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Sample records for inhibitors induce peroxisome

  1. Proteasome inhibitors induce peroxisome proliferator-activated receptor transactivation through RXR accumulation and a protein kinase C-dependent pathway

    SciTech Connect

    Tsao, W.-C.; Wu, H.-M.; Chi, K.-H.; Chang, Y.-H.; Lin, W.-W. . E-mail: wwl@ha.mc.ntu.edu.tw

    2005-03-10

    Peroxisome proliferator-activated receptor {gamma} (PPAR{gamma}), a member of nuclear hormone receptors, forms a heterodimeric DNA binding complex with retinoid X receptor (RXR) and serves as a transcriptional regulator of gene expression. In this study, using luciferase assay of a reporter gene containing PPAR response element (PPRE), we found PPRE transactivity was additively induced by PPAR{gamma} activator (15dPGJ{sub 2}) and RXR activator (9-cis retinoic acid, 9-cis RA). Proteasome inhibitors MG132 and MG262 also stimulate PPRE transactivity in a concentration-dependent manner, and this effect is synergistic to 15dPGJ{sub 2} and 9-cis RA. PKC activation by 12-myristate 13-acetate (PMA) and ingenol 3,20-dibenzoate (IDB) also led to an increased PPRE activation, and this action was additive to PPAR{gamma} activators and 9-cis RA, but not to proteasome inhibitors. Results indicate that the PPAR{gamma} enhancing effect of proteasome inhibitors was attributed to redox-sensitive PKC activation. Western blot analysis showed that the protein level of RXR{alpha}, but not PPAR{gamma}, RXR{beta}, or PKC isoforms, was accumulated in the presence of proteasome inhibitors. Taken together, we conclude that proteasome inhibitors can upregulate PPRE activity through RXR{alpha} accumulation and a PKC-dependent pathway. The former is due to inhibition of RXR{alpha} degradation through ubiquitin-dependent proteasome system, while the latter is mediated by reactive oxygen species (ROS) production.

  2. HYDROGEN PEROXIDE INDUCED OXIDATION OF PEROXISOMAL MALATE SYNTHASE AND CATALASE.

    E-print Network

    Simha, Rahul

    Page 1 HYDROGEN PEROXIDE INDUCED OXIDATION OF PEROXISOMAL MALATE SYNTHASE AND CATALASE. Pria Anand1 locations within the proteins' structures. Key words: biotin hydrazide, carbonylation, catalase, hydrogen peroxide, glyoxysome, MALDI- TOF MS, malate synthase, mass spectroscopy, peroxisome, protein oxidation

  3. Inhibition of peroxisome proliferator-activated receptor (PPAR)-mediated keratinocyte differentiation by lipoxygenase inhibitors.

    PubMed Central

    Thuillier, Philippe; Brash, Alan R; Kehrer, James P; Stimmel, Julie B; Leesnitzer, Lisa M; Yang, Peiying; Newman, Robert A; Fischer, Susan M

    2002-01-01

    Lipoxygenase (LOX) metabolites from arachidonic acid and linoleic acid have been implicated in atherosclerosis, inflammation, keratinocyte differentiation and tumour progression. We previously showed that peroxisome proliferator-activated receptors (PPARs) play a role in keratinocyte differentiation and that the PPARalpha ligand 8S-hydroxyeicosatetraenoic acid is important in this process. We hypothesized that blocking LOX activity would block PPAR-mediated keratinocyte differentiation. Three LOX inhibitors, nordihydroguaiaretic acid, quercetin and morin, were studied for their effects on primary keratinocyte differentiation and PPAR activity. All three LOX inhibitors blocked calcium-induced expression of the differentiation marker keratin 1. In addition, activity of a PPAR-responsive element was inhibited in the presence of all three inhibitors, and this effect was mediated primarily through PPARalpha and PPARgamma. LOX inhibitors decreased the activity of a chimaeric PPAR-Gal4-ligand-binding domain reporter system and this effect was reversed by addition of PPAR ligands. Ligand-binding studies revealed that the LOX inhibitors bind directly to PPARs and demonstrate a novel mechanism for these inhibitors in altering PPAR-mediated gene expression. PMID:12069687

  4. Dysfunction of peroxisomes in twitcher mice brain: A possible mechanism of psychosine-induced disease

    SciTech Connect

    Haq, Ehtishamul; Contreras, Miguel A.; Giri, Shailendra; Singh, Inderjit; Singh, Avtar K. . E-mail: singha@musc.edu

    2006-04-28

    Psychosine (galactosylsphingosine) accumulates in Brain of Krabbe disease (KD) patients as well as twitcher mice, a murine model of KD, resulting in loss of oligodendrocytes and myelin. This study documents progressive loss of peroxisomal proteins/functions and induction of expression of inflammatory cytokine TNF-{alpha} in twitcher brain. The observed decrease in peroxisomal proteins was accompanied by decreased level of peroxisome proliferator-activated receptor-alpha (PPAR-{alpha}), one of the transcription factors required for expression of peroxisomal protein genes. The role of psychosine in down-regulation of PPAR-{alpha} activity was further supported by decreased PPAR-{alpha} mediated PPRE transcriptional activity in cells transfected with PPAR-{alpha} and PPRE reporters. The psychosine-induced down-regulation of PPAR activity and cell death was attenuated by sPLA{sub 2} inhibitor. Therefore, this study provides First evidence of peroxisomal abnormality in a lysosomal disorder, suggesting that such dysfunction of peroxisomes may play a role in the pathogenesis of Krabbe disease.

  5. Peroxisomal fatty acid oxidation and inhibitors of the mitochondrial carnitine palmitoyltransferase I in isolated rat hepatocytes.

    PubMed Central

    Skorin, C; Necochea, C; Johow, V; Soto, U; Grau, A M; Bremer, J; Leighton, F

    1992-01-01

    Fatty acid oxidation was studied in the presence of inhibitors of carnitine palmitoyltransferase I (CPT I), in normal and in peroxisome-proliferated rat hepatocytes. The oxidation decreased in mitochondria, as expected, but in peroxisomes it increased. These two effects were seen, in variable proportions, with (+)-decanoylcarnitine, 2-tetradecylglycidic acid (TDGA) and etomoxir. The decrease in mitochondrial oxidation (ketogenesis) affected saturated fatty acids with 12 or more carbon atoms, whereas the increase in peroxisomal oxidation (H2O2 production) affected saturated fatty acids with 8 or more carbon atoms. The peroxisomal increase was sensitive to chlorpromazine, a peroxisomal inhibitor. To study possible mechanisms, palmitoyl-, octanoyl- and acetyl-carnitine acyltransferase activities were measured, in homogenates and in subcellular fractions from control and TDGA-treated cells. The palmitoylcarnitine acyltransferase was inhibited, as expected, but the octanoyltransferase activity also decreased. The CoA derivative of TDGA was synthesized and tentatively identified as being responsible for inhibition of the octanoylcarnitine acyltransferase. These results show that inhibitors of the mitochondrial CPT I may also inhibit the peroxisomal octanoyl transferase; they also support the hypothesis that the octanoyltransferase has the capacity to control or regulate peroxisomal fatty acid oxidation. PMID:1736904

  6. Expression level of methanol-inducible peroxisomal proteins and peroxisome morphology are affected by oxygen conditions and mitochondrial respiratory pathway function in the methylotrophic yeast Candida boidinii.

    PubMed

    Fujimura, Shuki; Yurimoto, Hiroya; Kurimoto, Shota; Matsufuji, Yoshimi; Ito, Takashi; Hayakawa, Takashi; Tomizuka, Noboru; Sakai, Yasuyoshi; Nakagawa, Tomoyuki

    2013-06-01

    In the methylotrophic yeast, Candida boidinii, methanol-inducible peroxisomal proteins, for example alcohol oxidase (AOD), dihydroxyacetone synthase (DAS), and peroxisomal glutathione peroxidase (Pmp20), were induced only under aerobic conditions, while expression of PMP47 encoding peroxisomal integral membrane protein Pmp47 was independent of oxygen conditions. Expression of the methanol-inducible peroxisomal enzymes was repressed by inhibition of the mitochondrial respiratory chain. In the respiratory-deficient (?0) mutant strain, their induction was at very low levels despite the presence of oxygen, whereas the expression of PMP47 was unaffected. Taken together, these facts indicate that C. boidinii can sense oxygen conditions, and that mitochondrial respiratory function may have a profound effect on induction of methanol-inducible gene expression of peroxisomal proteins. Peroxisome morphology was also affected by oxygen conditions and respiratory function. Under hypoxic conditions or respiration-inhibited conditions, cells induced by methanol contained small peroxisomes, indicating that peroxisome biogenesis and the protein import machinery were not affected by oxygen conditions but that peroxisome morphology was dependent on induction of peroxisomal matrix proteins. PMID:23448597

  7. Molecular basis of non-responsiveness to peroxisome proliferators: the guinea-pig PPARalpha is functional and mediates peroxisome proliferator-induced hypolipidaemia.

    PubMed Central

    Bell, A R; Savory, R; Horley, N J; Choudhury, A I; Dickins, M; Gray, T J; Salter, A M; Bell, D R

    1998-01-01

    The guinea pig does not undergo peroxisome proliferation in response to peroxisome proliferators, in contrast with other rodents. To understand the molecular basis of this phenotype, the peroxisome proliferator activated receptor alpha (PPARalpha) from guinea-pig liver was cloned; it encodes a protein of 467 amino acid residues that is similar to rodent and human PPARalpha. The guinea-pig PPARalpha showed a high substitution rate: maximum likelihood analysis was consistent with rodent monophyly, but could not exclude rodent polyphyly (P approximately 0.06). The guinea-pig PPARalpha cDNA was expressed in 293 cells and mediated the induction of the luciferase reporter gene by the peroxisome proliferator, Wy-14,643, dependent on the presence of a peroxisome proliferator response element. Moreover the PPARalpha RNA and protein were expressed in guinea-pig liver, although at lower levels than in a species which is responsive to peroxisome proliferators, the mouse. To determine whether the guinea-pig PPARalpha mediated any physiological effects, guinea pigs were exposed to two selective PPARalpha agonists, Wy-14, 643 and methylclofenapate; both compounds induced hypolipidaemia. Thus the guinea pig is a useful model for human responses to peroxisome proliferators. PMID:9620871

  8. Activation of Peroxisome Proliferator-Activated Receptor by Substituted Urea-Derived Soluble Epoxide Hydrolase Inhibitors

    E-print Network

    Hammock, Bruce D.

    effectively inhibiting sEH, increased peroxisome proliferator- activated receptor (PPAR) activity. In PPAR-adamantanyl-N -dodecanoic acid urea (AUDA) produced 6- and 3-fold increases, respec- tively, in PPAR activation. Neither CUDA nor AUDA activated PPAR or PPAR directly, indicating selectivity for PPAR . CUDA did not alter

  9. Peroxisome proliferator-activated receptor {alpha} agonist-induced down-regulation of hepatic glucocorticoid receptor expression in SD rats

    SciTech Connect

    Chen Xiang; Li Ming; Sun Weiping; Bi Yan; Cai Mengyin; Liang Hua; Yu Qiuqiong; He Xiaoying; Weng Jianping

    2008-04-18

    It was reported that glucocorticoid production was inhibited by fenofibrate through suppression of type-1 11{beta}-hydroxysteroid dehydrogenase gene expression in liver. The inhibition might be a negative-feedback regulation of glucocorticoid receptor (GR) activity by peroxisome proliferator-activated receptor alpha (PPAR{alpha}), which is quickly induced by glucocorticoid in the liver. However, it is not clear if GR expression is changed by fenofibrate-induced PPAR{alpha} activation. In this study, we tested this possibility in the liver of Sprague-Dawley rats. GR expression was reduced by fenofibrate in a time- and does-dependent manner. The inhibition was observed in liver, but not in fat and muscle. The corticosterone level in the blood was increased significantly by fenofibrate. These effects of fenofibrate were abolished by PPAR{alpha} inhibitor MK886, suggesting that fenofibrate activated through PPAR{alpha}. In conclusion, inhibition of GR expression may represent a new molecular mechanism for the negative feedback regulation of GR activity by PPAR{alpha}.

  10. Protective Role for Tissue Inhibitor of Metalloproteinase-4, a Novel Peroxisome Proliferator-Activated Receptor-? Target Gene, in Smooth Muscle in Deoxycorticosterone Acetate-Salt Hypertension.

    PubMed

    Ketsawatsomkron, Pimonrat; Keen, Henry L; Davis, Deborah R; Lu, Ko-Ting; Stump, Madeliene; De Silva, T Michael; Hilzendeger, Aline M; Grobe, Justin L; Faraci, Frank M; Sigmund, Curt D

    2016-01-01

    Loss of peroxisome proliferator-activated receptor-? (PPAR?) function causes hypertension, whereas its activation lowers blood pressure. Evidence suggests that these effects may be attributable to PPAR? activity in the vasculature. However, the specific transcriptional targets of PPAR? in vessels remain largely unidentified. In this study, we examined the role of smooth muscle PPAR? during salt-sensitive hypertension and investigated its transcriptional targets and functional effect. Transgenic mice expressing dominant-negative PPAR? (S-P467L) in smooth muscle cells were more prone to deoxycorticosterone acetate-salt-induced hypertension and mesenteric arterial dysfunction compared with nontransgenic controls. Despite similar morphometry at baseline, vascular remodeling in conduit and small arteries was enhanced in S-P467L after deoxycorticosterone acetate-salt treatment. Gene expression profiling in aorta and mesenteric arteries revealed significantly decreased expression of tissue inhibitor of metalloproteinase-4 (TIMP-4) in S-P467L. Expression of TIMP-4 was increased by deoxycorticosterone acetate-salt treatment, but this increase was ablated in S-P467L. Interference with PPAR? activity either by treatment with a PPAR? inhibitor, GW9662, or by expressing P467L PPAR? markedly suppressed TIMP-4 in primary smooth muscle cells. PPAR? binds to a PPAR response element (PPRE) in chromatin close to the TIMP-4 gene in smooth muscle cells, suggesting that TIMP-4 is a novel target of PPAR?. The interference with PPAR? and decrease in TIMP-4 were accompanied by an increase in total matrix metalloproteinase activity. PPAR?-mediated loss of TIMP-4 increased, whereas overexpression of TIMP-4 decreased smooth muscle cell migration in a scratch assay. Our findings highlight a protective mechanism induced by PPAR? in deoxycorticosterone acetate-salt treatment, establishing a novel mechanistic link between PPAR? and TIMP-4. PMID:26597823

  11. Histone deacetylase inhibitor upregulates peroxisomal fatty acid oxidation and inhibits apoptotic cell death in abcd1-deficient glial cells.

    PubMed

    Singh, Jaspreet; Khan, Mushfiquddin; Pujol, Aurora; Baarine, Mauhamad; Singh, Inderjit

    2013-01-01

    In X-ALD, mutation/deletion of ALD gene (ABCD1) and the resultant very long chain fatty acid (VLCFA) derangement has dramatically opposing effects in astrocytes and oligodendrocytes. While loss of Abcd1 in astrocytes produces a robust inflammatory response, the oligodendrocytes undergo cell death leading to demyelination in X-linked adrenoleukodystrophy (X-ALD). The mechanisms of these distinct pathways in the two cell types are not well understood. Here, we investigated the effects of Abcd1-knockdown and the subsequent alteration in VLCFA metabolism in human U87 astrocytes and rat B12 oligodendrocytes. Loss of Abcd1 inhibited peroxisomal ?-oxidation activity and increased expression of VLCFA synthesizing enzymes, elongase of very long chain fatty acids (ELOVLs) (1 and 3) in both cell types. However, higher induction of ELOVL's in Abcd1-deficient B12 oligodendrocytes than astrocytes suggests that ELOVL pathway may play a prominent role in oligodendrocytes in X-ALD. While astrocytes are able to maintain the cellular homeostasis of anti-apoptotic proteins, Abcd1-deletion in B12 oligodendrocytes downregulated the anti-apototic (Bcl-2 and Bcl-xL) and cell survival (phospho-Erk1/2) proteins, and upregulated the pro-apoptotic proteins (Bad, Bim, Bax and Bid) leading to cell loss. These observations provide insights into different cellular signaling mechanisms in response to Abcd1-deletion in two different cell types of CNS. The apoptotic responses were accompanied by activation of caspase-3 and caspase-9 suggesting the involvement of mitochondrial-caspase-9-dependent mechanism in Abcd1-deficient oligodendrocytes. Treatment with histone deacetylase (HDAC) inhibitor suberoylanilide hydroxamic acid (SAHA) corrected the VLCFA derangement both in vitro and in vivo, and inhibited the oligodendrocytes loss. These observations provide a proof-of principle that HDAC inhibitor SAHA may have a therapeutic potential for X-ALD. PMID:23923017

  12. Clofibrate treatment in pigs: Effects on parameters critical with respect to peroxisome proliferator-induced hepatocarcinogenesis in rodents

    PubMed Central

    Luci, Sebastian; Giemsa, Beatrice; Hause, Gerd; Kluge, Holger; Eder, Klaus

    2007-01-01

    Background In rodents treatment with fibrates causes hepatocarcinogenesis, probably as a result of oxidative stress and an impaired balance between apoptosis and cell proliferation in the liver. There is some debate whether fibrates could also induce liver cancer in species not responsive to peroxisome proliferation. In this study the effect of clofibrate treatment on peroxisome proliferation, production of oxidative stress, gene expression of pro- and anti-apoptotic genes and proto-oncogenes was investigated in the liver of pigs, a non-proliferating species. Results Pigs treated with clofibrate had heavier livers (+16%), higher peroxisome counts (+61%), higher mRNA concentration of acyl-CoA oxidase (+66%), a higher activity of catalase (+41%) but lower concentrations of hydrogen peroxide (-32%) in the liver than control pigs (P < 0.05); concentrations of lipid peroxidation products (thiobarbituric acid-reactive substances, conjugated dienes) and total and reduced glutathione in the liver did not differ between both groups. Clofibrate treated pigs also had higher hepatic mRNA concentrations of bax and the proto-oncogenes c-myc and c-jun and a lower mRNA concentration of bcl-XL than control pigs (P < 0.05). Conclusion The data of this study show that clofibrate treatment induces moderate peroxisome proliferation but does not cause oxidative stress in the liver of pigs. Gene expression analysis indicates that clofibrate treatment did not inhibit but rather stimulated apoptosis in the liver of these animals. It is also shown that clofibrate increases the expression of the proto-oncogenes c-myc and c-jun in the liver, an event which could be critical with respect to carcinogenesis. As the extent of peroxisome proliferation by clofibrate was similar to that observed in humans, the pig can be regarded as a useful model for investigating the effects of peroxisome proliferators on liver function and hepatocarcinogenesis. PMID:17437637

  13. Fenofibrate, a peroxisome proliferator-activated receptor ? ligand, prevents abnormal liver function induced by a fasting–refeeding process

    SciTech Connect

    Lee, Joon No; Dutta, Raghbendra Kumar; Kim, Seul-Gi; Lim, Jae-Young; Kim, Se-Jin; Choe, Seong-Kyu; Yoo, Kyeong-Won; Immune-network Pioneer Research Center, Department of Biochemistry, College of Medicine, Dong-A University, Busan ; Song, Seung Ryel; Park, Do-Sim; Department of Laboratory of Medicine, School of Medicine, Wonkwang University, Iksan ; So, Hong-Seob; Park, Raekil

    2013-12-06

    Highlights: •A fasting–refeeding high fat diet (HDF) model mimics irregular eating habit. •A fasting–refeeding HFD induces liver ballooning injury. •A fasting–refeeding HDF process elicits hepatic triglyceride accumulation. •Fenofibrate, PPAR? ligand, prevents liver damage induced by refeeding HFD. -- Abstract: Fenofibrate, a peroxisome proliferator-activated receptor ? (PPAR?) agonist, is an anti-hyperlipidemic agent that has been widely used in the treatment of dyslipidemia. In this study, we examined the effect of fenofibrate on liver damage caused by refeeding a high-fat diet (HFD) in mice after 24 h fasting. Here, we showed that refeeding HFD after fasting causes liver damage in mice determined by liver morphology and liver cell death. A detailed analysis revealed that hepatic lipid droplet formation is enhanced and triglyceride levels in liver are increased by refeeding HFD after starvation for 24 h. Also, NF-?B is activated and consequently induces the expression of TNF-?, IL1-?, COX-2, and NOS2. However, treating with fenofibrate attenuates the liver damage and triglyceride accumulation caused by the fasting–refeeding HFD process. Fenofibrate reduces the expression of NF-?B target genes but induces genes for peroxisomal fatty acid oxidation, peroxisome biogenesis and mitochondrial fatty acid oxidation. These results strongly suggest that the treatment of fenofibrate ameliorates the liver damage induced by fasting–refeeding HFD, possibly through the activation of fatty acid oxidation.

  14. Activation of peroxisome proliferator-activated receptor ? ameliorates monocrotaline-induced pulmonary arterial hypertension in rats

    PubMed Central

    XIE, XINMING; WANG, GUIZUO; ZHANG, DEXIN; ZHANG, YONGHONG; ZHU, YANTING; LI, FANGWEI; LI, SHAOJUN; LI, MANXIANG

    2015-01-01

    Activation of peroxisome proliferator-activated receptor ? (PPAR?) suppresses the proliferation of pulmonary artery smooth muscle cells (PASMCs) and vascular remodeling in rats and humans, and therefore improves the development of pulmonary arterial hypertension (PAH). However, molecular mechanisms underlying these effects have not been completely understood. In the present study, the effects of PPAR? activation in monocrotaline (MCT)-induced pulmonary artery remodeling in rats were investigated. Eighteen Sprague-Dawley (SD) rats were randomly assigned into three groups (n=6): Control (Con), PAH and PAH treated with rosiglitazone (MCT + Rosi). The right ventricular systolic pressure (RVSP), the ratio of the right to left ventricle plus septum weight [RV/(LV + S)], the percentage of medial wall thickness (%MT) and wall area (%WA) were used to evaluate the development of PAH. Tissue morphology was measured using hematoxylin and eosin staining. The protein levels of the phosphatase and tensin homologue deleted on chromosome ten (PTEN), Akt (ser473) phosphorylation (p-Akt) and total Akt in intrapulmonary arteries were determined by western blot analysis. MCT treatment significantly increased the RVSP, which was reduced by rosiglitazone treatment. The ratio of RV/(LV + S), %MT and %WA induced by MCT were similarly inhibited, which was associated with the increase of PTEN expression and the inhibition of Akt phosphorylation levels by rosiglitazone. In conclusion, activation of PPAR? ameliorates the proliferation of PASMCs and vascular remodeling by regulating the PTEN/PI3K/Akt pathway, suggesting that the activation of PPAR? has potential benefits for PAH. PMID:26171162

  15. Peroxisomal disorders.

    PubMed

    Aubourg, Patrick; Wanders, Ronald

    2013-01-01

    The peroxisomal disorders represent a group of genetic diseases in man in which there is an impairment in one or more peroxisomal functions. The peroxisomal disorders are subdivided into three subgroups comprising: (1) the peroxisome biogenesis disorders (PBDs); (2) the single peroxisomal (enzyme-) protein deficiencies; and (3) the single peroxisomal substrate transport deficiencies. The PBD group comprises four different disorders that include Zellweger syndrome (ZS), neonatal adrenoleukodystrophy (NALD), infantile Refsum disease (IRD), and rhizomelic chondrodysplasia punctata (RCDP). ZS, NALD, and IRD are clearly distinct from RCDP and are usually referred to as the Zellweger spectrum with ZS being the most severe, and IRD the less severe disorder, with sometimes onset in adulthood. The single peroxisomal enzyme deficiency group comprises seven different disorders, of which D-bifunctional protein and phytanoyl-CoA hydroxylase (adult Refsum disease) deficiencies are the most frequent. The single peroxisomal substrate transport deficiency group consists of only one disease, X-linked adrenoleukodystrophy. It is the purpose of this chapter to describe the current state of knowledge about the clinical, biochemical, cellular, and molecular aspects of peroxisomal diseases, and to provide guidelines for their post- and prenatal diagnosis. Therapeutic interventions are mostly limited to X-linked adrenoleukodystrophy. PMID:23622381

  16. Identification of transcriptional networks involved in peroxisome proliferator chemical-induced hepatocyte proliferation

    EPA Science Inventory

    Peroxisome proliferator chemical (PPC) exposure leads to increases in rodent liver tumors through a non-genotoxic mode of action (MOA). The PPC MOA includes increased oxidative stress, hepatocyte proliferation and decreased apoptosis. We investigated the putative genetic regulato...

  17. Bezafibrate induces a mitochondrial derangement in human cell lines: a PPAR-independent mechanism for a peroxisome proliferator.

    PubMed

    Scatena, R; Bottoni, P; Vincenzoni, F; Messana, I; Martorana, G E; Nocca, G; De Sole, P; Maggiano, N; Castagnola, M; Giardina, B

    2003-11-01

    Bezafibrate is a hypolipidemic drug that belongs to the group of peroxisome proliferators because it binds to peroxisome proliferator-activated receptors type alpha (PPARs). Peroxisome proliferators produce a myriad of extraperoxisomal effects, which are not necessarily dependent on their interaction with PPARs. An investigation on the peculiar activities of bezafibrate could clarify some of the molecular events and the relationship with the biochemical and pharmacological properties of this class of compounds. In this view, the human acute promyelocytic leukemia HL-60 cell line and human rabdomiosarcoma TE-671 cell line were cultured in media containing bezafibrate and a number of observations such as spectrophotometric analysis of mitochondrial respiratory chain enzymes, NMR metabolite determinations, phosphofructokinase enzymatic analysis, and differentiation assays were carried on. Bezafibrate induced a derangement of NADH cytochrome c reductase activity accompanied by metabolic alterations, mainly a shift to anaerobic glycolysis and an increase of fatty acid oxidation, as shown by NMR analysis of culture supernatants where acetate, lactate, and alanine levels increased. On the whole, the present results suggest a biochemical profile and a therapeutic role of this class of PPARs ligands more complex than those previously proposed. PMID:14615970

  18. MECHANISMS OF PEROXISOME PROLIFERATOR-INDUCED CARCINOGENESIS: HISTORICAL PERSPECTIVES AND CURRENT STATUS

    EPA Science Inventory

    This report is a comprehensive review of past and current thinking, as reflected in the scientific literature, on the mechanisms by which peroxisome proliferating agents are thought to act as carcinogens. he report is divided into four main sections: (1) background information on...

  19. White tea extract induces apoptosis in non-small cell lung cancer cells: the role of peroxisome proliferator-activated receptor-{gamma} and 15-lipoxygenases.

    PubMed

    Mao, Jenny T; Nie, Wen-Xian; Tsu, I-Hsien; Jin, Yu-Sheng; Rao, Jian Yu; Lu, Qing-Yi; Zhang, Zuo-Feng; Go, Vay Liang W; Serio, Kenneth J

    2010-09-01

    Emerging preclinical data suggests that tea possess anticarcinogenic and antimutagenic properties. We therefore hypothesize that white tea extract (WTE) is capable of favorably modulating apoptosis, a mechanism associated with lung tumorigenesis. We examined the effects of physiologically relevant doses of WTE on the induction of apoptosis in non-small cell lung cancer cell lines A549 (adenocarcinoma) and H520 (squamous cell carcinoma) cells. We further characterized the molecular mechanisms responsible for WTE-induced apoptosis, including the induction of peroxisome proliferator-activated receptor-gamma (PPAR-gamma) and the 15-lipoxygenase (15-LOX) signaling pathways. We found that WTE was effective in inducing apoptosis in both A549 and H520 cells, and inhibition of PPAR-gamma with GW9662 partially reversed WTE-induced apoptosis. We further show that WTE increased PPAR-gamma activation and mRNA expression, concomitantly increased 15(S)-hydroxy-eicosatetraenoic acid release, and upregulated 15-LOX-1 and 15-LOX-2 mRNA expression by A549 cells. Inhibition of 15-LOX with nordihydroguaiaretic acid (NGDA), as well as caffeic acid, abrogated WTE-induced PPAR-gamma activation and upregulation of PPAR-gamma mRNA expression in A549 cells. WTE also induced cyclin-dependent kinase inhibitor 1A mRNA expression and activated caspase-3. Inhibition of caspase-3 abrogated WTE-induced apoptosis. Our findings indicate that WTE is capable of inducing apoptosis in non-small cell lung cancer cell lines. The induction of apoptosis seems to be mediated, in part, through the upregulation of the PPAR-gamma and 15-LOX signaling pathways, with enhanced activation of caspase-3. Our findings support the future investigation of WTE as an antineoplastic and chemopreventive agent for lung cancer. PMID:20668019

  20. Identification of a peroxisome proliferator responsive element (PPRE)-like cis-element in mouse plasminogen activator inhibitor-1 gene promoter

    SciTech Connect

    Chen Jiegen; Li Xi; Huang Haiyan; Liu Honglei; Liu Deguo; Song Tanjing; Ma Chungu; Ma Duan; Song Houyan; Tang Qiqun . E-mail: qqtang@shmu.edu.cn

    2006-09-01

    PAI-1 is expressed and secreted by adipose tissue which may mediate the pathogenesis of obesity-associated cardiovascular complications. Evidence is presented in this report that PAI-1 is not expressed by preadipocyte, but significantly induced during 3T3-L1 adipocyte differentiation and the PAI-1 expression correlates with the induction of peroxisome proliferator-activated receptor {gamma} (PPAR{gamma}). A peroxisome proliferator responsive element (PPRE)-like cis-element (-206TCCCCCATGCCCT-194) is identified in the mouse PAI-1 gene promoter by electrophoretic mobility shift assay (EMSA) combined with transient transfection experiments; the PPRE-like cis-element forms a specific DNA-protein complex only with adipocyte nuclear extracts, not with preadipocyte nuclear extracts; the DNA-protein complex can be totally competed away by non-labeled consensus PPRE, and can be supershifted with PPAR{gamma} antibody. Mutation of this PPRE-like cis-element can abolish the transactivation of mouse PAI-1 promoter mediated by PPAR{gamma}. Specific PPAR{gamma} ligand Pioglitazone can significantly induce the PAI-1 expression, and stimulate the secretion of PAI-1 into medium.

  1. The role of peroxisome proliferator-activated receptor-{beta}/{delta} in epidermal growth factor-induced HaCaT cell proliferation

    SciTech Connect

    Liang Pengfei; Jiang Bimei; Yang Xinghua; Xiao Xianzhong Huang Xu; Long Jianhong; Zhang Pihong; Zhang Minghua; Xiao Muzhang; Xie Tinghong; Huang Xiaoyuan

    2008-10-15

    Epidermal growth factor (EGF) has been shown to be a potent mitogen for epidermal cells both in vitro and in vivo, thus contributing to the development of an organism. It has recently become clear that peroxisome proliferator-activated receptor-{beta}/{delta} (PPAR{beta}/{delta}) expression and activation is involved in the cell proliferation. However, little is known about the role of PPAR{beta}/{delta} in EGF-induced proliferation of HaCaT keratinocytes. In this study, HaCaT cells were cultured in the presence and absence of EGF and we identified that EGF induced an increase of PPAR{beta}/{delta} mRNA and protein level expression in time-dependent and dose-dependent manner, and AG1487, an EGF receptor (EGFR) special inhibitor, caused attenuation of PPAR{beta}/{delta} protein expression. Electrophoretic mobility shift assay (EMSA) revealed that EGF significantly increased PPAR{beta}/{delta} binding activity in HaCaT keratinocytes. Antisense phosphorothioate oligonucleotides (asODNs) against PPAR{beta}/{delta} caused selectively inhibition of PPAR{beta}/{delta} protein content induced by EGF and significantly attenuated EGF-mediated cell proliferation. Treatment of the cells with L165041, a specific synthetic ligand for PPAR{beta}/{delta}, significantly enhanced EGF-mediated cell proliferation. Finally, c-Jun ablation inhibited PPAR{beta}/{delta} up-regulation induced by EGF, and chromatin immunoprecipitation (ChIP) showed that c-Jun bound to the PPAR{beta}/{delta} promoter and the binding increased in EGF-stimulated cells. These results demonstrate that EGF induces PPAR{beta}/{delta} expression in a c-Jun-dependent manner and PPAR{beta}/{delta} plays a vital role in EGF-stimulated proliferation of HaCaT cells.

  2. 4-Hydroxydocosahexaenoic acid, a potent peroxisome proliferator-activated receptor {gamma} agonist alleviates the symptoms of DSS-induced colitis

    SciTech Connect

    Yamamoto, Keiko; Ninomiya, Yuichi; Iseki, Mioko; Nakachi, Yutaka; Kanesaki-Yatsuka, Yukiko; Yamanoue, Yu; Itoh, Toshimasa; Nishii, Yasuho; Petrovsky, Nikolai; Okazaki, Yasushi

    2008-03-14

    (5E,7Z,10Z,13Z,16Z,19Z)-4-Hydroxy-5,7,10,13,16,19-docosahexaenoic acid (4-OHDHA) is a potential agonist of peroxisome proliferator-activated receptor-{gamma} (PPAR{gamma}) and antidiabetic agent as has been previously reported. As PPAR{gamma} agonists may also have anti-inflammatory functions, in this study, we investigated whether 4-OHDHA has an inhibitory effect on expression of inflammatory genes in vitro and whether 4-OHDHA could relieve the symptoms of dextran sodium sulfate (DSS)-induced colitis in a murine model of inflammatory bowel disease. 4-OHDHA inhibited production of nitric oxide and expression of a subset of inflammatory genes including inducible nitric oxide synthase (Nos2/iNOS) and interleukin 6 (Il6) by lipopolysaccharide (LPS)-activated macrophages. In addition, 4-OHDHA-treated mice when compared to control mice not receiving treatment recovered better from the weight loss caused by DSS-induced colitis. Changes in disease activity index (DAI) of 4-OHDHA-treated mice were also more favorable than for control mice and were comparable with mice treated with a typical anti-inflammatory-drug, 5-aminosalichylic acid (5-ASA). These results suggest that 4-OHDHA has potentially clinically useful anti-inflammatory effects mediated by suppression of inflammatory gene expression.

  3. A Peroxisome Proliferator-Activated Receptor-? Activator Induces Renal CYP2C23 Activity and Protects from Angiotensin II-Induced Renal Injury

    PubMed Central

    Muller, Dominik N.; Theuer, Juergen; Shagdarsuren, Erdenechimeg; Kaergel, Eva; Honeck, Horst; Park, Joon-Keun; Markovic, Marija; Barbosa-Sicard, Eduardo; Dechend, Ralf; Wellner, Maren; Kirsch, Torsten; Fiebeler, Anette; Rothe, Michael; Haller, Hermann; Luft, Friedrich C.; Schunck, Wolf-Hagen

    2004-01-01

    Cytochrome P450 (CYP)-dependent arachidonic acid (AA) metabolites are involved in the regulation of renal vascular tone and salt excretion. The epoxygenation product 11,12-epoxyeicosatrienoic acid (EET) is anti-inflammatory and inhibits nuclear factor-?B activation. We tested the hypothesis that the peroxisome proliferator-activated receptor-?-activator fenofibrate (Feno) induces CYP isoforms, AA hydroxylation, and epoxygenation activity, and protects against inflammatory organ damage. Double-transgenic rats (dTGRs) overexpressing human renin and angiotensinogen genes were treated with Feno. Feno normalized blood pressure, albuminuria, reduced nuclear factor-?B activity, and renal leukocyte infiltration. Renal epoxygenase activity was lower in dTGRs compared to nontransgenic rats. Feno strongly induced renal CYP2C23 protein and AA-epoxygenase activity under pathological and nonpathological conditions. In both cases, CYP2C23 was themajor isoform responsible for 11,12-EET formation. Moreover, we describe a novel CYP2C23-dependent pathway leading to hydroxy-EETs (HEETs), which may serve as endogenous peroxisome proliferator-activated receptor-? activators. The capacity to produce HEETs via CYP2C23-dependent epoxygenation of 20-HETE and CYP4A-dependent hydroxylation of EETs was reduced in dTGR kidneys and induced by Feno. These results demonstrate that Feno protects against angiotensin II-induced renal damage and acts as inducer of CYP2C23-mediated epoxygenase activities. We propose that CYP-dependent EET/HEET production may serve as an anti-inflammatory control mechanism. PMID:14742258

  4. Dipeptidyl peptidase-4 inhibitor for steroid-induced diabetes

    PubMed Central

    Yanai, Hidekatsu; Masui, Yoshinori; Yoshikawa, Reo; Kunimatsu, Junwa; Kaneko, Hiroshi

    2010-01-01

    The addition of the dipeptidyl peptidase-4 (DDP-4) inhibitor has been reported to achieve greater improvements in glucose metabolism with fewer adverse events compared to increasing the metformin dose in type 2 diabetic patients. We present a patient with steroid-induced diabetes whose blood glucose levels were ameliorated by the use of the DPP-4 inhibitor, showing that the DPP-4 inhibitors may be an effective and safe oral anti-diabetic drug for steroid-induced diabetes. PMID:21537433

  5. Decorin Induces Mitophagy in Breast Carcinoma Cells via Peroxisome Proliferator-activated Receptor ? Coactivator-1? (PGC-1?) and Mitostatin*

    PubMed Central

    Neill, Thomas; Torres, Annabel; Buraschi, Simone; Owens, Rick T.; Hoek, Jan B.; Baffa, Raffaele; Iozzo, Renato V.

    2014-01-01

    Tumor cell mitochondria are key biosynthetic hubs that provide macromolecules for cancer progression and angiogenesis. Soluble decorin protein core, hereafter referred to as decorin, potently attenuated mitochondrial respiratory complexes and mitochondrial DNA (mtDNA) in MDA-MB-231 breast carcinoma cells. We found a rapid and dynamic interplay between peroxisome proliferator-activated receptor ? coactivator-1? (PGC-1?) and the decorin-induced tumor suppressor gene, mitostatin. This interaction stabilized mitostatin mRNA with concurrent accumulation of mitostatin protein. In contrast, siRNA-mediated abrogation of PGC-1?-blocked decorin-evoked stabilization of mitostatin. Mechanistically, PGC-1? bound MITOSTATIN mRNA to achieve rapid stabilization. These processes were orchestrated by the decorin/Met axis, as blocking the Met-tyrosine kinase or knockdown of Met abrogated these responses. Furthermore, depletion of mitostatin blocked decorin- or rapamycin-evoked mitophagy, increased vascular endothelial growth factor A (VEGFA) production, and compromised decorin-evoked VEGFA suppression. Collectively, our findings underscore the complexity of PGC-1?-mediated mitochondrial homeostasis and establish mitostatin as a key regulator of tumor cell mitophagy and angiostasis. PMID:24403067

  6. Decorin induces mitophagy in breast carcinoma cells via peroxisome proliferator-activated receptor ? coactivator-1? (PGC-1?) and mitostatin.

    PubMed

    Neill, Thomas; Torres, Annabel; Buraschi, Simone; Owens, Rick T; Hoek, Jan B; Baffa, Raffaele; Iozzo, Renato V

    2014-02-21

    Tumor cell mitochondria are key biosynthetic hubs that provide macromolecules for cancer progression and angiogenesis. Soluble decorin protein core, hereafter referred to as decorin, potently attenuated mitochondrial respiratory complexes and mitochondrial DNA (mtDNA) in MDA-MB-231 breast carcinoma cells. We found a rapid and dynamic interplay between peroxisome proliferator-activated receptor ? coactivator-1? (PGC-1?) and the decorin-induced tumor suppressor gene, mitostatin. This interaction stabilized mitostatin mRNA with concurrent accumulation of mitostatin protein. In contrast, siRNA-mediated abrogation of PGC-1?-blocked decorin-evoked stabilization of mitostatin. Mechanistically, PGC-1? bound MITOSTATIN mRNA to achieve rapid stabilization. These processes were orchestrated by the decorin/Met axis, as blocking the Met-tyrosine kinase or knockdown of Met abrogated these responses. Furthermore, depletion of mitostatin blocked decorin- or rapamycin-evoked mitophagy, increased vascular endothelial growth factor A (VEGFA) production, and compromised decorin-evoked VEGFA suppression. Collectively, our findings underscore the complexity of PGC-1?-mediated mitochondrial homeostasis and establish mitostatin as a key regulator of tumor cell mitophagy and angiostasis. PMID:24403067

  7. Proton pump inhibitor-induced hypomagnesemic hypoparathyroidism

    PubMed Central

    Swaminathan, Krishnan

    2015-01-01

    Proton pump inhibitors are the one of the most widely used drugs in the world. Hypomagnesemic hypoparathyroidism has been reported with different proton pump inhibitors with prolonged oral use. We report the first reported case of possible such effect with intravenous preparation of proton pump inhibitor. This case report raises awareness among physicians worldwide of this often unknown association, as life-threatening cardiac and neuromuscular complications can arise with unrecognized hypocalcemia and hypomagnesemia with proton pump inhibitors. PMID:26069375

  8. The methylotrophic yeast Hansenula polymorpha contains an inducible import pathway for peroxisomal matrix proteins with an N-terminal targeting signal (PTS2 proteins).

    PubMed Central

    Faber, K N; Haima, P; Gietl, C; Harder, W; Ab, G; Veenhuis, M

    1994-01-01

    Two main types of peroxisomal targeting signals have been identified that reside either at the extreme C terminus (PTS1) or the N terminus (PTS2) of the protein. In the methylotrophic yeast Hansenula polymorpha the majority of peroxisomal matrix proteins are of the PTS1 type. Thus far, for H. polymorpha only amine oxidase (AMO) has been shown to contain a PTS2 type signal. In the present study we expressed H. polymorpha AMO under control of the strong endogenous alcohol oxidase promoter. Partial import of AMO into peroxisomes was observed in cells grown in methanol/(NH4)2SO4-containing medium. However, complete import of AMO occurred if the cells were grown under conditions that induce expression of the endogenous AMO gene. Similar results were obtained when the heterologous PTS2 proteins, glyoxysomal malate dehydrogenase from watermelon and thiolase from Saccharomyces cerevisiae, were synthesized in H. polymorpha. The import of PTS1 proteins, however, was not affected by the growth conditions. These results indicate that the reduced rate of AMO import in (NH4)2SO4-grown cells is not due to competition with PTS1 proteins for the same import pathway. Apparently, AMO is imported via a separate pathway that is induced by amines and functions for PTS2 proteins in general. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 Fig. 5 PMID:7809160

  9. The methylotrophic yeast Hansenula polymorpha contains an inducible import pathway for peroxisomal matrix proteins with an N-terminal targeting signal (PTS2 proteins).

    PubMed

    Faber, K N; Haima, P; Gietl, C; Harder, W; Ab, G; Veenhuis, M

    1994-12-20

    Two main types of peroxisomal targeting signals have been identified that reside either at the extreme C terminus (PTS1) or the N terminus (PTS2) of the protein. In the methylotrophic yeast Hansenula polymorpha the majority of peroxisomal matrix proteins are of the PTS1 type. Thus far, for H. polymorpha only amine oxidase (AMO) has been shown to contain a PTS2 type signal. In the present study we expressed H. polymorpha AMO under control of the strong endogenous alcohol oxidase promoter. Partial import of AMO into peroxisomes was observed in cells grown in methanol/(NH4)2SO4-containing medium. However, complete import of AMO occurred if the cells were grown under conditions that induce expression of the endogenous AMO gene. Similar results were obtained when the heterologous PTS2 proteins, glyoxysomal malate dehydrogenase from watermelon and thiolase from Saccharomyces cerevisiae, were synthesized in H. polymorpha. The import of PTS1 proteins, however, was not affected by the growth conditions. These results indicate that the reduced rate of AMO import in (NH4)2SO4-grown cells is not due to competition with PTS1 proteins for the same import pathway. Apparently, AMO is imported via a separate pathway that is induced by amines and functions for PTS2 proteins in general. PMID:7809160

  10. The Hansenula polymorpha PEX14 gene encodes a novel peroxisomal membrane protein essential for peroxisome biogenesis.

    PubMed Central

    Komori, M; Rasmussen, S W; Kiel, J A; Baerends, R J; Cregg, J M; van der Klei, I J; Veenhuis, M

    1997-01-01

    We have cloned the Hansenula polymorpha PEX14 gene by functional complementation of the chemically induced pex14-1 mutant, which lacked normal peroxisomes. The sequence of the PEX14 gene predicts a novel protein product (Pex14p) of 39 kDa which showed no similarity to any known protein and lacked either of the two known peroxisomal targeting signals. Biochemical and electron microscopical analysis indicated that Pex14p is a component of the peroxisomal membrane. The synthesis of Pex14p is induced by peroxisome-inducing growth conditions. In cells of both pex14-1 and a PEX14 disruption mutant, peroxisomal membrane remnants were evident; these contained the H.polymorpha peroxisomal membrane protein Pex3p together with a small amount of the major peroxisomal matrix proteins alcohol oxidase, catalase and dihydroxyacetone synthase, the bulk of which resided in the cytosol. Unexpectedly, overproduction of Pex14p in wild-type H. polymorpha cells resulted in a peroxisome-deficient phenotype typified by the presence of numerous small vesicles which lacked matrix proteins; these were localized in the cytosol. Apparently, the stoichiometry of Pex14p relative to one or more other components of the peroxisome biogenesis machinery appears to be critical for protein import. PMID:9009266

  11. Crosstalk between mitochondria and peroxisomes.

    PubMed

    Demarquoy, Jean; Le Borgne, Françoise

    2015-11-26

    Mitochondria and peroxisomes are small ubiquitous organelles. They both play major roles in cell metabolism, especially in terms of fatty acid metabolism, reactive oxygen species (ROS) production, and ROS scavenging, and it is now clear that they metabolically interact with each other. These two organelles share some properties, such as great plasticity and high potency to adapt their form and number according to cell requirements. Their functions are connected, and any alteration in the function of mitochondria may induce changes in peroxisomal physiology. The objective of this paper was to highlight the interconnection and the crosstalk existing between mitochondria and peroxisomes. Special emphasis was placed on the best known connections between these organelles: origin, structure, and metabolic interconnections. PMID:26629313

  12. Crosstalk between mitochondria and peroxisomes

    PubMed Central

    Demarquoy, Jean; Le Borgne, Françoise

    2015-01-01

    Mitochondria and peroxisomes are small ubiquitous organelles. They both play major roles in cell metabolism, especially in terms of fatty acid metabolism, reactive oxygen species (ROS) production, and ROS scavenging, and it is now clear that they metabolically interact with each other. These two organelles share some properties, such as great plasticity and high potency to adapt their form and number according to cell requirements. Their functions are connected, and any alteration in the function of mitochondria may induce changes in peroxisomal physiology. The objective of this paper was to highlight the interconnection and the crosstalk existing between mitochondria and peroxisomes. Special emphasis was placed on the best known connections between these organelles: origin, structure, and metabolic interconnections. PMID:26629313

  13. ABCD2 Alters Peroxisome Proliferator-Activated Receptor ? Signaling In Vitro, but Does Not Impair Responses to Fenofibrate Therapy in a Mouse Model of Diet-Induced Obesity

    PubMed Central

    Liu, Xiaoxi; Liu, Jingjing; Liang, Shuang; Schlüter, Agatha; Fourcade, Stephane; Aslibekyan, Stella; Pujol, Aurora

    2014-01-01

    Fenofibrate is a peroxisome proliferator-activated receptor (PPAR) ? ligand that has been widely used as a lipid-lowering agent in the treatment of hypertriglyceridemia. ABCD2 (D2) is a peroxisomal long-chain acyl-CoA transporter that is highly induced by fenofibrate in the livers of mice. To determine whether D2 is a modifier of fibrate responses, wild-type and D2-deficient mice were treated with fenofibrate for 14 days. The absence of D2 altered expression of gene clusters associated with lipid metabolism, including PPAR? signaling. Using 3T3-L1 adipocytes, which express high levels of D2, we confirmed that knockdown of D2 modified genomic responses to fibrate treatment. We next evaluated the impact of D2 on effects of fibrates in a mouse model of diet-induced obesity. Fenofibrate treatment opposed the development of obesity, hypertriglyceridemia, and insulin resistance. However, these effects were unaffected by D2 genotype. We concluded that D2 can modulate genomic responses to fibrates, but that these effects are not sufficiently robust to alter the effects of fibrates on diet-induced obesity phenotypes. PMID:25123288

  14. Wound-induced Proteinase Inhibitor in Tomato Leaves

    PubMed Central

    Green, T. R.; Ryan, C. A.

    1973-01-01

    Wounding of single leaflets of young tomato (Lycopersicum esculentum var. Bonnie Best) plants causes the release of a proteinase inhibitor inducing factor. This factor is rapidly transported throughout the plant where it causes accumulation of inhibitor I, a potent inhibitor of several serine proteinases from both animals and microorganisms. The wound-induced accumulation of inhibitor I is both light- and temperature-dependent. In total darkness no accumulation results from wounding. The accumulation exhibits a linear dependence upon light up to 300 foot candles. At 600 foot candles and above, the response is maximal. In light the wound response possesses an unusual temperature dependence with an optimum rate of accumulation near 36 C. Below 20 C no accumulation occurs. The over-all process contains two light- and temperature-dependent steps, one involving wounding and transport, the other involving accumulation. PMID:16658283

  15. Peroxisome diversity and evolution

    PubMed Central

    Gabaldón, Toni

    2010-01-01

    Peroxisomes are organelles bounded by a single membrane that can be found in all major groups of eukaryotes. A single evolutionary origin of this cellular compartment is supported by the presence, in diverse organisms, of a common set of proteins implicated in peroxisome biogenesis and maintenance. Their enzymatic content, however, can vary substantially across species, indicating a high level of evolutionary plasticity. Proteomic analyses have greatly expanded our knowledge on peroxisomes in some model organisms, including plants, mammals and yeasts. However, we still have a limited knowledge about the distribution and functionalities of peroxisomes in the vast majority of groups of microbial eukaryotes. Here, I review recent advances in our understanding of peroxisome diversity and evolution, with a special emphasis on peroxisomes in microbial eukaryotes. PMID:20124343

  16. Peroxisome Biogenesis and Function

    PubMed Central

    Kaur, Navneet; Reumann, Sigrun; Hu, Jianping

    2009-01-01

    Peroxisomes are small and single membrane-delimited organelles that execute numerous metabolic reactions and have pivotal roles in plant growth and development. In recent years, forward and reverse genetic studies along with biochemical and cell biological analyses in Arabidopsis have enabled researchers to identify many peroxisome proteins and elucidate their functions. This review focuses on the advances in our understanding of peroxisome biogenesis and metabolism, and further explores the contribution of large-scale analysis, such as in sillco predictions and proteomics, in augmenting our knowledge of peroxisome function In Arabidopsis. PMID:22303249

  17. Peroxisomal APX knockdown triggers antioxidant mechanisms favourable for coping with high photorespiratory H2 O2 induced by CAT deficiency in rice.

