibitor, U0126, increased E-cadherin or claudin 4 mRNA and protein expression, and potently inhibited cell invasion. Because troglitazone down-regulates MEK-ERK signaling and inhibit cell invasion in PK-1 as shown in our previous study, these results suggest that troglitazone increases expression of E-cadherin and claudin 4 possibly through inhibition of MEK-ERK signaling in pancreatic cancer cells, which might be involved in the troglitazone-induced inhibition of cell invasive activity.

Rosiglitazone reduces cell invasiveness by inducing MKP-1 in human U87MG glioma cells

We sought to investigate the molecular mechanisms by which rosiglitazone (RGZ) inhibits cell invasion in human glioma cells. In this study, we found that RGZ attenuated MMP-2 protein levels, MMP-2 gelatinolytic activity, and cell invasiveness through a PPAR-γ independent pathway. RGZ increased mitogen activated protein kinase phosphatase-1 (MKP-1) expression. The addition of triptolide (a diterpenoid triepoxide, which blocked MKP-1 induction) abolished the inhibitory effects by RGZ. Furthermore, we demonstrated that the knock down of MKP-1 by MKP-1 specific small interference RNA reversed the reduction of MMP-2 secretion, and of cell invasiveness by RGZ. In contrast, the stable expression of MKP-1 in glioma cell lines decreased MMP-2 activity and cell invasiveness. These results suggest that RGZ may mediate the inhibitory effects through MKP-1 induction. Thus, MKP-1 could be a potential target in glioma therapy.

Rosiglitazone inhibits α4 nicotinic acetylcholine receptor expression in human lung carcinoma cells through peroxisome proliferator-activated receptor γ-independent signals

We and others have shown previously that nicotine, a major component of tobacco, stimulates non-small cell lung carcinoma (NSCLC) proliferation through nicotinic acetylcholine receptor (nAChR)-mediated signals. Activation of peroxisome proliferator-activated receptor gamma (PPARgamma) has been shown to inhibit NSCLC cell growth, but the exact mechanisms responsible for this effect remain incompletely defined. Herein, we show that nicotine induces NSCLC cell proliferation in part through alpha4 nAChR, prompting us to explore the effects of rosiglitazone, a synthetic PPARgamma ligand, on the expression of this receptor. Rosiglitazone inhibited the expression of alpha4 nAChR, but this effect was through a PPARgamma-independent pathway, because GW9662, an antagonist of PPARgamma, and the transfection of cells with PPARgamma small interfering RNA failed to abolish the response. The inhibitory effect of rosiglitazone on alpha4 nAChR expression was accompanied by phosphorylation of p38 mitogen-activated protein kinase and extracellular signal-regulated kinase 1/2 and down-regulation of Akt phosphorylation. These signals mediated the inhibitory effects of rosiglitazone on alpha4 nAChR expression because chemical inhibitors prevented the effect. Rosiglitazone was also found to stimulate p53, a tumor suppressor known to mediate some of the effects of nicotine. Interestingly, p53 up-regulation was needed for rosiglitazone-induced inhibition of alpha4 nAChR. Thus, rosiglitazone inhibits alpha4 nAChR expression in NSCLC cells through activation of extracellular signal-regulated kinase and p38 mitogen-activated protein kinase, which triggers induction of p53. Finally, like others, we found that nicotine stimulated the expression of alpha4 nAChR. This process was also inhibited by rosiglitazone through similar pathways.

Loss of XIAP sensitizes colon cancer cells to PPARγ independent antitumor effects of troglitazone and 15-PGJ2

We investigated whether the anticancer effect of a combination of XIAP down-regulation and PPAR γ activation on colon cancer is PPARγ receptor dependent. HCT116-XIAP +/+ cells and HCT116-XIAP −/− cells were treated with troglitazone or 15-deoxy-Δ 12,14-prostaglandin J2 (15-PGJ2) with or without prior exposure to PPARγ inhibitor GW9662. Cell proliferation and apoptosis was evaluated. Athymic mice carrying HCT116-XIAP −/− cells-derived tumors were treated with troglitazone in the presence or absence of GW9662. Inhibition of cell proliferation and induction of apoptosis by troglitazone and 15-PGJ2 were more prominent in HCT116-XIAP −/− cells. PPARγ ligand-induced growth inhibition, apoptosis, caspase and PARP cleavage could not be blocked by GW9662. Troglitazone significantly retarded growth of xenograft tumors and this effect was not blocked by GW9662. Marked apoptosis and an up-regulation of E-cadherin were observed in xenograft tumor tissues, and GW9662 did not affect these effects. Thus, a combination of XIAP down-regulation and PPARγ ligands exert a significant anticancer effect in colon cancer via a PPARγ independent pathway.