    PubMed

    Sousa, Rachel H V; Carvalho, Fabricio E L; Ribeiro, Carol W; Passaia, Gisele; Cunha, Juliana R; Lima-Melo, Yugo; Margis-Pinheiro, Márcia; Silveira, Joaquim A G

    2015-03-01

    The physiological role of peroxisomal ascorbate peroxidases (pAPX) is unknown; therefore, we utilized pAPX4 knockdown rice and catalase (CAT) inhibition to assess its role in CAT compensation under high photorespiration. pAPX4 knockdown induced co-suppression in the expression of pAPX3. The rice mutants exhibited metabolic changes such as lower CAT and glycolate oxidase (GO) activities and reduced glyoxylate content; however, APX activity was not altered. CAT inhibition triggered different changes in the expression of CAT, APX and glutathione peroxidase (GPX) isoforms between non-transformed (NT) and silenced plants. These responses were associated with alterations in APX, GPX and GO activities, suggesting redox homeostasis differences. The glutathione oxidation-reduction states were modulated differently in mutants, and the ascorbate redox state was greatly affected in both genotypes. The pAPX suffered less oxidative stress and photosystem II (PSII) damage and displayed higher photosynthesis than the NT plants. The improved acclimation exhibited by the pAPX plants was indicated by lower H2 O2 accumulation, which was associated with lower GO activity and glyoxylate content. The suppression of both pAPXs and/or its downstream metabolic and molecular effects may trigger favourable antioxidant and compensatory mechanisms to cope with CAT deficiency. This physiological acclimation may involve signalling by peroxisomal H2 O2 , which minimized the photorespiration. PMID:25039271

  18. Epigenetic Activity of Peroxisome Proliferator-Activated Receptor Gamma Agonists Increases the Anticancer Effect of Histone Deacetylase Inhibitors on Multiple Myeloma Cells

    PubMed Central

    Aouali, Nassera; Broukou, Angeliki; Bosseler, Manon; Keunen, Olivier; Schlesser, Vincent; Janji, Bassam; Palissot, Valerie; Stordeur, Philippe; Berchem, Guy

    2015-01-01

    Epigenetic modifications play a major role in the development of multiple myeloma. We have previously reported that the PPAR? agonist pioglitazone (PIO) enhances, in-vitro, the cytotoxic effect of the Histone deacetylase inhibitor (HDACi), valproic acid (VPA), on multiple myeloma cells. Here, we described the development of a new multiple myeloma mouse model using MOLP8 cells, in order to evaluate the effect of VPA/PIO combination on the progression of myeloma cells, by analyzing the proliferation of bone marrow plasma cells. We showed that VPA/PIO delays the progression of the disease and the invasion of myeloma cells in the bone marrow. Mechanistically, we demonstrated that VPA/PIO increases the cleavage of caspase 3 and PARP, and induces the acetylation of Histone 3 (H3). Furthermore, we provided evidence that PPAR? agonist is able to enhance the action of other HDACi such as Vorinostat or Mocetinostat. Using PPAR? antagonist or siPPAR?, we strongly suggest that, as described during adipogenesis, PIO behaves as an epigenetic regulator by improving the activity of HDACi. This study highlights the therapeutic benefit of PIO/VPA combination, compared to VPA treatment as a single-arm therapy on multiple myeloma and further highlights that such combination may constitute a new promising treatment strategy which should be supported by clinical trials. PMID:26091518

  19. ORIGINAL PAPER Discovery of potent inhibitors for interleukin-2-inducible

    E-print Network

    Lee, Keun Woo

    ORIGINAL PAPER Discovery of potent inhibitors for interleukin-2-inducible T-cell kinase: structure Structure-based hypothesis PTK Protein tyrosine kinase PH Pleckstrin homology SH3 Src homology3 SH2 Src (cytoplasmic) kinases. Src and eight related molecules formed the largest family of cytoplasmic PTKs, while

  20. AtPex14p maintains peroxisomal functions by determining protein targeting to three kinds of plant peroxisomes

    PubMed Central

    Hayashi, Makoto; Nito, Kazumasa; Toriyama-Kato, Kanako; Kondo, Maki; Yamaya, Tomoyuki; Nishimura, Mikio

    2000-01-01

    We previously isolated an Arabidopsis peroxisome-deficient ped2 mutant by its resistance to 2,4-dichlorophenoxybutyric acid. Here, we describe the isolation of a gene responsible for this deficiency, called the PED2 gene, by positional cloning and confirmed its identity by complementation analysis. The amino acid sequence of the predicted protein product is similar to that of human Pex14p, which is a key component of the peroxisomal protein import machinery. Therefore, we decided to call it AtPex14p. Analyses of the ped2 mutant revealed that AtPex14p controls intracellular transport of both peroxisome targeting signal (PTS)1- and PTS2-containing proteins into three different types of peroxisomes, namely glyoxysomes, leaf peroxisomes and unspecialized peroxisomes. Mutation in the PED2 gene results in reduction of enzymes in all of these functionally differentiated peroxisomes. The reduction in these enzymes induces pleiotropic defects, such as fatty acid degradation, photorespiration and the morphology of peroxisomes. These data suggest that the AtPex14p has a common role in maintaining physiological functions of each of these three kinds of plant peroxisomes by determining peroxisomal protein targeting. PMID:11060021

  1. Sodium transport inhibitors in pregnancy-induced hypertension.

    PubMed

    Poston, L

    1990-03-01

    In blacks and whites of similar socioeconomic background, the incidence of pregnancy-induced hypertension (PIH) is probably the same. In underdeveloped countries, however. PIH is often a life-threatening complication of pregnancy. Recent theories as to the etiology of PIH include the suggestion that vascular tone may be increased as a result of inhibition of active sodium transport in vascular smooth muscle. This may be the result of an inhibitor of sodium transport present in the serum. The literature concerning the demonstration of endogenous sodium transport inhibitors and endogenous digoxinlike immunoreactivity (EDLI) in PIH is reviewed and discussed. PMID:2176808

  2. Activation of peroxisome proliferator-activated receptor-{gamma} (PPAR{gamma}) induces cell death through MAPK-dependent mechanism in osteoblastic cells

    SciTech Connect

    Kim, Sung Hun; Yoo, Chong Il; Kim, Hui Taek; Park, Ji Yeon; Kwon, Chae Hwa; Keun Kim, Yong . E-mail: kim430@pusan.ac.kr

    2006-09-01

    The present study was undertaken to determine the role of the mitogen-activated protein kinase (MAPK) subfamilies in cell death induced by PPAR{gamma} agonists in osteoblastic cells. Ciglitazone and troglitazone, PPAR{gamma} agonists, resulted in a concentration- and time-dependent cell death, which was largely attributed to apoptosis. But a PPAR{alpha} agonist ciprofibrate did not affect the cell death. Ciglitazone caused reactive oxygen species (ROS) generation and ciglitazone-induced cell death was prevented by antioxidants, suggesting an important role of ROS generation in the ciglitazone-induced cell death. ROS generation and cell death induced by ciglitazone were inhibited by the PPAR{gamma} antagonist GW9662. Ciglitazone treatment caused activation of extracellular signal-regulated kinase (ERK) and p38. Activation of ERK was dependent on epidermal growth factor receptor (EGFR) and that of p38 was independent. Ciglitazone-induced cell death was significantly prevented by PD98059, an inhibitor of ERK upstream kinase MEK1/2, and SB203580, a p38 inhibitor. Ciglitazone treatment increased Bax expression and caused a loss of mitochondrial membrane potential, and its effect was prevented by N-acetylcysteine, PD98059, and SB203580. Ciglitazone induced caspase activation, which was prevented by PD98059 and SB203580. The general caspase inhibitor z-DEVD-FMK and the specific inhibitor of caspases-3 DEVD-CHO exerted the protective effect against the ciglitazone-induced cell death. The EGFR inhibitors AG1478 and suramin protected against the ciglitazone-induced cell death. Taken together, these findings suggest that the MAPK signaling pathways play an active role in mediating the ciglitazone-induced cell death of osteoblasts and function upstream of a mitochondria-dependent mechanism. These data may provide a novel insight into potential therapeutic strategies for treatment of osteoporosis.

  3. Expression regulation and targeting of the peroxisome proliferator-activated receptor ? following electrically-induced status epilepticus.

    PubMed

    Boes, Katharina; Russmann, Vera; Ongerth, Tanja; Licko, Thomas; Salvamoser, Josephine D; Siegl, Claudia; Potschka, Heidrun

    2015-09-14

    The neuroprotective and anti-inflammatory effects of the peroxisome proliferator-activated receptor ? (PPAR?) agonist rosiglitazone are of particular interest for disease-modifying and antiepileptogenic approaches. We studied the expression of PPAR? and the impact of rosiglitazone on the consequences of status epilepticus (SE) in a rat post-SE model. Immunohistochemical analysis revealed a selective overexpression of PPAR? in the piriform cortex of rats with spontaneous seizures. Rosiglitazone administration initiated following SE failed to exert relevant effects on the development of spontaneous seizures and neuronal cell loss. Whereas spatial learning in the Morris water maze was delayed in SE animals with vehicle administration, the learning curve of rosiglitazone-treated SE rats showed no significant difference to that of controls. The study provides first evidence arguing against a robust antiepileptogenic effect. However, the findings in the spatial learning paradigm indicate disease-modifying effects. PMID:26259695

  4. Danhong inhibits oxidized low-density lipoprotein-induced immune maturation of dentritic cells via a peroxisome proliferator activated receptor ?-mediated pathway.

    PubMed

    Liu, Hongying; Wang, Shijun; Sun, Aijun; Huang, Dong; Wang, Wei; Zhang, Chunyu; Shi, Dazhuo; Chen, Keji; Zou, Yunzeng; Ge, Junbo

    2012-01-01

    Danhong injection (DHI), a Chinese Materia Medica standardized product extracted from Radix Salviae miltiorrhizae and Flos Carthami tinctorii, is effective in the treatment of atherosclerosis (AS)-related diseases. It is widely recognized that AS is a complex inflammatory disease of the arterial wall and the dendritic cells (DCs) is a major player in the pathogenesis of AS via mediating atherosclerotic antigen presenting and T lymphocytes. Here, we determined the effect and possible mechanism of DHI on oxidized low-density lipoprotein (ox-LDL)-induced maturation and immune function of DCs. Human monocyte-derived DCs were incubated with DHI or ciglitazone and were subsequently stimulated with ox-LDL to induce maturation. Similar to ciglitazone, a peroxisome proliferator activated receptor (PPAR) ? agonist, DHI, could significantly reduce ox-LDL-induced expressions of mature markers, enhance the endocytotic function, and inhibit secretions of cytokine on DCs. These effects of DHI could be partly reversed by silencing the PPAR?. In conclusion, DHI could inhibit ox-LDL-induced maturation of DCs partly through activating a PPAR?-mediated signaling pathway. PMID:22739234

  5. Peroxisome proliferator-activated receptor ?/? (PPAR?/?) protects against ceramide-induced cellular toxicity in rat brain astrocytes and neurons by activation of ceramide kinase.

    PubMed

    Aleshin, Stepan; Reiser, Georg

    2014-03-01

    Peroxisome proliferator-activated receptors (PPARs) are important members of the nuclear receptor superfamily. Ligands of these nuclear receptors (PPAR?, ?/? and ?) belong to a wide range of lipophilic substances. In spite of the proven neuroprotective efficacy of PPAR?/? in models of neurological diseases, the biology of PPAR?/? in the brain has been much less investigated than that of PPAR? and PPAR?. In the present study, we test the hypothesis that neuroprotection induced by PPAR?/? could rely on the regulation of ceramide metabolism. We found that preincubation of neural cells with the PPAR?/? agonist L-165041 exerts significant protection against ceramide-induced cell death. Most importantly, L-165041 protects against ceramide-induced cell death not only before the insult, but also after the onset of the insult. To identify the mechanism of protection, we show that L-165041 upregulates ceramide kinase (CerK) expression levels in neural cells. Consistent with that, we detected that pharmacological inhibition of CerK reduces the protective effects of L-165041. To further decipher the mechanism of protection, gene knockdown in astrocytes was studied. Knockdown of PPAR?/? and CerK in astrocytes was used to verify that the protective effects of L-165041 are CerK- and PPAR?/?-dependent. We demonstrate that in CerK- or PPAR?/?-knockdown astrocytes, addition of L-165041 has no protective effect. Thus, we conclude that PPAR?/? protects neural cells against ceramide-induced cell death via induction and activation of CerK. PMID:24513118

  6. Peroxisomes Get Loud: A Redox Antidote to Hearing Loss.

    PubMed

    Mardones, Pablo; Hetz, Claudio

    2015-11-01

    Pejvakin (PJVK), a protein originally identified in Persian families with sensorineural hearing loss, regulates peroxisomal dynamics and the antioxidant defense triggered by noise exposure in hair cells and auditory neurons of the inner ear. These findings bring peroxisomes to the forefront of noise-induced hearing loss research. PMID:26544930

  7. [Adverse skin reactions induced by BRAF inhibitors: a systematic review].

    PubMed

    Sibaud, V; Lamant, L; Maisongrosse, V; Delord, J-P

    2013-01-01

    Recent developments and therapeutic use of selective BRAF inhibitors (e.g. dabrafenib and vemurafenib) have significantly improved overall survival and disease-free survival of patients with BRAF V600 mutation-positive metastatic melanoma. Despite their survival benefits, small-molecule inhibitors of BRAF are associated with significant and sometimes severe treatment-related dermatological toxicity. The most common adverse skin reactions include photosensitivity, induced malignant lesions of the skin such as keratoacanthomas, squamous cell carcinoma and new primary melanomas, as well as keratinocyte proliferation and differentiation dysfunctions that can manifest as skin papillomas, hand-foot skin reaction, keratosis pilaris-like rash, acantholytic dyskeratosis and cysts of the milia type. In this article, we describe the clinical and histological features of the cutaneous manifestations induced by vemurafenib and dabrafenib on the basis of our clinical experience and a literature review. The crucial role of dermatologists in patient management is also highlighted. PMID:24034635

  8. Potentially lethal ACE-inhibitor-induced angioedema in a child

    PubMed Central

    Bukhari, Esraa; Safdar, Osama Y; Shalaby, Mohammed; AlSharif, Shafiqa MJ; Alsufiany, Khoulod; Kari, Jameela A

    2015-01-01

    Key Clinical Message We report a case of a 9-year-old female with known end-stage kidney disease who presented with sudden onset tongue swelling. A diagnosis of angiotensin-converting enzyme inhibitor-induced angioedema related to bradykinin accumulation was made. Her symptoms resolved shortly after discontinuation of captopril. Early diagnosis can save patients from severe upper airway obstruction. PMID:26185642

  9. Ginger extract prevents high-fat diet-induced obesity in mice via activation of the peroxisome proliferator-activated receptor ? pathway.

    PubMed

    Misawa, Koichi; Hashizume, Kojiro; Yamamoto, Masaki; Minegishi, Yoshihiko; Hase, Tadashi; Shimotoyodome, Akira

    2015-10-01

    The initiation of obesity entails an imbalance wherein energy intake exceeds expenditure. Obesity is increasing in prevalence and is now a worldwide health problem. Food-derived peroxisome proliferator-activated receptor ? (PPAR?) stimulators represent potential treatment options for obesity. Ginger (Zingiber officinale Roscoe) was previously shown to regulate the PPAR? signaling pathway in adipocytes. In this study, we investigated the antiobesity effects of ginger in vivo and the mechanism of action in vitro. Energy expenditure was increased, and diet-induced obesity was attenuated in C57BL/6J mice treated with dietary ginger extract (GE). GE also increased the number of Type I muscle fibers, improved running endurance capacity and upregulated PPAR?-targeted gene expression in skeletal muscle and the liver. 6-Shogaol and 6-gingerol acted as specific PPAR? ligands and stimulated PPAR?-dependent gene expression in cultured human skeletal muscle myotubes. An analysis of cellular respiration revealed that pretreating cultured skeletal muscle myotubes with GE increased palmitate-induced oxygen consumption rate, which suggested an increase in cellular fatty acid catabolism. These results demonstrated that sustained activation of the PPAR? pathway with GE attenuated diet-induced obesity and improved exercise endurance capacity by increasing skeletal muscle fat catabolism. 6-Shogaol and 6-gingerol may be responsible for the regulatory effects of dietary ginger on PPAR? signaling. PMID:26101135

  10. Hypoxia-inducible Lipid Droplet-associated (HILPDA) Is a Novel Peroxisome Proliferator-activated Receptor (PPAR) Target Involved in Hepatic Triglyceride Secretion*

    PubMed Central

    Mattijssen, Frits; Georgiadi, Anastasia; Andasarie, Tresty; Szalowska, Ewa; Zota, Annika; Krones-Herzig, Anja; Heier, Christoph; Ratman, Dariusz; De Bosscher, Karolien; Qi, Ling; Zechner, Rudolf; Herzig, Stephan; Kersten, Sander

    2014-01-01

    Peroxisome proliferator-activated receptors (PPARs) play major roles in the regulation of hepatic lipid metabolism through the control of numerous genes involved in processes such as lipid uptake and fatty acid oxidation. Here we identify hypoxia-inducible lipid droplet-associated (Hilpda/Hig2) as a novel PPAR target gene and demonstrate its involvement in hepatic lipid metabolism. Microarray analysis revealed that Hilpda is one of the most highly induced genes by the PPAR? agonist Wy14643 in mouse precision cut liver slices. Induction of Hilpda mRNA by Wy14643 was confirmed in mouse and human hepatocytes. Oral dosing with Wy14643 similarly induced Hilpda mRNA levels in livers of wild-type mice but not Ppara?/? mice. Transactivation studies and chromatin immunoprecipitation showed that Hilpda is a direct PPAR? target gene via a conserved PPAR response element located 1200 base pairs upstream of the transcription start site. Hepatic overexpression of HILPDA in mice via adeno-associated virus led to a 4-fold increase in liver triglyceride storage, without any changes in key genes involved in de novo lipogenesis, ?-oxidation, or lipolysis. Moreover, intracellular lipase activity was not affected by HILPDA overexpression. Strikingly, HILPDA overexpression significantly impaired hepatic triglyceride secretion. Taken together, our data uncover HILPDA as a novel PPAR target that raises hepatic triglyceride storage via regulation of triglyceride secretion. PMID:24876382

  11. Relationship between peroxisome proliferator-activated receptors (PPAR? and PPAR?) and endothelium-dependent relaxation in streptozotocin-induced diabetic rats

    PubMed Central

    Kanie, Noriyasu; Matsumoto, Takayuki; Kobayashi, Tsuneo; Kamata, Katsuo

    2003-01-01

    The aim of the present study was to investigate the causal relationship between peroxisome proliferator-activated receptor (PPAR) and endothelium-dependent relaxation in streptozotocin (STZ)-induced diabetic rats. Acetylcholine (ACh)-induced endothelium-dependent relaxation was significantly weaker in diabetic rats than in age-matched controls. The decreased relaxation in diabetes was improved by the chronic administration of bezafibrate (30 mg kg?1, p.o., 4 weeks). The expressions of the mRNAs for PPAR? and PPAR? were significantly decreased in STZ-induced diabetic rats (compared with the controls) and this decrease was restored partially, but not completely, by the chronic administration of bezafibrate. Superoxide dismutase activity in the aorta was not significantly different between diabetic rats and bezafibrate-treated diabetic rats. The expression of the mRNA for the p22phox subunit of NAD(P)H oxidase was significantly higher in diabetics than in controls, but it was lower in bezafibrate-treated diabetic rats than in nontreated diabetic rats. Although the expression of the mRNA for prepro ET-1 (ppET-1) was markedly increased in diabetic rats (compared with controls), this increase was prevented to a significant extent by the chronic administration of bezafibrate. These results suggest that downregulations of PPAR? and PPAR? may lead to an increased expression of ppET-1 mRNA in diabetic states and this increment may trigger endothelial dysfunction. PMID:12967931

  12. Administration of the peroxisomal proliferator-activated receptor {gamma} agonist pioglitazone during fractionated brain irradiation prevents radiation-induced cognitive impairment

    SciTech Connect

    Zhao Weiling; Payne, Valerie; Tommasi, Ellen; Diz, Debra I.; Hsu, F.-C.; Robbins, Mike E. . E-mail: mrobbins@wfubmc.edu

    2007-01-01

    Purpose: We hypothesized that administration of the anti-inflammatory peroxisomal proliferator-activated receptor {gamma} (PPAR{gamma}) agonist pioglitazone (Pio) to adult male rats would inhibit radiation-induced cognitive impairment. Methods and Materials: Young adult male F344 rats received one of the following: (1) fractionated whole brain irradiation (WBI); 40 or 45 Gy {gamma}-rays in 4 or 4.5 weeks, respectively, two fractions per week and normal diet; (2) sham-irradiation and normal diet; (3) WBI plus Pio (120 ppm) before, during, and for 4 or 54 weeks postirradiation; (4) sham-irradiation plus Pio; or (5) WBI plus Pio starting 24h after completion of WBI. Results: Administration of Pio before, during, and for 4 or 54 weeks after WBI prevented Radiation-induced cognitive impairment. Administration of Pio for 54 weeks starting after completion of fractionated WBI substantially but not significantly reduced Radiation-induced cognitive impairment. Conclusions: These findings offer the promise of improving the quality of life and increasing the therapeutic window for brain tumor patients.

  13. Endocrine disruption induced by organotin compounds; organotins function as a powerful agonist for nuclear receptors rather than an aromatase inhibitor.

    PubMed

    Nakanishi, Tsuyoshi

    2008-08-01

    Organotin compounds have been widely used as antifouling biocides for ships and fishing nets, agricultural fungicides and rodent repellents. These widespread uses have resulted in the release of increasing amounts of organotins into the environment. In aquatic invertebrates, particularly marine gastropods, organotin compounds, such as tributyltin (TBT) and triphenyltin (TPT), induce irreversible sexual abnormality in females which is termed "imposex" at very low concentrations. Although it has been theorized that these compounds act as potential competitive inhibitors of aromatase, which converts androgen to estrogen, and then increase levels of unconverted androgens in gastropods, their effective concentrations for aromatase inhibition are high. In addition to wildlife, organotins may have various undesirable effects on human health. Contrary to the theory of organotin-induced aromatase inhibition in gastropods, in human choriocarcinoma cells, these compounds markedly enhance estradiol biosynthesis along with the increase of both aromatase activity and 17beta-hydroxysteroid dehydrogenase type I (17beta-HSD I) activity, which converts low-activity estrogen estrone to the biologically more active form estradiol, at the same low concentrations. Although there are many reports describing the potential toxicity of organotins in human and mammals, the critical target molecules for the toxicity of organotin compounds remain unclear. Recently, organotin compounds including TBT and TPT were identified as nanomolar agonists for retinoid X receptor (RXR) and peroxisome proliferator-activated receptor (PPAR) gamma, which are members of the nuclear receptor superfamily. Here, we review the potential genetics action and subsequent toxicity induced by organotins via these nuclear receptors. PMID:18670157

  14. PEROXISOME PROLIFERATOR-ACTIVATED RECEPTOR? (PPAR?) AGONISTS DIFFERENTIALLY REGULATE INHIBITOR OF DNA BINDING (ID2) EXPRESSION IN RODENTS AND HUMAN CELLS

    EPA Science Inventory

    Abstract Inhibitor of DNA binding (Id2) is a member of the helix-loop-helix (HLH) transcription factor family whose members play important roles in cell differentiation and proliferation. Id2 has been linked to the development of cardiovascular diseases since thiazolidinediones,...

  15. Cell death is induced by ciglitazone, a peroxisome proliferator-activated receptor {gamma} (PPAR{gamma}) agonist, independently of PPAR{gamma} in human glioma cells

    SciTech Connect

    Lee, Myoung Woo; Kim, Dae Seong; Kim, Hye Ryung; Kim, Hye Jin; Yang, Jin Mo; Ryu, Somi; Noh, Yoo Hun; Lee, Soo Hyun; Son, Meong Hi; Jung, Hye Lim; Yoo, Keon Hee; Koo, Hong Hoe; Sung, Ki Woong

    2012-01-06

    Highlights: Black-Right-Pointing-Pointer Greater than 30 {mu}M ciglitazone induces cell death in glioma cells. Black-Right-Pointing-Pointer Cell death by ciglitazone is independent of PPAR{gamma} in glioma cells. Black-Right-Pointing-Pointer CGZ induces cell death by the loss of MMP via decreased Akt. -- Abstract: Peroxisome proliferator-activated receptor {gamma} (PPAR{gamma}) regulates multiple signaling pathways, and its agonists induce apoptosis in various cancer cells. However, their role in cell death is unclear. In this study, the relationship between ciglitazone (CGZ) and PPAR{gamma} in CGZ-induced cell death was examined. At concentrations of greater than 30 {mu}M, CGZ, a synthetic PPAR{gamma} agonist, activated caspase-3 and induced apoptosis in T98G cells. Treatment of T98G cells with less than 30 {mu}M CGZ effectively induced cell death after pretreatment with 30 {mu}M of the PPAR{gamma} antagonist GW9662, although GW9662 alone did not induce cell death. This cell death was also observed when cells were co-treated with CGZ and GW9662, but was not observed when cells were treated with CGZ prior to GW9662. In cells in which PPAR{gamma} was down-regulated cells by siRNA, lower concentrations of CGZ (<30 {mu}M) were sufficient to induce cell death, although higher concentrations of CGZ ( Greater-Than-Or-Slanted-Equal-To 30 {mu}M) were required to induce cell death in control T98G cells, indicating that CGZ effectively induces cell death in T98G cells independently of PPAR{gamma}. Treatment with GW9662 followed by CGZ resulted in a down-regulation of Akt activity and the loss of mitochondrial membrane potential (MMP), which was accompanied by a decrease in Bcl-2 expression and an increase in Bid cleavage. These data suggest that CGZ is capable of inducing apoptotic cell death independently of PPAR{gamma} in glioma cells, by down-regulating Akt activity and inducing MMP collapse.

  16. A Selective Novel Peroxisome Proliferator–Activated Receptor (PPAR)-? Antagonist Induces Apoptosis and Inhibits Proliferation of CLL Cells In Vitro and In Vivo

    PubMed Central

    Messmer, Davorka; Lorrain, Kymmy; Stebbins, Karin; Bravo, Yalda; Stock, Nicholas; Cabrera, Geraldine; Correa, Lucia; Chen, Austin; Jacintho, Jason; Chiorazzi, Nicholas; Yan, Xiao Jie; Spaner, David; Prasit, Peppi; Lorrain, Daniel

    2015-01-01

    Tumor-specific metabolic changes can reveal new therapeutic targets. Our findings implicate a supporting role for fatty acid metabolism in chronic lymphocytic leukemia (CLL) cell survival. Peroxisome proliferator–activated receptor (PPAR)-?, a major transcriptional regulator of fatty acid oxidation, was recently shown to be upregulated in CLL. To evaluate PPAR? as a potential therapeutic target, we developed a highly selective, potent small molecule antagonist of PPAR?, NXT629. NXT629 inhibited agonist-induced transcription of PPAR?-regulated genes, demonstrating target engagement in CLL cells. Furthermore, NXT629 induced apoptosis of CLL cells even in the presence of a protective microenvironment. To mimic the proliferative lymphoid compartment of CLL, we examined the activity of NXT629 on CLL cells that were stimulated to proliferate in vitro. NXT629 reduced the number of leukemia cells undergoing cell division. In addition, in two xenograft mouse models of CLL (one a model for nondividing and one for dividing CLL), NXT629 reduced the number of viable CLL cells in vivo. Overall, these results suggest that fatty acid metabolism promotes survival and proliferation of primary CLL cells and that inhibiting PPAR? gene regulation could be a new therapeutic approach to treating CLL. PMID:26070013

  17. Peroxisome proliferator activated receptor-? agonists protect oligodendrocyte progenitors against tumor necrosis factor-alpha-induced damage: Effects on mitochondrial functions and differentiation.

    PubMed

    De Nuccio, C; Bernardo, A; Cruciani, C; De Simone, R; Visentin, S; Minghetti, L

    2015-09-01

    The activation of the nuclear receptor peroxisome proliferator-activated receptor-? (PPAR-?) is known to exert anti-inflammatory and neuroprotective effects and PPAR-? agonists are considered potential therapeutic agents in brain diseases including those affecting myelin. In demyelinating diseases such as multiple sclerosis (MS), inflammation is one of the causes of myelin and axonal damage. Oligodendrocyte (OL) differentiation is highly dependent on mitochondria, which are major targets of inflammatory insult. Here we show that PPAR-? agonists protect OL progenitors against the maturational arrest induced by the inflammatory cytokine TNF-? by affecting mitochondrial functions. We demonstrate that the inhibition of OL differentiation by TNF-? is associated with i) increased mitochondrial superoxide production; ii) decreased mitochondrial membrane potential (mMP); and iii) decreased ADP-induced Ca(2+) oscillations, which we previously showed to be dependent on efficient mitochondria. The TNF-? effects were comparable to those of the mitochondrial toxin rotenone, further suggesting that TNF-? damage is mediated by mitochondrial function impairment. PPAR-? agonists protected OL progenitors against the inhibitory activities of both TNF-? and rotenone on mMP, mitochondrial ROS production, Ca(2+) oscillations and OL differentiation. Finally, the PPAR-? agonist pioglitazone increased the expression of PGC-1? (a mitochondrial biogenesis master regulator), UCP2 (a mitochondrial protein known to reduce ROS production), and cytochrome oxidase subunit COX1. These findings confirm the central role of mitochondria in OL differentiation and point to mitochondria as major targets of PPAR-? agonist protection against TNF-? damage. PMID:26210873

  18. Peroxisome proliferator-activated receptor {alpha}-independent peroxisome proliferation

    SciTech Connect

    Zhang Xiuguo; Tanaka, Naoki . E-mail: naopi@hsp.md.shinshu-u.ac.jp; Nakajima, Takero; Kamijo, Yuji; Gonzalez, Frank J.; Aoyama, Toshifumi

    2006-08-11

    Hepatic peroxisome proliferation, increases in the numerical and volume density of peroxisomes, is believed to be closely related to peroxisome proliferator-activated receptor {alpha} (PPAR{alpha}) activation; however, it remains unknown whether peroxisome proliferation depends absolutely on this activation. To verify occurrence of PPAR{alpha}-independent peroxisome proliferation, fenofibrate treatment was used, which was expected to significantly enhance PPAR{alpha} dependence in the assay system. Surprisingly, a novel type of PPAR{alpha}-independent peroxisome proliferation and enlargement was uncovered in PPAR{alpha}-null mice. The increased expression of dynamin-like protein 1, but not peroxisome biogenesis factor 11{alpha}, might be associated with the PPAR{alpha}-independent peroxisome proliferation at least in part.

  19. Evaluating Chemical CDK Inhibitors as Cell Death Inducers.

    PubMed

    Hirai, Hiroshi; Nakatsuru, Yoko

    2016-01-01

    The cell cycle of eukaryotic cells is regulated by a family of protein kinases called cyclin-dependent kinases (Cdks). We have reported the identification and biological characterization of a highly potent, small-molecule pan-Cdk inhibitor, which inhibited Cdk1, 2, 4, 5, 6, and 9 with equal potency in the nM range. This compound inhibited multiple events in the cell cycle and induced cell death in human cancer cell lines as well as in peripheral blood or purified resting lymphocytes ex vivo. We describe the materials and methods to determine antitumor efficacy in vivo xenograft models. Pharmacodynamic marker assays that have been performed using tumors and normal tissues are explained. Moreover, we briefly describe methods for determining the effects of chemical Cdk inhibitors on peripheral blood cells or lymphocytes ex vivo. PMID:26231716

  20. Over-expression of C/EBP-{alpha} induces apoptosis in cultured rat hepatic stellate cells depending on p53 and peroxisome proliferator-activated receptor-{gamma}

    SciTech Connect

    Wang Xueqing; Huang Guangcun; Mei Shuang; Qian Jin; Ji Juling; Zhang Jinsheng

    2009-03-06

    Hepatic stellate cells (HSCs) play a key role in the pathogenesis of hepatic fibrosis. In our previous studies, CCAAT enhancer binding protein-{alpha} (C/EBP-{alpha}) has been shown to be involved in the activation of HSCs and to have a repression effect on hepatic fibrosis in vivo. However, the mechanisms are largely unknown. In this study, we show that the infection of adenovirus vector expressing C/EBP-{alpha} gene (Ad-C/EBP-{alpha}) could induce HSCs apoptosis in a dose- and time-dependent manner by Annexin V/PI staining, caspase-3 activation assay, and flow cytometry. Also, over-expression of C/EBP-{alpha} resulted in the up-regulation of peroxisome proliferator-activated receptor-{gamma} (PPAR-{gamma}) and P53, while P53 expression was regulated by PPAR-{gamma}. In addition, Fas, FasL, DR4, DR5, and TRAIL were studied. The results indicated that the death receptor pathway was mainly involved and regulated by PPAR-{gamma} and p53 in the process of apoptosis triggered by C/EBP-{alpha} in HSCs.

  1. Antagonist of peroxisome proliferator-activated receptor {gamma} induces cerebellar amyloid-{beta} levels and motor dysfunction in APP/PS1 transgenic mice

    SciTech Connect

    Du, Jing; Sun, Bing; Chen, Kui; Fan, Li; Cardiovascular Research, Starr Academic Center, Providence Heart and Vascular Institute, Portland, OR 97225 ; Wang, Zhao

    2009-07-03

    Recent evidences show that peroxisome proliferator-activated receptor {gamma} (PPAR{gamma}) is involved in the modulation of the amyloid-{beta} (A{beta}) cascade causing Alzheimer's disease (AD) and treatment with PPAR{gamma} agonists protects against AD pathology. However, the function of PPAR{gamma} steady-state activity in A{beta} cascade and AD pathology remains unclear. In this study, an antagonist of PPAR{gamma}, GW9662, was injected into the fourth ventricle of APP/PS1 transgenic mice to inhibit PPAR{gamma} activity in cerebellum. The results show that inhibition of PPAR{gamma} significantly induced A{beta} levels in cerebellum and caused cerebellar motor dysfunction in APP/PS1 transgenic mice. Moreover, GW9662 treatment markedly decreased the cerebellar levels of insulin-degrading enzyme (IDE), which is responsible for the cellular degradation of A{beta}. Since cerebellum is spared from significant A{beta} accumulation and neurotoxicity in AD patients and animal models, these findings suggest a crucial role of PPAR{gamma} steady-state activity in protection of cerebellum against AD pathology.

  2. Peroxisome proliferator-activated receptor-? ligands induce heme oxygenase-1 in lung fibroblasts by a PPAR?-independent, glutathione-dependent mechanism

    PubMed Central

    Ferguson, Heather E.; Thatcher, Thomas H.; Olsen, Keith C.; Garcia-Bates, Tatiana M.; Baglole, Carolyn J.; Kottmann, R. M.; Strong, Emily R.; Phipps, Richard P.

    2009-01-01

    Oxidative stress plays an important role in the pathogenesis of pulmonary fibrosis. Heme oxygenase-1 (HO-1) is a key antioxidant enzyme, and overexpression of HO-1 significantly decreases lung inflammation and fibrosis in animal models. Peroxisome proliferator-activated receptor-? (PPAR?) is a transcription factor that regulates adipogenesis, insulin sensitization, and inflammation. We report here that the PPAR? ligands 15d-PGJ2 and 2-cyano-3,12-dioxoolean-1,9-dien-28-oic acid (CDDO), which have potent antifibrotic effects in vitro, also strongly induce HO-1 expression in primary human lung fibroblasts. Pharmacological and genetic approaches are used to demonstrate that induction of HO-1 is PPAR? independent. Upregulation of HO-1 coincides with decreased intracellular glutathione (GSH) levels and can be inhibited by N-acetyl cysteine (NAC), a thiol antioxidant and GSH precursor. Upregulation of HO-1 is not inhibited by Trolox, a non-thiol antioxidant, and does not involve the transcription factors AP-1 or Nrf2. CDDO and 15d-PGJ2 contain an ?/? unsaturated ketone that acts as an electrophilic center that can form covalent bonds with free reduced thiols. Rosiglitazone, a PPAR? ligand that lacks an electrophilic center, does not induce HO-1. These data suggest that in human lung fibroblasts, 15d-PGJ2 and CDDO induce HO-1 via a GSH-dependent mechanism involving the formation of covalent bonds between 15d-PGJ2 or CDDO and GSH. Inhibiting HO-1 upregulation with NAC has only a small effect on the antifibrotic properties of 15d-PGJ2 and CDDO in vitro. These results suggest that CDDO and similar electrophilic PPAR? ligands may have great clinical potential as antifibrotic agents, not only through direct effects on fibroblast differentiation and function, but indirectly by bolstering antioxidant defenses. PMID:19734319

  3. Peroxisome proliferator-activated receptor-gamma ligands induce heme oxygenase-1 in lung fibroblasts by a PPARgamma-independent, glutathione-dependent mechanism.

    PubMed

    Ferguson, Heather E; Thatcher, Thomas H; Olsen, Keith C; Garcia-Bates, Tatiana M; Baglole, Carolyn J; Kottmann, R M; Strong, Emily R; Phipps, Richard P; Sime, Patricia J

    2009-11-01

    Oxidative stress plays an important role in the pathogenesis of pulmonary fibrosis. Heme oxygenase-1 (HO-1) is a key antioxidant enzyme, and overexpression of HO-1 significantly decreases lung inflammation and fibrosis in animal models. Peroxisome proliferator-activated receptor-gamma (PPARgamma) is a transcription factor that regulates adipogenesis, insulin sensitization, and inflammation. We report here that the PPARgamma ligands 15d-PGJ2 and 2-cyano-3,12-dioxoolean-1,9-dien-28-oic acid (CDDO), which have potent antifibrotic effects in vitro, also strongly induce HO-1 expression in primary human lung fibroblasts. Pharmacological and genetic approaches are used to demonstrate that induction of HO-1 is PPARgamma independent. Upregulation of HO-1 coincides with decreased intracellular glutathione (GSH) levels and can be inhibited by N-acetyl cysteine (NAC), a thiol antioxidant and GSH precursor. Upregulation of HO-1 is not inhibited by Trolox, a non-thiol antioxidant, and does not involve the transcription factors AP-1 or Nrf2. CDDO and 15d-PGJ2 contain an alpha/beta unsaturated ketone that acts as an electrophilic center that can form covalent bonds with free reduced thiols. Rosiglitazone, a PPARgamma ligand that lacks an electrophilic center, does not induce HO-1. These data suggest that in human lung fibroblasts, 15d-PGJ2 and CDDO induce HO-1 via a GSH-dependent mechanism involving the formation of covalent bonds between 15d-PGJ2 or CDDO and GSH. Inhibiting HO-1 upregulation with NAC has only a small effect on the antifibrotic properties of 15d-PGJ2 and CDDO in vitro. These results suggest that CDDO and similar electrophilic PPARgamma ligands may have great clinical potential as antifibrotic agents, not only through direct effects on fibroblast differentiation and function, but indirectly by bolstering antioxidant defenses. PMID:19734319

  4. Peroxisome Proliferator-Activated Receptors-Alpha and Gamma Are Targets to Treat Offspring from Maternal Diet-Induced Obesity in Mice

    PubMed Central

    Magliano, D'Angelo Carlo; Bargut, Thereza Cristina Lonzetti; de Carvalho, Simone Nunes; Aguila, Marcia Barbosa; Mandarim-de-Lacerda, Carlos Alberto; Souza-Mello, Vanessa

    2013-01-01

    Aim The aim of the present study was to evaluate whether activation of peroxisome proliferator-activated receptor (PPAR)alpha and PPARgamma by Bezafibrate (BZ) could attenuate hepatic and white adipose tissue (WAT) abnormalities in male offspring from diet-induced obese dams. Materials and Methods C57BL/6 female mice were fed a standard chow (SC; 10% lipids) diet or a high-fat (HF; 49% lipids) diet for 8 weeks before mating and during gestation and lactation periods. Male offspring received SC diet at weaning and were subdivided into four groups: SC, SC/BZ, HF and HF/BZ. Treatment with BZ (100 mg/Kg diet) started at 12 weeks of age and was maintained for three weeks. Results The HF diet resulted in an overweight phenotype and an increase in oral glucose intolerance and fasting glucose of dams. The HF offspring showed increased body mass, higher levels of plasmatic and hepatic triglycerides, higher levels of pro-inflammatory and lower levels of anti-inflammatory adipokines, impairment of glucose metabolism, abnormal fat pad mass distribution, higher number of larger adipocytes, hepatic steatosis, higher expression of lipogenic proteins concomitant to decreased expression of PPARalpha and carnitine palmitoyltransferase I (CPT-1) in liver, and diminished expression of PPARgamma and adiponectin in WAT. Treatment with BZ ameliorated the hepatic and WAT abnormalities generated by diet-induced maternal obesity, with improvements observed in the structural, biochemical and molecular characteristics of the animals' livers and epididymal fat. Conclusion Diet-induced maternal obesity lead to alterations in metabolism, hepatic lipotoxicity and adverse liver and WAT remodeling in the offspring. Targeting PPAR with Bezafibrate has beneficial effects reducing the alterations, mainly through reduction of WAT inflammatory state through PPARgamma activation and enhanced hepatic beta-oxidation due to increased PPARalpha/PPARgamma ratio in liver. PMID:23700465

  5. Curcumin protects neurons against oxygen-glucose deprivation/reoxygenation-induced injury through activation of peroxisome proliferator-activated receptor-? function.

    PubMed

    Liu, Zun-Jing; Liu, Hong-Qiang; Xiao, Cheng; Fan, Hui-Zhen; Huang, Qing; Liu, Yun-Hai; Wang, Yu

    2014-11-01

    The turmeric derivative curcumin protects against cerebral ischemic injury. We previously demonstrated that curcumin activates peroxisome proliferator-activated receptor-? (PPAR?), a ligand-activated transcription factor involved in both neuroprotective and anti-inflammatory signaling pathways. This study tested whether the neuroprotective effects of curcumin against oxygen-glucose deprivation/reoxygenation (OGD/R)-induced injury of rat cortical neurons are mediated (at least in part) by PPAR?. Curcumin (10 ?M) potently enhanced PPAR? expression and transcriptional activity following OGD/R. In addition, curcumin markedly increased neuronal viability, as evidenced by decreased lactate dehydrogenase release and reduced nitric oxide production, caspase-3 activity, and apoptosis. These protective effects were suppressed by coadministration of the PPAR? antagonist 2-chloro-5-nitrobenzanilide (GW9662) and by prior transfection of a small-interfering RNA (siRNA) targeting PPAR?, treatments that had no toxic effects on healthy neurons. Curcumin reduced OGD/R-induced accumulation of reactive oxygen species and inhibited the mitochondrial apoptosis pathway, as indicated by reduced release of cytochrome c and apoptosis-inducing factor and maintenance of both the mitochondrial membrane potential and the Bax/Bcl-2 ratio. Again, GW9662 or PPAR? siRNA transfection mitigated the protective effects of curcumin on mitochondrial function. Curcumin suppressed I?B kinase phosphorylation and I?B degradation, thereby inhibiting nuclear factor-? B (NF-?B) nuclear translocation, effects also blocked by GW9662 or PPAR? siRNA. Immunoprecipitation experiments revealed that PPAR? interacted with NF-?B p65 and inhibited NF-?B activation. The present study provides strong evidence that at least some of the neuroprotective effects of curcumin against OGD/R are mediated by PPAR? activation. PMID:24975470

  6. Study on cow ghee versus soybean oil on 7,12-dimethylbenz(a)-anthracene induced mammary carcinogenesis & expression of cyclooxygenase-2 & peroxisome proliferators activated receptor- ? in rats

    PubMed Central

    Rani, Rita; Kansal, Vinod K.

    2011-01-01

    Background & objectives Breast cancer is a leading cause of cancer death in women; dietary fat is the one of the factors that influences its incidence. In the present study we investigated the effect of feeding cow ghee versus soybean oil on 7,12-dimethylbenz(a)anthracene (DMBA) induced mammary cancer in rat and expression of cyclooxygenase-2 and peroxisome proliferators activated receptor- ? (PPAR-?) in mammary gland. Methods Two groups of 21 day old female rats (30 each) were fed for 44 wk diet containing cow ghee or soybean oil (10%). The animals were given DMBA (30mg/kg body weight) through oral intubation after 5 wk feeding. Another two groups (8 each) fed similarly but not given DMBA served as control for the gene expression study. Results In DMBA treated groups, the animal fed soybean oil had higher tumour incidence (65.4%), tumour weight (6.18 g) and tumour volume (6285 mm3) compared to those fed cow ghee (26.6%, 1.67 g, 1925 mm3, respectively). Tumour latency period was 23 wk on soybean oil compared to 27 wk on cow ghee. Histological analysis of tumours showed that the progression of carcinogenesis was more rapid on soybean oil than on cow ghee. The expression of cyclooxygenase-2 was observed only in DMBA treated rats and it was significantly less on cow ghee than on soybean oil. The expression of PPAR-? was significantly more on cow ghee than on soybean oil. Interpretation & conclusions Our results show that dietary cow ghee opposed to soybean oil attenuates mammary carcinogenesis induced by DMBA; and the effect is mediated by decreased expression of cyclooxygenase-2 and increased expression of PPAR-? in the former group. PMID:21623034

  7. Rosiglitazone, a Peroxisome Proliferator-Activated Receptor (PPAR)-? Agonist, Attenuates Inflammation Via NF-?B Inhibition in Lipopolysaccharide-Induced Peritonitis.

    PubMed

    Zhang, Yun-Fang; Zou, Xun-Liang; Wu, Jun; Yu, Xue-Qing; Yang, Xiao

    2015-12-01

    We assessed the anti-inflammatory effect of peroxisome proliferator-activated receptor (PPAR)-? agonist, rosiglitazone, in a lipopolysaccharide (LPS)-induced peritonitis rat model. LPS was intraperitoneally injected into rats to establish peritonitis model. Male Sprague-Dawley (SD) rats were assigned to normal saline (the solvent of LPS), LPS, rosiglitazone plus LPS, and rosiglitazone alone. A simple peritoneal equilibrium test was performed with 20 ml 4.25 % peritoneal dialysis fluid. We measured the leukocyte count in dialysate and ultrafiltration volume. Peritoneal membrane histochemical staining was performed, and peritoneal thickness was assessed. CD40 and intercellular adhesion molecule-1 messenger RNA (ICAM-1 mRNA) levels in rat visceral peritoneum were detected by reverse transcription (RT)-PCR. IL-6 in rat peritoneal dialysis effluent was measured using enzyme-linked immunosorbent assay. The phosphorylation of NF-?B-p65 and I?B? was analyzed by Western blot. LPS administration resulted in increased peritoneal thickness and decreased ultrafiltration volume. Rosiglitazone pretreatment significantly decreased peritoneal thickness. In addition to CD40 and ICAM-1 mRNA expression, the IL-6, p-p65, and p-I?B? protein expressions were enhanced in LPS-administered animals. Rosiglitazone pretreatment significantly decreased ICAM-1 mRNA upregulation, secretion of IL-6 protein, and phosphorylation of NF-?B-p65 and I?B? without decreasing CD40 mRNA expression. Rosiglitazone has a protective effect in peritonitis, simultaneously decreasing NF-?B phosphorylation, suggesting that NF-?B signaling pathway mediated peritoneal inflammation induced by LPS. PPAR-? might be considered a potential therapeutic target against peritonitis. PMID:26047949

  8. Inhibitors

    MedlinePLUS

    ... caregivers to learn how to care for the device. CDC Research CDC is interested in learning more about why some people develop inhibitors and how they could be prevented. The Inhibitor Project began in 2005 and was implemented to explore ...

  9. How Peroxisomes Affect Aflatoxin Biosynthesis in Aspergillus Flavus

    PubMed Central

    Reverberi, Massimo; Punelli, Marta; Smith, Carrie A.; Zjalic, Slaven; Scarpari, Marzia; Scala, Valeria; Cardinali, Giorgia; Aspite, Nicaela; Pinzari, Flavia; Payne, Gary A.; Fabbri, Anna A.; Fanelli, Corrado

    2012-01-01

    In filamentous fungi, peroxisomes are crucial for the primary metabolism and play a pivotal role in the formation of some secondary metabolites. Further, peroxisomes are important site for fatty acids ?-oxidation, the formation of reactive oxygen species and for their scavenging through a complex of antioxidant activities. Oxidative stress is involved in different metabolic events in all organisms and it occurs during oxidative processes within the cell, including peroxisomal ?-oxidation of fatty acids. In Aspergillus flavus, an unbalance towards an hyper-oxidant status into the cell is a prerequisite for the onset of aflatoxin biosynthesis. In our preliminary results, the use of bezafibrate, inducer of both peroxisomal ?-oxidation and peroxisome proliferation in mammals, significantly enhanced the expression of pex11 and foxA and stimulated aflatoxin synthesis in A. flavus. This suggests the existence of a correlation among peroxisome proliferation, fatty acids ?-oxidation and aflatoxin biosynthesis. To investigate this correlation, A. flavus was transformed with a vector containing P33, a gene from Cymbidium ringspot virus able to induce peroxisome proliferation, under the control of the promoter of the Cu,Zn-sod gene of A. flavus. This transcriptional control closely relates the onset of the antioxidant response to ROS increase, with the proliferation of peroxisomes in A. flavus. The AfP33 transformant strain show an up-regulation of lipid metabolism and an higher content of both intracellular ROS and some oxylipins. The combined presence of a higher amount of substrates (fatty acids-derived), an hyper-oxidant cell environment and of hormone-like signals (oxylipins) enhances the synthesis of aflatoxins in the AfP33 strain. The results obtained demonstrated a close link between peroxisome metabolism and aflatoxin synthesis. PMID:23094106

  10. Hypervulnerability to Sound Exposure through Impaired Adaptive Proliferation of Peroxisomes.

    PubMed

    Delmaghani, Sedigheh; Defourny, Jean; Aghaie, Asadollah; Beurg, Maryline; Dulon, Didier; Thelen, Nicolas; Perfettini, Isabelle; Zelles, Tibor; Aller, Mate; Meyer, Anaïs; Emptoz, Alice; Giraudet, Fabrice; Leibovici, Michel; Dartevelle, Sylvie; Soubigou, Guillaume; Thiry, Marc; Vizi, E Sylvester; Safieddine, Saaid; Hardelin, Jean-Pierre; Avan, Paul; Petit, Christine

    2015-11-01

    A deficiency in pejvakin, a protein of unknown function, causes a strikingly heterogeneous form of human deafness. Pejvakin-deficient (Pjvk(-/-)) mice also exhibit variable auditory phenotypes. Correlation between their hearing thresholds and the number of pups per cage suggest a possible harmful effect of pup vocalizations. Direct sound or electrical stimulation show that the cochlear sensory hair cells and auditory pathway neurons of Pjvk(-/-) mice and patients are exceptionally vulnerable to sound. Subcellular analysis revealed that pejvakin is associated with peroxisomes and required for their oxidative-stress-induced proliferation. Pjvk(-/-) cochleas display features of marked oxidative stress and impaired antioxidant defenses, and peroxisomes in Pjvk(-/-) hair cells show structural abnormalities after the onset of hearing. Noise exposure rapidly upregulates Pjvk cochlear transcription in wild-type mice and triggers peroxisome proliferation in hair cells and primary auditory neurons. Our results reveal that the antioxidant activity of peroxisomes protects the auditory system against noise-induced damage. PMID:26544938

  11. Peroxisomal cholesterol biosynthesis and Smith-Lemli-Opitz syndrome

    SciTech Connect

    Weinhofer, Isabelle; Kunze, Markus; Stangl, Herbert; Porter, Forbes D.; Berger, Johannes . E-mail: johannes.berger@meduniwien.ac.at

    2006-06-23

    Smith-Lemli-Opitz syndrome (SLOS), caused by 7-dehydrocholesterol-reductase (DHCR7) deficiency, shows variable severity independent of DHCR7 genotype. To test whether peroxisomes are involved in alternative cholesterol synthesis, we used [1-{sup 14}C]C24:0 for peroxisomal {beta}-oxidation to generate [1-{sup 14}C]acetyl-CoA as cholesterol precursor inside peroxisomes. The HMG-CoA reductase inhibitor lovastatin suppressed cholesterol synthesis from [2-{sup 14}C]acetate and [1-{sup 14}C]C8:0 but not from [1-{sup 14}C]C24:0, implicating a peroxisomal, lovastatin-resistant HMG-CoA reductase. In SLOS fibroblasts lacking DHCR7 activity, no cholesterol was formed from [1-{sup 14}C]C24:0-derived [1-{sup 14}C]acetyl-CoA, indicating that the alternative peroxisomal pathway also requires this enzyme. Our results implicate peroxisomes in cholesterol biosynthesis but provide no link to phenotypic variation in SLOS.

  12. Phosphatidylcholine Supply to Peroxisomes of the Yeast Saccharomyces cerevisiae

    PubMed Central

    Ramprecht, Claudia; Zellnig, Günther; Leitner, Erich; Hermetter, Albin; Daum, Günther

    2015-01-01

    In the yeast Saccharomyces cerevisiae, phosphatidylcholine (PC), the major phospholipid (PL) of all organelle membranes, is synthesized via two different pathways. Methylation of phosphatidylethanolamine (PE) catalyzed by the methyl transferases Cho2p/Pem1p and Opi3p/Pem2p as well as incorporation of choline through the CDP (cytidine diphosphate)-choline branch of the Kennedy pathway lead to PC formation. To determine the contribution of these two pathways to the supply of PC to peroxisomes (PX), yeast mutants bearing defects in the two pathways were cultivated under peroxisome inducing conditions, i.e. in the presence of oleic acid, and subjected to biochemical and cell biological analyses. Phenotype studies revealed compromised growth of both the cho20?opi3? (mutations in the methylation pathway) and the cki1?dpl1?eki1? (mutations in the CDP-choline pathway) mutant when grown on oleic acid. Analysis of peroxisomes from the two mutant strains showed that both pathways produce PC for the supply to peroxisomes, although the CDP-choline pathway seemed to contribute with higher efficiency than the methylation pathway. Changes in the peroxisomal lipid pattern of mutants caused by defects in the PC biosynthetic pathways resulted in changes of membrane properties as shown by anisotropy measurements with fluorescent probes. In summary, our data define the origin of peroxisomal PC and demonstrate the importance of PC for peroxisome membrane formation and integrity. PMID:26241051

  13. Peroxisomes contribute to reactive oxygen species homeostasis and cell division induction in Arabidopsis protoplasts

    PubMed Central

    Tiew, Terence W.-Y.; Sheahan, Michael B.; Rose, Ray J.

    2015-01-01

    The ability to induce Arabidopsis protoplasts to dedifferentiate and divide provides a convenient system to analyze organelle dynamics in plant cells acquiring totipotency. Using peroxisome-targeted fluorescent proteins, we show that during protoplast culture, peroxisomes undergo massive proliferation and disperse uniformly around the cell before cell division. Peroxisome dispersion is influenced by the cytoskeleton, ensuring unbiased segregation during cell division. Considering their role in oxidative metabolism, we also investigated how peroxisomes influence homeostasis of reactive oxygen species (ROS). Protoplast isolation induces an oxidative burst, with mitochondria the likely major ROS producers. Subsequently ROS levels in protoplast cultures decline, correlating with the increase in peroxisomes, suggesting that peroxisome proliferation may also aid restoration of ROS homeostasis. Transcriptional profiling showed up-regulation of several peroxisome-localized antioxidant enzymes, most notably catalase (CAT). Analysis of antioxidant levels, CAT activity and CAT isoform 3 mutants (cat3) indicate that peroxisome-localized CAT plays a major role in restoring ROS homeostasis. Furthermore, protoplast cultures of pex11a, a peroxisome division mutant, and cat3 mutants show reduced induction of cell division. Taken together, the data indicate that peroxisome proliferation and CAT contribute to ROS homeostasis and subsequent protoplast division induction. PMID:26379686

  14. Mode of action framework analysis for receptor-mediated toxicity: the Peroxisome Proliferator-Activated Receptor alpha (PPAR?) as a case study

    EPA Science Inventory

    Therapeutic hypolipidemic agents and industrial chemicals that cause peroxisome proliferation and induce liver tumors in rodents activate the nuclear receptor peroxisome proliferator-activated receptor alpha (PPAR?). Research has elucidated the cellular and molecular events by w...

  15. Formation of the peroxisome lumen is abolished by loss of Pichia pastoris Pas7p, a zinc-binding integral membrane protein of the peroxisome.

    PubMed

    Kalish, J E; Theda, C; Morrell, J C; Berg, J M; Gould, S J

    1995-11-01

    We have cloned and sequenced PAS7, a gene required for peroxisome assembly in the yeast Pichia pastoris. The product of this gene, Pas7p, is a member of the C3HC4 superfamily of zinc-binding proteins. Point mutations that alter conserved residues of the C3HC4 motif abolish PAS7 activity and reduce zinc binding, suggesting that Pas7p binds zinc in vivo and that zinc binding is essential for PAS7 function. As with most pas mutants, pas7 cells exhibit a pronounced deficiency in import of peroxisomal matrix proteins that contain either the type 1 peroxisomal targeting signal (PTS1) or the type 2 PTS (PTS2). However, while other yeast and mammalian pas mutants accumulate ovoid, vesicular peroxisomal intermediates, loss of Pas7p leads to accumulation of membrane sheets and vesicles which lack a recognizable lumen. Thus, Pas7p appears to be essential for protein translocation into peroxisomes as well as formation of the lumen of the organelle. Consistent with these data, we find that Pas7p is an integral peroxisomal membrane protein which is entirely resistant to exogenous protease and thus appears to reside completely within the peroxisome. Our observations suggest that the function of Pas7p defines a previously unrecognized step in peroxisome assembly: formation of the peroxisome lumen. Furthermore, because the peroxisomal intermediates in the pas7 delta mutant proliferate in response to peroxisome-inducing environmental conditions, we conclude that Pas7p is not required for peroxisome proliferation. PMID:7565793

  16. Formation of the peroxisome lumen is abolished by loss of Pichia pastoris Pas7p, a zinc-binding integral membrane protein of the peroxisome.

    PubMed Central

    Kalish, J E; Theda, C; Morrell, J C; Berg, J M; Gould, S J

    1995-01-01

    We have cloned and sequenced PAS7, a gene required for peroxisome assembly in the yeast Pichia pastoris. The product of this gene, Pas7p, is a member of the C3HC4 superfamily of zinc-binding proteins. Point mutations that alter conserved residues of the C3HC4 motif abolish PAS7 activity and reduce zinc binding, suggesting that Pas7p binds zinc in vivo and that zinc binding is essential for PAS7 function. As with most pas mutants, pas7 cells exhibit a pronounced deficiency in import of peroxisomal matrix proteins that contain either the type 1 peroxisomal targeting signal (PTS1) or the type 2 PTS (PTS2). However, while other yeast and mammalian pas mutants accumulate ovoid, vesicular peroxisomal intermediates, loss of Pas7p leads to accumulation of membrane sheets and vesicles which lack a recognizable lumen. Thus, Pas7p appears to be essential for protein translocation into peroxisomes as well as formation of the lumen of the organelle. Consistent with these data, we find that Pas7p is an integral peroxisomal membrane protein which is entirely resistant to exogenous protease and thus appears to reside completely within the peroxisome. Our observations suggest that the function of Pas7p defines a previously unrecognized step in peroxisome assembly: formation of the peroxisome lumen. Furthermore, because the peroxisomal intermediates in the pas7 delta mutant proliferate in response to peroxisome-inducing environmental conditions, we conclude that Pas7p is not required for peroxisome proliferation. PMID:7565793

  17. Peroxisomes in Different Skeletal Cell Types during Intramembranous and Endochondral Ossification and Their Regulation during Osteoblast Differentiation by Distinct Peroxisome Proliferator-Activated Receptors

    PubMed Central

    Qian, Guofeng; Karnati, Srikanth; Baumgart-Vogt, Eveline

    2015-01-01

    Ossification defects leading to craniofacial dysmorphism or rhizomelia are typical phenotypes in patients and corresponding knockout mouse models with distinct peroxisomal disorders. Despite these obvious skeletal pathologies, to date no careful analysis exists on the distribution and function of peroxisomes in skeletal tissues and their alterations during ossification. Therefore, we analyzed the peroxisomal compartment in different cell types of mouse cartilage and bone as well as in primary cultures of calvarial osteoblasts. The peroxisome number and metabolism strongly increased in chondrocytes during endochondral ossification from the reserve to the hypertrophic zone, whereas in bone, metabolically active osteoblasts contained a higher numerical abundance of this organelle than osteocytes. The high abundance of peroxisomes in these skeletal cell types is reflected by high levels of Pex11? gene expression. During culture, calvarial pre-osteoblasts differentiated into secretory osteoblasts accompanied by peroxisome proliferation and increased levels of peroxisomal genes and proteins. Since many peroxisomal genes contain a PPAR-responsive element, we analyzed the gene expression of PPAR?/ß/? in calvarial osteoblasts and MC3T3-E1 cells, revealing higher levels for PPARß than for PPAR? and PPAR?. Treatment with different PPAR agonists and antagonists not only changed the peroxisomal compartment and associated gene expression, but also induced complex alterations of the gene expression patterns of the other PPAR family members. Studies in M3CT3-E1 cells showed that the PPARß agonist GW0742 activated the PPRE-mediated luciferase expression and up-regulated peroxisomal gene transcription (Pex11, Pex13, Pex14, Acox1 and Cat), whereas the PPARß antagonist GSK0660 led to repression of the PPRE and a decrease of the corresponding mRNA levels. In the same way, treatment of calvarial osteoblasts with GW0742 increased in peroxisome number and related gene expression and accelerated osteoblast differentiation. Taken together, our results suggest that PPARß regulates the numerical abundance and metabolic function of peroxisomes via Pex11ß in parallel to osteoblast differentiation. PMID:26630504

  18. Hypoxia-inducible factor 1 mediates hypoxia-induced cardiomyocyte lipid accumulation by reducing the DNA binding activity of peroxisome proliferator-activated receptor {alpha}/retinoid X receptor

    SciTech Connect

    Belanger, Adam J.; Luo Zhengyu; Vincent, Karen A.; Akita, Geoffrey Y.; Cheng, Seng H.; Gregory, Richard J.; Jiang Canwen

    2007-12-21

    In response to cellular hypoxia, cardiomyocytes adapt to consume less oxygen by shifting ATP production from mitochondrial fatty acid {beta}-oxidation to glycolysis. The transcriptional activation of glucose transporters and glycolytic enzymes by hypoxia is mediated by hypoxia-inducible factor 1 (HIF-1). In this study, we examined whether HIF-1 was involved in the suppression of mitochondrial fatty acid {beta}-oxidation in hypoxic cardiomyocytes. We showed that either hypoxia or adenovirus-mediated expression of a constitutively stable hybrid form (HIF-1{alpha}/VP16) suppressed mitochondrial fatty acid metabolism, as indicated by an accumulation of intracellular neutral lipid. Both treatments also reduced the mRNA levels of muscle carnitine palmitoyltransferase I which catalyzes the rate-limiting step in the mitochondrial import of fatty acids for {beta}-oxidation. Furthermore, adenovirus-mediated expression of HIF-1{alpha}/VP16 in cardiomyocytes under normoxic conditions also mimicked the reduction in the DNA binding activity of peroxisome proliferator-activated receptor {alpha} (PPAR{alpha})/retinoid X receptor (RXR), in the presence or absence of a PPAR{alpha} ligand. These results suggest that HIF-1 may be involved in hypoxia-induced suppression of fatty acid metabolism in cardiomyocytes by reducing the DNA binding activity of PPAR{alpha}/RXR.

  19. Selective Serotonin Reuptake Inhibitor-Induced Sexual Dysfunction in Adolescents: A Review.

    ERIC Educational Resources Information Center

    Scharko, Alexander M.

    2004-01-01

    Objective: To review the existing literature on selective serotonin reuptake inhibitor (SSRI)-induced sexual dysfunction in adolescents. Method: A literature review of SSRI-induced adverse effects in adolescents focusing on sexual dysfunction was done. Nonsexual SSRI-induced adverse effects were compared in adult and pediatric populations.…

  20. The peroxisome: still a mysterious organelle

    PubMed Central

    Fahimi, H. Dariush

    2008-01-01

    More than half a century of research on peroxisomes has revealed unique features of this ubiquitous subcellular organelle, which have often been in disagreement with existing dogmas in cell biology. About 50 peroxisomal enzymes have so far been identified, which contribute to several crucial metabolic processes such as ?-oxidation of fatty acids, biosynthesis of ether phospholipids and metabolism of reactive oxygen species, and render peroxisomes indispensable for human health and development. It became obvious that peroxisomes are highly dynamic organelles that rapidly assemble, multiply and degrade in response to metabolic needs. However, many aspects of peroxisome biology are still mysterious. This review addresses recent exciting discoveries on the biogenesis, formation and degradation of peroxisomes, on peroxisomal dynamics and division, as well as on the interaction and cross talk of peroxisomes with other subcellular compartments. Furthermore, recent advances on the role of peroxisomes in medicine and in the identification of novel peroxisomal proteins are discussed. PMID:18274771

  1. Arsenic toxicity induced endothelial dysfunction and dementia: Pharmacological interdiction by histone deacetylase and inducible nitric oxide synthase inhibitors

    SciTech Connect

    Sharma, Bhupesh Sharma, P.M.

    2013-11-15

    Arsenic toxicity has been reported to damage all the major organs including the brain and vasculature. Dementia including Alzheimer's disease (AD) and vascular dementia (VaD) are posing greater risk to the world population as it is now increasing at a faster rate. We have investigated the role of sodium butyrate, a selective histone deacetylase (HDAC) inhibitor and aminoguanidine, a selective inducible nitric oxide synthase (iNOS) inhibitor in pharmacological interdiction of arsenic toxicity induced vascular endothelial dysfunction and dementia in rats. Arsenic toxicity was done by administering arsenic drinking water to rats. Morris water-maze (MWM) test was used for assessment of learning and memory. Endothelial function was assessed using student physiograph. Oxidative stress (aortic superoxide anion, serum and brain thiobarbituric acid reactive species, brain glutathione) and nitric oxide levels (serum nitrite/nitrate) were also measured. Arsenic treated rats have shown impairment of endothelial function, learning and memory, reduction in serum nitrite/nitrate and brain GSH levels along with increase in serum and brain TBARS. Sodium butyrate as well as aminoguanidine significantly convalesce arsenic induced impairment of learning, memory, endothelial function, and alterations in various biochemical parameters. It may be concluded that arsenic induces endothelial dysfunction and dementia, whereas, sodium butyrate, a HDAC inhibitor as well as aminoguanidine, a selective iNOS inhibitor may be considered as potential agents for the management of arsenic induced endothelial dysfunction and dementia. - Highlights: • As has induced endothelial dysfunction (Edf) and vascular dementia (VaD). • As has increased oxidative stress, AChE activity and decreased serum NO. • Inhibitors of HDAC and iNOS have attenuated As induced Edf and VaD. • Both the inhibitors have attenuated As induced biochemical changes. • Inhibitor of HDAC and iNOS has shown good potential in As induced VaD.

  2. Cloning and characterization of PAS5: a gene required for peroxisome biogenesis in the methylotrophic yeast Pichia pastoris

    PubMed Central

    1993-01-01

    The biogenesis and maintenance of cellular organelles is of fundamental importance in all eukaryotic cells. One such organelle is the peroxisome. The establishment of a genetic system to study peroxisome biogenesis in the methylotrophic yeast Pichia pastoris has yielded many different complementation groups of peroxisomal assembly (pas) or peroxisome-deficient (per) mutants. Each appears to be deficient in functional peroxisomes. One of these mutants, pas5, has been characterized, complemented, and the gene sequenced. Ultrastructural studies show that normal peroxisomes are not present in pas5, but aberrant peroxisomal structures resembling "membranous ghosts" are frequently observed. The "peroxisome ghosts" appear to be induced and segregated to daughter cells normally. Biochemical fractionation analysis of organelles of the pas5 mutant reveals that peroxisomal matrix enzymes are induced normally but are found mostly in the cytosol. However, purification of peroxisome ghosts from the mutant shows that small amounts (< 5%) of matrix enzymes are imported. The PAS5 gene was cloned and found to encode a 127-kD protein, which contains a 200-amino acid-long region of homology with PAS1, NEM- sensitive factor (NSF), and other related ATPases. Weak homology to a yeast myosin was also observed. The gene is not essential for growth on glucose but is essential for growth on oleic acid and methanol. The role of PAS5 in peroxisome biogenesis is discussed. PMID:8227124

  3. Dual-Action Inhibitors of HIF Prolyl Hydroxylases That Induce Binding of a Second Iron Ion

    PubMed Central

    Thalhammer, Armin; Demetriades, Marina; Chowdhury, Rasheduzzaman; Tian, Ya-Min; Stolze, Ineke; McNeill, Luke A.; Lee, Myung Kyu; Woon, Esther C. Y.; Mackeen, Mukram M.; Kawamura, Akane; Ratcliffe, Peter J.; Mecinovi?, Jasmin; Schofield, Christopher J.

    2015-01-01

    Inhibition of the hypoxia-inducible factor (HIF) prolyl-hydroxylases (PHD or EGLN enzymes) is of interest for the treatment of anemia and ischemia-related diseases. Most PHD inhibitors work by binding to the single ferrous ion and competing with 2-oxoglutarate (2OG) co-substrate for binding at the PHD active site. Non-specific iron chelators also inhibit the PHDs, both in vitro and in cells. We report the identification of dual action PHD inhibitors, which bind to the active site iron and also induce the binding of a second iron ion at the active site. Following analysis of small-molecule iron complexes and application of non-denaturing protein mass spectrometry to assess PHD2·iron·inhibitor stoichimetry, selected diacylhydrazines were identified as PHD2 inhibitors that induce the binding of a second iron ion. Some compounds were shown to inhibit the HIF hydroxylases in human hepatoma and renal carcinoma cell lines. PMID:23151668

  4. The Synergistic Enhancement of Cloning Efficiency in Individualized Human Pluripotent Stem Cells by Peroxisome Proliferative-activated Receptor-? (PPAR?) Activation and Rho-associated Kinase (ROCK) Inhibition.

    PubMed

    Kajabadi, Nasim-Sadat; Ghoochani, Ali; Peymani, Maryam; Ghaedi, Kamran; Kiani-Esfahani, Abbas; Hashemi, Motahareh-Sadat; Nasr-Esfahani, Mohammad Hossein; Baharvand, Hossein

    2015-10-23

    Although human pluripotent stem cells (hPSCs) provide valuable sources for regenerative medicine, their applicability is dependent on obtaining both suitable up-scaled and cost effective cultures. The Rho-associated kinase (ROCK) inhibitor Y-27632 permits hPSC survival upon dissociation; however, cloning efficiency is often still low. Here we have shown that pioglitazone, a selective peroxisome proliferative-activated receptor-? agonist, along with Y-27632 synergistically diminished dissociation-induced apoptosis and increased cloning efficiency (2-3-fold versus Y-27632) without affecting pluripotency of hPSCs. Pioglitazone exerted its positive effect by inhibition of glycogen synthase kinase (GSK3) activity and enhancement of membranous ?-catenin and E-cadherin proteins. These effects were reversed by GW-9662, an irreversible peroxisome proliferative-activated receptor-? antagonist. This novel setting provided a step toward hPSC manipulation and its biomedical applications. PMID:26336103

  5. Maternal protein restriction induces early-onset glucose intolerance and alters hepatic genes expression in the peroxisome proliferator-activated receptor pathway in offspring

    PubMed Central

    Zheng, Jia; Xiao, Xinhua; Zhang, Qian; Yu, Miao; Xu, Jianping; Wang, Zhixin

    2015-01-01

    Aims/Introduction Maternal undernutrition during pregnancy and/or lactation can alter the offspring's response to environmental challenges, and thus increases the risk of the development of metabolic diseases at a later age. However, whether maternal protein restriction can modulate glucose metabolism in the early life of offspring is less understood. Furthermore, we explored the potential underlying mechanisms that illustrate this phenotype. Materials and Methods To test this hypothesis, we examined the offspring of C57BL/6J mice at weaning to determine the effects of feeding their mothers a low-protein diet or normal chow diet throughout pregnancy and lactation. Gene array experiments and quantitative real-time polymerase chain reaction were utilized to explore the altered hepatic genes expression. Results The offspring of dams fed a low-protein diet had a lower birthweight and bodyweight, impaired glucose tolerance, decreased insulin sensitivity, and decreased serum cholesterol at weaning. Using gene array experiments, 253 differentially expressed genes were identified in the liver tissues of the offspring between the two groups. Bioinformatic analyses showed that all differentially expressed genes were mapped to 11 pathways. We focused on the ‘peroxisome proliferator-activated receptor signaling pathway,’ because peroxisome proliferator-activated receptors have emerged as central regulators of glucose and lipid homeostasis. Quantitative real-time polymerase chain reaction was utilized for the validation of genes in the pathway. Conclusions A maternal low-protein diet during pregnancy and lactation promotes early-onset glucose intolerance in the offspring mice, and the altered hepatic genes expression in peroxisome proliferator-activated receptor signaling pathway could play role in regulating this phenomenon. PMID:25969711

  6. BET inhibitors induce apoptosis through a MYC independent mechanism and synergise with CDK inhibitors to kill osteosarcoma cells

    PubMed Central

    Baker, Emma K; Taylor, Scott; Gupte, Ankita; Sharp, Phillip P; Walia, Mannu; Walsh, Nicole C; Zannettino, Andrew CW; Chalk, Alistair M; Burns, Christopher J; Walkley, Carl R

    2015-01-01

    Osteosarcoma (OS) survival rates have plateaued in part due to a lack of new therapeutic options. Here we demonstrate that bromodomain inhibitors (BETi), JQ1, I-BET151, I-BET762, exert potent anti-tumour activity against primary and established OS cell lines, mediated by inhibition of BRD4. Strikingly, unlike previous observations in long-term established human OS cell lines, the antiproliferative activity of JQ1 in primary OS cells was driven by the induction of apoptosis, not cell cycle arrest. In further contrast, JQ1 activity in OS was mediated independently of MYC downregulation. We identified that JQ1 suppresses the transcription factor FOSL1 by displacement of BRD4 from its locus. Loss of FOSL1 phenocopied the antiproliferative effects of JQ1, identifying FOSL1 suppression as a potential novel therapeutic approach for OS. As a monotherapy JQ1 demonstrated significant anti-tumour activity in vivo in an OS graft model. Further, combinatorial treatment approaches showed that JQ1 increased the sensitivity of OS cells to doxorubicin and induced potent synergistic activity when rationally combined with CDK inhibitors. The greater level of activity achieved with the combination of BETi with CDK inhibitors demonstrates the efficacy of this combination therapy. Taken together, our studies show that BET inhibitors are a promising new therapeutic for OS. PMID:25944566

  7. Reevaluation of the role of Pex1 and dynamin-related proteins in peroxisome membrane biogenesis.

    PubMed

    Motley, Alison M; Galvin, Paul C; Ekal, Lakhan; Nuttall, James M; Hettema, Ewald H

    2015-12-01

    A recent model for peroxisome biogenesis postulates that peroxisomes form de novo continuously in wild-type cells by heterotypic fusion of endoplasmic reticulum-derived vesicles containing distinct sets of peroxisomal membrane proteins. This model proposes a role in vesicle fusion for the Pex1/Pex6 complex, which has an established role in matrix protein import. The growth and division model proposes that peroxisomes derive from existing peroxisomes. We tested these models by reexamining the role of Pex1/Pex6 and dynamin-related proteins in peroxisome biogenesis. We found that induced depletion of Pex1 blocks the import of matrix proteins but does not affect membrane protein delivery to peroxisomes; markers for the previously reported distinct vesicles colocalize in pex1 and pex6 cells; peroxisomes undergo continued growth if fission is blocked. Our data are compatible with the established primary role of the Pex1/Pex6 complex in matrix protein import and show that peroxisomes in Saccharomyces cerevisiae multiply mainly by growth and division. PMID:26644516

  8. The Pichia pastoris PER6 gene product is a peroxisomal integral membrane protein essential for peroxisome biogenesis and has sequence similarity to the Zellweger syndrome protein PAF-1.

    PubMed Central

    Waterham, H R; de Vries, Y; Russel, K A; Xie, W; Veenhuis, M; Cregg, J M

    1996-01-01

    We report the cloning of PER6, a gene essential for peroxisome biogenesis in the methylotrophic yeast Pichia pastoris. The PER6 sequence predicts that its product Per6p is a 52-kDa polypeptide with the cysteine-rich C3HC4 motif. Per6p has significant overall sequence similarity with the human peroxisome assembly factor PAF-1, a protein that is defective in certain patients suffering from the peroxisomal disorder Zellweger syndrome, and with car1, a protein required for peroxisome biogenesis and caryogamy in the filamentous fungus Podospora anserina. In addition, the C3HC4 motif and two of the three membrane-spanning segments predicted for Per6p align with the C3HC4 motifs and the two membrane-spanning segments predicted for PAF-1 and car1. Like PAF-1, Per6p is a peroxisomal integral membrane protein. In methanol- or oleic acid-induced cells of per6 mutants, morphologically recognizable peroxisomes are absent. Instead, peroxisomal remnants are observed. In addition, peroxisomal matrix proteins are synthesized but located in the cytosol. The similarities between Per6p and PAF-1 in amino acid sequence and biochemical properties, and between mutants defective in their respective genes, suggest that Per6p is the putative yeast homolog of PAF-1. PMID:8628321

  9. Drug-induced osteoporosis: from Fuller Albright to aromatase inhibitors.

    PubMed

    Byreddy, D V; Bouchonville Ii, M F; Lewiecki, E M

    2015-12-01

    Many commonly prescribed medications, such as selective serotonin reuptake inhibitors, proton pump inhibitors, thiazolidinediones, aromatase inhibitors, and androgen deprivation therapy, have been associated with adverse skeletal effects. The levels of evidence in support of a causal relationship between drug use and the development of bone loss and fractures are variable. For some drugs, a causal relationship is suspected (but not proven) based on observational studies, while in others causality is firmly established with randomized, controlled clinical trials. The mechanism of action for skeletal damage is poorly understood for some drugs and well known for others. Guidelines for managing bone health in patients taking some medications with potential skeletal toxicity have been developed using the best available evidence and expert opinion. This is a review of selected medications that have been associated with bone loss and fractures, with recommendations for clinical care. PMID:26488130

  10. Resistance to BRAF inhibitors induces glutamine dependency in melanoma cells.

    PubMed

    Baenke, Franziska; Chaneton, Barbara; Smith, Matthew; Van Den Broek, Niels; Hogan, Kate; Tang, Haoran; Viros, Amaya; Martin, Matthew; Galbraith, Laura; Girotti, Maria R; Dhomen, Nathalie; Gottlieb, Eyal; Marais, Richard

    2016-01-01

    BRAF inhibitors can extend progression-free and overall survival in melanoma patients whose tumors harbor mutations in BRAF. However, the majority of patients eventually develop resistance to these drugs. Here we show that BRAF mutant melanoma cells that have developed acquired resistance to BRAF inhibitors display increased oxidative metabolism and increased dependency on mitochondria for survival. Intriguingly, the increased oxidative metabolism is associated with a switch from glucose to glutamine metabolism and an increased dependence on glutamine over glucose for proliferation. We show that the resistant cells are more sensitive to mitochondrial poisons and to inhibitors of glutaminolysis, suggesting that targeting specific metabolic pathways may offer exciting therapeutic opportunities to treat resistant tumors, or to delay emergence of resistance in the first-line setting. PMID:26365896

  11. 5-Lipoxygenase Inhibitors Attenuate TNF-?-Induced Inflammation in Human Synovial Fibroblasts

    PubMed Central

    Lin, Han-Ching; Lin, Tzu-Hung; Wu, Ming-Yueh; Chiu, Yung-Cheng; Tang, Chih-Hsin; Hour, Mann-Jen; Liou, Houng-Chi; Tu, Huang-Ju; Yang, Rong-Sen; Fu, Wen-Mei

    2014-01-01

    The lipoxygenase isoform of 5-lipoxygenase (5-LOX) is reported to be overexpressed in human rheumatoid arthritis synovial tissue and involved in the progress of inflammatory arthritis. However, the detailed mechanism of how 5-lipoxygenase regulates the inflammatory response in arthritis synovial tissue is still unclear. The aim of this study was to investigate the involvement of lipoxygenase pathways in TNF-?-induced production of cytokines and chemokines. Human synovial fibroblasts from rheumatoid patients were used in this study. 5-LOX inhibitors and shRNA were used to examine the involvement of 5-LOX in TNF-?-induced cytokines and chemokines expression. The signaling pathways were examined by Western Blotting or immunofluorescence staining. The effect of 5-LOX inhibitor on TNF-?-induced chemokine expression and paw edema was also explored in vivo in C57BL/6 mice. Treatment with 5-LOX inhibitors significantly decreased TNF-?-induced pro-inflammatory mediators including interleukin-6 (IL-6) and monocyte chemo-attractant protein-1 (MCP-1) in human synovial fibroblasts. Knockdown of 5-LOX using shRNA exerted similar inhibitory effects. The abrogation of NF-?B activation was involved in the antagonizing effects of these inhibitors. Furthermore, 5-LOX inhibitor decreased TNF-?-induced up-regulation of serum MCP-1 level and paw edema in mouse model. Our results provide the evidence that the administration of 5-LOX inhibitors is able to ameliorate TNF-?-induced cytokine/chemokine release and paw edema, indicating that 5-LOX inhibitors may be developed for therapeutic treatment of inflammatory arthritis. PMID:25229347

  12. The histone deacetylase inhibitor entinostat (SNDX-275) induces apoptosis in Hodgkin lymphoma cells and synergizes with Bcl-2 family inhibitors

    PubMed Central

    Jóna, Ádám; Khaskhely, Noor; Buglio, Daniela; Shafer, Jessica A.; Derenzini, Enrico; Bollard, Catherine M.; Medeiros, L. Jeffrey; Illés, Árpád; Ji, Yuan; Younes, Anas

    2011-01-01

    Objective Based on promising in vitro and in vivo activity of several histone deacetylase inhibitors (HDACi’s) in HL (Hodgkin lymphoma), we investigated SNDX-275, an oral class 1 isoform–selective HDACi in HL-derived cell lines. Materials and Methods Proliferation and cell death were examined by MTS assay, Annexin-V/propidium iodide, and FACS analysis. Gene and protein expression were measured by RT-PCR, Western blotting, and immunohistochemical analysis. A multiplex assay was used to determine cytokines and chemokines. Results SNDX-275 induced cell death in a dose- and time-dependent manner with an IC50 at the sub- and lower micromolar range at 72 hours. At the molecular level, SNDX-275 increased histone H3 acetylation, up-regulated p21 expression, and activated the intrinsic apoptosis pathway by down-regulating the X-linked inhibitor of apoptosis protein (XIAP). SNDX-275 down-regulated the expression of antiapoptotic Bcl-2 and Bcl-xL proteins without altering Mcl-1 or Bax levels. Combination studies demonstrated that two Bcl-2 inhibitors (ABT-737 and obatoclax) significantly enhanced the effect of SNDX-275. SNDX-275 modulated the level of several cytokines and chemokines, including IL-12 p40-70, IP-10, RANTES, IL-13, IL-4, and TARC, and variably induced the cancer/testis antigen expression of MAGE-A4 and survivin in HL cell lines. Conclusions SNDX-275 has antiproliferative activity in HL cell lines, involving several mechanisms: induction of apoptosis, regulation of cytokines and chemokines, and alteration of CTAs. Clinical investigation of SNDX-275 alone or in combination with Bcl-2 inhibitors is warranted in patients with HL. Phase 2 studies with SNDX-275 in HL are ongoing, and future clinical studies should investigate combinations with SNDX-275. PMID:21767511

  13. Angiotensin-converting enzyme inhibitors-induced angioedema treated by C1 esterase inhibitor concentrate (Berinert®): about one case and review of the therapeutic arsenal.

    PubMed

    Lipski, Samuel Michael; Casimir, Georges; Vanlommel, Martine; Jeanmaire, Mathieu; Dolhen, Pierre

    2015-02-01

    C1 esterase inhibitor (Berinert®) is generally used to treat severe attack of hereditary angioedema. We describe here the case of a patient who presented with a severe angioedema induced by angiotensin-converting enzyme inhibitors (ACEIs) endangering her life. It could be successfully treated with that medicine. PMID:25767713

  14. Angiotensin-converting enzyme inhibitors-induced angioedema treated by C1 esterase inhibitor concentrate (Berinert®): about one case and review of the therapeutic arsenal

    PubMed Central

    Lipski, Samuel Michael; Casimir, Georges; Vanlommel, Martine; Jeanmaire, Mathieu; Dolhen, Pierre

    2015-01-01

    Key Clinical Message C1 esterase inhibitor (Berinert®) is generally used to treat severe attack of hereditary angioedema. We describe here the case of a patient who presented with a severe angioedema induced by angiotensin-converting enzyme inhibitors (ACEIs) endangering her life. It could be successfully treated with that medicine. PMID:25767713

  15. The peroxisome proliferator-activated receptor ?, an integrator of transcriptional repression and nuclear receptor signaling

    PubMed Central

    Shi, Yanhong; Hon, Michelle; Evans, Ronald M.

    2002-01-01

    The three PPAR (peroxisome proliferator-activated receptor) isoforms are critical regulators of lipid homeostasis by controlling the balance between the burning and storage of long chain fatty acids. Whereas PPAR? and PPAR? have been studied extensively, the function of PPAR? remains the most elusive. Intriguingly, in cotransfection experiments, PPAR? is a potent inhibitor of ligand-induced transcriptional activity of PPAR? and PPAR?. This inhibition is achieved, in part, by binding of PPAR? to a peroxisome proliferator response element and the association of nonliganded PPAR? with corepressors SMRT (silencing mediator for retinoid and thyroid hormone receptors), SHARP (SMRT and histone deacetylase-associated repressor protein), and class I histone deacetylases. Stable expression of PPAR? inhibits the expression of endogenous PPAR? target gene expression in 3T3-PPAR? cells, whereas a PPAR? mutant that does not interact with the corepressor SMRT loses its ability to repress the induction of PPAR? target gene. Similarly, stable expression of PPAR? in 3T3-PPAR? cells leads to inhibition of PPAR? target gene expression and PPAR?-mediated adipogenesis. Given the widespread expression of PPAR? and the restricted pattern for PPAR? and PPAR?, these results suggest a role for PPAR? as a gateway receptor whose relative levels of expression can be used to modulate PPAR? and PPAR? activity. PMID:11867749

  16. Combined Treatment With Peroxisome Proliferator-Activated Receptor (PPAR) Gamma Ligands and Gamma Radiation Induces Apoptosis by PPAR?-Independent Up-Regulation of Reactive Oxygen Species-Induced Deoxyribonucleic Acid Damage Signals in Non-Small Cell Lung Cancer Cells

    SciTech Connect

    Han, Eun Jong; Im, Chang-Nim; Park, Seon Hwa; Moon, Eun-Yi; Hong, Sung Hee

    2013-04-01

    Purpose: To investigate possible radiosensitizing activities of the well-known peroxisome proliferator-activated receptor (PPAR)? ligand ciglitazone and novel PPAR? ligands CAY10415 and CAY10506 in non-small cell lung cancer (NSCLC) cells. Methods and Materials: Radiosensitivity was assessed using a clonogenic cell survival assay. To investigate the mechanism underlying PPAR? ligand-induced radiosensitization, the subdiploid cellular DNA fraction was analyzed by flow cytometry. Activation of the caspase pathway by combined PPAR? ligands and ?-radiation treatment was detected by immunoblot analysis. Reactive oxygen species (ROS) were measured using 2,7-dichlorodihydrofluorescein diacetate and flow cytometry. Results: The 3 PPAR? ligands induced cell death and ROS generation in a PPAR?-independent manner, enhanced ?-radiation–induced apoptosis and caspase-3–mediated poly (ADP-ribose) polymerase (PARP) cleavage in vitro. The combined PPAR? ligand/?-radiation treatment triggered caspase-8 activation, and this initiator caspase played an important role in the combination-induced apoptosis. Peroxisome proliferator-activated receptor-? ligands may enhance the ?-radiation-induced DNA damage response, possibly by increasing ?-H2AX expression. Moreover, the combination treatment significantly increased ROS generation, and the ROS scavenger N-acetylcysteine inhibited the combined treatment-induced ROS generation and apoptotic cell death. Conclusions: Taken together, these results indicated that the combined treatment of PPAR? ligands and ?-radiation synergistically induced DNA damage and apoptosis, which was regulated by ROS.

  17. Acyldepsipeptide HDAC inhibitor production induced in Burkholderia thailandensis

    PubMed Central

    Biggins, John B.; Gleber, Conrad D.; Brady, Sean F.

    2011-01-01

    Natural product gene clusters are often tightly regulated, resulting in gene cluster silencing in laboratory fermentation studies. The systematic overexpression of transcription factors (TFs) associated with biosynthetic gene clusters found in the genome of Burkholderia thailandensis E264 identified a set of TFs that, when overexpressed, alter the secondary metabolome of this bacterium. The isolation and characterization of burkholdacs A and B, two new acyldepsitripeptide histone deacetylase inhibitors produced by B. thailandensis overexpressing the TF bhcM is reported. PMID:21348454

  18. Fis1, DLP1, and Pex11p coordinately regulate peroxisome morphogenesis

    SciTech Connect

    Kobayashi, Shinta; Tanaka, Atsushi; Fujiki, Yukio . E-mail: yfujiscb@mbox.nc.kyushu-u.ac.jp

    2007-05-01

    Dynamin-like protein 1 (DLP1) and Pex11p{beta} function in morphogenesis of peroxisomes. In the present work, we investigated whether Fis1 is involved in fission of peroxisomes. Endogenous Fis1 was morphologically detected in peroxisomes as well as mitochondria in wild-type CHO-K1 and DLP1-defective ZP121 cells. Subcellular fractionation studies also revealed the presence of Fis1 in peroxisomes. Peroxisomal Fis1 showed the same topology, i.e., C-tail anchored membrane protein, as the mitochondrial one. Furthermore, ectopic expression of FIS1 induced peroxisome proliferation in CHO-K1 cells, while the interference of FIS1 RNA resulted in tubulation of peroxisomes, hence reducing the number of peroxisomes. Fis1 interacted with Pex11p{beta}, by direct binding apparently involving the C-terminal region of Pex11p{beta} in the interaction. Pex11p{beta} also interacted with each other, whereas the binding of Pex11p{beta} to DLP1 was not detectable. Moreover, ternary complexes comprising Fis1, Pex11p{beta}, and DLP1 were detected by chemical cross-linking. We also showed that the highly conserved N-terminal domain of Pex11p{beta} was required for the homo-oligomerization of Pex11p{beta} and indispensable for the peroxisome-proliferating activity. Taken together, these findings indicate that Fis1 plays important roles in peroxisome division and maintenance of peroxisome morphology in mammalian cells, possibly in a concerted manner with Pex11p{beta} and DLP1.

  19. Icatibant in the Treatment of Angiotensin-Converting Enzyme Inhibitor-Induced Angioedema

    PubMed Central

    Crooks, Neil H.; Patel, Jaimin; Diwakar, Lavanya; Smith, Fang Gao

    2014-01-01

    We describe the case of a 75-year-old woman who presented with massive tongue and lip swelling secondary to angiotensin-converting enzyme inhibitor-induced angioedema. An awake fibre-optic intubation was performed because of impending airway obstruction. As there was no improvement in symptoms after 72 hours, the selective bradykinin B2 receptor antagonist icatibant (Firazyr) was administered and the patient's trachea was successfully extubated 36 hours later. To our knowledge this is the first documented case of icatibant being used for the treatment of angiotensin-converting enzyme inhibitor-induced angioedema in the United Kingdom and represents a novel therapeutic option in its management. PMID:25328718

  20. Peroxisomes and sexual development in fungi

    PubMed Central

    Peraza-Reyes, Leonardo; Berteaux-Lecellier, Véronique

    2013-01-01

    Peroxisomes are versatile and dynamic organelles that are essential for the development of most eukaryotic organisms. In fungi, many developmental processes, such as sexual development, require the activity of peroxisomes. Sexual reproduction in fungi involves the formation of meiotic-derived sexual spores, often takes place inside multicellular fruiting bodies and requires precise coordination between the differentiation of multiple cell types and the progression of karyogamy and meiosis. Different peroxisomal functions contribute to the orchestration of this complex developmental process. Peroxisomes are required to sustain the formation of fruiting bodies and the maturation and germination of sexual spores. They facilitate the mobilization of reserve compounds via fatty acid ?-oxidation and the glyoxylate cycle, allowing the generation of energy and biosynthetic precursors. Additionally, peroxisomes are implicated in the progression of meiotic development. During meiotic development in Podospora anserina, there is a precise modulation of peroxisome assembly and dynamics. This modulation includes changes in peroxisome size, number and localization, and involves a differential activity of the protein-machinery that drives the import of proteins into peroxisomes. Furthermore, karyogamy, entry into meiosis and sorting of meiotic-derived nuclei into sexual spores all require the activity of peroxisomes. These processes rely on different peroxisomal functions and likely depend on different pathways for peroxisome assembly. Indeed, emerging studies support the existence of distinct import channels for peroxisomal proteins that contribute to different developmental stages. PMID:24046747

  1. Peroxisomal ABC transporters: functions and mechanism

    PubMed Central

    Baker, Alison; Carrier, David J.; Schaedler, Theresia; Waterham, Hans R.; van Roermund, Carlo W.; Theodoulou, Frederica L.

    2015-01-01

    Peroxisomes are arguably the most biochemically versatile of all eukaryotic organelles. Their metabolic functions vary between different organisms, between different tissue types of the same organism and even between different developmental stages or in response to changed environmental conditions. New functions for peroxisomes are still being discovered and their importance is underscored by the severe phenotypes that can arise as a result of peroxisome dysfunction. The ?-oxidation pathway is central to peroxisomal metabolism, but the substrates processed are very diverse, reflecting the diversity of peroxisomes across species. Substrates for ?-oxidation enter peroxisomes via ATP-binding cassette (ABC) transporters of subfamily D; (ABCD) and are activated by specific acyl CoA synthetases for further metabolism. Humans have three peroxisomal ABCD family members, which are half transporters that homodimerize and have distinct but partially overlapping substrate specificity; Saccharomyces cerevisiae has two half transporters that heterodimerize and plants have a single peroxisomal ABC transporter that is a fused heterodimer and which appears to be the single entry point into peroxisomes for a very wide variety of ?-oxidation substrates. Our studies suggest that the Arabidopsis peroxisomal ABC transporter AtABCD1 accepts acyl CoA substrates, cleaves them before or during transport followed by reactivation by peroxisomal synthetases. We propose that this is a general mechanism to provide specificity to this class of transporters and by which amphipathic compounds are moved across peroxisome membranes. PMID:26517910

  2. Proton pump inhibitor-induced hypomagnesemia: A new challenge.

    PubMed

    Florentin, Matilda; Elisaf, Moses S

    2012-12-01

    Proton pump inhibitors (PPIs) are commonly used in clinical practice for the prevention and treatment of peptic ulcer, gastritis, esophagitis and gastroesophageal reflux. Hypomagnesemia has recently been recognized as a side effect of PPIs. Low magnesium levels may cause symptoms from several systems, some of which being potentially serious, such as tetany, seizures and arrhythmias. It seems that PPIs affect the gastrointestinal absorption of magnesium. Clinicians should be vigilant in order to timely consider and prevent or reverse hypomagnesemia in patients who take PPIs, especially if they are prone to this electrolyte disorder. PMID:24175253

  3. Apoptosis induced by inhibitors of the plasma membrane NADH-oxidase involves Bcl-2 and calcineurin.

    PubMed

    Wolvetang, E J; Larm, J A; Moutsoulas, P; Lawen, A

    1996-10-01

    Activation of the plasma membrane NADH-oxidoreductase (PMOR) system by addition of growth factors or extracellular electron acceptors stimulates cellular proliferation. We now show that the vanilloids capsaicin, dihydrocapsaicin, and resiniferatoxin are inhibitors of the NADH-oxidase activity of the PMOR system and that both these and two previously identified PMOR inhibitors (chloroquine and retinoic acid) induce apoptosis in human B-cell and mouse myeloid cell lines. At the optimal concentration, PMOR inhibitors can induce between 50 and 70% of apoptosis in mouse myeloid and human B-cell lines within 8-12 h, provided these cell lines do not express Bcl-2. The immunosuppressants cyclosporin A and fujimycin (tacrolimus) inhibit PMOR inhibitor-induced apoptosis. By using combinations of these immunosuppressants and excess amounts of their nonimmunosuppressive analogues, we demonstrate that in human B-cell lines the Bcl-2-sensitive apoptotic pathway triggered by PMOR inhibitors involves signaling through the protein phosphatase calcineurin. We suggest that the PMOR system is a redox sensor that can, depending on the ambient redox environment and the availability of growth factors, regulate plasma membrane calcium fluxes and signal for apoptosis through calcineurin. Bcl-2, a protein that is thought to inhibit apoptosis by regulating reactive oxygen species and calcium fluxes in the cell, inhibits this apoptotic pathway. PMID:8891335

  4. UK114, a YjgF/Yer057p/UK114 family protein highly conserved from bacteria to mammals, is localized in rat liver peroxisomes

    SciTech Connect

    Antonenkov, Vasily D. . E-mail: vasily.antonenkov@oulu.fi; Ohlmeier, Steffen; Sormunen, Raija T.; Hiltunen, J. Kalervo

    2007-05-25

    Mammalian UK114 belongs to a highly conserved family of proteins with unknown functions. Although it is believed that UK114 is a cytosolic or mitochondrial protein there is no detailed study of its intracellular localization. Using analytical subcellular fractionation, electron microscopic colloidal gold technique, and two-dimensional gel electrophoresis of peroxisomal matrix proteins combined with mass spectrometric analysis we show here that a large portion of UK114 is present in rat liver peroxisomes. The peroxisomal UK114 is a soluble matrix protein and it is not inducible by the peroxisomal proliferator clofibrate. The data predict involvement of UK114 in peroxisomal metabolism.

  5. AMP-Activated Protein Kinase Interacts with the Peroxisome Proliferator-Activated Receptor Delta to Induce Genes Affecting Fatty Acid Oxidation in Human Macrophages

    PubMed Central

    Kemmerer, Marina; Finkernagel, Florian; Cavalcante, Marcela Frota; Abdalla, Dulcineia Saes Parra; Müller, Rolf; Brüne, Bernhard; Namgaladze, Dmitry

    2015-01-01

    AMP-activated protein kinase (AMPK) maintains energy homeostasis by suppressing cellular ATP-consuming processes and activating catabolic, ATP-producing pathways such as fatty acid oxidation (FAO). The transcription factor peroxisome proliferator-activated receptor ? (PPAR?) also affects fatty acid metabolism, stimulating the expression of genes involved in FAO. To question the interplay of AMPK and PPAR? in human macrophages we transduced primary human macrophages with lentiviral particles encoding for the constitutively active AMPK?1 catalytic subunit, followed by microarray expression analysis after treatment with the PPAR? agonist GW501516. Microarray analysis showed that co-activation of AMPK and PPAR? increased expression of FAO genes, which were validated by quantitative PCR. Induction of these FAO-associated genes was also observed upon infecting macrophages with an adenovirus coding for AMPK?1 regulatory subunit carrying an activating R70Q mutation. The pharmacological AMPK activator A-769662 increased expression of several FAO genes in a PPAR?- and AMPK-dependent manner. Although GW501516 significantly increased FAO and reduced the triglyceride amount in very low density lipoproteins (VLDL)-loaded foam cells, AMPK activation failed to potentiate this effect, suggesting that increased expression of fatty acid catabolic genes alone may be not sufficient to prevent macrophage lipid overload. PMID:26098914

  6. DNA Methyltransferase Inhibitors Coordinately Induce Expression of the Human Reelin and Glutamic Acid

    E-print Network

    Champagne, Frances A.