Deoxyelephantopin inhibits cancer cell proliferation and functions as a selective partial agonist against PPARγ

Deoxyelephantopin (ESD) was reported to potentiate apoptosis, inhibit invasion and abolish osteoclastogenesis but no target protein was disclosed. Here, we discovered that ESD could significantly inhibit the proliferation of different cancer cells and induce apoptosis and cell cycle arrest at G2/M phase in HeLa cell. Moreover, biochemical and biophysical assays revealed that ESD acted as a specific partial agonist against PPARγ. Molecular docking with site-directed mutagenesis analyses indicated that ESD functioned as a partial agonist of PPARγ by adopting a distinct binding mode to PPARγ compared with rosiglitazone. The PPARγ knockdown results indicated that the inhibition of ESD against the cancer cell proliferation is more possibly through PPARγ-independent pathway and our findings might supply potent binding features for ESD/PPARγ interaction at atomic level, and shed light on the potential acting target information for this natural compound.

Differentiation of Human Circulating Fibrocytes as Mediated by Transforming Growth Factor-β and Peroxisome Proliferator-activated Receptor γ

Fibrocytes are a distinct population of fibroblast-like progenitor cells in peripheral blood that have recently been shown to possess plasticity to differentiate along mesenchymal lineages, including commitment to myofibroblast and adipocyte cells. Here, we demonstrated that transforming growth factor (TGF) beta1 drives fibrocyte-to-myofibroblast differentiation through activating Smad2/3 and SAPK/JNK MAPK pathways, which in turn stimulates alpha-smooth muscle actin expression. We determined that SAPK/JNK signaling acts in a positive feedback loop to modulate Smad2/3 nuclear availability and Smad2/3-dependent transcription. Conversely, fibrocyte-to-adipocyte differentiation is driven by the peroxisome proliferator-activated receptor (PPAR) gamma agonist troglitazone, which is associated with cytoplasmic lipid accumulation and induction of aP2. Treatment with troglitazone also disrupted TGF beta 1-activated SAPK/JNK signaling, leading to decreased Smad2/3 transactivation activity and alpha-smooth muscle actin expression. Interestingly, TGF beta 1 was demonstrated to have reciprocal inhibition on fibrocyte differentiation to adipocytes. By activating SAPK/JNK signaling, which is normally suppressed during adipogenesis, PPARgamma-dependent transactivation activity and induction of aP2 expression were disrupted. Taken together, within the context of the local microenvironmental niche, the delicate balance of PPARgamma and TGF beta 1 activation drives the selection of an adipocyte or myofibroblast differentiation pathway through SAPK/JNK signaling.

Stimulation of the peroxisome proliferator-activated receptor γ (PPARγ) and the expression of selected blood monocyte cytokine genes in diabetic macroangiopathy

Monocytes and macrophages play a key role in the progression of atheromatous changes. The peroxisome proliferator-activated receptor γ (PPARγ) can limit macroangiopathy through the control of cytokine transcription. The objectives of this study were to examine the influence of PPARγ and its agonist (rosiglitazone) on the TNFα, IL-6, IL-8 and IL-10 gene expression in monocytes of patients with diabetic macroangiopathy and to analyse obtained results in context of selected atherogenic factors ant direct indicators of endothelial lesion. TNFα, IL-6, IL-8, IL-10 and PPARγ gene expression was assessed in peripheral blood monocytes in 45 patients with type 2 diabetes before and following 22 weeks of rosiglitazone therapy (real-time PCR [Applied Biosystems]). As indicators of endothelial lesion, concentration of thrombomodulin (immunoassay [Diagnostica Stago ®]) and amount of circulating blood endothelial cells (immunofluorescence method with MoAb CLB-HEC19) were determined. Following rosiglitazone therapy, a statistically significant downward tendency of TNFα ( p = 0.026) and IL-8 ( p = 0.008) gene expression was noted. Before and following rosiglitazone treatment, PPARγ, IL-6 and IL-10 gene expression was undetectable in studied monocytes in vivo. In conclusion, TNFα and IL-8 play an important role in monocyte atherogenic activity. Rosiglitazone reduces monocyte proinflammatory readiness by influencing the expression of selected atherogenic cytokines (PPARγ-independent pathway).