    DNA Methyltransferase Inhibitors Coordinately Induce Expression of the Human Reelin and Glutamic activity and DNA methyltransferase 1 (DNMT1) pro- tein levels. Time course Western blot analysis showed., 2005). These findings, together with studies using condi- tional DNA methyltransferase 1 (DNMT1) mutant

  7. Insect and wound induced GUS gene expression from a Beta vulgaris proteinase inhibitor gene promoter

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Inducible gene promoters that are specifically activated by pathogen invasion or insect pest attack are needed for effective expression of resistance genes to control plant diseases. In the present study, a promoter from a serine proteinase inhibitor gene (BvSTI) shown to be up-regulated in resist...

  8. [Effects of the 5-lipoxygenase inhibitor phenidone on antigen-induced release of histamine in vitro].

    PubMed

    Sauer, R R; Sauer, C I; Merk, H; Steigleder, G K

    1987-08-01

    The 5-lipoxygenase inhibitor Phenidone inhibits the antigen-induced release of histamine from human basophiles at concentrations of 10(-4) up to 10(-5) mol/l. These findings indicate once more that products of the arachidonic acid metabolism are involved in the induction of histamine release from human basophiles. PMID:2445116

  9. Proteasome inhibitors induce apoptosis and reduce viral replication in primary effusion lymphoma cells

    SciTech Connect

    Saji, Chiaki; Higashi, Chizuka; Niinaka, Yasufumi; Yamada, Koji; Noguchi, Kohji; Fujimuro, Masahiro

    2011-12-02

    Highlights: Black-Right-Pointing-Pointer Constitutive NF-{kappa}B signaling is essential for the survival and growth of PEL cells. Black-Right-Pointing-Pointer NF-{kappa}B signaling is upregulated by the proteasome-dependent degradation of I{kappa}B{alpha}. Black-Right-Pointing-Pointer Proteasome inhibitors suppress NF-{kappa}B signaling and induce apoptosis in PEL cells through stabilization of I{kappa}B{alpha}. Black-Right-Pointing-Pointer Proteasome inhibitors suppress viral replication in PEL cells during lytic KSHV infection. -- Abstract: Primary effusion lymphoma (PEL) is an aggressive neoplasm caused by Kaposi's sarcoma-associated herpesvirus (KSHV). This study provides evidence that proteasomal activity is required for both survival of PEL cells stably harboring the KSHV genome and viral replication of KSHV. We evaluated the cytotoxic effects of proteasome inhibitors on PEL cells. The proteasome inhibitors MG132, lactacystin, and proteasome inhibitor I dramatically inhibited cell proliferation and induced apoptosis of PEL cells through the accumulation of p21 and p27. Furthermore, proteasome inhibitors induced the stabilization of NF-{kappa}B inhibitory molecule (I{kappa}B{alpha}) and suppressed the transcriptional activity of NF-{kappa}B in PEL cells. The NF-{kappa}B specific inhibitor BAY11-7082 also induced apoptosis in PEL cells. The constitutive activation of NF-{kappa}B signaling is essential for the survival and growth of B cell lymphoma cells, including PEL cells. NF-{kappa}B signaling is upregulated by proteasome-dependent degradation of I{kappa}B{alpha}. The suppression of NF-{kappa}B signaling by proteasome inhibitors may contribute to the induction of apoptosis in PEL cells. In addition, proteasome activity is required for KSHV replication in KSHV latently infected PEL cells. MG132 reduced the production of progeny virus from PEL cells at low concentrations, which do not affect PEL cell growth. These findings suggest that proteasome inhibitors may represent a novel strategy for the treatment of KSHV infection and KSHV-associated lymphomas.

  10. Localization of peroxisomal matrix proteins by photobleaching

    SciTech Connect

    Buch, Charlotta; Soedertoerns University, Life Sciences, SE-141 89 Huddinge ; Hunt, Mary C.; Alexson, Stefan E.H.; Hallberg, Einar

    2009-10-16

    The distribution of some enzymes between peroxisomes and cytosol, or a dual localization in both these compartments, can be difficult to reconcile. We have used photobleaching in live cells expressing green fluorescent protein (GFP)-fusion proteins to show that imported bona fide peroxisomal matrix proteins are retained in the peroxisome. The high mobility of the GFP-fusion proteins in the cytosol and absence of peroxisomal escape makes it possible to eliminate the cytosolic fluorescence by photobleaching, to distinguish between exclusively cytosolic proteins and proteins that are also present at low levels in peroxisomes. Using this technique we found that GFP tagged bile acid-CoA:amino acid N-acyltransferase (BAAT) was exclusively localized in the cytosol in HeLa cells. We conclude that the cytosolic localization was due to its carboxyterminal non-consensus peroxisomal targeting signal (-SQL) since mutation of the -SQL to -SKL resulted in BAAT being efficiently imported into peroxisomes.

  11. Combining BET and HDAC inhibitors synergistically induces apoptosis of melanoma and suppresses AKT and YAP signaling

    PubMed Central

    Heinemann, Anja; Cullinane, Carleen; De Paoli-Iseppi, Ricardo; Wilmott, James S.; Gunatilake, Dilini; Madore, Jason; Strbenac, Dario; Yang, Jean Y.; Gowrishankar, Kavitha; Tiffen, Jessamy C.; Prinjha, Rab K.; Smithers, Nicholas; McArthur, Grant A.; Hersey, Peter; Gallagher, Stuart J.

    2015-01-01

    Histone acetylation marks have an important role in controlling gene expression and are removed by histone deacetylases (HDACs). These marks are read by bromodomain and extra-terminal (BET) proteins and novel inhibitiors of these proteins are currently in clinical development. Inhibitors of HDAC and BET proteins have individually been shown to cause apoptosis and reduce growth of melanoma cells. Here we show that combining the HDAC inhibitor LBH589 and BET inhibitor I-BET151 synergistically induce apoptosis of melanoma cells but not of melanocytes. Induction of apoptosis proceeded through the mitochondrial pathway, was caspase dependent and involved upregulation of the BH3 pro-apoptotic protein BIM. Analysis of signal pathways in melanoma cell lines resistant to BRAF inhibitors revealed that treatment with the combination strongly downregulated anti-apoptotic proteins and proteins in the AKT and Hippo/YAP signaling pathways. Xenograft studies showed that the combination of inhibitors was more effective than single drug treatment and confirmed upregulation of BIM and downregulation of XIAP as seen in vitro. These results support the combination of these two classes of epigenetic regulators in treatment of melanoma including those resistant to BRAF inhibitors. PMID:26087189

  12. Combining BET and HDAC inhibitors synergistically induces apoptosis of melanoma and suppresses AKT and YAP signaling.

    PubMed

    Heinemann, Anja; Cullinane, Carleen; De Paoli-Iseppi, Ricardo; Wilmott, James S; Gunatilake, Dilini; Madore, Jason; Strbenac, Dario; Yang, Jean Y; Gowrishankar, Kavitha; Tiffen, Jessamy C; Prinjha, Rab K; Smithers, Nicholas; McArthur, Grant A; Hersey, Peter; Gallagher, Stuart J

    2015-08-28

    Histone acetylation marks have an important role in controlling gene expression and are removed by histone deacetylases (HDACs). These marks are read by bromodomain and extra-terminal (BET) proteins and novel inhibitiors of these proteins are currently in clinical development. Inhibitors of HDAC and BET proteins have individually been shown to cause apoptosis and reduce growth of melanoma cells. Here we show that combining the HDAC inhibitor LBH589 and BET inhibitor I-BET151 synergistically induce apoptosis of melanoma cells but not of melanocytes. Induction of apoptosis proceeded through the mitochondrial pathway, was caspase dependent and involved upregulation of the BH3 pro-apoptotic protein BIM. Analysis of signal pathways in melanoma cell lines resistant to BRAF inhibitors revealed that treatment with the combination strongly downregulated anti-apoptotic proteins and proteins in the AKT and Hippo/YAP signaling pathways. Xenograft studies showed that the combination of inhibitors was more effective than single drug treatment and confirmed upregulation of BIM and downregulation of XIAP as seen in vitro. These results support the combination of these two classes of epigenetic regulators in treatment of melanoma including those resistant to BRAF inhibitors. PMID:26087189

  13. Encapsulation-Induced Stress Helps Saccharomyces cerevisiae Resist Convertible Lignocellulose Derived Inhibitors

    PubMed Central

    Westman, Johan O.; Manikondu, Ramesh Babu; Franzén, Carl Johan; Taherzadeh, Mohammad J.

    2012-01-01

    The ability of macroencapsulated Saccharomyces cerevisiae CBS8066 to withstand readily and not readily in situ convertible lignocellulose-derived inhibitors was investigated in anaerobic batch cultivations. It was shown that encapsulation increased the tolerance against readily convertible furan aldehyde inhibitors and to dilute acid spruce hydrolysate, but not to organic acid inhibitors that cannot be metabolized anaerobically. Gene expression analysis showed that the protective effect arising from the encapsulation is evident also on the transcriptome level, as the expression of the stress-related genes YAP1, ATR1 and FLR1 was induced upon encapsulation. The transcript levels were increased due to encapsulation already in the medium without added inhibitors, indicating that the cells sensed low stress level arising from the encapsulation itself. We present a model, where the stress response is induced by nutrient limitation, that this helps the cells to cope with the increased stress added by a toxic medium, and that superficial cells in the capsules degrade convertible inhibitors, alleviating the inhibition for the cells deeper in the capsule. PMID:23109889

  14. The EMT-activator ZEB1 induces bone metastasis associated genes including BMP-inhibitors

    PubMed Central

    Mock, Kerstin; Preca, Bogdan-Tiberius; Brummer, Tilman; Brabletz, Simone; Stemmler, Marc P.; Brabletz, Thomas

    2015-01-01

    Tumor cell invasion, dissemination and metastasis is triggered by an aberrant activation of epithelial-to-mesenchymal transition (EMT), often mediated by the transcription factor ZEB1. Disseminating tumor cells must acquire specific features that allow them to colonize at different organ sites. Here we identify a set of genes that is highly expressed in breast cancer bone metastasis and activated by ZEB1. This gene set includes various secreted factors, e.g. the BMP-inhibitor FST, that are described to reorganize the bone microenvironment. By inactivating BMP-signaling, BMP-inhibitors are well-known to induce osteolysis in development and disease. We here demonstrate that the expression of ZEB1 and BMP-inhibitors is correlated with bone metastasis, but not with brain or lung metastasis of breast cancer patients. In addition, we show that this correlated expression pattern is causally linked, as ZEB1 induces the expression of the BMP-inhibitors NOG, FST and CHRDL1 both by directly increasing their gene transcription, as well as by indirectly suppressing their reduction via miR-200 family members. Consequently, ZEB1 stimulates BMP-inhibitor mediated osteoclast differentiation. These findings suggest that ZEB1 is not only driving EMT, but also contributes to the formation of osteolytic bone metastases in breast cancer. PMID:25973542

  15. The interaction between Helminthosporium carbonum and maize: Induced resistance and the role of an inhibitor

    SciTech Connect

    Cantone, F.A.

    1989-01-01

    Helminthosporium carbonum race 1 produces large, necrotic lesions on susceptible leaves of maize, whereas race 2 causes small, chlorotic flecks. Resistance to race 1 on susceptible leaves was induced when race 2 was inoculated for at least 10 h prior to a challenge inoculation with the pathogen and was manifest as a decrease in the number of appressoria and reduced penetration by race 1 conidia. Induced resistance was prevented or reversed when HC-toxin was added to challenge race 1 inoculum. The basis for protection appears to be a volatile, inhibitory compound produced by the host. This inhibitor was always associated with treatments that resulted in resistance, whereas no inhibitory activity was detected in diffusates from susceptible reactions. The appearance of inhibitor in diffusates coincided with the appearance of protection on the leaf. In addition to race 2 of H. carbonum, other fungi (H. victoriae, H. turcicum, and Alternaria) also induced production of the inhibitor as well as resistance to race 1. The inhibitor prevented the germination of conidia of all fungi tested. The growth of two phytopathogenic bacteria was also completely inhibited. Incorporation of {sup 3}H-leucine and {sup 14}C-uridine into protein and RNA, respectively, by conidia of H. carbonum was prevented within 15 min of exposure to inhibitor. In addition, respiration of conidia in inhibitor was reduced within 90 min to just 25% of the rate of conidia germinated in water. However, inhibitory activity of the diffusates was readily reversed when conidia were rinsed with water or when organic or amino acids were added to inhibited conidia. The addition of sodium acetate to race 2 and race 1 inocula resulted in lesion enlargement and also nullified inhibitory activity in vitro.

  16. The association of peroxisomes with the developing cell plate in dividing onion root cells depends on actin microfilaments and myosin.

    PubMed

    Collings, David A; Harper, John D I; Vaughn, Kevin C

    2003-12-01

    We have investigated changes in the distribution of peroxisomes through the cell cycle in onion ( Allium cepa L.) root meristem cells with immunofluorescence and electron microscopy, and in leek ( Allium porrum L.) epidermal cells with immunofluorescence and peroxisomal-targeted green fluorescent protein. During interphase and mitosis, peroxisomes distribute randomly throughout the cytoplasm, but beginning late in anaphase, they accumulate at the division plane. Initially, peroxisomes occur within the microtubule phragmoplast in two zones on either side of the developing cell plate. However, as the phragmoplast expands outwards to form an annulus, peroxisomes redistribute into a ring immediately inside the location of the microtubules. Peroxisome aggregation depends on actin microfilaments and myosin. Peroxisomes first accumulate in the division plane prior to the formation of the microtubule phragmoplast, and throughout cytokinesis, always co-localise with microfilaments. Microfilament-disrupting drugs (cytochalasin and latrunculin), and a putative inhibitor of myosin (2,3-butanedione monoxime), inhibit aggregation. We propose that aggregated peroxisomes function in the formation of the cell plate, either by regulating hydrogen peroxide production within the developing cell plate, or by their involvement in recycling of excess membranes from secretory vesicles via the beta-oxidation pathway. Differences in aggregation, a phenomenon which occurs in onion, some other monocots and to a lesser extent in tobacco BY-2 suspension cells, but which is not obvious in the roots of Arabidopsis thaliana (L.) Heynh., may reflect differences within the primary cell walls of these plants. PMID:12937986

  17. Ciprofloxacin-Induced Antibacterial Activity Is Attenuated by Phosphodiesterase Inhibitors

    PubMed Central

    Masadeh, Majed M.; Alzoubi, Karem H.; Khabour, Omar F.; Al-Azzam, Sayer I.

    2014-01-01

    Background Ciprofloxacin is a commonly used antibiotic for urinary tract infection that interacts with bacterial topoisomerases leading to oxidative radicals generation and bacterial cell death. Phosphodiesterase inhibitors (PDEis), on the other hand, are commonly used drugs for the management of erectile dysfunction. The group includes agents such as sildenafil, vardenafil, and tadalafil. Objectives We investigated whether PDEi could interfere with the antibacterial activity of ciprofloxacin. Methods PDEis were tested in several reference bacteria, including Escherichia coli, Staphylococcus aureus, Pseudomonas aeruginosa, Staphylococcus epidermidis, Acinetobacter baumannii, Proteus mirabilis, and Klebsiella pneumoniae utilizing a standard disc diffusion method and measuring both zones of inhibition and MIC. Results Results from both assays indicated that ciprofloxacin demonstrates potent activity against the tested reference bacteria. Additionally, when bacteria were treated with a combination of ciprofloxacin and sildenafil, tadalafil, or vardenafil, the zones of the combination inhibition were significantly reduced, whereas the MIC values were significantly greater than those of ciprofloxacin alone for all tested bacterial strains. In an attempt to examine the mechanism by which PDEis interfere with the action of ciprofloxacin, we utilized the in vitro E coli DNA gyrase cleavage assay. The results showed that PDEi drugs had no effect on ciprofloxacin’s inhibition of E coli gyrase activity. Conclusions Pretreatment of various reference bacterial cells with PDEis largely inhibited the antibacterial activity of ciprofloxacin.

  18. NOVEL ATYPICAL PKC INHIBITORS PREVENT VASCULAR ENDOTHELIAL GROWTH FACTOR-INDUCED BLOOD-RETINAL BARRIER DYSFUNCTION

    PubMed Central

    Titchenell, Paul M.; Lin, Cheng-Mao; Keil, Jason M.; Sundstrom, Jeffrey M.; Smith, Charles D.; Antonetti, David A.

    2013-01-01

    SYNOPSIS Pro-inflammatory cytokines and growth factors such as vascular endothelial growth factor (VEGF) contribute to the loss of the blood-retinal barrier (BRB) and subsequent macular edema in various retinal pathologies. VEGF signaling requires conventional PKC (PKC?) activity; however, PKC? inhibition only partially prevents VEGF-induced endothelial permeability and does not affect pro-inflammatory cytokine-induced permeability suggesting the involvement of alternative signaling pathways. Here, we provide evidence for the involvement of atypical protein kinase C (aPKC) signaling in VEGF-induced endothelial permeability and identify a novel class of inhibitors of aPKC that prevent BRB breakdown in vivo. Genetic and pharmacological manipulations of aPKC isoforms were used to assess their contribution to endothelial permeability in culture. A chemical library was screened using an in vitro kinase assay to identify novel small molecule inhibitors and further medicinal chemistry was performed to delineate a novel pharmacophore. We demonstrate that aPKC isoforms are both sufficient and required for VEGF-induced endothelial permeability. Furthermore, these specific, potent, non-competitive, small molecule inhibitors prevented VEGF-induced tight junction internalization and retinal endothelial permeability in response to VEGF in both primary culture and in rodent retina. These data suggest that aPKC inhibition with 2-amino-4-phenyl-thiophene derivatives may be developed to preserve the BRB in retinal diseases such as diabetic retinopathy or uveitis and the blood-brain barrier (BBB) in the presence of brain tumors. PMID:22721706

  19. Neutral endopeptidase inhibitors potentiate substance P- and capsaicin-induced cough in awake guinea pigs.

    PubMed Central

    Kohrogi, H; Graf, P D; Sekizawa, K; Borson, D B; Nadel, J A

    1988-01-01

    To study the roles of substance P and endogenous neutral endopeptidase in mediating cough, we measured cough responses in awake guinea pigs in response to exogenous substance P and capsaicin aerosols in the presence and absence of the neutral endopeptidase inhibitors leucine-thiorphan and phosphoramidon. Substance P stimulated cough in very low concentrations (10(-17)-10(-16) M). In a second study where the investigator did not know whether substance P or diluent alone was aerosolized, substance P (10(-16) M) caused cough. Leucine-thiorphan (10(-5) M) and phosphoramidon (10(-5) M) potentiated substance P-induced cough; NEP inhibitors also potentiated capsaicin-induced cough significantly. These findings suggest that substance P is a potent stimulator of cough responses, that capsaicin-induced cough is mediated by substance P or another similar neuropeptide, and that cough responses are modulated by endogenous neutral endopeptidase. PMID:2461967

  20. Upregulation of DR5 by proteasome inhibitors potently sensitizes glioma cells to TRAIL-induced apoptosis.

    PubMed

    Hetschko, Holger; Voss, Valerie; Seifert, Volker; Prehn, Jochen H M; Kögel, Donat

    2008-04-01

    This study was undertaken to explore the potential of new therapeutic approaches designed to reactivate cell death pathways in apoptosis-refractory gliomas and to characterize the underlying molecular mechanisms of this reactivation. Here we investigated the sensitivity of a panel of glioma cell lines (U87, U251, U343, U373, MZ-54, and MZ-18) to apoptosis induced by the death receptor ligand tumor necrosis factor-related apoptosis-inducing ligand (TRAIL), TRAIL in combination with gamma irradiation, and TRAIL in combination with proteasome inhibitors (MG132 and epoxomicin). Analysis of these six glioma cell lines revealed drastic differences in their sensitivity to these treatments, with two of the six cell lines revealing no significant induction of cell death in response to TRAIL alone. Interestingly, the proteasome inhibitors MG132 and epoxomicin were capable of potentiating TRAIL-induced apoptosis in TRAIL-sensitive U87 and U251 cells and of reactivating apoptosis in TRAIL-resistant U343 and U373 cells. In contrast, gamma irradiation had no synergistic effects with TRAIL in the two TRAIL-resistant cell lines. RNA interference against death receptor 5 (DR5) revealed that reactivation of TRAIL-induced apoptosis by proteasome inhibitors depended on enhanced transcription and surface expression of DR5. Transient knockdown of the transcription factor GADD153/C/EBP homologous protein and application of the synthetic c-Jun N-terminal kinase inhibitor SP600125 indicated that enhanced DR5 expression occurred independently of GADD153/C/EBP homologous protein, but required activation of the c-Jun N-terminal kinase/c-Jun signaling pathway. Novel therapeutic approaches using TRAIL or agonistic TRAIL receptor antibodies in combination with proteasome inhibitors may represent a promising approach to reactivate apoptosis in therapy-resistant high-grade gliomas. PMID:18341587

  1. Aromatase Inhibitor-Induced Erythrocytosis in a Patient Undergoing Hormonal Treatment for Breast Cancer

    PubMed Central

    Yeruva, Sri Lakshmi Hyndavi; Ogbonna, Onyekachi Henry; Oneal, Patricia

    2015-01-01

    Aromatase inhibitors (AIs) are most commonly used for breast cancer patients with hormone receptor positive disease. Although the side effect profile of aromatase inhibitors is well known, including common side effects like arthralgia, bone pain, arthritis, hot flashes, and more serious problems like osteoporosis, we present a case of an uncommon side effect of these medications. We report the case of a postmenopausal woman on adjuvant hormonal therapy with anastrozole after completing definitive therapy for stage IIIB estrogen receptor-positive breast cancer, who was referred to hematology service for evaluation of persistent erythrocytosis. Primary and known secondary causes of polycythemia were ruled out. On further evaluation, we found that her erythrocytosis began after initiation of anastrozole and resolved after it was discontinued. We discuss the pathophysiology of aromatase inhibitor-induced erythrocytosis and reference of similar cases reported in the literature. PMID:26137331

  2. A novel histone deacetylase inhibitor Chidamide induces apoptosis of human colon cancer cells

    SciTech Connect

    Liu, Lin; Chen, Baoan; Qin, Shukui; Li, Suyi; He, Xiangming; Qiu, Shaomin; Zhao, Wei; Zhao, Hong

    2010-02-05

    Many studies have demonstrated that histone deacetylase (HDAC) inhibitors induce various tumor cells to undergo apoptosis, and such inhibitors have been used in different clinical trials against different human cancers. In this study, we designed and synthesized a novel HDAC inhibitor, Chidamide. We showed that Chidamide was able to increase the acetylation levels of histone H3 and to inhibit the PI3K/Akt and MAPK/Ras signaling pathways, which resulted in arresting colon cancer cells at the G1 phase of the cell cycle and promoting apoptosis. As a result, the proliferation of colon cancer cells was suppressed in vitro. Our data support the potential application of Chidamide as an anticancer agent in treating colon cancer. Future studies are needed to demonstrate its in vivo efficacy.

  3. Localization of a portion of the liver isoform of fatty-acid-binding protein (L-FABP) to peroxisomes

    PubMed Central

    Antonenkov, Vasily D.; Sormunen, Raija T.; Ohlmeier, Steffen; Amery, Leen; Fransen, Marc; Mannaerts, Guy P.; Hiltunen, J. Kalervo

    2005-01-01

    The liver isoform of fatty-acid-binding protein (L-FABP) facilitates the cellular uptake, transport and metabolism of fatty acids and is also involved in the regulation of gene expressions and cell differentiation. Consistent with these functions, L-FABP is predominantly present in the cytoplasm and to a lesser extent in the nucleus; however, a significant portion of this protein has also been detected in fractions containing different organelles. More recent observations, notably on L-FABP-deficient mice, indicated a possible direct involvement of L-FABP in the peroxisomal oxidation of long-chain fatty acids. In order to clarify the links between L-FABP and peroxisomal lipid metabolism, we reinvestigated the subcellular distribution of the protein. Analytical subcellular fractionation by a method preserving the intactness of isolated peroxisomes, two-dimensional gel electrophoresis of peroxisomal matrix proteins combined with MS analysis, and immunoelectron microscopy of liver sections demonstrate the presence of L-FABP in the matrix of peroxisomes as a soluble protein. Peroxisomal L-FABP was highly inducible by clofibrate. The induction of L-FABP was accompanied by a marked increase in the binding capacity of peroxisomal matrix proteins for oleic acid and cis-parinaric acid. The peroxisomal ?-oxidation of palmitoyl-CoA and acyl-CoA thioesterase activity were stimulated by L-FABP, indicating that the protein modulates the function of peroxisomal lipid-metabolizing enzymes. The possible role of intraperoxisomal L-FABP in lipid metabolism is discussed. PMID:16262600

  4. Involvement of caspase family proteases in FPT inhibitor III-induced apoptosis in human ovarian cancer cells.

    PubMed

    Hung, W C; Chuang, L Y

    1998-06-01

    We have previously demonstrated that a new farnesyltransferase inhibitor, FPT inhibitor III, triggers apoptosis in human ovarian cancer cells. Here, we report that induction of apoptotic cell death in PA-1 ovarian cancer cells by FPT inhibitor III was accompanied by the activation of interleukin-1 #-converting enzyme (ICE)-like proteases, which have recently been renamed as caspases. The caspase inhibitor, ZVAD-FMK, which inhibits a number of caspase family proteases, blocked FPT inhibitor III-induced apoptotic cell death in a dose-dependent manner. Additionally, cleavage of poly(ADP-ribose) polymerase, an identified in vivo substrate for caspase family proteases, in FPT inhibitor III-treated cells was confirmed by immunoblotting. Our results suggest that the caspase family proteases are involved in the induction of apoptosis triggered by FPT inhibitor III. PMID:9592196

  5. Thyromimetic mode of action of peroxisome proliferators: activation of 'malic' enzyme gene transcription.

    PubMed Central

    Hertz, R; Nikodem, V; Ben-Ishai, A; Berman, I; Bar-Tana, J

    1996-01-01

    Peroxisome proliferators induce thyroid-hormone-dependent liver activities, e.g. 'malic' enzyme, mitochondrial glycerol-3-phosphate dehydrogenase, glucose-6-phosphate dehydrogenase, S14[Hertz, Aurbach, Hashimoto and Bar-Tana (1991) Biochem. J. 274, 745-751]. Here we report that the thyromimetic effect of peroxisome proliferators with respect to 'malic' enzyme result from transcriptional activation of the 'malic' enzyme gene, mediated by binding of the peroxisome proliferator activated receptor (PPAR alpha)/retinoid X receptor (RXR alpha) heterodimer to a 5'-flanking enhancer of the 'malic' enzyme promoter. The enhancer involved is distinct from the thyroid hormone response element of the 'malic' enzyme promoter and is partly homologous with that which mediates transcriptional activation of peroxisomal acyl-CoA oxidase by peroxisome proliferators. Hence transcriptional activation of thyroid-hormone-dependent liver genes by xenobiotic or endogenous amphipathic carboxylates collectively defined as peroxisome proliferators is mediated by a transduction pathway similar to that involved in transcriptional activation of peroxisomal beta-oxidative genes and distinct from that which mediates thyroid hormone action. PMID:8870674

  6. Effect of inhibitors of eicosanoid metabolism in murine collagen-induced arthritis.

    PubMed

    Griswold, D E; Hillegass, L M; Meunier, P C; DiMartino, M J; Hanna, N

    1988-11-01

    The dual inhibitors of arachidonic acid metabolism, Smith Kline & French (SK&F) 86002, SK&F 104351, and phenidone; the corticosteroid, dexamethasone; and the selective cyclooxygenase inhibitors, ibuprofen, indomethacin, naproxen, and piroxicam were evaluated for their antiarthritic potency in the murine, collagen-induced arthritis model. The ability of these compounds to alter the severity of arthritic lesions and to reduce serum levels of the acute-phase reactant, serum amyloid P component (SAP) were monitored. Serum concentrations of SAP were found to correlate strongly (r = 0.985) with disease severity at day 35 postimmunization. Treatment with SK&F 86002, SK&F 104351, phenidone, or dexamethasone significantly reduced disease severity, as judged by clinical score (55%, 72%, 41%, and 45% inhibition, respectively) and SAP levels (62%, 94%, 52%, and 94% inhibition, respectively) in arthritic mice. This profile of activity was not shared by the selective cyclooxygenase inhibitors, which did not uniformly inhibit disease activity by both parameters. The results suggest that dual inhibitors of 5-lipoxygenase and cyclooxygenase may prove more effective than selective cyclooxygenase inhibitors as anti-arthritic agents. PMID:3142488

  7. Peroxisomal Disorders: A Review on Cerebellar Pathologies.

    PubMed

    De Munter, Stephanie; Verheijden, Simon; Régal, Luc; Baes, Myriam

    2015-11-01

    Peroxisomes are organelles with diverse metabolic tasks including essential roles in lipid metabolism. They are of utmost importance for the normal functioning of the nervous system as most peroxisomal disorders are accompanied with neurological symptoms. Remarkably, the cerebellum exquisitely depends on intact peroxisomal function both during development and adulthood. In this review, we cover all aspects of cerebellar pathology that were reported in peroxisome biogenesis disorders and in diseases caused by dysfunction of the peroxisomal ?-oxidation, ?-oxidation or ether lipid synthesis pathways. We also discuss the phenotypes of mouse models in which cerebellar pathologies were recapitulated and search for connections with the metabolic abnormalities. It becomes increasingly clear that besides the most severe forms of peroxisome dysfunction that are associated with developmental cerebellar defects, milder impairments can give rise to ataxia later in life. PMID:26201894

  8. Pseudoceramide stimulates peroxisome proliferator-activated receptor-? expression in a murine model of atopic dermatitis: molecular basis underlying the anti-inflammatory effect and the preventive effect against steroid-induced barrier impairment.

    PubMed

    Lee, Sang Eun; Jung, Min Kyung; Oh, Seung Joon; Jeong, Se Kyoo; Lee, Seung Hun

    2015-11-01

    Topical pseudoceramides are successfully used in skin barrier repair therapy for atopic dermatitis (AD) and demonstrated to reduce the adverse effects of topical glucocorticoids (GC). However, the molecular mechanisms involved are not fully understood. We investigated whether PC-9S (myristoyl/palmitoyloxostearamide/arachamide MEA, Neopharm, Daejeon, Korea), one of the synthetic pseudoceramides, could stimulate peroxisome proliferator-activated receptor (PPAR)? expression in a hapten [oxazolone (oxa)]-induced AD murine model (oxa-AD mice) and subsequently improved permeability barrier, reduced inflammation, and increased antimicrobial peptides (AMPs) expression. Normal hairless mice and oxa-AD mice were topically treated twice daily with either PC-9S-containing physiologic lipid mixture (PLM), vehicle (PLM), or PPAR? agonist for 4 days. Topical PC-9S significantly increased PPAR? expression in mouse epidermis in vivo and in oxa-AD mice skin comparable with PPAR? agonist. Topical PC-9S-containing PLM significantly reduced basal trans-epidermal water loss (TEWL), surface pH, and mast cell infiltrates and prevented the decline of AMPs expression in oxa-AD mice, which were abrogated by PPAR? antagonist. Then, oxa-AD mice were treated with super-potent topical GC twice daily for 4 days with or without PC-9S co-applications. Co-treatment with PC-9S-containing PLM suppressed GC-induced increase in basal TEWL, epidermal thinning, reduced loricrin expression, and impaired barrier recovery and these effects were attenuated by PPAR? antagonist. Collectively, our findings suggest that pseudoceramide PC-9S-induced stimulation of PPAR? expression provides a new mechanism by which pseudoceramides show anti-inflammatory property, improve the permeability and antimicrobial barrier function, and prevent the negative effects of topical GC. PMID:26121942

  9. Inhibitors of hypoxia-inducible factor 1 block breast cancer metastatic niche formation and lung metastasis.

    PubMed

    Wong, Carmen Chak-Lui; Zhang, Huafeng; Gilkes, Daniele M; Chen, Jasper; Wei, Hong; Chaturvedi, Pallavi; Hubbi, Maimon E; Semenza, Gregg L

    2012-07-01

    Intratumoral hypoxia, a frequent finding in metastatic cancer, results in the activation of hypoxia-inducible factors (HIFs). HIFs are implicated in many steps of breast cancer metastasis, including metastatic niche formation through increased expression of lysyl oxidase (LOX) and lysyl oxidase-like (LOXL) proteins, enzymes that remodel collagen at the metastatic site and recruit bone marrow-derived cells (BMDCs) to the metastatic niche. We investigated the effect of two chemically and mechanistically distinct HIF inhibitors, digoxin and acriflavine, on breast cancer metastatic niche formation. Both drugs blocked the hypoxia-induced expression of LOX and LOXL proteins, collagen cross-linking, CD11b? BMDC recruitment, and lung metastasis in an orthotopic breast cancer model. Patients with HIF-1 ?-overexpressing breast cancers are at increased risk of metastasis and mortality and our results suggest that such patients may benefit from aggressive therapy that includes a HIF inhibitor. PMID:22231744

  10. Inhibition of leukotriene-induced contraction of guinea pig trachea by 5-lipoxygenase inhibitors.

    PubMed

    Schwalm, S F; Lewis, A J; Hand, J M

    1987-01-01

    The effects of the 5-lipoxygenase inhibitors nordihydroguiaretic acid (NDGA), 5, 8, 11, 14-eicosatetraynoic acid (ETYA), 1-phenyl-3-pyrazolidone (phenidone) and BW-755c, on the contractile response to LTC4 or LTD4 were examined on the isolated guinea pig trachea. Responses to either LTC4 or LTD4 were obtained on indomethacin treated tissues, in the presence of either L-serine-borate complex or L-cysteine, respectively, to inhibit metabolic conversion of the leukotrienes. NDGA (30 microM) and ETYA (100 microM) produced a selective competitive antagonism of LTD4-induced contractions, while phenidone antagonized both LTC4- and LTD4-induced responses in a non-competitive manner. In contrast, BW-755c (30 microM) did not significantly antagonize LTC4 or LTD4 concentration-response curves. The results suggest that leukotriene antagonism may be produced by large concentrations of some 5-lipoxygenase inhibitors. PMID:3108959

  11. The STAT3 inhibitor WP1066 reverses the resistance of chronic lymphocytic leukemia cells to histone deacetylase inhibitors induced by interleukin-6.

    PubMed

    Lu, Kang; Fang, Xiao-sheng; Feng, Li-li; Jiang, Yu-jie; Zhou, Xiang-xiang; Liu, Xin; Li, Pei-pei; Chen, Na; Ding, Mei; Wang, Na; Zhang, Jie; Wang, Xin

    2015-04-10

    Interleukin-6 (IL-6) is a pleiotropic cytokine produced by a variety of cell types, including fibroblasts, endothelial cells, lymphocytes, and bone marrow stromal cells (BMSCs). Levels of IL-6 are increased in serum of CLL patients and correlated with adverse clinical features and short survival. In our study, we observed that IL-6 induced the resistance of CLL cells to pan-histone deacetylase (HDAC) inhibitors vorinostat (SAHA) and panobinostat (LBH589). Furthermore, low concentrations of SAHA and LBH589 enhanced the activation of the signal transducer and activator of transcription 3 (STAT3) signaling pathway induced by IL-6 in CLL cells. All of these effects were blocked by the STAT3-selective inhibitor, WP1066. Meanwhile, WP1066 decreased the expressions of Mcl-1 and Bcl-xL protein induced by IL-6 with or without low concentrations of HDAC inhibitors. Co-culture of CLL cells with BMSCs could also facilitate the activation of STAT3 and protected CLL cells from apoptosis when treated with HDAC inhibitors, and this cytoprotection was reversed by WP1066. The present study indicated that IL-6 or co-culture with BMSCs prevented HDAC inhibitor-induced apoptosis of CLL cells. This prevention was mediated by activation of the STAT3 signaling pathway. Moreover, WP1066 reversed the resistance of CLL cells to SAHA and LBH589 induced by either IL-6 or co-culture with BMSCs. Our findings suggest that targeting the STAT3 pathway may be a novel way to improve the efficacy of the HDAC inhibitor in CLL patients by overcoming antiapoptotic signaling of the microenvironment. PMID:25636517

  12. Bifunctional Peptide Inhibitors Suppress Interleukin-6 Proliferation and Ameliorates Murine Collagen-Induced Arthritis

    E-print Network

    Buyuktimkin, Barlas; Kiptoo, Paul Kipkemboi; Siahaan, Teruna J.

    2014-11-01

    Bifunctional Peptide Inhibitors Suppress Interleukin-6 Proliferation and Ameliorates Murine Collagen-Induced Arthritis Barlas Büyüktimkin, Paul Kiptoo and Teruna J Siahaan * Department of Pharmaceutical Chemistry, The University of Kansas, Lawrence, KS 66047... permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Abstract The objective of this study is to evaluate the efficacy and potential mechanism of action of type-II collagen...

  13. The Calpain Inhibitor MDL28170 Induces the Expression of Apoptotic Markers in Leishmania amazonensis Promastigotes

    PubMed Central

    Marinho, Fernanda A.; Gonçalves, Keyla C. S.; Oliveira, Simone S. C.; Gonçalves, Diego S.; Matteoli, Filipe P.; Seabra, Sergio H.; Oliveira, Ana Carolina S.; Bellio, Maria; Oliveira, Selma S.; Souto-Padrón, Thaïs; d'Avila-Levy, Claudia M.; Santos, André L. S.; Branquinha, Marta H.

    2014-01-01

    Background Human cutaneous leishmaniasis is caused by distinct species, including Leishmania amazonensis. Treatment of cutaneous leishmaniasis is far from satisfactory due to increases in drug resistance and relapses, and toxicity of compounds to the host. As a consequence for this situation, the development of new leishmanicidal drugs and the search of new targets in the parasite biology are important goals. Methodology/Principal Findings In this study, we investigated the mechanism of death pathway induced by the calpain inhibitor MDL28170 on Leishmania amazonensis promastigote forms. The combined use of different techniques was applied to contemplate this goal. MDL28170 treatment with IC50 (15 µM) and two times the IC50 doses induced loss of parasite viability, as verified by resazurin assay, as well as depolarization of the mitochondrial membrane, which was quantified by JC-1 staining. Scanning and transmission electron microscopic images revealed drastic alterations on the parasite morphology, some of them resembling apoptotic-like death, including cell shrinking, surface membrane blebs and altered chromatin condensation pattern. The lipid rearrangement of the plasma membrane was detected by Annexin-V labeling. The inhibitor also induced a significant increase in the proportion of cells in the sub-G0/G1 phase, as quantified by propidium iodide staining, as well as genomic DNA fragmentation, detected by TUNEL assay. In cells treated with MDL28170 at two times the IC50 dose, it was also possible to observe an oligonucleossomal DNA fragmentation by agarose gel electrophoresis. Conclusions/Significance The data presented in the current study suggest that MDL28170 induces apoptotic marker expression in promastigotes of L. amazonensis. Altogether, the results described in the present work not only provide a rationale for further exploration of the mechanism of action of calpain inhibitors against trypanosomatids, but may also widen the investigation of the potential clinical utility of calpain inhibitors in the chemotherapy of leishmaniases. PMID:24498160

  14. Proteins and enzymes of the peroxisomal membrane in mammals.

    PubMed

    Causeret, C; Bentejac, M; Bugaut, M

    1993-01-01

    Proteins of the peroxisomal membrane can be schematically divided into two groups, one being made up of more or less characterized proteins with generally unknown functions and the other consisting of enzyme activities of which the corresponding proteins have not been characterized. In the present report, these proteins and enzymes are described with the addition of unpublished results regarding their induction by peroxisome proliferators at the post-transcriptional level. Integral membrane proteins (IMPs) can be isolated using an alkaline solution of sodium carbonate. A dozen of preponderant IMPs can be seen on sodium dodecyl sulfate polyacrylamide gel electrophoresis, and the major band corresponds to a 70 kDa IMP, of which the corresponding rat cDNA is known. Some IMPs have been characterized by immunoblot analysis. Recently, a cDNA has been cloned for a peroxisome assembly factor (35 kDa IMP). Functions have also been proposed for some IMPs but are not yet firmly settled. Some IMPs (450/520, 70 and 26 kDa) are strongly induced by peroxisome proliferators. Our results extend to cipro- and fenofibrate the observation that the 70 kDa IMP mRNA level is strongly increased in di(2-ethylhexyl)phtalate-treated rats. All the enzyme activities associated with the peroxisomal membrane are involved in lipid metabolism: activation of substrates (fatty acids), ether lipid biosynthesis, and formation of precursors (fatty alcohols). It is believed that the same long-chain acyl-CoA synthetase occurs in the peroxisome as well as in the outer mitochondrial membrane and the endoplasmic reticulum. However, two highly homologous but different cDNAs encoding rat liver and brain long-chain acyl-CoA synthetases have been isolated recently. Evidence has been accumulated for a distinct synthetase that specifically activates very-long chain fatty acids. The first two steps of ether lipid biosynthesis require dihydroxyacetone-phosphate (DHAP) acyltransferase and alkyl-DHAP synthetase, the active sites of which are located on the inner surface of the membrane. In contrast, the catalytic site of the acyl/alkyl-DHAP reductase, which generates sn-1-alkyl-glycerol-3-phosphate, is located on the outer surface. Long-chain fatty alcohols, which are obligate precursors of ether lipids and wax esters, are biosynthetized by the reduction of the corresponding acyl-CoAs via the action of an acyl-CoA reductase. Peroxisome proliferators do not appear to stimulate these enzyme activities specifically. However, we report that feno- and ciprofibrate treatments increase six-fold the palmitoyl-CoA synthetase mRNA level in the rat liver. PMID:8518748

  15. Interferon-? and cyclooxygenase-2 inhibitor cooperatively mediates TRAIL-induced apoptosis in hepatocellular carcinoma.

    PubMed

    Zuo, Chaohui; Qiu, Xiaoxin; Liu, Nianli; Yang, Darong; Xia, Man; Liu, Jingshi; Wang, Xiaohong; Zhu, Haizhen; Xie, Hailong; Dan, Hanguo; Li, Qinglong; Wu, Qunfeng; Burns, Michael; Liu, Chen

    2015-05-01

    Hepatocellular carcinoma (HCC) is a leading cause of cancer-related mortality worldwide. Interferon-alpha (IFN-?) has recently been recognized to harbor therapeutic potential in the prevention and treatment of HCC, but it remains controversial as to whether IFN-? exerts direct cytotoxicity against HCC. Cyclooxygenase-2 (COX-2) is overexpressed in HCC and is considered to play a role in hepatocarcinogenesis. Therefore, we aimed to elucidate the combined effect of a COX-2 inhibitor, celecoxib, and IFN-? on in vitro growth suppression of HCC using the hepatoma cell line HLCZ01 and the in vivo nude mouse xenotransplantation model using HLCZ01 cells. Treatment with celecoxib and IFN-? synergistically inhibited cell proliferation in a dose- and time-dependent manner. Apoptosis was identified by 4?,6-diamidino-2-phenylindole dihydrochloride and fluorescent staining. IFN-? upregulated the expression of TRAIL, while celecoxib increased the expression of TRAIL receptors. The combined regimen with celecoxib and IFN-? reduced the growth of xenotransplanted HCCs in nude mice. The regulation of IFN-?- and COX-2 inhibitor-induced cell death is impaired in a subset of TRAIL-resistant cells. The molecular mechanisms of HCC cells resistant to TRAIL-induced apoptosis were explored using molecular biological and immunological methods. Interferon-? and the COX-2 inhibitor celecoxib synergistically increased TRAIL-induced apoptosis in hepatocellular carcinoma. These data suggest that IFN-? and celecoxib may offer a novel role with important implications in designing new therapeutics for TRAIL-resistant tumors. PMID:25724899

  16. ACE inhibitor-induced intestinal angio-oedema: rare adverse effect of a common drug.

    PubMed

    Shahani, Lokesh

    2013-01-01

    ACE inhibitors are the leading cause of drug-induced angio-oedema in the USA. ACE inhibitor-induced intestinal angio-oedema, a much rarer complication of this medication, has been reported. The author reports a patient presenting with a 1-day history of severe abdominal pain. The patient was started on lisinopril 2 days prior to this presentation. Computer axial tomography (CAT) scan of the abdomen demonstrated extensive and marked thickening, and oedema involving the duodenum and proximal jejunum associated with significant mesenteric oedema. Concerns for visceral angio-oedema and a possible association with lisinopril according to the Naranjo algorithm were raised. Lisinopril was discontinued and the patient was treated with antihistamines. The patient improved clinically in the next 24 h and discharged home with education and documentation of this serious allergy. ACE inhibitor-induced visceral angio-oedema is under-reported and most often missed resulting in waste of hospital resources towards working up this clinical diagnosis. PMID:23878294

  17. Interplant communication: airborne methyl jasmonate induces synthesis of proteinase inhibitors in plant leaves.

    PubMed Central

    Farmer, E E; Ryan, C A

    1990-01-01

    Inducible defensive responses in plants are known to be activated locally and systemically by signaling molecules that are produced at sites of pathogen or insect attacks, but only one chemical signal, ethylene, is known to travel through the atmosphere to activate plant defensive genes. Methyl jasmonate, a common plant secondary compound, when applied to surfaces of tomato plants, induces the synthesis of defensive proteinase inhibitor proteins in the treated plants and in nearby plants as well. The presence of methyl jasmonate in the atmosphere of chambers containing plants from three species of two families, Solanaceae and Fabaceae, results in the accumulation of proteinase inhibitors in leaves of all three species. When sagebrush, Artemisia tridentata, a plant shown to possess methyl jasmonate in leaf surface structures, is incubated in chambers with tomato plants, proteinase inhibitor accumulation is induced in the tomato leaves, demonstrating that interplant communication can occur from leaves of one species of plant to leaves of another species to activate the expression of defensive genes. PMID:11607107

  18. Depressor effect of chymase inhibitor in mice with high salt-induced moderate hypertension.

    PubMed

    Devarajan, Sankar; Yahiro, Eiji; Uehara, Yoshinari; Habe, Shigehisa; Nishiyama, Akira; Miura, Shin-Ichiro; Saku, Keijiro; Urata, Hidenori

    2015-12-01

    The aim of the present study was to determine whether long-term high salt intake in the drinking water induces hypertension in wild-type (WT) mice and whether a chymase inhibitor or other antihypertensive drugs could reverse the increase of blood pressure. Eight-week-old male WT mice were supplied with drinking water containing 2% salt for 12 wk (high-salt group) or high-salt drinking water plus an oral chymase inhibitor (TPC-806) at four different doses (25, 50, 75, or 100 mg/kg), captopril (75 mg/kg), losartan (100 mg/kg), hydrochlorothiazide (3 mg/kg), eplerenone (200 mg/kg), or amlodipine (6 mg/kg). Control groups were given normal water with or without the chymase inhibitor. Blood pressure and heart rate gradually showed a significant increase in the high-salt group, whereas a dose-dependent depressor effect of the chymase inhibitor was observed. There was also partial improvement of hypertension in the losartan- and eplerenone-treated groups but not in the captopril-, hydrochlorothiazide-, and amlodipine-treated groups. A high salt load significantly increased chymase-dependent ANG II-forming activity in the alimentary tract. In addition, the relative contribution of chymase to ANG II formation, but not actual average activity, showed a significant increase in skin and skeletal muscle, whereas angiotensin-converting enzyme-dependent ANG II-forming activity and its relative contribution were reduced by high salt intake. Plasma and urinary renin-angiotensin system components were significantly increased in the high-salt group but were significantly suppressed in the chymase inhibitor-treated group. In conclusion, 2% salt water drinking for 12 wk caused moderate hypertension and activated the renin-angiotensin system in WT mice. A chymase inhibitor suppressed both the elevation of blood pressure and heart rate, indicating a definite involvement of chymase in salt-sensitive hypertension. PMID:26432844

  19. Effects of inhibitors on 1-methyladenine induced maturation of starfish oocytes

    NASA Astrophysics Data System (ADS)

    Lee, Harold H.; Xu, Quanhan

    1986-12-01

    1-methladenine (1-MA) induces starfish oocytes maturation via surface reaction followed by the appearance of a cytoplasmic maturation factor which in turn induces germinal vesicle breakdown (GVBD) to resume meiosis. Cellular mechanisms involved in GVBD were investigated by microinjection of metabolic inhibitors. Colchicine (Co) inhibited maturation, cytochalasin-B (CB) delayed GVBD and actinomycin-D-(Act-D) and puromycin (Pu) had no effect. It appears that the microtubule and the microfilament systems are associated with the nuclear membrane dissolution during the process of oocyte maturation of starfish.

  20. Discovery of Indenopyrazoles as a New Class of Hypoxia Inducible Factor (HIF)-1 Inhibitors

    PubMed Central

    2013-01-01

    The indenopyrazole framework was investigated as a new class of HIF-1? inhibitors. Indenopyrazole 2l was found to most strongly inhibit the hypoxia-induced HIF-1? transcriptional activity (IC50 = 0.014 ?M) among all of the known compounds having relatively simple structures, unlike manassantins. Indenopyrazole 2l suppressed HIF-1? transcriptional activity without affecting both HIF-1? protein accumulation and HIF-1?/HIF-1? heterodimerization in nuclei under the hypoxic conditions, suggesting that 2l probably affected the transcriptional pathway induced by the HIF-1?/HIF-1? heterodimer. PMID:24900662

  1. Further studies of auxin and ACC induced feminization in the cucumber plant using ethylene inhibitors

    NASA Technical Reports Server (NTRS)

    Takahashi, H.; Jaffe, M. J.

    1984-01-01

    The present study was designed to establish the role of an essential hormone controlling sex expression in cucumber. A potent anti-ethylene agent, AgNO3, completely inhibited pistillate flower formation caused by IAA, ACC or ethephon. Inhibitors of ethylene biosynthesis, AVG and CoCl2 also suppressed feminization due to exogenous IAA or ACC. Though AVG also suppressed ethephon-induced feminization, this may be due to the second effect of AVG rather than the effect on ACC biosynthesis. These results confirm that ethylene is a major factor regulating feminization and that exogenous auxin induces pistillate flower formation through its stimulation of ethylene production, rather than ACC production.

  2. Inhibition of Interleukin-1?-Induced Group IIA Secretory Phospholipase A2 Expression by Peroxisome Proliferator-Activated Receptors (PPARs) in Rat Vascular Smooth Muscle Cells: Cooperation between PPAR? and the Proto-Oncogene BCL-6?

    PubMed Central

    Ravaux, Lucas; Denoyelle, Chantal; Monne, Claire; Limon, Isabelle; Raymondjean, Michel; El Hadri, Khadija

    2007-01-01

    The inflammation that occurs during atherosclerosis is characterized by the release of large amounts of group IIA secretory phospholipase A2 (sPLA2-IIA). This study was designed to define the function of the three peroxisome proliferator-activated receptors (PPARs) on sPLA2 expression in vascular smooth muscle cells (VSMCs). We found that PPAR ligands decreased sPLA2-IIA activity and inhibited mRNA accumulation under inflammatory conditions. Furthermore, interleukin-1?-induced sPLA2-IIA promoter activity was inhibited by the three PPAR ligands and in a similar way when cells were cotransfected with PPAR?, PPAR?, or PPAR?, plus retinoid X receptor ? (RXR?). Our study revealed that the regulation of sPLA2-IIA gene transcription by PPAR?/RXR and PPAR?/RXR heterodimers requires an interaction with a PPAR response element (PPRE) of the sPLA2-IIA promoter. In contrast, PPAR? operates through a PPRE-independent mechanism. In addition, we demonstrated that VSMCs expressed the transcriptional repressor BCL-6. Overexpression of BCL-6 markedly reduced sPLA2-IIA promoter activity in VSMCs, while a dominant negative form of BCL-6 abrogated sPLA2 repression by PPAR?. The PPAR? agonist induced a BCL-6 binding to the sPLA2 promoter in VSMCs under inflammatory conditions. The knockdown of BCL-6 by short interfering RNA abolished the inhibitory effect of the PPAR? ligand on sPLA2 activity and prostaglandin E2 release. Thus, the inhibition of sPLA2-IIA activity by PPAR? agonists may provide a promising approach to impacting the initiation and progression of atherosclerosis. PMID:17908795

  3. Proton pump inhibitor alone vs proton pump inhibitor plus mucosal protective agents for endoscopic submucosal dissection-induced ulcer: a systematic review and meta-analysis.

    PubMed

    Nishizawa, Toshihiro; Suzuki, Hidekazu; Kanai, Takanori; Yahagi, Naohisa

    2015-03-01

    Mucosal protective agents may improve healing of patients with endoscopic submucosal dissection-induced ulcers. The present study systematically evaluated published clinical trials to determine whether combined therapeutic use of mucosal protective agents and proton pump inhibitors can improve the outcome of patients with endoscopic submucosal dissection-induced ulcers compared to treatment with proton pump inhibitors alone. PubMed, the Cochrane Library, and the Igaku-Chuo-Zasshi database were searched to identify eligible randomized trials for systematic review. We identified 11 randomized trials for inclusion in our study (1,160 patients). Pooled endoscopic submucosal dissection-induced ulcer healing rates were 45.8% and 34.4% for patients with or without mucosal protective agents, respectively. The odds ratio was 2.28 (95% confidence interval, 1.57-3.31) with no significant study heterogeneity. In conclusion, the systematic review and meta-analysis showed that the combined therapeutic use of proton pump inhibitors and mucosal protective agents improved healing rates of endoscopic submucosal dissection-induced ulcers compared to treatment with proton pump inhibitor monotherapy. PMID:25759512

  4. Proton pump inhibitor alone vs proton pump inhibitor plus mucosal protective agents for endoscopic submucosal dissection-induced ulcer: a systematic review and meta-analysis

    PubMed Central

    Nishizawa, Toshihiro; Suzuki, Hidekazu; Kanai, Takanori; Yahagi, Naohisa

    2015-01-01

    Mucosal protective agents may improve healing of patients with endoscopic submucosal dissection-induced ulcers. The present study systematically evaluated published clinical trials to determine whether combined therapeutic use of mucosal protective agents and proton pump inhibitors can improve the outcome of patients with endoscopic submucosal dissection-induced ulcers compared to treatment with proton pump inhibitors alone. PubMed, the Cochrane Library, and the Igaku-Chuo-Zasshi database were searched to identify eligible randomized trials for systematic review. We identified 11 randomized trials for inclusion in our study (1,160 patients). Pooled endoscopic submucosal dissection-induced ulcer healing rates were 45.8% and 34.4% for patients with or without mucosal protective agents, respectively. The odds ratio was 2.28 (95% confidence interval, 1.57–3.31) with no significant study heterogeneity. In conclusion, the systematic review and meta-analysis showed that the combined therapeutic use of proton pump inhibitors and mucosal protective agents improved healing rates of endoscopic submucosal dissection-induced ulcers compared to treatment with proton pump inhibitor monotherapy. PMID:25759512

  5. NPI-0052, a novel proteasome inhibitor, induces caspase-8 and ROS-dependent apoptosis alone and in combination with HDAC inhibitors in leukemia cells.

    PubMed

    Miller, Claudia P; Ban, Kechen; Dujka, Melanie E; McConkey, David J; Munsell, Mark; Palladino, Michael; Chandra, Joya

    2007-07-01

    The proteasome has been successfully targeted for the treatment of multiple myeloma and mantle cell lymphoma; however, in other hematologic malignancies, bortezomib has been less effective as a single agent. Here, we describe effects of NPI-0052, a novel proteasome inhibitor, in leukemia model systems. In cell lines, NPI-0052 inhibits all 3 proteolytic activities associated with the proteasome: chymotrypsin-, trypsin-, and caspase-like. NPI-0052 also induces DNA fragmentation in leukemia lines and in mononuclear cells from a Ph + acute lymphoblastic leukemia (ALL) patient. Caspase-3 activation by NPI-0052 was seen in wild-type Jurkat cells, but was significantly lessened in Fas-associated death domain (FADD)-deficient or caspase-8-deficient counterparts. NPI-0052-induced apoptosis was further probed using caspase-8 inhibitors, which were more protective than caspase-9 inhibitors. N-acetyl cysteine (NAC) also conferred protection against NPI-0052-induced apoptosis, indicating a role for oxidative stress by NPI-0052. In support of the drug's in vitro activities, biweekly treatment with NPI-0052 lessened total white blood cell (WBC) burden over 35 days in leukemic mice. Interestingly, combining NPI-0052 with either MS-275 or valproic acid (VPA) induced greater levels of cell death than the combination of bortezomib with these histone deacetylase inhibitors (HDACi). These effects of NPI-0052, alone and in combination with HDACi, warrant further testing to determine the compound's clinical efficacy in leukemia. PMID:17356134

  6. Cell Death Inducing Microbial Protein Phosphatase Inhibitors—Mechanisms of Action

    PubMed Central

    Kleppe, Rune; Herfindal, Lars; Døskeland, Stein Ove

    2015-01-01

    Okadaic acid (OA) and microcystin (MC) as well as several other microbial toxins like nodularin and calyculinA are known as tumor promoters as well as inducers of apoptotic cell death. Their intracellular targets are the major serine/threonine protein phosphatases. This review summarizes mechanisms believed to be responsible for the death induction and tumor promotion with focus on the interdependent production of reactive oxygen species (ROS) and activation of Ca2+/calmodulin kinase II (CaM-KII). New data are presented using inhibitors of specific ROS producing enzymes to curb nodularin/MC-induced liver cell (hepatocyte) death. They indicate that enzymes of the arachidonic acid pathway, notably phospholipase A2, 5-lipoxygenase, and cyclooxygenases, may be required for nodularin/MC-induced (and presumably OA-induced) cell death, suggesting new ways to overcome at least some aspects of OA and MC toxicity. PMID:26506362

  7. Novel peroxisome clustering mutants and peroxisome biogenesis mutants of Saccharomyces cerevisiae

    PubMed Central

    1993-01-01

    The goal of this research is to identify and characterize the protein machinery that functions in the intracellular translocation and assembly of peroxisomal proteins in Saccharomyces cerevisiae. Several genes encoding proteins that are essential for this process have been identified previously by Kunau and collaborators, but the mutant collection was incomplete. We have devised a positive selection procedure that identifies new mutants lacking peroxisomes or peroxisomal function. Immunofluorescence procedures for yeast were simplified so that these mutants could be rapidly and efficiently screened for those in which peroxisome biogenesis is impaired. With these tools, we have identified four complementation groups of peroxisome biogenesis mutants, and one group that appears to express reduced amounts of peroxisomal proteins. Two of our mutants lack recognizable peroxisomes, although they might contain peroxisomal membrane ghosts like those found in Zellweger syndrome. Two are selectively defective in packaging peroxisomal proteins and moreover show striking intracellular clustering of the peroxisomes. The distribution of mutants among complementation groups implies that the collection of peroxisome biogenesis mutants is still incomplete. With the procedures described, it should prove straightforward to isolate mutants from additional complementation groups. PMID:7902359

  8. Characterization of a novel component of the peroxisomal protein import apparatus using fluorescent peroxisomal proteins.

    PubMed Central

    Kalish, J E; Keller, G A; Morrell, J C; Mihalik, S J; Smith, B; Cregg, J M; Gould, S J

    1996-01-01

    Fluorescent peroxisomal probes were developed by fusing green fluorescent protein (GFP) to the matrix peroxisomal targeting signals PTS1 and PTS2, as well as to an integral peroxisomal membrane protein (IPMP). These proteins were used to identify and characterize novel peroxisome assembly (pas) mutants in the yeast Pichia pastoris. Mutant cells lacking the PAS10 gene mislocalized both PTS1-GFP and PTS2-GFP to the cytoplasm but did incorporate IPMP-GFP into peroxisome membranes. Similar distributions were observed for endogenous peroxisomal matrix and membrane proteins. While peroxisomes from translocation-competent pas mutants sediment in sucrose gradients at the density of normal peroxisomes, >98% of peroxisomes from pas10 cells migrated to a much lower density and had an extremely low ratio of matrix:membrane protein. These data indicate that Pas10p plays an important role in protein translocation across the peroxisome membrane. Consistent with this hypothesis, we find that Pas10p is an integral protein of the peroxisome membrane. In addition, Pas10p contains a cytoplasmically-oriented C3HC4 zinc binding domain that is essential for its biological activity. Images PMID:8670828

  9. Nifedipine inhibits advanced glycation end products (AGEs) and their receptor (RAGE) interaction-mediated proximal tubular cell injury via peroxisome proliferator-activated receptor-gamma activation

    SciTech Connect

    Matsui, Takanori; Yamagishi, Sho-ichi; Takeuchi, Masayoshi; Ueda, Seiji; Fukami, Kei; Okuda, Seiya

    2010-07-23

    Research highlights: {yields} Nifedipine inhibited the AGE-induced up-regulation of RAGE mRNA levels in tubular cells, which was prevented by GW9662, an inhibitor of peroxisome proliferator-activated receptor-{gamma}. {yields} GW9662 treatment alone increased RAGE mRNA levels in tubular cells. {yields} Nifedipine inhibited the AGE-induced reactive oxygen species generation, NF-{kappa}B activation and increases in intercellular adhesion molecule-1 and transforming growth factor-{beta} gene expression in tubular cells, all of which were blocked by GW9662. -- Abstract: There is a growing body of evidence that advanced glycation end products (AGEs) and their receptor (RAGE) interaction evokes oxidative stress generation and subsequently elicits inflammatory and fibrogenic reactions, thereby contributing to the development and progression of diabetic nephropathy. We have previously found that nifedipine, a calcium-channel blocker (CCB), inhibits the AGE-induced mesangial cell damage in vitro. However, effects of nifedipine on proximal tubular cell injury remain unknown. We examined here whether and how nifedipine blocked the AGE-induced tubular cell damage. Nifedipine, but not amlodipine, a control CCB, inhibited the AGE-induced up-regulation of RAGE mRNA levels in tubular cells, which was prevented by the simultaneous treatment of GW9662, an inhibitor of peroxisome proliferator-activated receptor-{gamma} (PPAR{gamma}). GW9662 treatment alone was found to increase RAGE mRNA levels in tubular cells. Further, nifedipine inhibited the AGE-induced reactive oxygen species generation, NF-{kappa}B activation and increases in intercellular adhesion molecule-1 and transforming growth factor-beta gene expression in tubular cells, all of which were blocked by GW9662. Our present study provides a unique beneficial aspect of nifedipine on diabetic nephropathy; it could work as an anti-oxidative and anti-inflammatory agent against AGEs in tubular cells by suppressing RAGE expression via PPAR{gamma} activation.

  10. Calpain Inhibitor PD150606 Attenuates Glutamate Induced Spiral Ganglion Neuron Apoptosis through Apoptosis Inducing Factor Pathway In Vitro

    PubMed Central

    Song, Yong-Li; Chen, Xiao-Dong; Mi, Wen-Juan; Wang, Jian; Lin, Ying; Chen, Fu-Quan; Qiu, Jian-Hua

    2015-01-01

    Objective This research aimed to investigate whether glutamate induced spiral ganglion neurons (SGNs) apoptosis through apoptosis inducing factor (AIF) pathway. And verify whether PD150606, a calpain inhibitor could prevent apoptosis by inhibiting cleaving and releasing AIF in mitochondrion. Methods SGNs of postnatal days 0-3 were harvested and cultured in dishes. 20 mM Glu, the caspase inhibitor Z-VAD-FMK and calpain inhibitor PD150606 were added into cultured dishes separately. We used optical microscope and immunofluoresence staining to observe cell morphology and AIF distribution, RT-PCR and Westernblot to analyse AIF and calpain expression in SGNs. TUNEL assay was used to test cell apoptosis. Results Cell morphology and nuclear translocation of AIF were altered in SGNs by 20 mM Glu treated in vitro. The axon of SGN shortened, more apoptosis SGN were observed and the expression of AIF and calpain were up-regulated in Glu-treated group than the normal one (P<0.05). The same experiments were conducted in 20 mM+PD150606 treated group and 20 mM+Z-VAD-FMK group. Obviously AIF were located from cytoplasm to the nuclear and the expressions of AIF and calpain were down-regulated by PD150606 (P<0.05). Positive cells in TUNEL staining decreased after PD150606 treating. However, Z-VAD-FMK had no influence on AIF, calpain expression or cell apoptosis. Conclusion The AIF-related apoptosis pathway is involved in the process of Glu-induced SGN injury. Furthermore, the inhibition of calpain can prevent AIF from releasing the nuclear or inducing SGN apoptosis. PMID:25874633

  11. HSP90 inhibitor CH5164840 induces micronuclei in TK6 cells via an aneugenic mechanism.

    PubMed

    Matsuzaki, Kaori; Harada, Asako; Tanaka, Kenji; Takeiri, Akira; Mishima, Masayuki

    2014-10-01

    Heat-shock protein 90 (HSP90) is a promising druggable target for therapy of conditions including cancer, renal disease, and chronic neurodegenerative diseases. Despite the possible beneficial effects of HSP90 inhibitors, some of these agents present a genotoxicity liability. We have examined the mode of action of micronucleus formation in TK6 cells by a novel and highly specific HSP90 inhibitor, CH5164840, by means of an in vitro micronucleus test with fluorescence in situ hybridization (FISH), ?H2AX staining to detect DNA damage, and microscopic observation of chromosomal alignment in mitotic cells. The percentage of centromere-positive micronuclei induced by CH5164840 (FISH analysis) was significant, but the percentage of centromere-negative ones was not, suggesting that induction of micronuclei was due to a mechanism of aneugenicity rather than DNA reactivity. This conclusion was further supported by the result of co-staining ?H2AX and the apoptosis marker caspase-3; the predominant elevation of apoptotic ?H2AX rather than non-apoptotic ?H2AX indicated little involvement of DNA-reactivity mechanisms. Microscopic observation revealed asymmetric spindle microtubules and chromosomal misalignment of metaphase cells. These data indicated that CH5164840 causes spindle dysfunction that induces micronuclei. The risk/benefit ratio must be considered in the development of HSP90 inhibitors. PMID:25308700

  12. Diverse intracellular pathogens activate Type III Interferon expression from peroxisomes

    PubMed Central

    Odendall, Charlotte; Dixit, Evelyn; Stavru, Fabrizia; Bierne, Helene; Franz, Kate M.; Fiegen, Ann; Boulant, Steeve; Gehrke, Lee; Cossart, Pascale; Kagan, Jonathan C.

    2014-01-01

    Type I Interferon (IFN) responses are considered the primary means by which viral infections are controlled in mammals. Despite this view, several pathogens activate antiviral responses in the absence of Type I IFNs. The mechanisms controlling Type I IFN-independent responses are undefined. We have found that RIG-I like Receptors (RLRs) induce Type III IFN expression in a variety of human cell types, and identified factors that differentially regulate Type I and III IFN expression. We identified peroxisomes as a primary site that initiates Type III IFN expression, and revealed that the process of intestinal epithelial cell differentiation upregulates peroxisome biogenesis and promotes robust Type III IFN responses in human cells. These findings highlight the interconnections between innate immunity and cell biology. PMID:24952503

  13. Factors Affecting Development of Peroxisomes and Glycolate Metabolism among Algae of Different Evolutionary Lines of the Prasinophyceae.

    PubMed Central

    Kehlenbeck, P.; Goyal, A.; Tolbert, N. E.

    1995-01-01

    Leaf-type peroxisomes are not present in the primitive unicellular Prasinophycean line of algae but are present in the multicellular algae Mougeotia, Chara, and Nitella, which are in the one evolutionary line, Charophyceae, that led to higher plants. Processes related to glycolate metabolism that may have been modified or induced with the appearance of peroxisomes have been examined. The algal dissolved inorganic carbon-concentrating mechanism and alkalization of the medium during photosynthesis were not lost when peroxisomes appeared in the members of the Charophycean line of algae. Therefore, it is unlikely that lowering of the CO2 concentration in the environment was a major factor in the evolutionary appearance of peroxisomes. Multicellular Mougeotia, early members of the Charophycean line of algae, have peroxisomes, but they excrete excess glycolate into the medium. The cytosolic pyruvate reductase for D-lactate synthesis and the glycolate dehydrogenase activity almost disappeared when peroxisomal glycolate oxidase, which also oxidizes L-lactate, appeared. These biochemical changes do not indicate what caused the induction of leaf-type peroxisomes in this evolutionary line of algae. The oxygenase activity of ribulose-1,5-bisphosphate carboxylase/oxygenase and glycolate oxidase require about 200 to 400 [mu]M O2 for 0.5 Vmax. These high-O2-requiring steps in glycolate metabolism would have functioned faster with increasing atmospheric O2, which might have been the causative factor in the induction of peroxisomes. PMID:12228674

  14. Suppression of VEGF-induced angiogenesis by the protein tyrosine kinase inhibitor, lavendustin A.

    PubMed Central

    Hu, D E; Fan, T P

    1995-01-01

    1. Vascular endothelial growth factor (VEGF) is a heparin-binding angiogenic factor which specifically acts on endothelial cells via distinct membrane-spanning tyrosine kinase receptors. Here we used the rat sponge implant model to test the hypothesis that the angiogenic activity of VEGF can be suppressed by protein tyrosine kinase (PTK) inhibitors. 2. Neovascular responses in subcutaneous sponge implants were determined by measurements of relative sponge blood flow by use of a 133Xe clearance technique, and confirmed by histological studies and morphometric analysis. 3. Daily local administration of 250 ng VEGF165 accelerated the rate of 133Xe clearance from the sponges and induced an intense neovascularisation. This VEGF165-induced angiogenesis was inhibited by daily co-administration of the selective PTK inhibitor, lavendustin A (10 micrograms), but not its negative control, lavendustin B (10 micrograms). Blood flow measurements and morphometric analysis of 8-day-old sponges showed that lavendustin A reduced the 133Xe clearance of VEGF165-treated sponges from 32.9 +/- 1.5% to 20.9 +/- 1.6% and the total fibrovascular growth area from 62.4 +/- 6.1% to 21.6 +/- 6.8% (n = 12, P < 0.05). 4. Co-injection of suramin (3 mg), an inhibitor of heparin-binding growth factors, also suppressed the VEGF165-elicited neovascular response. In contrast, neither lavendustin A nor suramin produced any effect on the basal sponge-induced angiogenesis. 5. When given alone, low doses of VEGF165 (25 ng) or basic fibroblast growth factor (bFGF; 10 ng) did not modify the basal sponge-induced neovascularisation.(ABSTRACT TRUNCATED AT 250 WORDS) Images Figure 2 Figure 2 PMID:7533611

  15. Risk assessment and management of anthracycline and HER2 receptor inhibitor-induced cardiomyopathy.

    PubMed

    Jahangir, Eiman; Shah, Sangeeta; Shum, Kelly; Baxter, Caitlin; Fitzpatrick, Jill D; Cole, John; Gilliland, Yvonne; Polin, Nichole M

    2015-02-01

    With the advent and increased use of chemotherapeutic agents and radiation therapy, cancer survival rates have increased. With increased survival, both acute and chronic cardiotoxic adverse effects have emerged. The growing need for managing the treatment of individuals with chemotherapy-induced cardiotoxicity has led to the formation of cardio-oncology programs throughout the United States. These programs concentrate on many aspects of cardiac disease in the oncology patient. Of these, the cardiotoxic effects (particularly cardiomyopathy) of anthracyclines and HER2 receptor inhibitors are a large focus of cardio-oncology practice. Despite the increasing availability of these programs, no consensus guidelines have been established to provide a framework for treating these patients. This review describes the initial evaluation, risk assessment, and management of individuals receiving anthracycline and HER2 receptor inhibitor therapy for cardiomyopathy. These recommendations are supported by the current literature in this field. PMID:25688890

  16. Hypoxia inducible factor 1? expression and effects of its inhibitors in canine lymphoma

    PubMed Central

    KAMBAYASHI, Satoshi; IGASE, Masaya; KOBAYASHI, Kosuke; KIMURA, Ayana; SHIMOKAWA MIYAMA, Takako; BABA, Kenji; NOGUCHI, Shunsuke; MIZUNO, Takuya; OKUDA, Masaru

    2015-01-01

    Hypoxic conditions in various cancers are believed to relate with their malignancy, and hypoxia inducible factor-1? (HIF-1?) has been shown to be a major regulator of the response to low oxygen. In this study, we examined HIF-1? expression in canine lymphoma using cell lines and clinical samples and found that these cells expressed HIF-1?. Moreover, the HIF-1? inhibitors, echinomycin, YC-1 and 2-methoxyestradiol, suppressed the proliferation of canine lymphoma cell lines. In a xenograft model using NOD/scid mice, echinomycin treatment resulted in a dose-dependent regression of the tumor. Our results suggest that HIF-1? contributes to the proliferation and/or survival of canine lymphoma cells. Therefore, HIF-1? inhibitors may be potential agents to treat canine lymphoma. PMID:26050843

  17. Neuroprotective effects of peroxisome proliferator-activated receptor alpha and gamma agonists in model of parkinsonism induced by intranigral 1-methyl-4-phenyl-1,2,3,6-tetrahyropyridine.

    PubMed

    Barbiero, Janaína K; Santiago, Ronise M; Persike, Daniele Suzete; da Silva Fernandes, Maria José; Tonin, Fernanda S; da Cunha, Claudio; Lucio Boschen, Suelen; Lima, Marcelo M S; Vital, Maria A B F

    2014-11-01

    A large body of evidence suggests that peroxisome proliferator-activated receptor (PPAR) agonists may improve some of the pathological features of Parkinson's disease (PD). In the present study, we evaluated the effects of the PPAR-? agonist fenofibrate (100mg/kg) and PPAR-? agonist pioglitazone (30mg/kg) in a rat model of parkinsonism induced by intranigral 1-methyl-4-phenyl-1,2,3,6-tetrahyropyridine (MPTP). Male Wistar rats were pretreated with both drugs for 5 days and received an infusion of MPTP. The experiments were divided into two parts. First, 1, 7, 14, and 21 days after surgery, the animals were submitted to the open field test. On days 21 and 22, the rats were subjected to the forced swim test and two-way active avoidance task. In the second part of the study, 24h after neurotoxin administration, immunohistochemistry was performed to assess tyrosine hydroxylase activity. The levels of dopamine and its metabolites in the striatum were determined using high-performance liquid chromatography, and fluorescence detection was used to assess caspase-3 activation in the substantia nigra pars compacta (SNpc). Both fenofibrate as pioglitazone protected against hypolocomotion, depressive-like behavior, impairment of learning and memory, and dopaminergic neurodegeneration caused by MPTP, with dopaminergic neuron loss of approximately 33%. Fenofibrate and pioglitazone also protected against the increased activation of caspase-3, an effector enzyme of the apoptosis cascade that is considered one of the pathological features of PD. Thus, PPAR agonists may contribute to therapeutic strategies in PD. PMID:25127682

  18. Proteasome inhibitor-induced autophagy in PC12 cells overexpressing A53T mutant ?-synuclein.

    PubMed

    Lan, Danmei; Wang, Wenzhao; Zhuang, Jianhua; Zhao, Zhongxin

    2015-03-01

    The aim of the present study was to examine the effects of proteasome inhibitor (PI)?induced autophagy on PC12 cells overexpressing A53T mutant ??synuclein (??syn) by detecting alterations in the levels of microtubule?associated protein 1A/1B light chain (LC3)+ autophagosomes and the lysotracker?positive autolysosomes using immunofluorescence, the expression of LC3?II using western blot analysis and the morphology of PC12 cells using transmission electron microscopy. It was found that the addition of MG132 (500 nmol/l) significantly increased the number of autophagosomes and autolysosomes and upregulated the expression of LC3?II. The autophagy inhibitor 3?methyladenine (3?MA) completely inhibited the autophagy induced by MG132 (500 nmol/l). The autophagy enhancer trehalose significantly increased the number of autophagosomes and autolysosomes and improved the protein level of LC3?II induced by MG132. To examine the effect of PI?induced autophagy on the degradation of A53T mutant ??syn, the expression of ??syn was detected by western blot analysis. It was revealed that MG132 increased the expression of A53T ??syn and trehalose counteracted the increase of A53T ??syn induced by MG132. Combined inhibition of 3?MA and PI significantly increased the accumulation of A53T ??syn as compared with treatment using either single agent. In addition, combination of MG132 (500 nmol/l) with trehalose (50 mmol/l) or 3?MA (2 mmol/l) markedly decreased the cell viability as compared with treatment using either single agent individually as demonstrated using a 3?(4,5?dimethylthiazol?2?yl)?2,5?diphenyltetrazolium bromide assay. These results suggest that the PI, MG132, could induce autophagy in PC12 cells overexpressing A53T mutant ??syn and this autophagy could be completely inhibited by 3?MA, indicating that PI?induced autophagy is mediated by the upregulation of the macroautophagy class III PI3K pathway. PI?induced autophagy may act as a compensatory degradation system for degradation of A53T ??syn when the ubiquitin?proteasome system is impaired. Autophagy activation may directly contribute to the survival of PC12 cells treated with proteasome inhibitors. The present study may assist in illuminating the association between PI and autophagy in the pathogenesis of Parkinson's disease. PMID:25434876

  19. p53 modulates the AMPK inhibitor compound C induced apoptosis in human skin cancer cells

    SciTech Connect

    Huang, Shi-Wei; Wu, Chun-Ying; Wang, Yen-Ting; Kao, Jun-Kai; Lin, Chi-Chen; Chang, Chia-Che; Mu, Szu-Wei; Chen, Yu-Yu; Chiu, Husan-Wen; Chang, Chuan-Hsun; Liang, Shu-Mei; Chen, Yi-Ju; Huang, Jau-Ling; Shieh, Jeng-Jer

    2013-02-15

    Compound C, a well-known inhibitor of the intracellular energy sensor AMP-activated protein kinase (AMPK), has been reported to cause apoptotic cell death in myeloma, breast cancer cells and glioma cells. In this study, we have demonstrated that compound C not only induced autophagy in all tested skin cancer cell lines but also caused more apoptosis in p53 wildtype skin cancer cells than in p53-mutant skin cancer cells. Compound C can induce upregulation, phosphorylation and nuclear translocalization of the p53 protein and upregulate expression of p53 target genes in wildtype p53-expressing skin basal cell carcinoma (BCC) cells. The changes of p53 status were dependent on DNA damage which was caused by compound C induced reactive oxygen species (ROS) generation and associated with activated ataxia-telangiectasia mutated (ATM) protein. Using the wildtype p53-expressing BCC cells versus stable p53-knockdown BCC sublines, we present evidence that p53-knockdown cancer cells were much less sensitive to compound C treatment with significant G2/M cell cycle arrest and attenuated the compound C-induced apoptosis but not autophagy. The compound C induced G2/M arrest in p53-knockdown BCC cells was associated with the sustained inactive Tyr15 phosphor-Cdc2 expression. Overall, our results established that compound C-induced apoptosis in skin cancer cells was dependent on the cell's p53 status. - Highlights: ? Compound C caused more apoptosis in p53 wildtype than p53-mutant skin cancer cells. ? Compound C can upregulate p53 expression and induce p53 activation. ? Compound C induced p53 effects were dependent on ROS induced DNA damage pathway. ? p53-knockdown attenuated compound C-induced apoptosis but not autophagy. ? Compound C-induced apoptosis in skin cancer cells was dependent on p53 status.

  20. The behavior of peroxisomes in vitro: mammalian peroxisomes are osmotically sensitive particles.

    PubMed

    Antonenkov, Vasily D; Sormunen, Raija T; Hiltunen, J Kalervo

    2004-12-01

    It has been known for a long time that mammalian peroxisomes are extremely fragile in vitro. Changes in the morphological appearance and leakage of proteins from purified particles demonstrate that peroxisomes are damaged during isolation. However, some properties of purified peroxisomes, e.g., the latency of catalase, imply that their membranes are not disrupted. In the current study, we tried to ascertain the mechanism of this unusual behavior of peroxisomes in vitro. Biochemical and morphological examination of isolated peroxisomes subjected to sonication or to freezing and thawing showed that the membrane of the particles seals after disruption, restoring permeability properties. Transient damage of the membrane leads to the formation of peroxisomal "ghosts" containing nucleoid but nearly devoid of matrix proteins. The rate of leakage of matrix proteins from broken particles depended inversely on their molecular size. The effect of polyethylene glycols on peroxisomal integrity indicated that these particles are osmotically sensitive. Peroxisomes suffered an osmotic lysis during isolation that was resistant to commonly used low-molecular-mass osmoprotectors, e.g., sucrose. Damage to peroxisomes was partially prevented by applying more "bulky" osmoprotectors, e.g., polyethylene glycol 1500. A method was developed for the isolation of highly purified and nearly intact peroxisomes from rat liver by using polyethylene glycol 1500 as an osmoprotector. PMID:15306541

  1. The role of peroxisomal fatty acyl-CoA beta-oxidation in bile acid biosynthesis

    SciTech Connect

    Hayashi, H.; Miwa, A. )

    1989-11-01

    The physiological role of the peroxisomal fatty acyl-CoA beta-oxidizing system (FAOS) is not yet established. We speculated that there might be a relationship between peroxisomal degradation of long-chain fatty acids in the liver and the biosynthesis of bile acids. This was investigated using (1-{sup 14}C)butyric acid and (1-{sup 14}C)lignoceric acid as substrates of FAOS in mitochondria and peroxisomes, respectively. The incorporation of ({sup 14}C)lignoceric acid into primary bile acids was approximately four times higher than that of ({sup 14}C)butyric acid (in terms of C-2 units). The pools of these two fatty acids in the liver were exceedingly small. The incorporations of radioactivity into the primary bile acids were strongly inhibited by administration of aminotriazole, which is a specific inhibitor of peroxisomal FAOS in vivo. Aminotriazole inhibited preferentially the formation of cholate, the major primary bile acid, from both ({sup 14}C)lignoceric acid and ({sup 14}C)butyric acid, rather than the formation of chenodeoxycholate. The former inhibition was about 70% and the latter was approximately 40-50%. In view of reports that cholate is biosynthesized from endogenous cholesterol, the above results indicate that peroxisomal FAOS may have an anabolic function, supplying acetyl CoA for bile acid biosynthesis.

  2. The farnesyltransferase inhibitor, FPT inhibitor III upregulates Bax and Bcl-xs expression and induces apoptosis in human ovarian cancer cells.

    PubMed

    Hung, W C; Chaung, L Y

    1998-01-01

    Deregulation in the ras oncogene is a common event in many types of human cancer. Our previous study clearly demonstrated that genetic alterations of ras oncogene are frequently found in human epithelial ovarian cancer. Recent reports have indicated that farnesyltransferase is involved in the regulation of post-translational modification and biological function of Ras proteins. Here, we report that a newly synthesized farnesyltransferase inhibitor, FPT inhibitor III, upregulates Bax and Bcl-xs expression and induces apoptosis in human ovarian cancer cells. This is a critical finding that farnesyltransferase inhibitors may directly activate apoptotic signaling pathways in cancer cells and may help to provide a new strategy in the treatment of human cancer. PMID:9454897

  3. An efficient screen for peroxisome-deficient mutants of Pichia pastoris.

    PubMed Central

    Liu, H; Tan, X; Veenhuis, M; McCollum, D; Cregg, J M

    1992-01-01

    We describe a rapid and efficient screen for peroxisome-deficient (per) mutants in the yeast Pichia pastoris. The screen relies on the unusual ability of P. pastoris to grow on two carbon sources, methanol and oleic acid, both of which absolutely require peroxisomes to be metabolized. A collection of 280 methanol utilization-defective (Mut-) P. pastoris mutants was isolated, organized into 46 complementation groups, and tested for those that were also oleate-utilization defective (Out-) but still capable of growth on ethanol and glucose. Mutants in 10 groups met this phenotypic description, and 8 of these were observed by electron microscopy to be peroxisome deficient (Per-). In each per mutant, Mut-, Out-, and Per- phenotypes were tightly linked and therefore were most likely due to a mutation at a single locus. Subcellular fractionation experiments indicated that the peroxisomal marker enzyme catalase was mislocalized to the cytosol in both methanol- and oleate-induced cultures of the mutants. In contrast, alcohol oxidase, a peroxisomal methanol utilization pathway enzyme, was virtually absent from per mutant cells. The relative ease of per mutant isolation in P. pastoris, in conjunction with well-developed procedures for its molecular and genetic manipulation, makes this organism an attractive system for studies on peroxisome biogenesis. Images PMID:1629154

  4. Peroxisomal Import Reduces the Proapoptotic Activity of Deubiquitinating Enzyme USP2

    PubMed Central

    Reglinski, Katharina; Keil, Marina; Altendorf, Sabrina; Waithe, Dominic; Eggeling, Christian; Schliebs, Wolfgang; Erdmann, Ralf

    2015-01-01

    The human deubiquitinating enzyme ubiquitin-specific protease 2 (USP2) regulates multiple cellular pathways, including cell proliferation and apoptosis. As a result of alternative splicing four USP2 isoenzymes are expressed in human cells of which all contain a weak peroxisome targeting signal of type 1 (PTS1) at their C-termini. Here, we systematically analyzed apoptotic effects induced by overexpression and intracellular localization for each isoform. All isoforms exhibit proapoptotic activity and are post-translationally imported into the matrix of peroxisomes in a PEX5-dependent manner. However, a significant fraction of the USP2 pool resides in the cytosol due to a weaker PTS1 and thus low affinity to the PTS receptor PEX5. Blocking of peroxisomal import did not interfere with the proapoptotic activity of USP2, suggesting that the enzyme performs its critical function outside of this compartment. Instead, increase of the efficiency of USP2 import into peroxisomes either by optimization of its peroxisomal targeting signal or by overexpression of the PTS1 receptor did result in a reduction of the apoptotic rate of transfected cells. Our studies suggest that peroxisomal import of USP2 provides additional control over the proapoptotic activity of cytosolic USP2 by spatial separation of the deubiquitinating enzymes from their interaction partners in the cytosol and nucleus. PMID:26484888

  5. Association between cardiac changes and stress, and the effect of peroxisome proliferator-activated receptor-? on stress-induced myocardial injury in mice.

    PubMed

    Gao, Jin-liao; Xue, Qiao; Wang, Shi-wen; Gao, Li-fei; Lan, Yun-feng; Fang, Zhou; Fu, Yi-cheng; Liu, Yan; Li, Yang; Fan, Li

    2015-02-01

    This study was aimed to investigate the effect of stress induced by high-intensity exercises on the cardiovascular system. In the epidemiological investigation, 200 subjects (test group) engaged in special high-intensity exercises, and 97 who lived and worked in the same environment and conditions as those in the test group, but did not participate in the exercises served as controls. In the second part of the study, 50 mice were randomly divided into control group, exhaustive swimming group, white noise group, exhaustive swimming plus white noise group, and pioglitazone intervention group. The results showed that the plasma concentrations of the myocardial injury markers heart fatty acid-binding protein (H-FABP), C-reactive protein (CRP), ?-endorphin (?-EP) and levels of psychological stress were significantly increased in test group as compared with control group; special high-intensity exercises resulted in a significant elevation of the incidence of cardiac arrhythmias. Animal experiments showed that the plasma levels of corticosterone (CORT) and troponin I (TnI) were raised while the level of SOD was reduced in exhaustive swimming group, white noise group, and exhaustive swimming plus white noise group. The expression levels of PPAR? mRNA and protein were decreased in myocardial tissues in these groups as well. HE staining showed no remarkable change in myocardial tissues in all the groups. Treatment with pioglitazone significantly decreased the plasma levels of TnI and CORT, while increased the level of SOD and the expression levels of PPAR? mRNA and protein. It was concluded that the high-intensity exercises may induce a heavy physical and psychological stress and predispose the subjects to accumulated fatigue and sleep deprivation; high-intensity exercises also increases the incidence of arrhythmias and myocardial injury. PPAR? may be involved in the physical and psychological changes induced by high-intensity exercises. PMID:25673189

  6. Mitochondrial division inhibitor-1 induces mitochondrial hyperfusion and sensitizes human cancer cells to TRAIL-induced apoptosis.

    PubMed

    Akita, Mamoru; Suzuki-Karasaki, Miki; Fujiwara, Kyoko; Nakagawa, Chinatsu; Soma, Masayoshi; Yoshida, Yukihiro; Ochiai, Toyoko; Tokuhashi, Yasuaki; Suzuki-Karasaki, Yoshihiro

    2014-11-01

    Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is a promising candidate for cancer treatment, but some cancer cell types are resistant to TRAIL cytotoxicity. Therefore, overcoming this resistance is necessary for effective TRAIL therapy. Mitochondrial morphology is important for the maintenance of cell function and survival, and is regulated by the delicate balance between fission and fusion. However, the role of mitochondrial morphology dynamics in TRAIL-induced apoptosis is unknown. Here we show that mitochondrial division inhibitor-1 (mdivi-1), an inhibitor of dynamin-related protein1 (Drp1), modulates mitochondrial morphology and TRAIL-induced apoptosis in human cancer cells. mdivi-1 treatment (?12.5 µM) caused dose- and time?dependent cell death in malignant melanoma, lung cancer and osteosarcoma cells, while sparing normal cells. mdivi-1 also sensitized cancer cells to TRAIL-induced apoptosis. This potentiation of apoptosis occurred through a caspase-depependent mechanism including the mitochondrial and endoplasmic reticulum (ER) stress pathways. Mdivi-1 potentiated mitochondrial oxidative stress, a major cause of mitochondrial and ER stresses, as evidenced by increases in mitochondrial reactive oxygen species levels, mitochondrial mass, and cardiolipin oxidation. Live cell fluorescence imaging using MitoTracker Red CMXRos revealed that Mdivi-1 caused substantial mitochondrial hyperfusion. Moreover, silencing of Drp1 expression also caused mitochondrial hyperfusion and sensitized cancer cells to TRAIL-induced apoptosis. Our results suggest that cancer cells are more vulnerable than normal cells to a perturbation in mitochondrial morphology dynamics and that this higher susceptibility can be exploited to selectively kill cancer cells and sensitize to TRAIL. PMID:25174275

  7. Peroxisome Biogenesis Disorders: Biological, Clinical and Pathophysiological Perspectives

    ERIC Educational Resources Information Center

    Braverman, Nancy E.; D'Agostino, Maria Daniela; MacLean, Gillian E.

    2013-01-01

    The peroxisome biogenesis disorders (PBD) are a heterogeneous group of autosomal recessive disorders in which peroxisome assembly is impaired, leading to multiple peroxisome enzyme deficiencies, complex developmental sequelae and progressive disabilities. Mammalian peroxisome assembly involves the protein products of 16 "PEX" genes;…

  8. Histone deacetylase inhibitor treatment induces 'BRCAness' and synergistic lethality with PARP inhibitor and cisplatin against human triple negative breast cancer cells.

    PubMed

    Ha, Kyungsoo; Fiskus, Warren; Choi, Dong Soon; Bhaskara, Srividya; Cerchietti, Leandro; Devaraj, Santhana G T; Shah, Bhavin; Sharma, Sunil; Chang, Jenny C; Melnick, Ari M; Hiebert, Scott; Bhalla, Kapil N

    2014-07-30

    There is an unmet need to develop new, more effective and safe therapies for the aggressive forms of triple negative breast cancers (TNBCs). While up to 20% of women under 50 years of age with TNBC harbor germline mutations in BRCA1, and these tumors are sensitive to treatment with poly(ADP) ribose polymerase inhibitors, a majority of TNBCs lack BRCA1 mutations or loss of expression. Findings presented here demonstrate that by attenuating the levels of DNA damage response and homologous recombination proteins, pan-histone deacetylase inhibitor (HDI) treatment induces 'BRCAness' and sensitizes TNBC cells lacking BRCA1 to lethal effects of PARP inhibitor or cisplatin. Treatment with HDI also induced hyperacetylation of nuclear hsp90. Similar effects were observed following shRNA-mediated depletion of HDAC3, confirming its role as the deacetylase for nuclear HSP90. Furthermore, cotreatment with HDI and ABT-888 induced significantly more DNA strand breaks than either agent alone, and synergistically induced apoptosis of TNBC cells. Notably, co-treatment with HDI and ABT-888 significantly reduced in vivo tumor growth and markedly improved the survival of mice bearing TNBC cell xenografts. These findings support the rationale to interrogate the clinical activity of this novel combination against human TNBC, irrespective of its expression of mutant BRCA1. PMID:25026298

  9. Histone deacetylase inhibitor treatment induces ‘BRCAness’ and synergistic lethality with PARP inhibitor and cisplatin against human triple negative breast cancer cells

    PubMed Central

    Ha, Kyungsoo; Bhaskara, Srividya; Cerchietti, Leandro; Devaraj, Santhana G. T.; Shah, Bhavin; Sharma, Sunil; Chang, Jenny C.; Melnick, Ari M.; Hiebert, Scott; Bhalla, Kapil N.

    2014-01-01

    There is an unmet need to develop new, more effective and safe therapies for the aggressive forms of triple negative breast cancers (TNBCs). While up to 20% of women under 50 years of age with TNBC harbor germline mutations in BRCA1, and these tumors are sensitive to treatment with poly(ADP) ribose polymerase inhibitors, a majority of TNBCs lack BRCA1 mutations or loss of expression. Findings presented here demonstrate that by attenuating the levels of DNA damage response and homologous recombination proteins, pan-histone deacetylase inhibitor (HDI) treatment induces ‘BRCAness’ and sensitizes TNBC cells lacking BRCA1 to lethal effects of PARP inhibitor or cisplatin. Treatment with HDI also induced hyperacetylation of nuclear hsp90. Similar effects were observed following shRNA-mediated depletion of HDAC3, confirming its role as the deacetylase for nuclear HSP90. Furthermore, cotreatment with HDI and ABT-888 induced significantly more DNA strand breaks than either agent alone, and synergistically induced apoptosis of TNBC cells. Notably, co-treatment with HDI and ABT-888 significantly reduced in vivo tumor growth and markedly improved the survival of mice bearing TNBC cell xenografts. These findings support the rationale to interrogate the clinical activity of this novel combination against human TNBC, irrespective of its expression of mutant BRCA1. PMID:25026298

  10. Effects of lipoxygenase inhibitors in a model of lens-induced uveitis in dogs.

    PubMed

    Dziezyc, J; Millichamp, N J; Rohde, B H; Baker, J S; Chiou, G C

    1989-11-01

    Uveitis was induced in dogs by intracameral injection of canine lens protein. The lipoxygenase inhibitors phenidone and norhydroguaiaretic acid, and dimethyl sulfoxide decreased fibrin production at 0.5 and 1 hour after induction of uveitis. Phenidone and norhydroguaiaretic acid also inhibited the initial increase in intraocular pressure early in the course of inflammation. Leukotriene B4 in the aqueous was measured by use of radioimmunoassay at 1 hour after inflammation. In control dogs, 230 to 1,700 pg of leukotriene B4/ml was measured; in dogs treated with phenidone, leukotriene B4 was not measured. PMID:2515781

  11. Heat shock protein inhibitors, 17-DMAG and KNK437, enhance arsenic trioxide-induced mitotic apoptosis

    SciTech Connect

    Wu Yichen; Yen Wenyen; Lee, T.-C. Yih, L.-H.

    2009-04-15

    Arsenic trioxide (ATO) has recently emerged as a promising therapeutic agent in leukemia because of its ability to induce apoptosis. However, there is no sufficient evidence to support its therapeutic use for other types of cancers. In this study, we investigated if, and how, 17-dimethylaminoethylamino-17-demethoxy-geldanamycin (17-DMAG), an antagonist of heat shock protein 90 (HSP90), and KNK437, a HSP synthesis inhibitor, potentiated the cytotoxic effect of ATO. Our results showed that cotreatment with ATO and either 17-DMAG or KNK437 significantly increased ATO-induced cell death and apoptosis. siRNA-mediated attenuation of the expression of the inducible isoform of HSP70 (HSP70i) or HSP90{alpha}/{beta} also enhanced ATO-induced apoptosis. In addition, cotreatment with ATO and 17-DMAG or KNK437 significantly increased ATO-induced mitotic arrest and ATO-induced BUBR1 phosphorylation and PDS1 accumulation. Cotreatment also significantly increased the percentage of mitotic cells with abnormal mitotic spindles and promoted metaphase arrest as compared to ATO treatment alone. These results indicated that 17-DMAG or KNK437 may enhance ATO cytotoxicity by potentiating mitotic arrest and mitotic apoptosis possibly through increased activation of the spindle checkpoint.

  12. VEGF-induced HUVEC migration and proliferation are decreased by PDE2 and PDE4 inhibitors.

    PubMed

    Favot, Laure; Keravis, Thérèse; Holl, Vincent; Le Bec, Alain; Lugnier, Claire

    2003-08-01

    Migration and proliferation of endothelial cells in response to VEGF play an important role in angiogenesis associated to pathologies such as atherosclerosis, diabetes and tumor development. Elevation of cAMP in endothelial cells has been shown to inhibit growth factor-induced proliferation. Our hypothesis was that inactivation of cAMP-specific phosphodiesterases (PDEs) would inhibit angiogenesis. The purpose of this study was to evaluate the effect of PDE inhibitors on in vitro and in vivo angiogenesis, using human umbilical vein endothelial cell (HUVEC) and chick chorioallantoic membrane (CAM) models respectively. Here, we report that: 1) PDE2, PDE3, PDE4 and PDE5 are expressed in HUVEC; 2) EHNA (20 microM), PDE2 selective inhibitor, and RP73401 (10 microM), PDE4 selective inhibitor, are able to increase the intracellular cAMP level in HUVEC; 3) EHNA and RP73401 are able to inhibit proliferation, cell cycle progression and migration of HUVEC stimulated by VEGF; 4) these in vitro effects can be mimic by treating HUVEC with the cAMP analogue, 8-Br-cAMP (600 microM); 5) only the association of EHNA and RP73401 inhibits in vivo angiogenesis, indicating that both migration and proliferation must be inhibited. These data strongly suggest that PDE2 and PDE4 represent new potential therapeutic targets in pathological angiogenesis. PMID:12888882

  13. Binding-induced, turn-on fluorescence of the EGFR/ERBB kinase inhibitor, lapatinib.

    PubMed

    Wilson, James N; Liu, Wenjun; Brown, Adrienne S; Landgraf, Ralf

    2015-05-01

    We report the photophysical properties, binding-induced turn-on emission, and fluorescence imaging of the cellular uptake and distribution of lapatinib, an EGFR/ERBB inhibitor. Lapatinib, a type II, i.e. inactive state, inhibitor that targets the ATP binding pocket of the EGFR family of receptor tyrosine kinases. DFT calculations predict that the 6-furanylquinazoline core of lapatinib should exhibit an excited state with charge transfer character and an S0 to S1 transition energy of 3.4 eV. Absorption confirms an optical transition in the near UV to violet, while fluorescence spectroscopy shows that photoemission is highly sensitive to solvent polarity. The hydrophobicity of lapatinib leads to fluorescent aggregates in solution, however, binding to the lipid-carrier protein, BSA or to the kinase domain of ERBB2, produces spectroscopically distinct photoemission. Confocal fluorescence microscopy imaging of lapatinib uptake in ERBB2-overexpressing MCF7 and BT474 cells reveals pools of intracellular inhibitor with emission profiles consistent with aggregated lapatinib. PMID:25820099

  14. Identification of an Allosteric Small Molecule Inhibitor Selective for Inducible Form of Heat Shock Protein 70

    PubMed Central

    Howe, Matthew K.; Bodoor, Khaldon; Carlson, David A.; Hughes, Philip F.; Alwarawrah, Yazan; Loiselle, David R.; Jaeger, Alex M.; Darr, David B.; Jordan, Jamie L.; Hunter, Lucas M.; Molzberger, Eileen T.; Gobillot, Theodore A.; Thiele, Dennis J.; Brodsky, Jeffrey L.; Spector, Neil L.; Haystead, Timothy A. J.