A Peroxisome Proliferator-activated Receptor-γ Agonist, Troglitazone, Facilitates Caspase-8 and -9 Activities by Increasing the Enzymatic Activity of Protein-tyrosine Phosphatase-1B on Human Glioma Cells

Despite dramatic advances in adjuvant therapies, patients with malignant glioma face a bleak prognosis. Because many adjuvant therapies seek to induce glioma apoptosis, strategies that lower thresholds for the induction of apoptosis may improve patient outcomes. Therefore, elucidation of the biological mechanisms that underlie resistance to current therapies is needed to develop new therapeutic strategies. Here we proposed a novel mechanism of proapoptotic effect induced by a pharmacological peroxisome proliferator-activated receptor-gamma (PPARgamma) agonist, troglitazone, that facilitates caspase signaling in human glioma cells. Troglitazone activates protein-tyrosine phosphatase (PTP)-1B, which subsequently reduces phosphotyrosine 705 STAT3 (pY705-STAT3) via a PPARgamma-independent pathway. Reduction of pY705-STAT3 in glioma cells caused down-regulation of FLIP (FADD-like IL-1beta-converting enzyme-inhibitory protein) and Bcl-2. Furthermore, troglitazone induced Ser-392 phosphorylation of p53 via a PPARgamma-dependent pathway and up-regulation of Bax in a p53 wild-type glioma. When given with tumor necrosis factor-related apoptosis-inducing ligand or caspase-dependent chemotherapeutic agents, such as etoposide and paclitaxel, troglitazone exhibited a synergistic effect by facilitating caspase-8/9 activities. A PPARgamma antagonist, GW9662, did not block this effect, although a PTP inhibitor abrogated it. Knockdown of STAT3 by STAT3-small interfering RNA negated the inhibitory effect of PTP inhibitor on troglitazone, indicating that troglitazone uses a STAT3 inactivation mechanism that makes caspase-8/9 activities susceptible to cytotoxic agents in glioma cells and that PTP1B plays a critical role in the down-regulation of activated STAT3, as well as FLIP and Bcl-2. When taken with caspase-dependent anti-neoplastic agents, troglitazone may be a promising drug for use against malignant gliomas because it facilitates the caspase cascade, thereby lowering thresholds for the apoptosis induction of glioma cells.

C-Jun N-terminal kinase is involved in the suppression of adiponectin expression by TNF-α in 3T3-L1 adipocytes

Adiponectin, one of adipokines that is secreted from adipocytes, plays an important role in the regulation of glucose and lipid metabolism. Paradoxically, serum concentrations of adiponectin are decreased in obese and type 2 diabetic patients, although it is produced in adipose tissue. On the other hand, plasma TNF-α levels are increased in such subjects. In the present study, the mechanism by which adiponectin is regulated by TNF-α was investigated. The decreased adiponectin mRNA levels by TNF-α were partially recovered by treatment with a c-Jun N-terminal kinase (JNK) inhibitor or the PPAR-γ agonist rosiglitazone in 3T3-L1 adipocytes. Interestingly, however, cotreatment with the JNK inhibitor and rosiglitazone led to a recovery of TNF-α-mediated adiponectin suppression to the control level. The JNK inhibitor regulated the expression of adiponectin by the increase of PPAR-γ DNA binding activity and the recovery of its mRNA expression while rosiglitazone acted via a PPAR-γ independent pathway which remains to be elucidated. These findings suggest that the JNK signaling pathway, activated by TNF-α, is involved in the regulation of adiponectin expression.