    2014-01-01

    Summary Inducible Hsp70 (Hsp70i) is overexpressed in a wide spectrum of human tumors and its expression correlates with metastasis, poor outcomes, and resistance to chemotherapy in patients. Identification of small molecule inhibitors selective for Hsp70i could provide new therapeutic tools for cancer treatment. In this work, we used fluorescence-linked enzyme chemoproteomic strategy (FLECS) to identify HS-72, an allosteric inhibitor selective for Hsp70i. HS-72 displays the hallmarks of Hsp70 inhibition in cells, promoting substrate protein degradation and growth inhibition. Importantly, HS-72 is selective for Hsp70i over the closely related constitutively active Hsc70. Studies with purified protein show HS-72 acts as an allosteric inhibitor, reducing ATP affinity. In vivo HS-72 is well-tolerated, showing bioavailability and efficacy, inhibiting tumor growth and promoting survival in a HER2+ model of breast cancer. The HS-72 scaffold is amenable to resynthesis and iteration, suggesting an ideal starting point for a new generation of anticancer therapeutics targeting Hsp70i. PMID:25500222

  15. mTOR inhibitors counteract tamoxifen-induced activation of breast cancer stem cells.

    PubMed

    Karthik, Govindasamy-Muralidharan; Ma, Ran; Lövrot, John; Kis, Lorand Levente; Lindh, Claes; Blomquist, Lennart; Fredriksson, Irma; Bergh, Jonas; Hartman, Johan

    2015-10-10

    Breast cancer cells with stem cell characteristics (CSC) are a distinct cell population with phenotypic similarities to mammary stem cells. CSCs are important drivers of tumorigenesis and the metastatic process. Tamoxifen is the most widely used hormonal therapy for estrogen receptor (ER) positive cancers. In our study, tamoxifen was effective in reducing proliferation of ER?+?adherent cancer cells, but not their CSC population. We isolated, expanded and incubated CSC from seven breast cancers with or without tamoxifen. By genome-wide transcriptional analysis we identified tamoxifen-induced transcriptional pathways associated with ribosomal biogenesis and mRNA translation, both regulated by the mTOR-pathway. We observed induction of the key mTOR downstream targets S6K1, S6RP and 4E-BP1 in-patient derived CSCs by tamoxifen on protein level. Using the mTOR inhibitors rapamycin, everolimus and PF-04691502 (a dual PI3K/mTOR inhibitor) and in combination with tamoxifen, significant reduction in mammosphere formation was observed. Hence, we suggest that the CSC population play a significant role during endocrine resistance through activity of the mTOR pathway. In addition, tamoxifen further stimulates the mTOR-pathway but can be antagonized using mTOR-inhibitors. PMID:26208432

  16. Topically applied Hsp90 inhibitor 17AAG inhibits ultraviolet radiation-induced cutaneous squamous cell carcinomas

    PubMed Central

    Singh, Anupama; Singh, Ashok; Sand, Jordan M.; Bauer, Samuel J.; Hafeez, Bilal Bin; Meske, Louise; Verma, Ajit K.

    2014-01-01

    We present here that Heat shock protein 90 (Hsp90) inhibitor 17-(allylamino)-17-demethoxygeldanamycin (17AAG), when topically applied to mouse skin, inhibits ultraviolet radiation (UVR)-induced development of cutaneous squamous cell carcinoma (SCC). In these experiments, DMSO:acetone (1:40 v/v) solution of 17AAG (500nmol) was applied topically to mouse skin in conjunction with each UVR exposure (1.8 kJ/m2). The UVR source was Kodacel-filtered FS-40 sun lamps (approximately 60% UVB and 40% UVA). In independent experiments with three separate mouse lines (SKH-1 hairless mice, wild-type FVB, and PKC? overexpressing transgenic FVB mice), 17AAG treatment increased the latency and decreased both the incidence and multiplicity of UVR-induced SCC. Topical 17AAG alone or in conjunction with UVR treatments elicited neither skin nor systemic toxicity. 17AAG-caused inhibition of SCC induction was accompanied by decrease in UVR-induced: 1) hyperplasia, 2) Hsp90?-PKC? interaction, 3) expression levels of Hsp90?, Stat3, pStat3Ser727, pStat3Tyr705, pAktSer473 and matrix metalloproteinase (MMPs). The results presented here indicate that topical Hsp90 inhibitor 17AAG is effective in prevention of UVR-induced epidermal hyperplasia and SCC. One may conclude from the preclinical data presented here that topical 17AAG may be useful for prevention of UVR-induced inflammation and cutaneous SCC either developed in UVR exposed or organ transplant population. PMID:25337691

  17. Use of a soluble epoxide hydrolase inhibitor in smoke-induced chronic obstructive pulmonary disease.

    PubMed

    Wang, Lei; Yang, Jun; Guo, Lei; Uyeminami, Dale; Dong, Hua; Hammock, Bruce D; Pinkerton, Kent E

    2012-05-01

    Tobacco smoke-induced chronic obstructive pulmonary disease (COPD) is a prolonged inflammatory condition of the lungs characterized by progressive and largely irreversible airflow limitation attributable to a number of pathologic mechanisms, including bronchitis, bronchiolitis, emphysema, mucus plugging, pulmonary hypertension, and small-airway obstruction. Soluble epoxide hydrolase inhibitors (sEHIs) demonstrated anti-inflammatory properties in a rat model after acute exposure to tobacco smoke. We compared the efficacy of sEHI t-TUCB (trans-4-{4-[3-(4-trifluoromethoxy-phenyl)-ureido]-cyclohexyloxy}-benzoic acid) and the phosphodiesterase-4 (PDE4) inhibitor Rolipram (Biomol International, Enzo Life Sciences, Farmingdale, NY) to reduce lung injury and inflammation after subacute exposure to tobacco smoke over a period of 4 weeks. Pulmonary physiology, bronchoalveolar lavage, cytokine production, and histopathology were analyzed to determine the efficacy of sEHI and Rolipram to ameliorate tobacco smoke-induced inflammation and injury in the spontaneously hypertensive rat. Both t-TUCB and Rolipram inhibited neutrophil elevation in bronchoalveolar lavage. sEHI t-TUCB suppressed IFN-?, while improving lung function by reducing tobacco smoke-induced total respiratory resistance and tissue damping (small-airway and peripheral tissue resistance). Increases in tobacco smoke-induced alveolar airspace size were attenuated by t-TUCB. Rolipram inhibited the production of airway mucus. Both t-TUCB and Rolipram inhibited vascular remodeling-related growth factor. These findings suggest that sEHI t-TUCB has therapeutic potential for treating COPD by improving lung function and attenuating the lung inflammation and emphysematous changes caused by tobacco smoke. To the best of our knowledge, this is the first report to demonstrate that sEHI exerts significant protective effects after repeated, subacute tobacco smoke-induced lung injury in a rat model of COPD. PMID:22180869

  18. Cytoprotective effect of selective small-molecule caspase inhibitors against staurosporine-induced apoptosis

    PubMed Central

    Wu, Jianghong; Wang, Yuren; Liang, Shuguang; Ma, Haiching

    2014-01-01

    Caspases are currently known as the central executioners of the apoptotic pathways. Inhibition of apoptosis and promotion of normal cell survival by caspase inhibitors would be a tremendous benefit for reducing the side effects of cancer therapy and for control of neurodegenerative disorders such as Parkinson’s, Alzheimer’s, and Huntington’s diseases. The objective of this study was to discover small-molecule caspase inhibitors with which to achieve cytoprotective effect. We completed the high-throughput screening of Bionet’s 37,500-compound library (Key Organics Limited, Camelford, Cornwall, UK) against caspase-1, -3, and -9 and successfully identified 43 initial hit compounds. The 43 hit compounds were further tested for cytoprotective activity against staurosporine-induced cell death in NIH3T3 cells. Nineteen compounds were found to have significant cytoprotective effects in cell viability assays. One of the compounds, RBC1023, was demonstrated to protect NIH3T3 cells from staurosporine-induced caspase-3 cleavage and activation. RBC1023 was also shown to protect against staurosporine-induced impairment of mitochondrial membrane potential. DNA microarray analysis demonstrated that staurosporine treatment induced broad global gene expression alterations, and RBC1023 co-treatment significantly restored these changes, especially of the genes that are related to cell growth and survival signaling such as Egr1, Cdc25c, cdkn3, Rhob, Nek2, and Taok1. Collectively, RBC1023 protects NIH3T3 cells against staurosporine-induced apoptosis via inhibiting caspase activity, restoring mitochondrial membrane potential, and possibly upregulating some cell survival-related gene expressions and pathways. PMID:24920883

  19. Topically applied Hsp90 inhibitor 17AAG inhibits UVR-induced cutaneous squamous cell carcinomas.

    PubMed

    Singh, Anupama; Singh, Ashok; Sand, Jordan M; Bauer, Samuel J; Hafeez, Bilal Bin; Meske, Louise; Verma, Ajit K

    2015-04-01

    We present here that heat-shock protein 90 (Hsp90) inhibitor 17-(allylamino)-17-demethoxygeldanamycin (17AAG), when topically applied to mouse skin, inhibits UVR-induced development of cutaneous squamous cell carcinoma (SCC). In these experiments, DMSO:acetone (1:40 v/v) solution of 17AAG (500?nmol) was applied topically to mouse skin in conjunction with each UVR exposure (1.8?kJ?m(-2)). The UVR source was Kodacel-filtered FS-40 sun lamps (approximately 60% UVB and 40% UVA). In independent experiments with three separate mouse lines (SKH-1 hairless mice, wild-type FVB, and protein kinase C epsilon (PKC?)-overexpressing transgenic FVB mice), 17AAG treatment increased the latency and decreased both the incidence and multiplicity of UVR-induced SCC. Topical 17AAG alone or in conjunction with UVR treatments elicited neither skin nor systemic toxicity. 17AAG-caused inhibition of SCC induction was accompanied by a decrease in UVR-induced (1) hyperplasia, (2) Hsp90?-PKC? interaction, and (3) expression levels of Hsp90?, Stat3, pStat3Ser727, pStat3Tyr705, pAktSer473, and matrix metalloproteinase (MMP). The results presented here indicate that topical Hsp90 inhibitor 17AAG is effective in prevention of UVR-induced epidermal hyperplasia and SCC. One may conclude from the preclinical data presented here that topical 17AAG may be useful for prevention of UVR-induced inflammation and cutaneous SCC either developed in UVR-exposed or organ transplant population. PMID:25337691

  20. Chronic activation of peroxisome proliferator-activated receptor-alpha with fenofibrate prevents alterations in cardiac metabolic phenotype without changing the onset of decompensation in pacing-induced heart failure

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Severe heart failure (HF) is characterized by profound alterations in cardiac metabolic phenotype, with down-regulation of the free fatty acid (FFA) oxidative pathway and marked increase in glucose oxidation. We tested whether fenofibrate, a pharmacological agonist of peroxisome proliferator-activat...

  1. Eribulin synergizes with Polo-like kinase 1 inhibitors to induce apoptosis in rhabdomyosarcoma.

    PubMed

    Stehle, Angelika; Hugle, Manuela; Fulda, Simone

    2015-08-28

    Eribulin, a novel microtubule-interfering drug, was recently shown to exhibit high antitumor activity in vivo against various pediatric cancers. Here, we identify a novel synthetic lethal interaction of Eribulin together with Polo-like kinase 1 (PLK1) inhibitors against rhabdomyosarcoma (RMS) in vitro and in vivo. Eribulin and the PLK1 inhibitor BI 2536 at subtoxic concentrations synergize to induce apoptosis in RMS cells as confirmed by calculation of combination index (CI). Also, Eribulin/BI 2536 co-treatment is significantly more effective than monotherapy to reduce cell viability and inhibit colony formation of RMS cells. Similarly, Eribulin and BI 2536 act in concert to trigger apoptosis in a primary, patient-derived ARMS culture, underscoring the clinical relevance of this combination. Importantly, Eribulin and BI 2536 cooperate to suppress tumor growth in an in vivo model of RMS. On molecular grounds, Eribulin/BI 2536 co-treatment causes profound mitotic arrest, which is critically required for synergism, since inhibition of mitotic arrest by CDK1 inhibitor RO-3306 abolishes Eribulin/BI 2536-mediated apoptosis. Eribulin and BI 2536 cooperate to activate caspase-9, -3 and -8, which is necessary for apoptosis induction, since the broad-range caspase inhibitor N-benzyloxycarbonyl-Val-Ala-Asp-fluoromethylketone (zVAD.fmk) reduces Eribulin/BI 2536-induced apoptosis significantly, yet partially. Intriguingly, knockdown of endonuclease G (ENDOG) also significantly inhibits Eribulin/BI 2536-triggered apoptosis, demonstrating the involvement of both caspase-dependent and -independent effector pathways. Synergistic induction of apoptosis is similarly found for Eribulin/BI 2536 co-treatment in neuroblastoma cells and for the combination of vincristine (another antimicrotubule chemotherapeutic) with Poloxin (another PLK1 inhibitor), thus pointing to a broader significance of this concomitant microtubule- and PLK1-targeting strategy for pediatric oncology. In conclusion, the identification of a novel synthetic lethality by dual targeting of mitosis using microtubule-interfering and PLK1-targeted drugs, i.e. Eribulin and BI 2536, has important implications for the development of more effective treatment strategies for RMS. PMID:25917079

  2. Systematically Studying Kinase Inhibitor Induced Signaling Network Signatures by Integrating Both Therapeutic and Side Effects

    PubMed Central

    Shao, Hongwei; Peng, Tao; Ji, Zhiwei; Su, Jing; Zhou, Xiaobo

    2013-01-01

    Substantial effort in recent years has been devoted to analyzing data based large-scale biological networks, which provide valuable insight into the topologies of complex biological networks but are rarely context specific and cannot be used to predict the responses of cell signaling proteins to specific ligands or compounds. In this work, we proposed a novel strategy to investigate kinase inhibitor induced pathway signatures by integrating multiplex data in Library of Integrated Network-based Cellular Signatures (LINCS), e.g. KINOMEscan data and cell proliferation/mitosis imaging data. Using this strategy, we first established a PC9 cell line specific pathway model to investigate the pathway signatures in PC9 cell line when perturbed by a small molecule kinase inhibitor GW843682. This specific pathway revealed the role of PI3K/AKT in modulating the cell proliferation process and the absence of two anti-proliferation links, which indicated a potential mechanism of abnormal expansion in PC9 cell number. Incorporating the pathway model for side effects on primary human hepatocytes, it was used to screen 27 kinase inhibitors in LINCS database and PF02341066, known as Crizotinib, was finally suggested with an optimal concentration 4.6 uM to suppress PC9 cancer cell expansion while avoiding severe damage to primary human hepatocytes. Drug combination analysis revealed that the synergistic effect region can be predicted straightforwardly based on a threshold which is an inherent property of each kinase inhibitor. Furthermore, this integration strategy can be easily extended to other specific cell lines to be a powerful tool for drug screen before clinical trials. PMID:24339888

  3. Induced resistance to methionyl-tRNA synthetase inhibitors in Trypanosoma brucei is due to overexpression of the target.

    PubMed

    Ranade, Ranae M; Gillespie, J Robert; Shibata, Sayaka; Verlinde, Christophe L M J; Fan, Erkang; Hol, Wim G J; Buckner, Frederick S

    2013-07-01

    New classes of antiparasitic drugs active against Trypanosoma brucei are needed to combat human African trypanosomiasis. Inhibitors of methionyl-tRNA synthetase (MetRS) have excellent potential to be developed for this purpose (S. Shibata, J. R. Gillespie, A. M. Kelley, A. J. Napuli, Z. Zhang, K. V. Kovzun, R. M. Pefley, J. Lam, F. H. Zucker, W. C. Van Voorhis, E. A. Merritt, W. G. Hol, C. L. Verlinde, E. Fan, and F. S. Buckner, Antimicrob. Agents Chemother. 55:1982-1989, 2011). In order to assess the potential for resistance to develop against this new class of inhibitors, T. brucei cultures were grown in the presence of MetRS inhibitors or comparison drugs. Resistance up to ?50 times the baseline 50% inhibitory concentration (IC50) was induced against a MetRS inhibitor after ?120 days. A similar level of resistance to the clinical drug eflornithine was induced after ?50 days and for pentamidine after ?80 days. Thus, resistance was induced more slowly against MetRS inhibitors than against clinically used drugs. The parasites resistant to the MetRS inhibitor were shown to overexpress MetRS mRNA by a factor of 35 over the parental strain. Southern analysis indicated that the MetRS gene was amplified in the genome by nearly 8-fold. When injected into mice, the MetRS inhibitor-resistant parasites caused a reduced level of infection, indicating that the changes associated with resistance attenuated their virulence. This finding and the fact that resistance to MetRS inhibitors developed relatively slowly are encouraging for further development of this class of compounds. Published studies on other antitrypanosomal drugs have primarily shown that alterations in membrane transporters were the mechanisms responsible for resistance. This is the first published report of induced drug resistance in the African trypanosome due to overexpression of the target enzyme. PMID:23587950

  4. Induced Resistance to Methionyl-tRNA Synthetase Inhibitors in Trypanosoma brucei Is Due to Overexpression of the Target

    PubMed Central

    Ranade, Ranae M.; Gillespie, J. Robert; Shibata, Sayaka; Verlinde, Christophe L. M. J.; Fan, Erkang; Hol, Wim G. J.

    2013-01-01

    New classes of antiparasitic drugs active against Trypanosoma brucei are needed to combat human African trypanosomiasis. Inhibitors of methionyl-tRNA synthetase (MetRS) have excellent potential to be developed for this purpose (S. Shibata, J. R. Gillespie, A. M. Kelley, A. J. Napuli, Z. Zhang, K. V. Kovzun, R. M. Pefley, J. Lam, F. H. Zucker, W. C. Van Voorhis, E. A. Merritt, W. G. Hol, C. L. Verlinde, E. Fan, and F. S. Buckner, Antimicrob. Agents Chemother. 55:1982–1989, 2011). In order to assess the potential for resistance to develop against this new class of inhibitors, T. brucei cultures were grown in the presence of MetRS inhibitors or comparison drugs. Resistance up to ?50 times the baseline 50% inhibitory concentration (IC50) was induced against a MetRS inhibitor after ?120 days. A similar level of resistance to the clinical drug eflornithine was induced after ?50 days and for pentamidine after ?80 days. Thus, resistance was induced more slowly against MetRS inhibitors than against clinically used drugs. The parasites resistant to the MetRS inhibitor were shown to overexpress MetRS mRNA by a factor of 35 over the parental strain. Southern analysis indicated that the MetRS gene was amplified in the genome by nearly 8-fold. When injected into mice, the MetRS inhibitor-resistant parasites caused a reduced level of infection, indicating that the changes associated with resistance attenuated their virulence. This finding and the fact that resistance to MetRS inhibitors developed relatively slowly are encouraging for further development of this class of compounds. Published studies on other antitrypanosomal drugs have primarily shown that alterations in membrane transporters were the mechanisms responsible for resistance. This is the first published report of induced drug resistance in the African trypanosome due to overexpression of the target enzyme. PMID:23587950

  5. Pharmacologic Profiling of Phosphoinositide 3-Kinase Inhibitors as Mitigators of Ionizing Radiation–Induced Cell Death

    PubMed Central

    Sharlow, Elizabeth R.; Epperly, Michael W.; Lira, Ana; Leimgruber, Stephanie; Skoda, Erin M.; Wipf, Peter; Greenberger, Joel S.

    2013-01-01

    Ionizing radiation (IR) induces genotoxic stress that triggers adaptive cellular responses, such as activation of the phosphoinositide 3-kinase (PI3K)/Akt signaling cascade. Pluripotent cells are the most important population affected by IR because they are required for cellular replenishment. Despite the clear danger to large population centers, we still lack safe and effective therapies to abrogate the life-threatening effects of any accidental or intentional IR exposure. Therefore, we computationally analyzed the chemical structural similarity of previously published small molecules that, when given after IR, mitigate cell death and found a chemical cluster that was populated with PI3K inhibitors. Subsequently, we evaluated structurally diverse PI3K inhibitors. It is remarkable that 9 of 14 PI3K inhibitors mitigated ?IR-induced death in pluripotent NCCIT cells as measured by caspase 3/7 activation. A single intraperitoneal dose of LY294002 [2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one], administered to mice at 4 or 24 hours, or PX-867 [(4S,4aR,5R,6aS,9aR,Z)-11-hydroxy-4-(methoxymethyl)-4a,6a-dimethyl-2,7,10-trioxo-1-(pyrrolidin-1-ylmethylene)-1,2,4,4a,5,6,6a,7,8,9,9a,10-dodecahydroindeno[4,5-H]isochromen-5-yl acetate (CID24798773)], administered 4 hours after a lethal dose of ?IR, statistically significantly (P < 0.02) enhanced in vivo survival. Because cell cycle checkpoints are important regulators of cell survival after IR, we examined cell cycle distribution in NCCIT cells after ?IR and PI3K inhibitor treatment. LY294002 and PX-867 treatment of nonirradiated cells produced a marked decrease in S phase cells with a concomitant increase in the G1 population. In irradiated cells, LY294002 and PX-867 treatment also decreased S phase and increased the G1 and G2 populations. Treatment with LY294002 or PX-867 decreased ?IR-induced DNA damage as measured by ?H2AX, suggesting reduced DNA damage. These results indicate pharmacologic inhibition of PI3K after IR abrogated cell death. PMID:24068833

  6. Three peroxisome protein packaging pathways suggested by selective permeabilization of yeast mutants defective in peroxisome biogenesis.

    PubMed Central

    Zhang, J W; Luckey, C; Lazarow, P B

    1993-01-01

    We have identified five complementation groups of peroxisome biogenesis (peb) mutants in Saccharomyces cerevisiae by a positive selection procedure. Three of these contained morphologically recognizable peroxisomes, and two appeared to lack the organelle altogether. The packaging of peroxisomal proteins in these mutants has been analyzed with a new gentle cell fractionation procedure. It employs digitonin titration for the selective permeabilization of yeast plasma and intracellular membranes. Proteins were measured by enzymatic assay or by quantitative chemiluminescent immunoblotting. With this gentle fractionation method, it was demonstrated that two mutants are selectively defective in assembling proteins into peroxisomes. Peb1-1 packages catalase and acyl-CoA oxidase within peroxisomes but not thiolase. Peb5-1 packages thiolase and acyl-CoA oxidase within peroxisomes but not catalase. The data suggest that the peroxisome biogenesis machinery contains components that are specific for each of three classes of peroxisomal proteins, represented by catalase, thiolase, and acyl-CoA oxidase. In the two mutants lacking morphologically recognizable peroxisomes, peb2-1 and peb4-1, all three enzymes were mislocalized to the cytosol. Images PMID:7909460

  7. Phenotypic Screening Identifies Protein Synthesis Inhibitors as H-Ras-Nanocluster-Increasing Tumor Growth Inducers.

    PubMed

    Najumudeen, Arafath K; Posada, Itziar M D; Lectez, Benoit; Zhou, Yong; Landor, Sebastian K-J; Fallarero, Adyary; Vuorela, Pia; Hancock, John; Abankwa, Daniel

    2015-12-15

    Ras isoforms H-, N-, and K-ras are each mutated in specific cancer types at varying frequencies and have different activities in cell fate control. On the plasma membrane, Ras proteins are laterally segregated into isoform-specific nanoscale signaling hubs, termed nanoclusters. As Ras nanoclusters are required for Ras signaling, chemical modulators of nanoclusters represent ideal candidates for the specific modulation of Ras activity in cancer drug development. We therefore conducted a chemical screen with commercial and in-house natural product libraries using a cell-based H-ras-nanoclustering FRET assay. Next to established Ras inhibitors, such as a statin and farnesyl-transferase inhibitor, we surprisingly identified five protein synthesis inhibitors as positive regulators. Using commonly employed cycloheximide as a representative compound, we show that protein synthesis inhibition increased nanoclustering and effector recruitment specifically of active H-ras but not of K-ras. Consistent with these data, cycloheximide treatment activated both Erk and Akt kinases and specifically promoted H-rasG12V-induced, but not K-rasG12V-induced, PC12 cell differentiation. Intriguingly, cycloheximide increased the number of mammospheres, which are enriched for cancer stem cells. Depletion of H-ras in combination with cycloheximide significantly reduced mammosphere formation, suggesting an exquisite synthetic lethality. The potential of cycloheximide to promote tumor cell growth was also reflected in its ability to increase breast cancer cell tumors grown in ovo. These results illustrate the possibility of identifying Ras-isoform-specific modulators using nanocluster-directed screening. They also suggest an unexpected feedback from protein synthesis inhibition to Ras signaling, which might present a vulnerability in certain tumor cell types. PMID:26568031

  8. Efficacy of Rho kinase inhibitor on cognitive impairment induced by chronic cerebral hypoperfusion in rats

    PubMed Central

    Zhang, Qiang; Zhang, Jun-Jian; Han, Zhong-Mou

    2015-01-01

    This work aims to explore the efficacy of Rho kinase inhibitor Fasudil on cognitive impairment induced by chronic cerebral hypoperfusion in rats. A total of 32 male adult Sprague Dawley (SD) rats were randomly divided into three groups: treatment group, control group and sham-operated group for severe carotid artery stenosis model. After two weeks, 8.35 mg/kg Fasudil and physiological saline were intraperitoneally applied twice per day in treatment group and control group, respectively. Morris water maze test was performed in each group to detect the changes of cognitive function and observe the hippocampal pathomorphology in rats after eight weeks. The average escape latency distinctly shortened (P < 0.01) and the percentage of swimming distance in the platform quadrant significantly increased (P < 0.01) in treatment group compared with those at corresponding time points in control group. The rate of carotid artery stenosis in rats had no statistical difference between treatment and control groups (P > 0.05). Fasudil effectively improved hippocampal pathomorphology. Rho kinase inhibitor obviously ameliorated cognitive impairment induced by chronic cerebral hypoperfusion in rats. PMID:25932185

  9. Farnesyltransferase Inhibitor, Tipifarnib, Prevents Galactosamine/Lipopolysaccharide-Induced Acute Liver Failure

    PubMed Central

    Shirozu, Kazuhiro; Hirai, Shuichi; Tanaka, Tomokazu; Hisaka, Shinsuke

    2014-01-01

    Acute liver failure (ALF) is a fatal syndrome associated with massive hepatocyte death. There is no cure for ALF except liver transplantation. Protein farnesylation is a lipid modification of cysteine residues that is catalyzed by farnesyltransferase (FTase) and has been proposed as an integral component of acute inflammation. Previously, we have demonstrated that FTase inhibitors improve survival in mouse models of endotoxemia and sepsis. Here we studied the effects of FTase inhibitor, tipifarnib, on galactosamine (GalN) /lipopolysaccharide (LPS)-induced acute liver failure. The effects of tipifarnib (10 mg/kg, IP) were studied in GalN (400 mg/kg, IP) and LPS (3 ?g/kg)-challenged mice by histological and biochemical analyses. GalN/LPS administration caused prominent liver injury characterized by the increased plasma alanine aminotransferase (ALT) and aspartic aminotransferase (AST) levels leading to significant mortality in mice. Tipifarnib inhibited GalN/LPS-induced caspase 3 activation, inflammatory cytokine production, and c-Jun N-terminal Kinase (JNK) phosphorylation in the liver. On the other hand, Tipifarnib upregulated anti-apoptotic protein, Bcl-xL, in the liver after GalN/LPS challenge. Tipifarnib also protected primary hepatocytes from GalN/tumor necrosis factor-? (TNF-?)-induced cell death by inhibiting caspase 3 activation and upregulating anti-apoptotic proteins. GalN/LPS-induced liver injury was associated with increased protein farnesylation in the liver. Tipifarnib prevented protein farnesylation in the liver and markedly attenuated liver injury and mortality in GalN/LPS-challenged mice. These results suggest that protein farnesylation is a novel potential molecular target to prevent hepatocyte death and acute inflammatory liver failure in fulminant hepatitis. PMID:25046541

  10. Effect of the Angiotensin I Converting Enzyme Inhibitor, MK-421, on Experimentally Induced Drinking

    NASA Technical Reports Server (NTRS)

    Fregley, Melvin J.; Fater, Dennis C.; Greenleaf, John E.

    1982-01-01

    MK-421, the ethyl ester maleate salt of N-(S)-1-(ethoxycarbonyl)-3-phenyl-propyl- Ala-L-Pro, is an angiotensin I converting enzyme inhibitor. An initial objective was to determine whether MK-421, administered at 0, 2.5, 5.0, 10.0, 20.0 and 40.0 mg/kg, ip to 96 female rats 15 min prior to administration of the beta-adrenergic agonist, isoproterenol (25 microgram/kg, ip), would inhibit the drinking induced by isoproterenol during 2 h after its administration. The water intake induced by isoproterenol was inhibited significantly by 2.5 mg MK-421/kg. When a similar experiment was performed using Angiotensin I (AI) (200 microgram/kg, ip) as the dipsogenic agent, MK-421 (5 mg/kg, ip), administered 15 min prior to AI, inhibited significantly both the dipsogenic and the diuretic effect of AI. However, administration of angiotensin II (AII, 200 microgram/kg, ip) 15 min after MK-421 (5mg/kg) was accompanied by a water intake that did not differ from AII alone. The drink induced by ip administration of 1.0 m NaCl solution (1% of body wt, ip) was not inhibited by administration of MK-421 (5 mg/kg) 15 min prior to allowing access to water while the drink induced by a 24 h dehydration was partially inhibited. Thus, the drinks induced by administraition of either isoproterenol or AI are dependent on formation of AII. That induced by dehydration is partially dependent, while that induced by hypertonic siilinc is independent of the formation of AII.

  11. A rat retinal damage model predicts for potential clinical visual disturbances induced by Hsp90 inhibitors

    SciTech Connect

    Zhou, Dan; Liu, Yuan; Ye, Josephine; Ying, Weiwen; Ogawa, Luisa Shin; Inoue, Takayo; Tatsuta, Noriaki; Wada, Yumiko; Koya, Keizo; Huang, Qin; Bates, Richard C.; Sonderfan, Andrew J.

    2013-12-01

    In human trials certain heat shock protein 90 (Hsp90) inhibitors, including 17-DMAG and NVP-AUY922, have caused visual disorders indicative of retinal dysfunction; others such as 17-AAG and ganetespib have not. To understand these safety profile differences we evaluated histopathological changes and exposure profiles of four Hsp90 inhibitors, with or without clinical reports of adverse ocular effects, using a rat retinal model. Retinal morphology, Hsp70 expression (a surrogate marker of Hsp90 inhibition), apoptotic induction and pharmacokinetic drug exposure analysis were examined in rats treated with the ansamycins 17-DMAG and 17-AAG, or with the second-generation compounds NVP-AUY922 and ganetespib. Both 17-DMAG and NVP-AUY922 induced strong yet restricted retinal Hsp70 up-regulation and promoted marked photoreceptor cell death 24 h after the final dose. In contrast, neither 17-AAG nor ganetespib elicited photoreceptor injury. When the relationship between drug distribution and photoreceptor degeneration was examined, 17-DMAG and NVP-AUY922 showed substantial retinal accumulation, with high retina/plasma (R/P) ratios and slow elimination rates, such that 51% of 17-DMAG and 65% of NVP-AUY922 present at 30 min post-injection were retained in the retina 6 h post-dose. For 17-AAG and ganetespib, retinal elimination was rapid (90% and 70% of drugs eliminated from the retina at 6 h, respectively) which correlated with lower R/P ratios. These findings indicate that prolonged inhibition of Hsp90 activity in the eye results in photoreceptor cell death. Moreover, the results suggest that the retina/plasma exposure ratio and retinal elimination rate profiles of Hsp90 inhibitors, irrespective of their chemical class, may predict for ocular toxicity potential. - Highlights: • In human trials some Hsp90 inhibitors cause visual disorders, others do not. • Prolonged inhibition of Hsp90 in the rat eye results in photoreceptor cell death. • Retina/plasma ratio and retinal elimination rate are linked to toxicity potential. • Rat retinotoxic responses to individual Hsp90 inhibitors reflect clinical profiles. • Rodent modeling may be used to assess ocular risks of targeted Hsp90 compounds.

  12. Microporation is an efficient method for siRNA-induced knockdown of PEX5 in HepG2 cells: evaluation of the transfection efficiency, the PEX5 mRNA and protein levels and induction of peroxisomal deficiency.

    PubMed

    Ahlemeyer, Barbara; Vogt, Julia-Franziska; Michel, Vera; Hahn-Kohlberger, Petra; Baumgart-Vogt, Eveline

    2014-11-01

    The pathomechanism of peroxisomal biogenesis disorders (PBDs), a group of inherited autosomal recessive diseases with mutations of peroxin (PEX) genes, is not yet fully understood. Therefore, several knockout models, e.g., the PEX5 knockout mouse, have been generated exhibiting a complete loss of peroxisomal function. In this study, we wanted to knockdown PEX5 using the siRNA technology (1) to mimic milder forms of PBDs in which the mutated peroxin has some residual function and (2) to analyze the cellular consequences of a reduction of the PEX5 protein without adaption during the development as it is the case in a knockout animal. First, we tried to optimize the transfection of the hepatoma cell line HepG2 with PEX5 siRNA using different commercially available liposomal and non-liposomal transfection reagents (Lipofectamine(®) 2000, FuGENE 6, HiPerFect(®), INTERFERin™, RiboJuice™) as well as microporation using the Neon™ Transfection system. Microporation was found to be superior to the transfection reagents with respect to the transfection efficiency (100 vs. 0-70%), to the reduction of PEX5 mRNA (by 90 vs. 0-50%) and PEX5 protein levels (by 70 vs. 0-50%). Interestingly, we detected that a part of the cleaved PEX5 mRNA still existed as 3' fragment (15%) 24 h after microporation. Using microporation, we further analyzed whether the reduced PEX5 protein level impaired peroxisomal function. We indeed detected a reduced targeting of SKL-tagged proteins into peroxisomes as well as an increased oxidative stress as found in PBD patients and respective knockout mouse models. Knockdown of the PEX5 protein and functional consequences were at a maximum 48 h after microporation. Thereafter, the PEX5 protein was resynthesized, which may allow the temporal analysis of the loss as well as the reconstitution of peroxisomes in the future. In conclusion, we propose microporation as an efficient and reproducible method to transfect HepG2 cells with PEX5 siRNA. We succeeded to transiently knockdown PEX5 mRNA and its protein level leading to functional consequences similar as observed in peroxisome deficiencies. PMID:25224142

  13. Inter-? inhibitor protein and its associated glycosaminoglycans protect against histone-induced injury.

    PubMed

    Chaaban, Hala; Keshari, Ravi S; Silasi-Mansat, Robert; Popescu, Narcis I; Mehta-D'Souza, Padmaja; Lim, Yow-Pin; Lupu, Florea

    2015-04-01

    Extracellular histones are mediators of tissue injury and organ dysfunction; therefore they constitute potential therapeutic targets in sepsis, inflammation, and thrombosis. Histone cytotoxicity in vitro decreases in the presence of plasma. Here, we demonstrate that plasma inter-? inhibitor protein (IAIP) neutralizes the cytotoxic effects of histones and decreases histone-induced platelet aggregation. These effects are mediated through the negatively charged glycosaminoglycans (GAGs) chondroitin sulfate and high-molecular-weight hyaluronan (HMW-HA) associated with IAIP. Cell surface anionic glycosaminoglycans heparan sulfate and HA protect the cells against histone-mediated damage in vitro. Surface plasmon resonance showed that both IAIP and HMW-HA directly bind to recombinant histone H4. In vivo neutralization of histones with IAIP and HMW-HA prevented histone-induced thrombocytopenia, bleeding, and lung microvascular thrombosis, decreased neutrophil activation, and averted histone-induced production of inflammatory cytokines and chemokines. IAIP and HMW-HA colocalized with histones in necrotic tissues and areas that displayed neutrophil extracellular traps. Increasing amounts of IAIP-histone complexes detected in the plasma of septic baboons correlated with increase in histones and/or nucleosomes and consumption of plasma IAIP. Our data suggest that IAIP, chondroitin sulfate, and HMW-HA are potential therapeutic agents to protect against histone-induced cytotoxicity, coagulopathy, systemic inflammation, and organ damage during inflammatory conditions such as sepsis and trauma. PMID:25631771

  14. Relaxation of rat thoracic aorta induced by the Ca(2+)-ATPase inhibitor, cyclopiazonic acid, possibly through nitric oxide formation.

    PubMed Central

    Moritoki, H.; Hisayama, T.; Takeuchi, S.; Kondoh, W.; Imagawa, M.

    1994-01-01

    1 The effect of the Ca(2+)-ATPase inhibitor, cyclopiazonic acid (CPA), was studied on rat thoracic aortic ring preparations. 2 At concentrations above 0.3 microM, CPA induced relaxation in the arteries precontracted with phenylephrine. Removal of the endothelium abolished CPA-induced relaxation. 3 The nitric oxide (NO) synthase inhibitor NG-nitro L-arginine (3-300 microM), the free radical scavenger haemoglobin (0.1-3 microM), the soluble guanylate cyclase inhibitor, LY83583 (0.1-10 microM), each inhibited the endothelium-dependent relaxation to CPA. The potassium channel blocker, glibenclamide (10 microM) and cyclo-oxygenase inhibitor, indomethacin (100 microM for 60 min and then washed out) did not alter the action of CPA. 4 The calmodulin inhibitors calmidazolium (3-10 microM) and W-7 (100 microM) also abolished CPA-induced relaxation. 5 CPA (10 microM) increased guanosine 3':5'-cyclic monophosphate (cyclic GMP) levels in arteries with an intact endothelium, without affecting adenosine 3':5'-cyclic monophosphate (cyclic AMP) levels. 6 The inhibitors of NO synthesis and actions, the calmodulin inhibitor and removal of the endothelium abolished the CPA-stimulated increase in the levels of cyclic GMP. 7 In Ca(2+)-free solution, CPA failed to induce relaxation or to stimulate cyclic GMP production. Relaxation to nitroprusside was not affected under these conditions. 8 These results suggest that CPA can stimulate NO synthesis, possibly by inhibiting a Ca(2+)-ATPase, which replenishes Ca2+ in the intracellular storage sites in endothelial cells. Depletion of the Ca2+ store in the endothelium may then trigger influx of extracellular Ca2+, contributing to an increase in free Ca2+ in the endothelial cells, which activates NO synthase and NO formation. PMID:7517325

  15. An inventory of peroxisomal proteins and pathways in Drosophila melanogaster

    PubMed Central

    Faust, Joseph E.; Verma, Avani; Peng, Chengwei; McNew, James A.

    2012-01-01

    Peroxisomes are ubiquitous organelles housing a variety of essential biochemical pathways. Peroxisome dysfunction causes a spectrum of human diseases known as peroxisome biogenesis disorders (PBD). While much is known regarding the mechanism of peroxisome biogenesis, it is still unclear how peroxisome dysfunction leads to the disease state. Several recent studies have shown that mutations in Drosophila peroxin genes cause phenotypes similar to those seen in humans with PBDs suggesting that Drosophila might be a useful system to model PBDs. We have analyzed the proteome of Drosophila to identify the proteins involved in peroxisomal biogenesis and homeostasis as well as metabolic enzymes that function within the organelle. The subcellular localization of five of these predicted peroxisomal proteins was confirmed. Similar to C. elegans, Drosophila appears to only utilize the peroxisome targeting signal (PTS) type 1 system for matrix protein import. This work will further our understanding of peroxisomes in Drosophila and add to the usefulness of this emerging model system. PMID:22758915

  16. Peroxisome proliferator-activated receptor gamma regulates expression of signal transducer and activator of transcription 5A

    SciTech Connect

    Olsen, Hanne; Haldosen, Lars-Arne . E-mail: Lars-Arne.Haldosen@mednut.ki.se

    2006-05-01

    Signal transducer and activator of transcription 5A (STAT5A) has been shown to be important for terminal differentiation of mammary epithelial cells. In order to understand regulation of expression of STAT5A, the 5' end of the mouse Stat5a gene was isolated. Putative regulatory elements was searched for and several peroxisome proliferator response elements (PPREs) were found, one with high (12/13 nucleotides) and three with less (8-10/13) similarity to the reported consensus sequence. Mouse mammary epithelial HC11 cells were treated with peroxisome proliferator-activated receptor {gamma} (PPAR{gamma}) ligand, the thiazolidinedione (TZD) troglitazone, and an increase in STAT5A protein expression was seen. The 5' flank of Stat5a gene was cloned in a luciferase reporter vector. A concentration dependent activation of the STAT5A-luciferase reporter was detected, when transiently transfected HC11 cells were treated with TZD. The activation could be inhibited by treatment with a PPAR{gamma} antagonist. It has earlier been shown that epidermal growth factor (EGF) induces MAPK phosphorylation of PPAR{gamma} resulting in a less transcriptionally active receptor. In HC11 cells, EGF inhibited TZD induced STAT5A-reporter activity suggesting that our previously reported EGF-mediated suppression of STAT5A expression is mediated in all or partly through inhibition of PPAR{gamma} activity. Furthermore, the MEK inhibitor PD98059 inhibited the EGF effect. All together, data presented suggest that PPAR{gamma} participates in regulation of STAT5A expression.

  17. Inhibition of chlorine-induced lung injury by the type 4 phosphodiesterase inhibitor rolipram

    SciTech Connect

    Chang, Weiyuan; Chen, Jing; Schlueter, Connie F.; Rando, Roy J.; Pathak, Yashwant V.; Hoyle, Gary W.

    2012-09-01

    Chlorine is a highly toxic respiratory irritant that when inhaled causes epithelial cell injury, alveolar-capillary barrier disruption, airway hyperreactivity, inflammation, and pulmonary edema. Chlorine is considered a chemical threat agent, and its release through accidental or intentional means has the potential to result in mass casualties from acute lung injury. The type 4 phosphodiesterase inhibitor rolipram was investigated as a rescue treatment for chlorine-induced lung injury. Rolipram inhibits degradation of the intracellular signaling molecule cyclic AMP. Potential beneficial effects of increased cyclic AMP levels include inhibition of pulmonary edema, inflammation, and airway hyperreactivity. Mice were exposed to chlorine (whole body exposure, 228–270 ppm for 1 h) and were treated with rolipram by intraperitoneal, intranasal, or intramuscular (either aqueous or nanoemulsion formulation) delivery starting 1 h after exposure. Rolipram administered intraperitoneally or intranasally inhibited chlorine-induced pulmonary edema. Minor or no effects were observed on lavage fluid IgM (indicative of plasma protein leakage), KC (Cxcl1, neutrophil chemoattractant), and neutrophils. All routes of administration inhibited chlorine-induced airway hyperreactivity assessed 1 day after exposure. The results of the study suggest that rolipram may be an effective rescue treatment for chlorine-induced lung injury and that both systemic and targeted administration to the respiratory tract were effective routes of delivery. -- Highlights: ? Chlorine causes lung injury when inhaled and is considered a chemical threat agent. ? Rolipram inhibited chlorine-induced pulmonary edema and airway hyperreactivity. ? Post-exposure rolipram treatments by both systemic and local delivery were effective. ? Rolipram shows promise as a rescue treatment for chlorine-induced lung injury.

  18. Alternative lengthening of telomeres is induced by telomerase inhibitors in Barrett's esophageal cells.

    PubMed

    Qin, Xiuguang; Qi, Bo; Zhao, Baosheng

    2013-04-01

    A crucial step in the path to the malignant transformation of cells and tumor formation is immortalization, which essentially depends on telomere maintenance. The aim of this study was to investigate the role of telomerase in the progression of Barrett's esophagus. Telomerase activity was measured in Barrett's cells using terminal restriction fragment (TRF) analysis. Telomere length was measured using Q-FISH analysis. Furthermore, the telomere recombination events were detected between sister chromatids using chromosome orientation FISH (CO-FISH). There was a reduction in telomerase activity in the CP-A cells transduced with MT-hTER/47A+siRNA, which led to an almost complete disappearance of telomerase activity. The telomere length of the CP-A cells transduced with MT-hTER/47A+siRNA was slightly shorter compared to that of the untransduced cells. The telomerase-inhibited cells were morphologically indistinguishable from those untransduced and WT-hter-transduced cells. In the control cells, the growth rate was between 0.9 to 1.1 with the population doubling per day. Although the transduction of the telomerase inhibitors in the CP-A cells did not cause a significant reduction in cell growth, these transduced cells grew generally slower compared with the control cells. The heterogeneous telomere length was also be detected in the telomerase-inhibited CP-A cells. However, the telomere length remained homogeneous in the control cells. The metaphase of the CP-A cells transduced with MT-hTER/47A+siRNA demonstrated 70% heterogeneous telomeres. In addition, no increased recombination was observed between sister chromatids in the transduced CP-A cells compared with the control cells. Our findings suggest that an alternative lengthening of telomeres (ALT) may be induced by telomerase inhibitors in CP-A cells. Therefore, telomerase inhibitors may exhibit high potency in the treatment of esophageal adenocarcinoma arising from Barrett's esophagus. PMID:23338150

  19. Angiotensin-converting enzyme inhibitor (enalapril maleate) accelerates recovery of mouse skin from UVB-induced wrinkles

    SciTech Connect

    Matsuura-Hachiya, Yuko; Arai, Koji Y.; Ozeki, Rieko; Kikuta, Ayako; Nishiyama, Toshio

    2013-12-06

    Highlights: •Angiotensin converting enzyme (ACE) increases in UVB-irradiated skin. •Administration of an ACE inhibitor improved UVB-induced skin wrinkle. •ACE inhibitor improved UVB-induced epidermal hypertrophy. •ACE inhibitor improved transepidermal water loss in the UVB-irradiated skin. -- Abstract: Angiotensin-converting enzyme (ACE) activity and angiotensin II signaling regulate cell proliferation, differentiation, and tissue remodeling, as well as blood pressure, while in skin, angiotensin II signaling is involved in wound healing, inflammation, and pathological scar formation. Therefore, we hypothesized that angiotensin II is also involved in photoaging of skin. In this study, we examined the effect of enalapril maleate, an ACE inhibitor, on recovery of wrinkled skin of hairless mice exposed to long-term UVB irradiation. Immunohistochemical observation revealed that expression of ACE, angiotensin II, and angiotensin II type 1 (AT1) and type 2 (AT2) receptors in the skin was increased after UVB irradiation (3 times/week at increasing intensities for 8 weeks). Administration of enalapril maleate (5 times/week for 6 weeks, starting 1 week after 10-week irradiation) accelerated recovery from UVB-induced wrinkles, epidermal hyperplasia and epidermal barrier dysfunction, as compared with the vehicle control. Our results indicate that ACE and angiotensin II activity are involved in skin photoaging, and suggest that ACE inhibitor such as enalapril maleate may have potential for improvement of photoaged skin.

  20. Optimized Plk1 PBD Inhibitors Based on Poloxin Induce Mitotic Arrest and Apoptosis in Tumor Cells.

    PubMed

    Scharow, Andrej; Raab, Monika; Saxena, Krishna; Sreeramulu, Sridhar; Kudlinzki, Denis; Gande, Santosh; Dötsch, Christina; Kurunci-Csacsko, Elisabeth; Klaeger, Susan; Kuster, Bernhard; Schwalbe, Harald; Strebhardt, Klaus; Berg, Thorsten

    2015-11-20

    Polo-like kinase 1 (Plk1) is a central regulator of mitosis and has been validated as a target for antitumor therapy. The polo-box domain (PBD) of Plk1 regulates its kinase activity and mediates the subcellular localization of Plk1 and its interactions with a subset of its substrates. Functional inhibition of the Plk1 PBD by low-molecular weight inhibitors has been shown to represent a viable strategy by which to inhibit the enzyme, while avoiding selectivity issues caused by the conserved nature of the ATP binding site. Here, we report structure-activity relationships and mechanistic analysis for the first reported Plk1 PBD inhibitor, Poloxin. We present the identification of the optimized analog Poloxin-2, displaying significantly improved potency and selectivity over Poloxin. Poloxin-2 induces mitotic arrest and apoptosis in cultured human tumor cells at low micromolar concentrations, highlighting it as a valuable tool compound for exploring the function of the Plk1 PBD in living cells. PMID:26279064

  1. Selectively targeting an inactive conformation of interleukin-2-inducible T-cell kinase by allosteric inhibitors.

    PubMed

    Han, Seungil; Czerwinski, Robert M; Caspers, Nicole L; Limburg, David C; Ding, WeiDong; Wang, Hong; Ohren, Jeffrey F; Rajamohan, Francis; McLellan, Thomas J; Unwalla, Ray; Choi, Chulho; Parikh, Mihir D; Seth, Nilufer; Edmonds, Jason; Phillips, Chris; Shakya, Subarna; Li, Xin; Spaulding, Vikki; Hughes, Samantha; Cook, Andrew; Robinson, Colin; Mathias, John P; Navratilova, Iva; Medley, Quintus G; Anderson, David R; Kurumbail, Ravi G; Aulabaugh, Ann

    2014-06-01

    ITK (interleukin-2-inducible T-cell kinase) is a critical component of signal transduction in T-cells and has a well-validated role in their proliferation, cytokine release and chemotaxis. ITK is an attractive target for the treatment of T-cell-mediated inflammatory diseases. In the present study we describe the discovery of kinase inhibitors that preferentially bind to an allosteric pocket of ITK. The novel ITK allosteric site was characterized by NMR, surface plasmon resonance, isothermal titration calorimetry, enzymology and X-ray crystallography. Initial screening hits bound to both the allosteric pocket and the ATP site. Successful lead optimization was achieved by improving the contribution of the allosteric component to the overall inhibition. NMR competition experiments demonstrated that the dual-site binders showed higher affinity for the allosteric site compared with the ATP site. Moreover, an optimized inhibitor displayed non-competitive inhibition with respect to ATP as shown by steady-state enzyme kinetics. The activity of the isolated kinase domain and auto-activation of the full-length enzyme were inhibited with similar potency. However, inhibition of the activated full-length enzyme was weaker, presumably because the allosteric site is altered when ITK becomes activated. An optimized lead showed exquisite kinome selectivity and is efficacious in human whole blood and proximal cell-based assays. PMID:24593284

  2. Reversal of boswellic acid analog BA145 induced caspase dependent apoptosis by PI3K inhibitor LY294002 and MEK inhibitor PD98059.

    PubMed

    Pathania, Anup S; Joshi, Amit; Kumar, Suresh; Guru, Santosh K; Bhushan, Shashi; Sharma, Parduman R; Bhat, Wajid W; Saxena, Ajit K; Singh, Jaswant; Shah, Bhahwal A; Andotra, Samar S; Taneja, Subhash C; Malik, Fayaz A; Kumar, Ajay

    2013-12-01

    PI3K/Akt and ERK pathways are important for growth and proliferation of many types of cancers. Therefore, PI3K inhibitor LY294002 (LY) and MEK1/2 inhibitor PD98059 (PD) are used to sensitize many types of cancer cell lines to chemotherapeutic agents, where AKT and ERK pathways are over activated. However, in this study, we show for the first time that PD could protect the leukemia cells independent of ERK pathway inhibition, besides, we also report a detailed mechanism for antiapoptotic effect of LY in HL-60 cells against the cytotoxicity induced by a boswellic acid analog BA145. Apoptosis induced by BA145 is accompanied by downregulation of PI3K/Akt and ERK pathways in human myelogenous leukemia HL-60 cells, having activating N-Ras mutation. Both LY and PD protected the cells against mitochondrial stress caused by BA145, and reduced the release of cytochrome c and consequent activation of caspase-9. LY and PD also diminished the activation of caspase-8 without affecting the death receptors. Besides, LY and PD also reversed the caspase dependent DNA damage induced by BA145. Further studies revealed that LY and PD significantly reversed the inhibitory effect of BA145 on cell cycle regulatory proteins by upregulating hyperphosphorylated retinoblastoma, pRB (S795) and downregulating p21 and cyclin E. More importantly, all these events were reversed by caspase inhibition by Z-VAD-fmk, suggesting that both LY and PD act at the level of caspases to diminish the apoptosis induced by BA145. These results indicate that inhibitors of PI3K/Akt and ERK pathways can play dual role and act against chemotherapeutic agents. PMID:23948751

  3. Redox interplay between mitochondria and peroxisomes.

    PubMed

    Lismont, Celien; Nordgren, Marcus; Van Veldhoven, Paul P; Fransen, Marc

    2015-01-01

    Reduction-oxidation or "redox" reactions are an integral part of a broad range of cellular processes such as gene expression, energy metabolism, protein import and folding, and autophagy. As many of these processes are intimately linked with cell fate decisions, transient or chronic changes in cellular redox equilibrium are likely to contribute to the initiation and progression of a plethora of human diseases. Since a long time, it is known that mitochondria are major players in redox regulation and signaling. More recently, it has become clear that also peroxisomes have the capacity to impact redox-linked physiological processes. To serve this function, peroxisomes cooperate with other organelles, including mitochondria. This review provides a comprehensive picture of what is currently known about the redox interplay between mitochondria and peroxisomes in mammals. We first outline the pro- and antioxidant systems of both organelles and how they may function as redox signaling nodes. Next, we critically review and discuss emerging evidence that peroxisomes and mitochondria share an intricate redox-sensitive relationship and cooperate in cell fate decisions. Key issues include possible physiological roles, messengers, and mechanisms. We also provide examples of how data mining of publicly-available datasets from "omics" technologies can be a powerful means to gain additional insights into potential redox signaling pathways between peroxisomes and mitochondria. Finally, we highlight the need for more studies that seek to clarify the mechanisms of how mitochondria may act as dynamic receivers, integrators, and transmitters of peroxisome-derived mediators of oxidative stress. The outcome of such studies may open up exciting new avenues for the community of researchers working on cellular responses to organelle-derived oxidative stress, a research field in which the role of peroxisomes is currently highly underestimated and an issue of discussion. PMID:26075204

  4. Redox interplay between mitochondria and peroxisomes

    PubMed Central

    Lismont, Celien; Nordgren, Marcus; Van Veldhoven, Paul P.; Fransen, Marc

    2015-01-01

    Reduction-oxidation or “redox” reactions are an integral part of a broad range of cellular processes such as gene expression, energy metabolism, protein import and folding, and autophagy. As many of these processes are intimately linked with cell fate decisions, transient or chronic changes in cellular redox equilibrium are likely to contribute to the initiation and progression of a plethora of human diseases. Since a long time, it is known that mitochondria are major players in redox regulation and signaling. More recently, it has become clear that also peroxisomes have the capacity to impact redox-linked physiological processes. To serve this function, peroxisomes cooperate with other organelles, including mitochondria. This review provides a comprehensive picture of what is currently known about the redox interplay between mitochondria and peroxisomes in mammals. We first outline the pro- and antioxidant systems of both organelles and how they may function as redox signaling nodes. Next, we critically review and discuss emerging evidence that peroxisomes and mitochondria share an intricate redox-sensitive relationship and cooperate in cell fate decisions. Key issues include possible physiological roles, messengers, and mechanisms. We also provide examples of how data mining of publicly-available datasets from “omics” technologies can be a powerful means to gain additional insights into potential redox signaling pathways between peroxisomes and mitochondria. Finally, we highlight the need for more studies that seek to clarify the mechanisms of how mitochondria may act as dynamic receivers, integrators, and transmitters of peroxisome-derived mediators of oxidative stress. The outcome of such studies may open up exciting new avenues for the community of researchers working on cellular responses to organelle-derived oxidative stress, a research field in which the role of peroxisomes is currently highly underestimated and an issue of discussion. PMID:26075204

  5. Prodrugs of Fluoro-Substituted Benzoates of EGC as Tumor Cellular Proteasome Inhibitors and Apoptosis Inducers

    PubMed Central

    Yu, Zhiyong; Qin, Xu Long; Gu, Yan Yan; Chen, Di; Cui, Qiuzhi Cindy; Jiang, Tao; Wan, Sheng Biao; Dou, Q. Ping

    2008-01-01

    The most potent catechin in green tea is (-)-epigallocatechin-3-gallate [(-)-EGCG], which, however, is unstable under physiological conditions. To discover more stable and more potent polyphenol proteasome inhibitors, we synthesized several novel fluoro-substituted (-)-EGCG analogs, named F-EGCG analogs, as well as their prodrug forms with all of -OH groups protected by acetate. We report that the prodrug form of one F-EGCG analog exhibited greater potency than the previously reported peracetate of (-)-EGCG to inhibit proteasomal activity, suppress cell proliferation, and induce apoptosis in human leukemia Jurkat T cells, demonstrating the potential of these compounds to be developed into novel anti-cancer and cancer-preventive agents. PMID:19325839

  6. Histone Deacetylase Inhibitor Trichostatin A Ameliorated Endotoxin-Induced Neuroinflammation and Cognitive Dysfunction.

    PubMed

    Hsing, Chung-Hsi; Hung, Shih-Kai; Chen, Yeong-Chang; Wei, Tsui-Shan; Sun, Ding-Ping; Wang, Jhi-Joung; Yeh, Ching-Hua

    2015-01-01

    Excessive production of cytokines by microglia may cause cognitive dysfunction and long-lasting behavioral changes. Activating the peripheral innate immune system stimulates cytokine secretion in the central nervous system, which modulates cognitive function. Histone deacetylases (HDACs) modulate cytokine synthesis and release. Trichostatin A (TSA), an HDAC inhibitor, is documented to be anti-inflammatory and neuroprotective. We investigated whether TSA reduces lipopolysaccharide- (LPS-) induced neuroinflammation and cognitive dysfunction. ICR mice were first intraperitoneally (i.p.) injected with vehicle or TSA (0.3?mg/kg). One hour later, they were injected (i.p.) with saline or Escherichia coli LPS (1?mg/kg). We analyzed the food and water intake, body weight loss, and sucrose preference of the injected mice and then determined the microglia activation and inflammatory cytokine expression in the brains of LPS-treated mice and LPS-treated BV-2 microglial cells. In the TSA-pretreated mice, microglial activation was lower, anhedonia did not occur, and LPS-induced cognitive dysfunction (anorexia, weight loss, and social withdrawal) was attenuated. Moreover, mRNA expression of HDAC2, HDAC5, indoleamine 2,3-dioxygenase (IDO), TNF-?, MCP-1, and IL-1? in the brain of LPS-challenged mice and in the LPS-treated BV-2 microglial cells was lower. TSA diminished LPS-induced inflammatory responses in the mouse brain and modulated the cytokine-associated changes in cognitive function, which might be specifically related to reducing HDAC2 and HDAC5 expression. PMID:26273133

  7. Histone Deacetylase Inhibitor Trichostatin A Ameliorated Endotoxin-Induced Neuroinflammation and Cognitive Dysfunction

    PubMed Central

    Hsing, Chung-Hsi; Hung, Shih-Kai; Chen, Yeong-Chang; Wei, Tsui-Shan; Sun, Ding-Ping; Wang, Jhi-Joung; Yeh, Ching-Hua

    2015-01-01

    Excessive production of cytokines by microglia may cause cognitive dysfunction and long-lasting behavioral changes. Activating the peripheral innate immune system stimulates cytokine secretion in the central nervous system, which modulates cognitive function. Histone deacetylases (HDACs) modulate cytokine synthesis and release. Trichostatin A (TSA), an HDAC inhibitor, is documented to be anti-inflammatory and neuroprotective. We investigated whether TSA reduces lipopolysaccharide- (LPS-) induced neuroinflammation and cognitive dysfunction. ICR mice were first intraperitoneally (i.p.) injected with vehicle or TSA (0.3?mg/kg). One hour later, they were injected (i.p.) with saline or Escherichia coli LPS (1?mg/kg). We analyzed the food and water intake, body weight loss, and sucrose preference of the injected mice and then determined the microglia activation and inflammatory cytokine expression in the brains of LPS-treated mice and LPS-treated BV-2 microglial cells. In the TSA-pretreated mice, microglial activation was lower, anhedonia did not occur, and LPS-induced cognitive dysfunction (anorexia, weight loss, and social withdrawal) was attenuated. Moreover, mRNA expression of HDAC2, HDAC5, indoleamine 2,3-dioxygenase (IDO), TNF-?, MCP-1, and IL-1? in the brain of LPS-challenged mice and in the LPS-treated BV-2 microglial cells was lower. TSA diminished LPS-induced inflammatory responses in the mouse brain and modulated the cytokine-associated changes in cognitive function, which might be specifically related to reducing HDAC2 and HDAC5 expression. PMID:26273133

  8. MLN4924 induces Noxa upregulation in acute myelogenous leukemia and synergizes with Bcl-2 inhibitors.

    PubMed

    Knorr, K L B; Schneider, P A; Meng, X W; Dai, H; Smith, B D; Hess, A D; Karp, J E; Kaufmann, S H

    2015-12-01

    MLN4924 (pevonedistat), an inhibitor of the Nedd8 activating enzyme (NAE), has exhibited promising clinical activity in acute myelogenous leukemia (AML). Here we demonstrate that MLN4924 induces apoptosis in AML cell lines and clinical samples via a mechanism distinct from those observed in other malignancies. Inactivation of E3 cullin ring ligases (CRLs) through NAE inhibition causes accumulation of the CRL substrate c-Myc, which transactivates the PMAIP1 gene encoding Noxa, leading to increased Noxa protein, Bax and Bak activation, and subsequent apoptotic changes. Importantly, c-Myc knockdown diminishes Noxa induction; and Noxa siRNA diminishes MLN4924-induced killing. Because Noxa also neutralizes Mcl-1, an anti-apoptotic Bcl-2 paralog often upregulated in resistant AML, further experiments have examined the effect of combining MLN4924 with BH3 mimetics that target other anti-apoptotic proteins. In combination with ABT-199 or ABT-263 (navitoclax), MLN4924 exerts a synergistic cytotoxic effect. Collectively, these results provide new insight into MLN4924-induced engagement of the apoptotic machinery that could help guide further exploration of MLN4924 for AML. PMID:26045051

  9. Mitigation and Treatment of Radiation-Induced Thoracic Injury With a Cyclooxygenase-2 Inhibitor, Celecoxib

    SciTech Connect

    Hunter, Nancy R.; Valdecanas, David; Liao Zhongxing; Milas, Luka; Thames, Howard D.; Mason, Kathy A.

    2013-02-01

    Purpose: To test whether a cyclooxygenase-2 inhibitor (celecoxib) could reduce mortality resulting from radiation-induced pneumonitis. Methods and Materials: Celecoxib was given to mice twice daily for 40 consecutive days starting on the day of local thoracic irradiation (LTI) or 40 or 80 days later. C3Hf/KamLaw mice were observed for morbidity, and time to death was determined. Results were analyzed using the Cox proportional hazards model. Results: Timing of celecoxib relative to LTI determined efficacy. A significant reduction in time to death was achieved only when celecoxib was started 80 days after LTI, corresponding to the time when pneumonitis is expressed. For these mice the reduction in mortality was quantified as a hazard ratio for mortality of treated vs untreated of 0.36 (95% confidence interval [CI] 0.24-0.53), thus significantly less than 1.0. Correspondingly, the median lethal dose for treated mice (12.9 Gy; 95% CI 12.55-13.25 Gy) was significantly (P=.026) higher than for untreated mice (12.4 Gy; 95% CI 12.2-12.65 Gy). Conclusions: Celecoxib significantly reduced lung toxicity when administered months after LTI when the deleterious effects of radiation were expressed. The schedule-dependent reduction in fatal pneumonitis suggests that celecoxib could be clinically useful by reintroduction of treatment months after completion of radiation therapy. These findings may be important for designing clinical trials using cyclooxygenase-2 inhibitors to treat radiation-induced lung toxicity as a complement to concurrent radiation therapy of lung cancers.

  10. USP7 inhibitor P22077 inhibits neuroblastoma growth via inducing p53-mediated apoptosis

    PubMed Central

    Fan, Y-H; Cheng, J; Vasudevan, S A; Dou, J; Zhang, H; Patel, R H; Ma, I T; Rojas, Y; Zhao, Y; Yu, Y; Zhang, H; Shohet, J M; Nuchtern, J G; Kim, E S; Yang, J

    2013-01-01

    Neuroblastoma (NB) is a common pediatric cancer and contributes to more than 15% of all pediatric cancer-related deaths. Unlike adult tumors, recurrent somatic mutations in NB, such as tumor protein 53 (p53) mutations, occur with relative paucity. In addition, p53 downstream function is intact in NB cells with wild-type p53, suggesting that reactivation of p53 may be a viable therapeutic strategy for NB treatment. Herein, we report that the ubiquitin-specific protease 7 (USP7) inhibitor, P22077, potently induces apoptosis in NB cells with an intact USP7-HDM2-p53 axis but not in NB cells with mutant p53 or without human homolog of MDM2 (HDM2) expression. In this study, we found that P22077 stabilized p53 by inducing HDM2 protein degradation in NB cells. P22077 also significantly augmented the cytotoxic effects of doxorubicin (Dox) and etoposide (VP-16) in NB cells with an intact USP7-HDM2-p53 axis. Moreover, P22077 was found to be able to sensitize chemoresistant LA-N-6 NB cells to chemotherapy. In an in vivo orthotopic NB mouse model, P22077 significantly inhibited the xenograft growth of three NB cell lines. Database analysis of NB patients shows that high expression of USP7 significantly predicts poor outcomes. Together, our data strongly suggest that targeting USP7 is a novel concept in the treatment of NB. USP7-specific inhibitors like P22077 may serve not only as a stand-alone therapy but also as an effective adjunct to current chemotherapeutic regimens for treating NB with an intact USP7-HDM2-p53 axis. PMID:24136231

  11. MDM2 Inhibitor, Nutlin 3a, Induces p53 Dependent Autophagy in Acute Leukemia by AMP Kinase Activation

    PubMed Central

    Borthakur, Gautam; Duvvuri, Seshagiri; Ruvolo, Vivian; Tripathi, Durga Nand; Piya, Sujan; Burks, Jared; Jacamo, Rodrigo; Kojima, Kensuke; Ruvolo, Peter; Fueyo-Margareto, Juan; Konopleva, Marina; Andreeff, Michael

    2015-01-01

    MDM2 (mouse double minute 2) inhibitors that activate p53 and induce apoptosis in a non-genotoxic manner are in clinical development for treatment of leukemias. P53 can modulate other programmed cell death pathways including autophagy both transcriptionally and non-transcriptionally. We investigated autophagy induction in acute leukemia by Nutlin 3a, a first-in-class MDM2 inhibitor. Nutlin 3a induced autophagy in a p53 dependent manner and transcriptional activation of AMP kinase (AMPK) is critical, as this effect is abrogated in AMPK -/- mouse embryonic fibroblasts. Nutlin 3a induced autophagy appears to be pro-apoptotic as pharmacological (bafilomycin) or genetic inhibition (BECLIN1 knockdown) of autophagy impairs apoptosis induced by Nutlin 3a. PMID:26440941

  12. Kallikrein inhibitors limit kinin B2 antagonist-induced progression of oedematous to haemorrhagic pancreatitis in rats

    PubMed Central

    Griesbacher, T; Rainer, I; Tiran, B; Peskar, B A

    2008-01-01

    Background and purpose: Exocrine hyperstimulation with caerulein is an established model for oedematous acute pancreatitis. Prevention of oedema formation by bradykinin B2 receptor antagonists induces a progression to a haemorrhagic course in this model. We have investigated whether increased kallikrein activity in the pancreas is responsible for vascular damage and whether this could be prevented by selective kallikrein inhibitors. Experimental approach: Caerulein was infused i.v. and vascular damage was assessed by histological evaluation and determination of haemoglobin accumulation in the tissue. In addition, oedema formation, tissue and plasma kallikrein (PK) activities and the endogenous kallikrein inhibitors ?1-antitrypsin (?1-AT) and ?2-macroglobulin (?2-M) were measured. Key results: Haemorrhagic lesions induced by icatibant in caerulein-induced pancreatitis were associated with a reduction in ?1-AT and ?2-M in the pancreas and a concomitant augmentation of tissue kallikrein (TK) activity. The TK inhibitor VA999024 (previously FE999024), or its combination with the PK inhibitor VA999026 (previously FE999026), inhibited oedema formation to the same extent but did not induce vascular damage. Furthermore, VA999024 inhibited TK activity. When icatibant was combined with VA999024 and VA999026, progression from oedematous to haemorrhagic pancreatitis was abolished. Conclusions and implications: Reduced oedema formation by B2 antagonists prevented influx of endogenous kallikrein inhibitors and led to an excessive activity of kallikrein in the pancreas leading to vascular damage. This can be prevented by a combined inhibition of both tissue-type and plasma-type kallikrein. Kallikrein inhibitors thus should be further evaluated for their therapeutic potential in preventing haemorrhagic lesions in acute pancreatitis. PMID:18695645

  13. The Hansenula polymorpha PER8 gene encodes a novel peroxisomal integral membrane protein involved in proliferation

    PubMed Central

    1995-01-01

    We previously described the isolation of mutants of the methylotrophic yeast Hansenula polymorpha that are defective in peroxisome biogenesis. Here, we describe the characterization of one of these mutants, per8, and the cloning of the PER8 gene. In either methanol or methylamine medium, conditions that normally induce the organelles, per8 cells contain no peroxisome-like structures and peroxisomal enzymes are located in the cytosol. The sequence of PER8 predicts that its product (Per8p) is a novel polypeptide of 34 kD, and antibodies against Per8p recognize a protein of 31 kD. Analysis of the primary sequence of Per8p revealed a 39-amino-acid cysteine-rich segment with similarity to the C3HC4 family of zinc-finger motifs. Overexpression of PER8 results in a markedly enhanced increase in peroxisome numbers. We show that Per8p is an integral membrane protein of the peroxisome and that it is concentrated in the membranes of newly formed organelles. We propose that Per8p is a component of the molecular machinery that controls the proliferation of this organelle. PMID:7844145

  14. ABCD1 deletion-induced mitochondrial dysfunction is corrected by SAHA: implication for adrenoleukodystrophy.

    PubMed

    Baarine, Mauhamad; Beeson, Craig; Singh, Avtar; Singh, Inderjit

    2015-05-01

    X-linked Adrenoleukodystrophy (X-ALD), an inherited peroxisomal metabolic neurodegenerative disorder, is caused by mutations/deletions in the ATP-binding cassette transporter (ABCD1) gene encoding peroxisomal ABC transporter adrenoleukodystrophy protein (ALDP). Metabolic dysfunction in X-ALD is characterized by the accumulation of very long chain fatty acids ? C22:0) in the tissues and plasma of patients. Here, we investigated the mitochondrial status following deletion of ABCD1 in B12 oligodendrocytes and U87 astrocytes. This study provides evidence that silencing of peroxisomal protein ABCD1 produces structural and functional perturbations in mitochondria. Activities of electron transport chain-related enzymes and of citric acid cycle (TCA cycle) were reduced; mitochondrial redox status was dysregulated and the mitochondrial membrane potential was disrupted following ABCD1 silencing. A greater reduction in ATP levels and citrate synthase activities was observed in oligodendrocytes as compared to astrocytes. Furthermore, most of the mitochondrial perturbations induced by ABCD1 silencing were corrected by treating cells with suberoylanilide hydroxamic acid, an Histone deacetylase inhibitor. These observations indicate a novel relationship between peroxisomes and mitochondria in cellular homeostasis and the importance of intact peroxisomes in relation to mitochondrial integrity and function in the cell types that participate in the pathobiology of X-ALD. These observations suggest suberoylanilide hydroxamic acid as a potential therapy for X-ALD. Schematic description of the effects of loss of peroxisomal ATP-binding cassette transporter D1 (ABCD1) gene on cellular Redox and mitochondrial activities and their correction by suberoylanilide hydroxamic acid (SAHA) treatment. Pathogenomic accumulation of very long chain fatty acids (VLCFA) as a result of loss of ABCD1 leads to dysfunctions of mitochondrial biogenesis and its activities. Treatment with SAHA corrects mitochondrial dysfunctions. These studies describe unique cooperation between mitochondria and peroxisome for cellular activities. PMID:25393703

  15. BIM Mediates EGFR Tyrosine Kinase Inhibitor-Induced Apoptosis in Lung Cancers with Oncogenic EGFR Mutations

    PubMed Central

    Costa, Daniel B; Halmos, Balázs; Kumar, Amit; Schumer, Susan T; Huberman, Mark S; Boggon, Titus J; Tenen, Daniel G; Kobayashi, Susumu

    2007-01-01

    Background Epidermal growth factor receptor (EGFR) mutations are present in the majority of patients with non-small cell lung cancer (NSCLC) responsive to the EGFR tyrosine kinase inhibitors (TKIs) gefitinib or erlotinib. These EGFR-dependent tumors eventually become TKI resistant, and the common secondary T790M mutation accounts for half the tumors with acquired resistance to gefitinib. However, the key proapoptotic proteins involved in TKI-induced cell death and other secondary mutations involved in resistance remain unclear. The objective of this study was to identify the mechanism of EGFR TKI-induced apoptosis and secondary resistant mutations that affect this process. Methods and Findings To study TKI-induced cell death and mechanisms of resistance, we used lung cancer cell lines (with or without EGFR mutations), Ba/F3 cells stably transfected with EGFR mutation constructs, and tumor samples from a gefitinib-resistant patient. Here we show that up-regulation of the BH3-only polypeptide BIM (also known as BCL2-like 11) correlated with gefitinib-induced apoptosis in gefitinib-sensitive EGFR-mutant lung cancer cells. The T790M mutation blocked gefitinib-induced up-regulation of BIM and apoptosis. This blockade was overcome by the irreversible TKI CL-387,785. Knockdown of BIM by small interfering RNA was able to attenuate apoptosis induced by EGFR TKIs. Furthermore, from a gefitinib-resistant patient carrying the activating L858R mutation, we identified a novel secondary resistant mutation, L747S in cis to the activating mutation, which attenuated the up-regulation of BIM and reduced apoptosis. Conclusions Our results provide evidence that BIM is involved in TKI-induced apoptosis in sensitive EGFR-mutant cells and that both attenuation of the up-regulation of BIM and resistance to gefitinib-induced apoptosis are seen in models that contain the common EGFR T790M and the novel L747S secondary resistance mutations. These findings also suggest that induction of BIM may have a role in the treatment of TKI-resistant tumors. PMID:17973572

  16. Diabetes or peroxisome proliferator-activated receptor alpha agonist increases mitochondrial thioesterase I activity in heart

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Peroxisome proliferator-activated receptor alpha (PPAR alpha) is a transcriptional regulator of the expression of mitochondrial thioesterase I (MTE-I) and uncoupling protein 3 (UCP3), which are induced in the heart at the mRNA level in response to diabetes. Little is known about the regulation of pr...

  17. PPARÁ-DEPENDENT GENE EXPRESSION CHANGES IN THE MOUSE LIVER AFTER EXPOSURE TO PEROXISOME PROLIFERATORS

    EPA Science Inventory

    Peroxisome proliferators (PP) are a large class of structurally diverse chemicals that mediate their effects in the liver mainly through the PP-activated receptor ¿ (PPAR?). Development of PP induced hepatocarcinogenesis in mouse liver is known to be dependent on PPAR&#...

  18. IDENTIFICATION OF EARLY MOLECULAR EVENTS AFTER PEROXISOME PROLIFERATOR EXPOSURE IN THE RODENT LIVER

    EPA Science Inventory

    Peroxisome proliferators (PP) are a large class of structurally diverse chemicals that mediate their effects in the liver mainly through the PP-activated receptor ?(PPAR?). Development of PP induced hepatocarcinogenesis in mouse liver is known to be dependent on PPAR? but do...

  19. , -Carotene Decreases Peroxisome Proliferator Receptor Activity and Reduces Lipid Storage Capacity of Adipocytes in

    E-print Network

    Palczewski, Krzysztof

    University, Cleveland, Ohio 44106 and the § Laboratory of Molecular Biology, Nutrition and Biotechnology provided for a connection between retinoid metabolism and the activity of peroxisome proliferator receptors but not all-trans-retinol induced reti- noid signaling and decreased Ppar expression in white adipose tissue

  20. Cell-based chemical genetic screen identifies damnacanthal as an inhibitor of HIV-1 Vpr induced cell death

    E-print Network

    Huang, Jing

    Cell-based chemical genetic screen identifies damnacanthal as an inhibitor of HIV-1 Vpr induced August 2006 Abstract Viral protein R (Vpr), one of the human immunodeficiency virus type 1 (HIV-1. The mechanisms of Vpr have been intensely studied because it is believed that they underlie HIV-1 pathogenesis

  1. Electrophilic Peroxisome Proliferator–Activated Receptor-? Ligands Have Potent Antifibrotic Effects in Human Lung Fibroblasts

    PubMed Central

    Ferguson, Heather E.; Kulkarni, Ajit; Lehmann, Geniece M.; Garcia-Bates, Tatiana M.; Thatcher, Thomas H.; Huxlin, Krystel R.; Phipps, Richard P.; Sime, Patricia J.

    2009-01-01

    Pulmonary fibrosis is a progressive scarring disease with no effective treatment. Transforming growth factor (TGF)-? is up-regulated in fibrotic diseases, where it stimulates differentiation of fibroblasts to myofibroblasts and production of excess extracellular matrix. Peroxisome proliferator–activated receptor (PPAR) ? is a transcription factor that regulates adipogenesis, insulin sensitization, and inflammation. We report here that a novel PPAR? ligand, 2-cyano-3,12-dioxoolean-1,9-dien-28-oic acid (CDDO), is a potent inhibitor of TGF-?–stimulated differentiation of human lung fibroblasts to myofibroblasts, and suppresses up-regulation of ?–smooth muscle actin, fibronectin, collagen, and the novel myofibroblast marker, calponin. The inhibitory concentration causing a 50% decrease in aSMA for CDDO was 20-fold lower than the endogenous PPAR? ligand, 15-deoxy-?12,14-prostaglandin J2 (15 d-PGJ2), and 400-fold lower than the synthetic ligand, rosiglitazone. Pharmacologic and genetic approaches were used to demonstrate that CDDO mediates its activity via a PPAR?-independent pathway. CDDO and 15 d-PGJ2 contain an ?/? unsaturated ketone, which acts as an electrophilic center that can form covalent bonds with cellular proteins. Prostaglandin A1 and diphenyl diselenide, both strong electrophiles, also inhibit myofibroblast differentiation, but a structural analog of 15 d-PGJ2 lacking the electrophilic center is much less potent. CDDO does not alter TGF-?–induced Smad or AP-1 signaling, but does inhibit acetylation of CREB binding protein/p300, a critical coactivator in the transcriptional regulation of TGF-?–responsive genes. Overall, these data indicate that certain PPAR? ligands, and other small molecules with electrophilic centers, are potent inhibitors of critical TGF-?–mediated profibrogenic activities through pathways independent of PPAR?. As the inhibitory concentration causing a 50% decrease in aSMA for CDDO is 400-fold lower than that in rosiglitazone, the translational potential of CDDO for treatment of fibrotic diseases is high. PMID:19286977

  2. Differential Activation of Peroxisome Proliferator-activated Receptors by Eicosanoids*

    E-print Network

    Brown, Myles

    Differential Activation of Peroxisome Proliferator-activated Receptors by Eicosanoids* (Received, Pennsylvania 19104 Peroxisome proliferator-activated receptors (PPARs) are nuclear hormone receptors occurring compounds activate each of the PPAR subtypes. To address this issue, a screening assay

  3. DNA Methyltransferase Inhibitor Zebularine Inhibits Human Hepatic Carcinoma Cells Proliferation and Induces Apoptosis

    PubMed Central

    Nakamura, Kazuaki; Aizawa, Kazuko; Nakabayashi, Kazuhiko; Kato, Natsuko; Yamauchi, Junji; Hata, Kenichiro; Tanoue, Akito

    2013-01-01

    Hepatocellular carcinoma is one of the most common cancers worldwide. During tumorigenesis, tumor suppressor and cancer-related genes are commonly silenced by aberrant DNA methylation in their promoter regions. Zebularine (1-(?-D-ribofuranosyl)-1,2-dihydropyrimidin-2-one) acts as an inhibitor of DNA methylation and exhibits chemical stability and minimal cytotoxicity both in vitro and in vivo. In this study, we explore the effect and possible mechanism of action of zebularine on hepatocellular carcinoma cell line HepG2. We demonstrate that zebularine exhibits antitumor activity on HepG2 cells by inhibiting cell proliferation and inducing apoptosis, however, it has little effect on DNA methylation in HepG2 cells. On the other hand, zebularine treatment downregulated CDK2 and the phosphorylation of retinoblastoma protein (Rb), and upregulated p21WAF/CIP1 and p53. We also found that zebularine treatment upregulated the phosphorylation of p44/42 mitogen-activated protein kinase (MAPK). These results suggest that the p44/42 MAPK pathway plays a role in zebularine-induced cell-cycle arrest by regulating the activity of p21WAF/CIP1 and Rb. Furthermore, although the proapoptotic protein Bax levels were not affected, the antiapoptotic protein Bcl-2 level was downregulated with zebularine treatment. In addition, the data in the present study indicate that inhibition of the double-stranded RNA-dependent protein kinase (PKR) is involved in inducing apoptosis with zebularine. These results suggest a novel mechanism of zebularine-induced cell growth arrest and apoptosis via a DNA methylation-independent pathway in hepatocellular carcinoma. PMID:23320119

  4. Clinical significance of plasminogen activator inhibitor activity in patients with exercise-induced ischemia

    SciTech Connect

    Sakata, K.; Kurata, C.; Taguchi, T.; Suzuki, S.; Kobayashi, A.; Yamazaki, N.; Rydzewski, A.; Takada, Y.; Takada, A. )

    1990-10-01

    To assess the fibrinolytic system in patients with exercise-induced ischemia and its relation to ischemia and severity of coronary artery disease (CAD), 47 patients with CAD confirmed by results of coronary angiography underwent symptom-limited multistage exercise thallium-201 emission computed tomography. All patients with CAD had exercise-induced ischemia as assessed from thallium-201 images. Pre- and peak exercise blood samples from each patient and preexercise blood samples from control subjects were assayed for several fibrinolytic components and were also assayed for plasma adrenaline. The extent of ischemia was defined as delta visual uptake score (total visual uptake score in delayed images minus total visual uptake score in initial images) and the severity of CAD as the number of diseased vessels. In the basal condition, plasminogen activator inhibitor (PAI) activity was significantly higher in patients with exercise-induced ischemia as compared to control subjects (p less than 0.01), although there were no significant differences in other fibrinolytic variables between the two groups. Moreover, PAI activity in the basal condition displayed a significantly positive correlation with the extent of ischemia (r = 0.47, p less than 0.01). Patients with exercise-induced ischemia were divided into two groups (24 with single-vessel disease and 23 with multivessel disease). There were no significant differences in coronary risk factors, hemodynamics, or plasma adrenaline levels during exercise between single-vessel and multivessel disease except that delta visual uptake score was significantly higher in multivessel disease (p less than 0.01).

  5. Etodolac, a cyclooxygenase-2 inhibitor, attenuates paclitaxel-induced peripheral neuropathy in a mouse model of mechanical allodynia.

    PubMed

    Ito, Sunao; Tajima, Koyuki; Nogawa, Masaki; Inoue, Naoki; Kyoi, Takashi; Takahashi, Yosuke; Sasagawa, Takahiro; Nakamura, Akio; Kotera, Takashi; Ueda, Makoto; Yamashita, Yasuhiro; Banno, Kouji

    2012-07-01

    The effect of the cyclooxygenase-2 (COX-2) inhibitor etodolac on the mechanical allodynia induced by paclitaxel was investigated in mice and compared with the effects of the nonselective COX inhibitors indomethacin and diclofenac, the selective COX-2 inhibitor celecoxib, the calcium channel ?(2)? subunit inhibitor pregabalin, the sodium channel blocker mexiletine, and the serotonin-norepinephrine reuptake inhibitor duloxetine. The decrease in the paw-withdrawal threshold induced by paclitaxel was reversed by oral administration of etodolac at 10 mg/kg but was not affected by indomethacin, diclofenac, or celecoxib. The antiallodynic effect of etodolac gradually increased during repeated administration, and after 2 weeks the paw-withdrawal threshold at the preadministration point was significantly increased. Pregabalin, duloxetine, and mexiletine also showed an antiallodynic effect in this model. Whereas pregabalin had a preadministration effect similar to that of etodolac during repeated administration, mexiletine or duloxetine had no such effect. There was almost no difference in the distribution of etodolac and diclofenac in nervous tissue, indicating that COX inhibition is unlikely to be involved in the antiallodynic effect of etodolac. Etodolac did not show a neuroprotective effect against morphological transformations such as the axonal degeneration induced by paclitaxel. Instead, etodolac probably acts at the level of functional changes accompanying paclitaxel treatment, such as alterations in the activation state of components of the pain transmission pathway. Our findings suggest that etodolac attenuates paclitaxel-induced peripheral neuropathy by a COX-independent pathway and that it might be useful for the treatment of paclitaxel-induced peripheral neuropathy. PMID:22460833

  6. The NADPH oxidase inhibitor apocynin induces nitric oxide synthesis via oxidative stress

    SciTech Connect

    Riganti, Chiara

    2008-05-01

    We have recently shown that apocynin elicits an oxidative stress in N11 mouse glial cells and other cell types. Here we report that apocynin increased the accumulation of nitrite, the stable derivative of nitric oxide (NO), in the extracellular medium of N11 cell cultures, and the NO synthase (NOS) activity in cell lysates. The increased synthesis of NO was associated with increased expression of inducible NOS (iNOS) mRNA, increased nuclear translocation of the redox-sensitive transcription factor NF-{kappa}B and decreased intracellular level of its inhibitor IkB{alpha}. These effects, accompanied by increased production of H{sub 2}O{sub 2}, were very similar to those observed after incubation with bacterial lipopolysaccharide (LPS) and were inhibited by catalase. These results suggest that apocynin, similarly to LPS, induces increased NO synthesis by eliciting a generation of reactive oxygen species (ROS), which in turn causes NF-{kappa}B activation and increased expression of iNOS. Therefore, the increased bioavailability of NO reported in the literature after in vivo or in vitro treatments with apocynin might depend, at least partly, on the drug-elicited induction of iNOS, and not only on the inhibition of NADPH oxidase and the subsequent decreased scavenging of NO by oxidase-derived ROS, as it is often supposed.

  7. CT-2576, an inhibitor of phospholipid signaling, suppresses constitutive and induced expression of human immunodeficiency virus.

    PubMed Central

    Leung, D W; Peterson, P K; Weeks, R; Gekker, G; Chao, C C; Kaplan, A H; Balantac, N; Tompkins, C; Underiner, G E; Bursten, S

    1995-01-01

    Viruses such as human immunodeficiency virus (HIV) require cellular activation for expression. Cellular activation in lymphoid cells is associated with augmented accumulation of certain phosphatidic acid (PA) species derived from the hydrolysis of glycan phosphatidylinositol (GPI). This suggests that activation of a phospholipid pathway may play a role in initiation of viral replication. To test this hypothesis, we examined the effect of tat gene expression on the production of cellular PA species, as the Tat protein is essential for HIV expression and has been implicated in activating the expression of multiple host cellular genes. Expression of tat increased the expression of PA. We then tested whether synthetic inhibitors of PA metabolism would inhibit activation of the HIV long terminal repeat by Tat and tumor necrosis factor alpha (TNF-alpha). CT-2576 suppressed both PA generation induced by Tat and HIV long terminal repeat-directed gene expression in response to Tat or TNF-alpha at a posttranscriptional step. CT-2576 also inhibited constitutive as well as TNF-alpha- and interleukin 6-induced expression of HIV p24 antigen in chronically infected U1 cells and in peripheral blood lymphocytes acutely infected with a clinical isolate of HIV. Pharmacological inhibition of synthesis of selected PA species may therefore provide a therapeutic approach to suppression of HIV replication. Images Fig. 1 Fig. 3 Fig. 5 PMID:7761405

  8. Novel Epidermal Growth Factor Receptor Inhibitor Attenuates Angiotensin II-Induced Kidney Fibrosis.

    PubMed

    Qian, Yuanyuan; Peng, Kesong; Qiu, Chenyu; Skibba, Melissa; Huang, Yi; Xu, Zheng; Zhang, Yali; Hu, Jie; Liang, Dandan; Zou, Chunpeng; Wang, Yi; Liang, Guang

    2016-01-01

    Chronic activation of renin-angiotensin system (RAS) greatly contributes to renal fibrosis and accelerates the progression of chronic kidney disease; however, the underlying molecular mechanism is poorly understood. Angiotensin II (Ang II), the central component of RAS, is a key regulator of renal fibrogenic destruction. Here we show that epidermal growth factor receptor (EGFR) plays an important role in Ang II-induced renal fibrosis. Inhibition of EGFR activation by novel small molecules or by short hairpin RNA knockdown in Ang II-treated SV40 mesangial cells in vitro suppresses protein kinase B and extracellular signal-related kinase signaling pathways and transforming growth factor-?/Sma- and Mad-related protein activation, and abolishes the accumulation of fibrotic markers such as connective tissue growth factor, collagen IV. The transactivation of EGFR by Ang II in SV40 cells depends on the phosphorylation of proto-oncogene tyrosine-protein kinase Src (c-Src) kinase. Further validation in vivo demonstrates that EGFR small molecule inhibitor successfully attenuates renal fibrosis and kidney dysfunction in a mouse model induced by Ang II infusion. These findings indicate a crucial role of EGFR in Ang II-dependent renal deterioration, and reveal EGFR inhibition as a new therapeutic strategy for preventing progression of chronic renal diseases. PMID:26514795

  9. Proteasome inhibitor-induced apoptosis of glioma cells involves the processing of multiple caspases and cytochrome c release.

    PubMed

    Wagenknecht, B; Hermisson, M; Groscurth, P; Liston, P; Krammer, P H; Weller, M

    2000-12-01

    The proteasome is a multiprotein complex that is involved in the intracellular protein degradation in eukaryotes. Here, we show that human malignant glioma cells are susceptible to apoptotic cell death induced by the proteasome inhibitors, MG132 and lactacystin. The execution of the apoptotic death program involves the processing of caspases 2, 3, 7, 8, and 9. Apoptosis is inhibited by ectopic expression of X-linked inhibitor of apoptosis (XIAP) and by coexposure to the broad-spectrum caspase inhibitor, benzoyl-VAD-fluoromethyl ketone (zVAD-fmk), but not by the preferential caspase 8 inhibitor, crm-A. It is interesting that specific morphological alterations induced by proteasome inhibition, such as dilated rough endoplasmic reticulum and the formation of cytoplasmic vacuoles and dense mitochondrial deposits, are unaffected by zVAD-fmk. Apoptosis is also inhibited by ectopic expression of Bcl-2 or by an inhibitor of protein synthesis, cycloheximide. Further, cytochrome c release and disruption of mitochondrial membrane potential are prominent features of apoptosis triggered by proteasome inhibition. Bcl-2 is a stronger inhibitor of cytochrome c release than zVAD-fmk. XIAP and crm-A fail to modulate cytochrome c release. These data place cytochrome c release downstream of Bcl-2 activity but upstream of XIAP- and crm-A-sensitive caspases. The partial inhibition of cytochrome c release by zVAD-fmk indicates a positive feedback loop that may involve cytochrome c release and zVAD-fmk-sensitive caspases. Finally, death ligand/receptor interactions, including the CD95/CD95 ligand system, do not mediate apoptosis induced by proteasome inhibition in human malignant glioma cells. PMID:11080180

  10. Dietary Inulin Fibers Prevent Proton-Pump Inhibitor (PPI)-Induced Hypocalcemia in Mice

    PubMed Central

    Hess, Mark W.; de Baaij, Jeroen H. F.; Gommers, Lisanne M. M.; Hoenderop, Joost G. J.; Bindels, René J. M.

    2015-01-01

    Background Proton-pump inhibitor-induced hypomagnesemia (PPIH) is the most recognized side effect of proton-pump inhibitors (PPIs). Additionally, PPIH is associated with hypocalcemia and hypokalemia. It is hypothesized that PPIs reduce epithelial proton secretion and thereby increase the pH in the colon, which may explain the reduced absorption of and Mg2+ and Ca2+. Fermentation of dietary oligofructose-enriched inulin fibers by the microflora leads to acidification of the intestinal lumen and by this enhances mineral uptake. This study aimed, therefore, to improve mineral absorption by application of dietary inulin to counteract PPIH. Methods Here, C57BL/J6 mice were supplemented with omeprazole and/or inulin. Subsequently, Mg2+ and Ca2+ homeostasis was assessed by means of serum, urine and fecal electrolyte measurements. Moreover, the mRNA levels of magnesiotropic and calciotropic genes were examined in the large intestine and kidney by real-time PCR. Results Treatment with omeprazole significantly reduced serum Mg2+ and Ca2+ levels. However, concomitant addition of dietary inulin fibers normalized serum Ca2+ but not serum Mg2+ concentrations. Inulin abolished enhanced expression of Trpv6 and S100g in the colon by omeprazole. Additionally, intestinal and renal mRNA levels of the Trpm6 gene were reduced after inulin intake. Conclusions This study suggests that dietary inulin counteracts reduced intestinal Ca2+ absorption upon PPI treatment. In contrast, inulin did not increase intestinal absorption of Mg2+ sufficiently to recover serum Mg2+. The clinical potential of dietary inulin treatment should be the subject of future studies. PMID:26397986

  11. MET inhibitor PHA-665752 suppresses the hepatocyte growth factor-induced cell proliferation and radioresistance in nasopharyngeal carcinoma cells.

    PubMed

    Liu, Tongxin; Li, Qi; Sun, Quanquan; Zhang, Yuqin; Yang, Hua; Wang, Rong; Chen, Longhua; Wang, Wei

    2014-06-20

    Although ionizing radiation (IR) has provided considerable improvements in nasopharyngeal carcinoma (NPC), in subsets of patients, radioresistance is still a major problem in the treatment. In this study, we demonstrated that irradiation induced MET overexpression and activation, and the aberrant MET signal mediated by hepatocyte growth factor (HGF) induced radioresistance. We also found that MET inhibitor PHA-665752 effectively suppressed HGF induced cell proliferation and radioresistance in NPC cells. Further investigation indicated that PHA-665752 suppressed the phosphorylation of the Akt, ERK1/2, and STAT3 proteins in a dose-dependent manner. Our data indicated that the combination of IR with a MET inhibitor, such as PHA-665752, might be a promising therapeutic strategy for NPC. PMID:24802404

  12. Direct thrombin inhibitors, but not the direct factor Xa inhibitor rivaroxaban, increase tissue factor-induced hypercoagulability in vitro and in vivo

    PubMed Central

    Perzborn, E; Heitmeier, S; Buetehorn, U; Laux, V

    2014-01-01

    Background Increased hypercoagulability has been reported with low doses of direct thrombin inhibitors but not with direct factor Xa inhibitors. Objectives To compare the effects of rivaroxaban with those of melagatran and dabigatran on thrombin generation (TG) and tissue factor-induced hypercoagulability and to explore the possible involvement of the thrombin–thrombomodulin/activated protein C system. Methods In normal human plasma and in protein C-deficient plasma, TG was investigated in vitro in the presence and absence of recombinant human soluble thrombomodulin (rhs-TM). TG was determined by calibrated automated thrombography and an ELISA for prothrombin fragments 1+2 (F1+2). In an in vivo rat model, hypercoagulability was induced by tissue factor; levels of thrombin–antithrombin (TAT) and fibrinogen and the platelet count were determined. Results Rivaroxaban inhibited TG in a concentration-dependent manner. In the absence of rhs-TM, melagatran and dabigatran also inhibited TG concentration dependently. However, in the presence of rhs-TM, lower concentrations of melagatran (119–474 nmol L–1) and dabigatran (68–545 nmol L?1) enhanced endogenous thrombin potential, peak TG, and F1+2 formation in normal plasma but not in protein C-deficient plasma. In vivo, rivaroxaban dose-dependently inhibited TAT generation, whereas melagatran showed a paradoxical effect, with an increase in TAT and a small decrease in fibrinogen and platelet count at lower doses. Conclusion Low concentrations of the direct thrombin inhibitors melagatran and dabigatran enhanced TG and hypercoagulability, possibly via inhibition of the protein C system. In contrast, rivaroxaban reduced TG and hypercoagulability under all conditions studied, suggesting that it does not suppress this negative-feedback system. PMID:24766850

  13. Trichostatin A, a histone deacetylase inhibitor, modulates unloaded-induced skeletal muscle atrophy.

    PubMed

    Dupré-Aucouturier, Sylvie; Castells, Josiane; Freyssenet, Damien; Desplanches, Dominique

    2015-08-15

    Skeletal muscle atrophy is commonly associated with immobilization, ageing, and catabolic diseases such as diabetes and cancer cachexia. Epigenetic regulation of gene expression resulting from chromatin remodeling through histone acetylation has been implicated in muscle disuse. The present work was designed to test the hypothesis that treatment with trichostatin A (TSA), a histone deacetylase inhibitor, would partly counteract unloading-induced muscle atrophy. Soleus muscle atrophy (-38%) induced by 14 days of rat hindlimb suspension was reduced to only 25% under TSA treatment. TSA partly prevented the loss of type I and IIa fiber size and reversed the transitions of slow-twitch to fast-twitch fibers in soleus muscle. Unloading or TSA treatment did not affect myostatin gene expression and follistatin protein. Soleus protein carbonyl content remained unchanged, whereas the decrease in glutathione vs. glutathione disulfide ratio and the increase in catalase activity (biomarkers of oxidative stress) observed after unloading were abolished by TSA treatment. The autophagy-lysosome pathway (Bnip3 and microtubule-associated protein 1 light chain 3 proteins, Atg5, Gabarapl1, Ulk1, and cathepsin B and L mRNA) was not activated by unloading or TSA treatment. However, TSA suppressed the rise in muscle-specific RING finger protein 1 (MuRF1) caused by unloading without affecting the forkhead box (Foxo3) transcription factor. Prevention of muscle atrophy by TSA might be due to the regulation of the skeletal muscle atrophy-related MuRF1 gene. Our findings suggest that TSA may provide a novel avenue to treat unloaded-induced muscle atrophy. PMID:26112243

  14. Flavone as PARP-1 inhibitor: its effect on lipopolysaccharide induced gene-expression.

    PubMed

    Geraets, Liesbeth; Moonen, Harald J J; Brauers, Karen; Gottschalk, Ralph W H; Wouters, Emiel F M; Bast, Aalt; Hageman, Geja J

    2007-11-14

    The nuclear enzyme poly(ADP-ribose) polymerase-1 (PARP-1) which was initially known for its role in the repair of oxidative stress-induced DNA damage, has also been reported to play a mediating role in the inflammatory response. Studies with PARP-1 knockout models have shown that PARP-1 is a co-activator of Nuclear Factor-kappa B (NF-kappaB), although this appears not to require its enzyme activity. In addition, drug-induced inhibition of the enzyme activity of PARP-1 was observed to reduce the production of pro-inflammatory mediators. In this study, the flavonoid compound flavone was demonstrated to significantly inhibit the enzyme activity of PARP-1. Further evaluation of flavone in N-methyl-N'-nitro-N-nitrosoguanidine (MNNG)-treated human pulmonary epithelial and vascular endothelial cells revealed that both the decrease in NAD(+) levels, as well as the formation of PAR-polymers was dose-dependently attenuated by flavone. In addition, flavone was found to reduce the lipopolysaccharide (LPS)-induced interleukin (IL)-8 production in pulmonary epithelial cells, which was confirmed by transcription analysis. Furthermore, the transcription Inhibitor kappa B alpha (of IkappaBalpha) was significantly increased by flavone. The results of the present study indicate that the flavonoid flavone could be a potential candidate for application in treatment of chronic inflammatory diseases. PARP-1 inhibition could have beneficial effects in such diseases as Chronic Obstructive Pulmonary Disease (COPD) and diabetes, by preservation of cellular NAD(+) levels and attenuating inflammatory conditions. PMID:17643414

  15. SLCO1B1 Variants and Angiotensin Converting Enzyme Inhibitor (Enalapril) -Induced Cough: a Pharmacogenetic Study

    PubMed Central

    Luo, Jian-Quan; He, Fa-Zhong; Wang, Zhen-Min; Sun, Ning-Ling; Wang, Lu-Yan; Tang, Gen-Fu; Liu, Mou-Ze; Li, Qing; Chen, Xiao-Ping; Liu, Zhao-Qian; Zhou, Hong-Hao; Zhang, Wei

    2015-01-01

    Clinical observations suggest that incidence of cough in Chinese taking angiotensin converting enzyme inhibitors is much higher than other racial groups. Cough is the most common adverse reaction of enalapril. We investigate whether SLCO1B1 genetic polymorphisms, previously reported to be important determinants of inter-individual variability in enalapril pharmacokinetics, are associated with the enalapril-induced cough. A cohort of 450 patients with essential hypertension taking 10?mg enalapril maleate were genotyped for the functional SLCO1B1 variants, 388A?>?G (Asn130Asp, rs2306283) and 521T?>?C (Val174Ala, rs4149056). The primary endpoint was cough, which was recorded when participants were bothered by cough and respiratory symptoms during enalapril treatment without an identifiable cause. SLCO1B1 521C allele conferred a 2-fold relative risk of enalapril-induced cough (95% confidence interval [CI]?=?1.34–3.04, P?=?6.2?×?10?4), and haplotype analysis suggested the relative risk of cough was 6.94-fold (95% CI?=?1.30–37.07, P?=?0.020) in SLCO1B1*15/*15 carriers. Furthermore, there was strong evidence for a gene-dose effect (percent with cough in those with 0, 1, or 2 copy of the 521C allele: 28.2%, 42.5%, and 71.4%, trend P?=?6.6?×?10?4). Our study highlights, for the first time, SLCO1B1 variants are strongly associated with an increased risk of enalapril-induced cough. The findings will be useful to provide pharmacogenetic markers for enalapril treatment. PMID:26607661

  16. Epilepsy and hippocampal neurodegeneration induced by glutamate decarboxylase inhibitors in awake rats.

    PubMed

    Salazar, Patricia; Tapia, Ricardo

    2015-10-01

    Glutamic acid decarboxylase (GAD), the enzyme responsible for GABA synthesis, requires pyridoxal phosphate (PLP) as a cofactor. Thiosemicarbazide (TSC) and ?-glutamyl-hydrazone (PLPGH) inhibit the free PLP-dependent isoform (GAD65) activity after systemic administration, leading to epilepsy in mice and in young, but not in adult rats. However, the competitive GAD inhibitor 3-mercaptopropionic acid (MPA) induces convulsions in both immature and adult rats. In the present study we tested comparatively the epileptogenic and neurotoxic effects of PLPGH, TSC and MPA, administered by microdialysis in the hippocampus of adult awake rats. Cortical EEG and motor behavior were analyzed during the next 2h, and aspartate, glutamate and GABA were measured by HPLC in the microdialysis-collected fractions. Twenty-four hours after drug administration rats were fixed for histological analysis of the hippocampus. PLPGH or TSC did not affect the motor behavior, EEG or cellular morphology, although the extracellular concentration of GABA was decreased. In contrast, MPA produced intense wet-dog shakes, EEG epileptiform discharges, a >75% reduction of extracellular GABA levels and remarkable neurodegeneration of the CA1 region, with >80% neuronal loss. The systemic administration of the NMDA glutamate receptor antagonist MK-801 30 min before MPA did not prevent the MPA-induced epilepsy but significantly protected against its neurotoxic effect, reducing neuronal loss to <30%. We conclude that in adult awake rats, drugs acting on PLP availability have only a weak effect on GABA neurotransmission, whereas direct GAD inhibition produced by MPA induces hyperexcitation leading to epilepsy and hippocampal neurodegeneration. Because this degeneration was prevented by the blockade of NMDA receptors, we conclude that it is due to glutamate-mediated excitotoxicity consequent to disinhibition of the hippocampal excitatory circuits. PMID:26354164

  17. SLCO1B1 Variants and Angiotensin Converting Enzyme Inhibitor (Enalapril) -Induced Cough: a Pharmacogenetic Study.

    PubMed

    Luo, Jian-Quan; He, Fa-Zhong; Wang, Zhen-Min; Sun, Ning-Ling; Wang, Lu-Yan; Tang, Gen-Fu; Liu, Mou-Ze; Li, Qing; Chen, Xiao-Ping; Liu, Zhao-Qian; Zhou, Hong-Hao; Zhang, Wei

    2015-01-01

    Clinical observations suggest that incidence of cough in Chinese taking angiotensin converting enzyme inhibitors is much higher than other racial groups. Cough is the most common adverse reaction of enalapril. We investigate whether SLCO1B1 genetic polymorphisms, previously reported to be important determinants of inter-individual variability in enalapril pharmacokinetics, are associated with the enalapril-induced cough. A cohort of 450 patients with essential hypertension taking 10?mg enalapril maleate were genotyped for the functional SLCO1B1 variants, 388A?>?G (Asn130Asp, rs2306283) and 521T?>?C (Val174Ala, rs4149056). The primary endpoint was cough, which was recorded when participants were bothered by cough and respiratory symptoms during enalapril treatment without an identifiable cause. SLCO1B1 521C allele conferred a 2-fold relative risk of enalapril-induced cough (95% confidence interval [CI]?=?1.34-3.04, P?=?6.2?×?10(-4)), and haplotype analysis suggested the relative risk of cough was 6.94-fold (95% CI?=?1.30-37.07, P?=?0.020) in SLCO1B1*15/*15 carriers. Furthermore, there was strong evidence for a gene-dose effect (percent with cough in those with 0, 1, or 2 copy of the 521C allele: 28.2%, 42.5%, and 71.4%, trend P?=?6.6?×?10(-4)). Our study highlights, for the first time, SLCO1B1 variants are strongly associated with an increased risk of enalapril-induced cough. The findings will be useful to provide pharmacogenetic markers for enalapril treatment. PMID:26607661

  18. Differential effects of glycolipid biosynthesis inhibitors on ceramide-induced cell death in neuroblastoma cells.

    PubMed

    Bieberich, E; Freischütz, B; Suzuki, M; Yu, R K

    1999-03-01

    An in vitro model of Gaucher's disease in murine neuroblastoma x rat glioma NG108-15 cells was used to investigate the physiological effects of two specific inhibitors of glucosylceramide synthase, d,l-threo-1-phenyl-2-decanoylamino-3-morpholino-1-propanol (d,l-PDMP) and N-butyldeoxynojirimycin (NB-DNJ), which have been suggested as agents for treatment of glycolipid storage disorders. Incubation of NG108-15 cells with conduritol-B-epoxide, a covalent inhibitor of glucosylceramidase, raised the intracellular concentration of glucosylceramide (GC) by more than fourfold, indicating a glycolipid composition equivalent to that of Gaucher's cells. The level of GC was decreased, and the cells were depleted of gangliosides by postincubation with d,l-PDMP or NB-DNJ. Treatment with d,l-PDMP, but not with NB-DNJ, resulted in a dose-dependent reduction of the growth rate and eventually caused cell death in NG108-15 cells on reaching confluency. An in situ detection assay using terminal nucleotidyltransferase indicated that cell degeneration was accompanied by apoptosis. Lipid analysis by high-performance TLC revealed that on incubation with d,l-PDMP, but not with NB-DNJ, the concentration of endogenous ceramide was elevated by threefold. Ceramide elevation and apoptosis were also observed when NG108-15 cells were incubated with daunorubicin, which was previously reported to induce programmed cell death by stimulation of ceramide synthesis. Structural characterization by HPLC and subsequent laser desorption mass spectrometry revealed that the endogenous ceramide contained fatty acids with chain lengths ranging from C14:0 to C24:0. The results indicate that elevation of levels of these ceramide species by incubation with d,l-PDMP or daunorubicin induces programmed cell death in NG108-15 cells. Because ceramide accumulation and cell death were not observed on incubation with NB-DNJ, its use is suggested to be less toxic than that of d,l-PDMP for treatment of Gaucher's disease and other sphingolipid storage disorders. PMID:10037475

  19. Transcriptional activation of peroxisome proliferator-activated receptor-{gamma} requires activation of both protein kinase A and Akt during adipocyte differentiation

    SciTech Connect

    Kim, Sang-pil; Ha, Jung Min; Yun, Sung Ji; Kim, Eun Kyoung; Chung, Sung Woon; Hong, Ki Whan; Kim, Chi Dae; Bae, Sun Sik

    2010-08-13

    Research highlights: {yields} Elevated cAMP activates both PKA and Epac. {yields} PKA activates CREB transcriptional factor and Epac activates PI3K/Akt pathway via Rap1. {yields} Akt modulates PPAR-{gamma} transcriptional activity in concert with CREB. -- Abstract: Peroxisome proliferator-activated receptor-{gamma} (PPAR-{gamma}) is required for the conversion of pre-adipocytes. However, the mechanism underlying activation of PPAR-{gamma} is unclear. Here we showed that cAMP-induced activation of protein kinase A (PKA) and Akt is essential for the transcriptional activation of PPAR-{gamma}. Hormonal induction of adipogenesis was blocked by a phosphatidylinositol 3-kinase (PI3K) inhibitor (LY294002), by a protein kinase A (PKA) inhibitor (H89), and by a Rap1 inhibitor (GGTI-298). Transcriptional activity of PPAR-{gamma} was markedly enhanced by 3-isobutyl-1-methylxanthine (IBMX), but not insulin and dexamethasone. In addition, IBMX-induced PPAR-{gamma} transcriptional activity was blocked by PI3K/Akt, PKA, or Rap1 inhibitors. 8-(4-Chlorophenylthio)-2'-O-methyl-cAMP (8-pCPT-2'-O-Me-cAMP) which is a specific agonist for exchanger protein directly activated by cAMP (Epac) significantly induced the activation of Akt. Furthermore, knock-down of Akt1 markedly attenuated PPAR-{gamma} transcriptional activity. These results indicate that both PKA and Akt signaling pathways are required for transcriptional activation of PPAR-{gamma}, suggesting post-translational activation of PPAR-{gamma} might be critical step for adipogenic gene expression.

  20. Modulation of curcumin-induced Akt phosphorylation and apoptosis by PI3K inhibitor in MCF-7 cells

    SciTech Connect

    Kizhakkayil, Jaleel; Thayyullathil, Faisal; Chathoth, Shahanas; Hago, Abdulkader; Patel, Mahendra; Galadari, Sehamuddin

    2010-04-09

    Curcumin has been shown to induce apoptosis in various malignant cancer cell lines. One mechanism of curcumin-induced apoptosis is through the PI3K/Akt signaling pathway. Akt, also known as protein kinase B (PKB), is a member of the family of phosphatidylinositol 3-OH-kinase regulated Ser/Thr kinases. The active Akt regulates cell survival and proliferation; and inhibits apoptosis. In this study we found that curcumin induces apoptotic cell death in MCF-7 cells, as assessed by MTT assay, DNA ladder formation, PARP cleavage, p53 and Bax induction. At apoptotic inducing concentration, curcumin induces a dramatic Akt phosphorylation, accompanied by an increased phosphorylation of glycogen synthase kinase 3{beta} (GSK3{beta}), which has been considered to be a pro-growth signaling molecule. Combining curcumin with PI3K inhibitor, LY290042, synergizes the apoptotic effect of curcumin. The inhibitor LY290042 was capable of attenuating curcumin-induced Akt phosphorylation and activation of GSK3{beta}. All together, our data suggest that blocking the PI3K/Akt survival pathway sensitizes the curcumin-induced apoptosis in MCF-7 cells.

  1. A Comparative Study of the Aneugenic and Polyploidy-inducing Effects of Fisetin and Two Model Aurora Kinase Inhibitors

    PubMed Central

    Gollapudi, P.; Hasegawa, L.S.; Eastmond, D.A.

    2014-01-01

    Fisetin, a plant flavonol commonly found in fruits, nuts and vegetables, is frequently added to nutritional supplements due to its reported cardioprotective, anti-carcinogenic and antioxidant properties. Earlier reports from our laboratory and others have indicated that fisetin has both aneugenic and clastogenic properties in cultured cells. More recently, fisetin has also been reported to target Aurora B kinase, a Ser/Thr kinase involved in ensuring proper microtubule attachment at the spindle assembly checkpoint, and an enzyme that is overexpressed in several types of cancer. Here we have further characterized the chromosome damage caused by fisetin and compared it with that induced by two known Aurora kinase inhibitors, VX-680 and ZM-447439, in cultured TK6 cells using the micronucleus assay with CREST staining as well as a flow cytometry-based assay that measures multiple types of numerical chromosomal aberrations. The three compounds were highly effective in inducing aneuploidy and polyploidy as evidenced by increases in kinetochore-positive micronuclei, hyperdiploidy, and polyploidy. With fisetin, however, the latter two effects were most significantly observed only after cells were allowed to overcome a cell cycle delay, and occurred at higher concentrations than those induced by the other Aurora kinase inhibitors. Modest increases in kinetochore-negative micronuclei were also seen with the model Aurora kinase inhibitors. These results indicate that fisetin induces multiple types of chromosome abnormalities in human cells, and indicate a need for a thorough investigation of fisetin-augmented dietary supplements. PMID:24680981

  2. Mechanisms of G1 cell cycle arrest and apoptosis in myeloma cells induced by hybrid-compound histone deacetylase inhibitor

    SciTech Connect

    Fujii, Seiko; Division of Maxillofacial Surgery, Kyushu Dental University ; Okinaga, Toshinori; Ariyoshi, Wataru; Oral Biology Research Center, Kyushu Dental University ; Takahashi, Osamu; Iwanaga, Kenjiro; Nishino, Norikazu; Tominaga, Kazuhiro; Nishihara, Tatsuji; Oral Biology Research Center, Kyushu Dental University

    2013-05-10

    Highlights: •Novel histone deacetylase inhibitor Ky-2, remarkably inhibits myeloma cell growth. •Ky-2 demonstrates no cytotoxicity against normal lymphocytic cells. •Ky-2 induces cell cycle arrest through the cell cycle-associated proteins. •Ky-2 induces Bcl-2-inhibitable apoptosis through a caspase-dependent cascade. -- Abstract: Objectives: Histone deacetylase (HDAC) inhibitors are new therapeutic agents, used to treat various types of malignant cancers. In the present study, we investigated the effects of Ky-2, a hybrid-compound HDAC inhibitor, on the growth of mouse myeloma cells. Materials and methods: Myeloma cells, HS-72, P3U1, and mouse normal cells were used in this study. Effect of HDAC inhibitors on cell viability was determined by WST-assay and trypan blue assay. Cell cycle was analyzed using flow cytometer. The expression of cell cycle regulatory and the apoptosis associated proteins were examined by Western blot analysis. Hoechst’s staining was used to detect apoptotic cells. Results: Our findings showed that Ky-2 decreased the levels of HDACs, while it enhanced acetylation of histone H3. Myeloma cell proliferation was inhibited by Ky-2 treatment. Interestingly, Ky-2 had no cytotoxic effects on mouse normal cells. Ky-2 treatment induced G1-phase cell cycle arrest and accumulation of a sub-G1 phase population, while Western blotting analysis revealed that expressions of the cell cycle-associated proteins were up-regulated. Also, Ky-2 enhanced the cleavage of caspase-9 and -3 in myeloma cells, followed by DNA fragmentation. In addition, Ky-2 was not found to induce apoptosis in bcl-2 overexpressing myeloma cells. Conclusion: These findings suggest that Ky-2 induces apoptosis via a caspase-dependent cascade and Bcl-2-inhibitable mechanism in myeloma cells.

  3. Investigation of the mechanism of protein denaturation by guanidine hydrochloride-induced dissociation of inhibitor-protease complexes.

    PubMed

    Qasim, Mohammad A; Taha, Mohammad

    2013-02-01

    In this communication we describe an approach in which guanidine hydrochloride-induced dissociation of a protein inhibitor-serine protease complex is used to explore the molecular basis of protein denaturation. The rationale behind this approach is that the inhibitor-protease complex is stabilized by the same types of non-covalent interactions that stabilize the native state of a protein. The dissociation of inhibitor-protease complex can be performed at concentrations of guanidine hydrochloride at which the inhibitor and the protease retain their native conformations. Here, we present our results on the effect of 0.1 M to 0.4 M guanidine hydrochloride concentrations on the association equilibrium constants (reciprocal of dissociation constant) of P?G, P?A, P?V, P?N, and P?S variants of turkey ovomucoid third domain with bovine ?-chymotrypsin. We use these results to calculate the free energy change in the dissociation of inhibitor-protease complexes (the m value) per mol of guanidine hydrochloride concentration. Our results agree with the general consensus that the denaturing effect of guanidine hydrochloride is due to its favorable interaction with the polar parts of proteins and that the non-polar side chains have no or little favorable interaction with guanidine hydrochloride. PMID:22894155

  4. PAS3, a Saccharomyces cerevisiae gene encoding a peroxisomal integral membrane protein essential for peroxisome biogenesis

    PubMed Central

    1991-01-01

    Saccharomyces cerevisiae pas3-mutants are described which conform the pas-phenotype recently reported for the peroxisomal assembly mutants pas1-1 and pas2 (Erdmann, R., M. Veenhuis, D. Mertens, and W.-H Kunau, 1989, Proc. Natl. Acad. Sci. USA. 86:5419-5423). The isolation of pas3- mutants enabled us to clone the PAS3 gene by functional complementation. DNA sequence analysis revealed a 50.6-kD protein with at least one domain of sufficient length and hydrophobicity to span a lipid bilayer. To verify these predictions antibodies were raised against a truncated portion of the PAS3 coding region overexpressed in E. coli. Pas3p was identified as a 48 kD peroxisomal integral membrane protein. It is shown that a lack of this protein causes the peroxisome- deficient phenotype and the cytosolic mislocalization of peroxisomal matrix enzymes. Based on protease digestion experiments Pas3p is discussed to be anchored in the peroxisomal membrane by its amino- terminus while the bulk of the molecule is exposed to the cytosol. These findings are consistent with the possibility that Pas3p is one component of the peroxisomal import machinery. PMID:1894692

  5. Catalase degradation in sunflower cotyledons during peroxisome transition from glyoxysomal to leaf peroxisomal function. [Helianthus annuus

    SciTech Connect

    Eising, R.; Gerhardt, B.

    1987-06-01

    First order rate constant for the degradation (degradation constants) of catalase in the cotyledons of sunflower (Helianthus annuus L.) were determined by measuring the loss of catalase containing /sup 14/C-labeled heme. During greening of the cotyledons, a period when peroxisomes change from glyoxysomal to leaf peroxisomal function, the degradation of glyoxysomal catalase is significantly slower than during all other stages of cotyledon development in light or darkness. The degradation constant during the transition stage of peroxisome function amounts to 0.205 day/sup -1/ in contrast to the constants ranging from 0.304 day/sup -1/ to 0.515 day/sup -1/ during the other developmental stages. Density labeling experiments comprising labeling of catalase with /sup 2/H/sub 2/O and its isopycnic centrifugation on CsCl gradients demonstrated that the determinations of the degradation constants were not substantially affected by reutilization of /sup 14/C-labeled compounds for catalase synthesis. The degradation constants for both glyoxysomal catalase and catalase synthesized during the transition of peroxisome function do not differ. This was shown by labeling the catalases with different isotopes and measuring the isotope ratio during the development of the cotyledons. The results are inconsistent with the concept that an accelerated and selective degradation of glyoxysomes underlies the change in peroxisome function. The data suggest that catalase degradation is at least partially due to an individual turnover of catalase and does not only result from a turnover of the whole peroxisomes.

  6. Suppression of Rat Oral Carcinogenesis by Agonists of Peroxisome Proliferator Activated Receptor ?

    PubMed Central

    McCormick, David L.; Horn, Thomas L.; Johnson, William D.; Peng, Xinjian; Lubet, Ronald A.; Steele, Vernon E.

    2015-01-01

    Peroxisome-proliferator-activated receptor ? (PPAR?) is a ligand-activated transcription factor that regulates cell proliferation, differentiation, and apoptosis. In vivo studies were performed to evaluate the activities of two thiazolidinedione PPAR? agonists, rosiglitazone and pioglitazone, as inhibitors of oral carcinogenesis in rats. Oral squamous cell carcinomas (OSCC) were induced in male F344 rats by 4-nitroquinoline-1-oxide (NQO; 20 ppm in the drinking water for 10 weeks). In each study, groups of 30 NQO-treated rats were exposed to a PPAR? agonist beginning at week 10 (one day after completion of NQO administration) or at week 17 (7 weeks post-NQO); chemopreventive agent exposure was continued until study termination at week 22 (rosiglitazone study) or week 24 (pioglitazone study). Administration of rosiglitazone (800 mg/kg diet) beginning at week 10 increased survival, reduced oral cancer incidence, and reduced oral cancer invasion score in comparison to dietary controls; however, chemopreventive activity was largely lost when rosiglitazone administration was delayed until week 17. Administration of pioglitazone (500 mg/kg diet beginning at week 10 or 1000 mg/kg diet beginning at week 17) induced significant reductions in oral cancer incidence without significant effects on OSCC invasion scores. Transcript levels of PPAR? and its three transcriptional variants (PPAR?v1, PPAR?v2, and PPAR?v3) were not significantly different in OSCC versus age- and site-matched phenotypically normal oral tissues from rats treated with NQO. These data suggest that PPAR? provides a useful molecular target for oral cancer chemoprevention, and that overexpression of PPAR? at the transcriptional level in neoplastic lesions is not essential for chemopreventive efficacy. PMID:26516762

  7. Defining the Plant Peroxisomal Proteome: From Arabidopsis to Rice

    PubMed Central

    Kaur, Navneet; Hu, Jianping

    2011-01-01

    Peroxisomes are small subcellular organelles mediating a multitude of processes in plants. Proteomics studies over the last several years have yielded much needed information on the composition of plant peroxisomes. In this review, the status of peroxisome proteomics studies in Arabidopsis and other plant species and the cumulative advances made through these studies are summarized. A reference Arabidopsis peroxisome proteome is generated, and some unique aspects of Arabidopsis peroxisomes that were uncovered through proteomics studies and hint at unanticipated peroxisomal functions are also highlighted. Knowledge gained from Arabidopsis was utilized to compile a tentative list of peroxisome proteins for the model monocot plant, rice. Differences in the peroxisomal proteome between these two model plants were drawn, and novel facets in rice were expounded upon. Finally, we discuss about the current limitations of experimental proteomics in decoding the complete and dynamic makeup of peroxisomes, and complementary and integrated approaches that would be beneficial to defining the peroxisomal metabolic and regulatory roadmaps. The synteny of genomes in the grass family makes rice an ideal model to study peroxisomes in cereal crops, in which these organelles have received much less attention, with the ultimate goal to improve crop yield. PMID:22645559

  8. Cholesterol transport through lysosome-peroxisome membrane contacts.

    PubMed

    Chu, Bei-Bei; Liao, Ya-Cheng; Qi, Wei; Xie, Chang; Du, Ximing; Wang, Jiang; Yang, Hongyuan; Miao, Hong-Hua; Li, Bo-Liang; Song, Bao-Liang

    2015-04-01

    Cholesterol is dynamically transported among organelles, which is essential for multiple cellular functions. However, the mechanism underlying intracellular cholesterol transport has remained largely unknown. We established an amphotericin B-based assay enabling a genome-wide shRNA screen for delayed LDL-cholesterol transport and identified 341 hits with particular enrichment of peroxisome genes, suggesting a previously unappreciated pathway for cholesterol transport. We show dynamic membrane contacts between peroxisome and lysosome, which are mediated by lysosomal Synaptotagmin VII binding to the lipid PI(4,5)P2 on peroxisomal membrane. LDL-cholesterol enhances such contacts, and cholesterol is transported from lysosome to peroxisome. Disruption of critical peroxisome genes leads to cholesterol accumulation in lysosome. Together, these findings reveal an unexpected role of peroxisome in intracellular cholesterol transport. We further demonstrate massive cholesterol accumulation in human patient cells and mouse model of peroxisomal disorders, suggesting a contribution of abnormal cholesterol accumulation to these diseases. PMID:25860611

  9. HDAC inhibitor misprocesses bantam oncomiRNA, but stimulates hid induced apoptotic pathway

    PubMed Central

    Bhadra, Utpal; Mondal, Tanmoy; Bag, Indira; Mukhopadhyay, Debasmita; Das, Paromita; Parida, Bibhuti B.; Mainkar, Prathama S.; Reddy, Chada Raji; Bhadra, Manika Pal

    2015-01-01

    Apoptosis or programmed cell death is critical for embryogenesis and tissue homeostasis. Uncontrolled apoptosis leads to different human disorders including immunodeficiency, autoimmune disorder and cancer. Several small molecules that control apoptosis have been identified. Here, we have shown the functional role of triazole derivative (DCPTN-PT) that acts as a potent HDAC inhibitor and mis-express proto onco microRNA (miRNA) bantam. To further understanding the mechanism of action of the molecule in apoptotic pathway, a series of experiments were also performed in Drosophila, a well known model organism in which the nature of human apoptosis is very analogous. DCPTN-PT mis processes bantam microRNA and alters its down regulatory target hid function and cleavage of Caspase-3 which in turn influence components of the mitochondrial apoptotic pathway in Drosophila. However regulatory microRNAs in other pro-apoptotic genes are not altered. Simultaneously, treatment of same molecule also affects the mitochondrial regulatory pathway in human tumour cell lines suggesting its conservative nature between fly and human. It is reasonable to propose that triazole derivative (DCPTN-PT) controls bantam oncomiRNA and increases hid induced apoptosis and is also able to influence mitochondrial apoptotic pathway. PMID:26442596

  10. Histone deacetylase inhibitors prevent activation-induced cell death and promote anti-tumor immunity

    PubMed Central

    Cao, K; Wang, G; Li, W; Zhang, L; Wang, R; Huang, Y; Du, L; Jiang, J; Wu, C; He, X; Roberts, A I; Li, F; Rabson, A B; Wang, Y; Shi, Y

    2015-01-01

    The poor efficacy of the in vivo anti-tumor immune response has been partially attributed to ineffective T-cell responses mounted against the tumor. Fas-FasL-dependent activation-induced cell death (AICD) of T cells is believed to be a major contributor to compromised anti-tumor immunity. The molecular mechanisms of AICD are well-investigated, yet the possibility of regulating AICD for cancer therapy remains to be explored. In this study, we show that histone deacetylase inhibitors (HDACIs) can inhibit apoptosis of CD4+ T cells within the tumor, thereby enhancing anti-tumor immune responses and suppressing melanoma growth. This inhibitory effect is specific for AICD through suppressing NFAT1-regulated FasL expression on activated CD4+ T cells. In gld/gld mice with mutation in FasL, the beneficial effect of HDACIs on AICD of infiltrating CD4+ T cells is not seen, confirming the critical role of FasL regulation in the anti-tumor effect of HDACIs. Importantly, we found that the co-administration of HDACIs and anti-CTLA4 could further enhance the infiltration of CD4+ T cells and achieve a synergistic therapeutic effect on tumor. Therefore, our study demonstrates that the modulation of AICD of tumor-infiltrating CD4+ T cells using HDACIs can enhance anti-tumor immune responses, uncovering a novel mechanism underlying the anti-tumor effect of HDACIs. PMID:25745993

  11. A New Generation Fatty Acid Amide Hydrolase Inhibitor Protects Against Kainate-Induced Excitotoxicity

    PubMed Central

    Naidoo, Vinogran; Nikas, Spyros P.; Karanian, David A.; Hwang, Jeannie; Zhao, Jianhong; Wood, JodiAnne T.; Alapafuja, Shakiru O.; Vadivel, Subramanian K.; Butler, David; Makriyannis, Alexandros; Bahr, Ben A.

    2010-01-01

    Endocannabinoids, including anandamide (AEA), have been implicated in neuroprotective on-demand responses. Related to such a response to injury, an excitotoxic kainic acid (KA) injection (i.p.) was found to increase AEA levels in the brain. To modulate the endocannabinoid response during events of excitotoxicity in vitro and in vivo, we utilized a new generation compound (AM5206) that selectively inhibits the AEA deactivating enzyme fatty acid amide hydrolase (FAAH). KA caused calpain-mediated spectrin breakdown, declines in synaptic markers, and disruption of neuronal integrity in cultured hippocampal slices. FAAH inhibition with AM5206 protected against the neurodegenerative cascade assessed in the slice model 24 h postinsult. In vivo, KA administration induced seizures and the same neurodegenerative events exhibited in vitro. When AM5206 was injected immediately after KA in rats, the seizure scores were markedly reduced as were levels of cytoskeletal damage and synaptic protein decline. The pre- and postsynaptic proteins were protected by the FAAH inhibitor to levels comparable to those found in healthy control brains. These data support the idea that endocannabinoids are released and converge on pro-survival pathways that prevent excitotoxic progression. PMID:21069475

  12. ROCK inhibitor and feeder cells induce the conditional reprogramming of epithelial cells.

    PubMed

    Liu, Xuefeng; Ory, Virginie; Chapman, Sandra; Yuan, Hang; Albanese, Chris; Kallakury, Bhaskar; Timofeeva, Olga A; Nealon, Caitlin; Dakic, Aleksandra; Simic, Vera; Haddad, Bassem R; Rhim, Johng S; Dritschilo, Anatoly; Riegel, Anna; McBride, Alison; Schlegel, Richard

    2012-02-01

    We demonstrate that a Rho kinase inhibitor (Y-27632), in combination with fibroblast feeder cells, induces normal and tumor epithelial cells from many tissues to proliferate indefinitely in vitro, without transduction of exogenous viral or cellular genes. Primary prostate and mammary cells, for example, are reprogrammed toward a basaloid, stem-like phenotype and form well-organized prostaspheres and mammospheres in Matrigel. However, in contrast to the selection of rare stem-like cells, the described growth conditions can generate 2 × 10(6) cells in 5 to 6 days from needle biopsies, and can generate cultures from cryopreserved tissue and from fewer than four viable cells. Continued cell proliferation is dependent on both feeder cells and Y-27632, and the conditionally reprogrammed cells (CRCs) retain a normal karyotype and remain nontumorigenic. This technique also efficiently establishes cell cultures from human and rodent tumors. For example, CRCs established from human prostate adenocarcinoma displayed instability of chromosome 13, proliferated abnormally in Matrigel, and formed tumors in mice with severe combined immunodeficiency. The ability to rapidly generate many tumor cells from small biopsy specimens and frozen tissue provides significant opportunities for cell-based diagnostics and therapeutics (including chemosensitivity testing) and greatly expands the value of biobanking. In addition, the CRC method allows for the genetic manipulation of epithelial cells ex vivo and their subsequent evaluation in vivo in the same host. PMID:22189618

  13. Modulation of oxazolone-induced hypersensitivity in mice by selective PDE inhibitors

    PubMed Central

    Moodley, I.; Sotsios, Y.

    1995-01-01

    The effects of PDE inhibitors on oxazolone-induced contact hypersensitivity (CS) were studied in mice. Rolipram, Ro 20-1724 and theophylline dose dependently inhibited CS but none caused >53% inhibition. ED30 values at 24 h before challenge for rolipram, Ro 20-1724 and theophylline were 2.1, 5.4 and 30.4 mg/kg, p.o., respectively. Milrinone and SKF 94836 at 30 mg/kg caused a small, but significant inhibition of 13% and 18%, respectively, although the inhibition (8%) caused by zaprinast was not significant. Betamethasone (10 mg/kg, p.o.) caused a marked inhibition (80%) as did indomethacin (65% at 5 mg/kg, p.o.). Rolipram and Ro 20-1724 inhibited proliferation of mouse lymphoblasts with IC50 values of 0.08 ?M and 0.83 ?M, respectively. In contrast, zaprinast caused only a weak inhibition (IC50 = 119 ?M) of lymphocyte proliferation, whereas SKF 94836 and theophylline failed to cause any significant inhibition at 100 ?M (26% and 2%, respectively). These findings suggest that PDE IV isozymes play a principal role in mediating CS by inhibiting lymphocyte activation. PMID:18475626

  14. Early events in vascular injury in the rat induced by the phosphodiesterase IV inhibitor SCH 351591.

    PubMed

    Weaver, James L; Zhang, Jun; Knapton, Alan; Miller, Terry; Espandiari, Parvaneh; Smith, Roger; Gu, Yi-Zhong; Snyder, Ronald D

    2010-08-01

    Treatment with drugs from multiple classes induces vascular injury with medial necrosis, hemorrhage, endothelial damage, and inflammation. Previous research has suggested early events might be occurring well in advance of the full lesions that appear forty-eight to seventy-two hours after dosing with SCH 351591, a PDE IV inhibitor. This study was performed to study early events in detail. Rats were dosed with 20 mg/kg of drug by gavage and sacrificed at times between fifteen and 240 minutes after dosing. Tissues were collected for histopathological analysis and gene expression studies. Serum was collected for biomarker analysis. The data from biomarker analysis showed a three-part response with an early phase that was maximal at fifteen to thirty minutes, a second phase from forty-five to 180 minutes, and the third phase that was starting to rise at four hours. The first phase included increases in lymphocytes, serum histamine, and serum nitrite. The second phase shows continued elevation of serum nitrite. The third phase was marked by an increase in serum GRO/CINC-1. At fifteen minutes, histopathology showed activation of mast cells, but not degranulation. Increases in endothelial activation and perivascular inflammatory cells were first apparent at thirty minutes and increased through 240 minutes. PMID:20585143

  15. Protease inhibitor reduces airway response and underlying inflammation in cockroach allergen-induced murine model.

    PubMed

    Saw, Sanjay; Arora, Naveen

    2015-04-01

    Protease(s) enhances airway inflammation and allergic cascade. In the present study, effect of a serine protease inhibitor was evaluated in mouse model of airway disease. Mice were sensitized with cockroach extract (CE) or Per a 10 and treated with 4-(2-aminoethyl) benzenesulfonyl fluoride hydrochloride (AEBSF) 1 h before or after challenge to measure airway response. Mice were euthanized to collect bronchoalveolar lavage fluid (BALF), blood, and lung to evaluate inflammation. AEBSF treatment significantly reduced the AHR in allergen-challenged mice in dose-dependent manner (p??0.01). IgE (p?0.05) and Th2 cytokines (p?0.05) were significantly reduced in treated mice. AEBSF treatment lowered total cell (p?0.05), eosinophil (p?0.05), and neutrophil (p?0.05) in BALF and lung tissue. Oxidative stress parameters were impaired on treatment in allergen-challenged mice (p?0.05). AEBSF had therapeutic effect in allergen-induced airway resistance and underling inflammation and had potential for combination or as add-on therapy for respiratory diseases. PMID:25052477

  16. The NF-?B inhibitor parthenolide interacts with histone deacetylase inhibitors to induce MKK7/JNK1-dependent apoptosis in human acute myeloid leukaemia cells

    PubMed Central

    Dai, Yun; Guzman, Monica L.; Chen, Shuang; Wang, Li; Yeung, Sin-Kei; Pei, Xin-Yan; Dent, Paul; Jordan, Craig T.; Grant, Steven

    2010-01-01

    Summary Interactions between the nuclear factor (NF)-?B inhibitor parthenolide and the pan-histone deacetylase inhibitors (HDACIs) vorinostat and LBH589 were investigated in human acute myeloid leukaemia (AML) cells, including primary AML blasts. Co-administration of parthenolide blocked HDACI-mediated phosphorylation/activation of IKK and RelA/p65 in association with increased JNK1 activation in various AML cell types. These events were accompanied by an increase in apoptosis in multiple AML cell lines (e.g. U937, HL-60, NB4, MV-4-11, and MOLM-13). Significantly, parthenolide also increased HDACI-mediated cell death in haematopoietic cells transduced with the MLL-MLLT1 fusion gene, which exhibit certain leukaemia-initiating cell characteristics, as well as primary AML blasts. Exposure to parthenolide/HDACI regimens clearly inhibited the growth of AML-colony-forming units but was relatively sparing toward normal haematopoietic progenitors. Notably, blockade of JNK signaling by either pharmacological inhibitors or genetic means (e.g., dominant-negative JNK1 or JNK1 shRNA) diminished parthenolide/HDACI-mediated lethality. Moreover, dominant-negative MKK7, but not dominant-negative MKK4/SEK1, blocked JNK1 activation and apoptosis induced by parthenolide/HDACI regimens. Together, these findings indicate that parthenolide potentiates HDACI lethality in human AML cells through a process involving NF-?B inhibition and subsequent MKK7-dependent activation of the SAPK/JNK pathway. They also raise the possibility that this strategy may target leukaemic progenitor cells. PMID:20701602

  17. A receptor tyrosine kinase inhibitor, Tyrphostin A9 induces cancer cell death through Drp1 dependent mitochondria fragmentation

    SciTech Connect

    Park, So Jung; Park, Young Jun; Shin, Ji Hyun; Kim, Eun Sung; Hwang, Jung Jin; Jin, Dong-Hoon; Kim, Jin Cheon; Cho, Dong-Hyung

    2011-05-13

    Highlights: {yields} We screened and identified Tyrphostin A9, a receptor tyrosine kinase inhibitor as a strong mitochondria fission inducer. {yields} Tyrphostin A9 treatment promotes mitochondria dysfunction and contributes to cytotoxicity in cancer cells. {yields} Tyrphostin A9 induces apoptotic cell death through a Drp1-mediated pathway. {yields} Our studies suggest that Tyrphostin A9 induces mitochondria fragmentation and apoptotic cell death via Drp1 dependently. -- Abstract: Mitochondria dynamics controls not only their morphology but also functions of mitochondria. Therefore, an imbalance of the dynamics eventually leads to mitochondria disruption and cell death. To identify specific regulators of mitochondria dynamics, we screened a bioactive chemical compound library and selected Tyrphostin A9, a tyrosine kinase inhibitor, as a potent inducer of mitochondrial fission. Tyrphostin A9 treatment resulted in the formation of fragmented mitochondria filament. In addition, cellular ATP level was decreased and the mitochondrial membrane potential was collapsed in Tyr A9-treated cells. Suppression of Drp1 activity by siRNA or over-expression of a dominant negative mutant of Drp1 inhibited both mitochondrial fragmentation and cell death induced by Tyrpohotin A9. Moreover, treatment of Tyrphostin A9 also evoked mitochondrial fragmentation in other cells including the neuroblastomas. Taken together, these results suggest that Tyrphostin A9 induces Drp1-mediated mitochondrial fission and apoptotic cell death.

  18. Chicago sky blue 6B, a vesicular glutamate transporters inhibitor, attenuates methamphetamine-induced hyperactivity and behavioral sensitization in mice.

    PubMed

    He, Zongsheng; Yan, Lingdi; Yong, Zheng; Dong, Zhaoqi; Dong, Huajin; Gong, Zehui

    2013-02-15

    Several lines of evidence demonstrate that glutamatergic system plays an important role in drug addiction. The present study was designed to investigate the effects of Chicago sky blue 6B (CSB6B), a vesicular glutamate transporters (VGLUTs) inhibitor, on methamphetamine (METH)-induced behaviors in mice. Mice were induced behavioral sensitization to METH by subcutaneous injection of 1mg/kg METH once daily for 7 days and then challenged with 1mg/kg METH in 14th day. Intracerebroventricular administration of CSB6B (7.5?g) 2.5h prior to METH was to observe its effects on METH -induced behavioral sensitization. Our results showed that the expressions of behavioral sensitization were significantly attenuated by intracerebroventricular administration of CSB6B 2.5h prior to METH either during the development period or before methamphetamine challenge in mice, while CSB6B itself had no effect on locomotor activity. Meanwhile, pretreatment of CSB6B also attenuated hyperactivity caused by a single injection of METH in mice. These results demonstrated that CSB6B, a VGLUTs inhibitor, attenuated acute METH-induced hyperactivity and chronic METH-induced behavioral sensitization, which indicated that VGLUTs were involved in the effect of chronic METH-induced behavioral sensitization and may be a new target against the addiction of METH. PMID:23159705

  19. An Efficient Positive Selection Procedure for the Isolation of Peroxisomal Import and Peroxisome Assembly Mutants of Saccharomyces Cerevisiae

    PubMed Central

    Elgersma, Y.; van-den-Berg, M.; Tabak, H. F.; Distel, B.

    1993-01-01

    To study peroxisome biogenesis, we developed a procedure to select for Saccharomyces cerevisiae mutants defective in peroxisomal protein import or peroxisome assembly. For this purpose, a chimeric gene was constructed encoding the bleomycin resistance protein linked to the peroxisomal protein luciferase. In wild-type cells this chimeric protein is imported into the peroxisome, which prevents the neutralizing interaction of the chimeric protein with its toxic phleomycin ligand. Peroxisomal import and peroxisome assembly mutants are unable to import this chimeric protein into their peroxisomes. This enables the bleomycin moiety of the chimeric protein to bind phleomycin, thereby preventing its toxicity. The selection is very efficient: upon mutagenesis, 84 (10%) of 800 phleomycin resistant colonies tested were unable to grow on oleic acid. This rate could be increased to 25% using more stringent selection conditions. The selection procedure is very specific; all oleic acid non utilizing (onu) mutants tested were disturbed in peroxisomal import and/or peroxisome assembly. The pas (peroxisome assembly) mutants that have been used for complementation analysis represent 12 complementation groups including three novel ones, designated pas20, pas21 and pas22. PMID:7507454

  20. The X-Linked Inhibitor of Apoptosis Protein Inhibitor Embelin Suppresses Inflammation and Bone Erosion in Collagen Antibody Induced Arthritis Mice

    PubMed Central

    Dharmapatni, Anak A. S. S. K.; Cantley, Melissa D.; Marino, Victor; Perilli, Egon; Crotti, Tania N.; Smith, Malcolm D.; Haynes, David R.

    2015-01-01

    Objective. To investigate the effect of Embelin, an inhibitor of X-Linked Inhibitor of Apoptosis Protein (XIAP), on inflammation and bone erosion in a collagen antibody induced arthritis (CAIA) in mice. Methods. Four groups of mice (n = 6 per group) were allocated: CAIA untreated mice, CAIA treated with Prednisolone (10?mg/kg/day), CAIA treated with low dose Embelin (30?mg/kg/day), and CAIA treated with high dose Embelin (50?mg/kg/day). Joint inflammation was evaluated using clinical paw score and histological assessments. Bone erosion was assessed using micro-CT, tartrate resistant acid phosphatase (TRAP) staining, and serum carboxy-terminal collagen crosslinks (CTX-1) ELISA. Immunohistochemistry was used to detect XIAP protein. TUNEL was performed to identify apoptotic cells. Results. Low dose, but not high dose Embelin, suppressed inflammation as reflected by lower paw scores (P < 0.05) and lower histological scores for inflammation. Low dose Embelin reduced serum CTX-1 (P < 0.05) and demonstrated lower histological score and TRAP counting, and slightly higher bone volume as compared to CAIA untreated mice. XIAP expression was not reduced but TUNEL positive cells were more abundant in Embelin treated CAIA mice. Conclusion. Low dose Embelin suppressed inflammation and serum CTX-1 in CAIA mice, indicating a potential use for Embelin to treat pathological bone loss. PMID:26347311

  1. NAAG peptidase inhibitors block cognitive deficit induced by MK-801 and motor activation induced by d-amphetamine in animal models of schizophrenia

    PubMed Central

    Olszewski, R T; Janczura, K J; Ball, S R; Madore, J C; Lavin, K M; Lee, J C-M; Lee, M J; Der, E K; Hark, T J; Farago, P R; Profaci, C P; Bzdega, T; Neale, J H

    2012-01-01

    The most widely validated animal models of the positive, negative and cognitive symptoms of schizophrenia involve administration of d-amphetamine or the open channel NMDA receptor blockers, dizocilpine (MK-801), phencyclidine (PCP) and ketamine. The drug ZJ43 potently inhibits glutamate carboxypeptidase II (GCPII), an enzyme that inactivates the peptide transmitter N-acetylaspartylglutamate (NAAG) and reduces positive and negative behaviors induced by PCP in several of these models. NAAG is an agonist at the metabotropic glutamate receptor 3 (mGluR3). Polymorphisms in this receptor have been associated with expression of schizophrenia. This study aimed to determine whether two different NAAG peptidase inhibitors are effective in dopamine models, whether their efficacy was eliminated in GCPII knockout mice and whether the efficacy of these inhibitors extended to MK-801-induced cognitive deficits as assessed using the novel object recognition test. ZJ43 blocked motor activation when given before or after d-amphetamine treatment. (R,S)-2-phosphono-methylpentanedioic acid (2-PMPA), another potent NAAG peptidase inhibitor, also reduced motor activation induced by PCP or d-amphetamine. 2-PMPA was not effective in GCPII knockout mice. ZJ43 and 2-PMPA also blocked MK-801-induced deficits in novel object recognition when given before, but not after, the acquisition trial. The group II mGluR antagonist LY341495 blocked the effects of NAAG peptidase inhibition in these studies. 2-PMPA was more potent than ZJ43 in a test of NAAG peptidase inhibition in vivo. By bridging the dopamine and glutamate theories of schizophrenia with two structurally different NAAG peptidase inhibitors and demonstrating their efficacy in blocking MK-801-induced memory deficits, these data advance the concept that NAAG peptidase inhibition represents a potentially novel antipsychotic therapy. PMID:22850437

  2. Efficacy of Cilostazol a selective phosphodiesterase-3 inhibitor in rat model of Streptozotocin diabetes induced vascular dementia.

    PubMed

    Kumar, Ashwani; Kumar, Amit; Jaggi, Amteshwar S; Singh, Nirmal

    2015-08-01

    The present study has been designed to investigate the potential of Cilostazol a phosphodiesterase-3 (PDE-3) inhibitor in diabetes-induced vascular dementia (Vad) employing Wistar rats. A single dose of Streptozotocin (STZ) was used for the induction of diabetes and subsequent Vad in rats. Memory and learning abilities of rats were evaluated with Morris water maze (MWM) test. Serum glucose, body weight, vascular endothelial function, serum nitrite/nitrate levels, brain oxidative stress levels (viz. brain thiobarbituric acid reactive species and reduced glutathione levels), inflammatory markers (viz. brain myeloperoxidase activity and neutrophil infiltration in the brain hippocampal area) and brain acetylcholinesterase activity were also tested. Donepezil was used as positive control. Streptozotocin treated animals showed poor performance on MWM indicating impairment of learning and memory abilities with a significant reduction in body weight, vascular endothelial function, serum nitrite/nitrate levels, along with an increase in serum glucose, brain oxidative stress levels, inflammatory changes and brain acetylcholinesterase activity. Treatment with selective PDE-3 inhibitor, Cilostazol significantly attenuated, diabetes-induced impairment of learning and memory; endothelial dysfunction, and changes in various biochemical parameters. It is concluded that selective PDE-3 inhibitor, Cilostazol may be considered as the potential pharmacological agent for the management of diabetes-induced vascular dementia. PMID:25987325

  3. Organic cadmium complexes as proteasome inhibitors and apoptosis inducers in human breast cancer cells

    PubMed Central

    Zhang, Zhen; Bi, Caifeng; Buac, Daniela; Fan, Yuhua; Zhang, Xia; Zuo, Jian; Zhang, Pengfei; Zhang, Nan; Dong, Lili; Dou, Q. Ping

    2013-01-01

    Although cadmium (Cd) is a widespread environmental contaminant and human carcinogen, our studies indicate an organic Cd complex to be a potent inhibitor of proteasomal chymotrypsin-like (CT-like) activity, further capable of inducing apoptosis in a cancer cell-specific manner. It has been reported that the ligands indole-3-butyric acid (L1) and indole-3-propionic acid (L2) have cancer-fighting effects when tested in a rat carcinoma model. In addition, 3, 5-diaminobenzoic acid o-vanillin Schiff bases (L3) have high antimicrobial activity and a large number of Schiff base complexes have been reported to have proteasome-inhibitory activity. We therefore hypothesized that synthetic forms of Cd in combination with L1, L2 and L3 may have proteasome-inhibitory and apoptosis-inducing activities, which would be cancer cell-specific. To test this hypothesis, we have synthesized three novel Cd-containing complexes: [Cd2(C12H12O2N)4(H2O)2]·2H2O (Cd1), [Cd2(C11H10O2N)4(H2O)2]·2H2O (Cd2) and [Cd(C7H4N2O2)(C8H6O2)2]·2H2O (Cd3), by using these three ligands. We sought out to characterize and assess the proteasome-inhibitory and anti-proliferative properties of these three Cd complexes in human breast cancer cells. Cd1, Cd2 and Cd3 were found to effectively inhibit the chymotrypsin-like activity of purified 20S proteasome with IC50 values of 2.6, 3.0 and 3.3 ?M, respectively. Moreover, inhibition of cancer cell proliferation also correlated with this effect. As a result of proteasomal shutdown, the accumulation of ubiquitinated proteins and the proteasome target I?B-? protein as well as induction of apoptosis were observed. To account for the cancer specificity of this effect, immortalized, non-tumorigenic breast MCF10A cells were used under the same experimental conditions. Our results indicate that MCF10A cells are much less sensitive to the Cd1, Cd2 and Cd3 complexes when compared to MDA MB 231 breast cancer cells. Therefore, our study suggests that these Cd organic complexes are capable of inhibiting tumor cellular proteasome activity and consequently induce cancer cell-specific apoptotic death. PMID:23499788

  4. Gyrase inhibitors induce an oxidative damage cellular death pathway in Escherichia coli

    E-print Network

    Collins, James J.

    by gyrase inhibitors. We show that superoxide-mediated oxidation of iron­sulfur clusters promotes inhibitors; hydroxyl radicals; iron­sulfur cluster; oxidative damage; systems biology Introduction Bacterial a breakdown of iron regulatory dynamics; in turn, iron misregulation drives the generation of highly

  5. The MAO-B inhibitor deprenyl reduces the oral tremor and the dopamine depletion induced by the VMAT-2 inhibitor tetrabenazine.

    PubMed

    Podurgiel, Samantha J; Yohn, Samantha E; Dortche, Kristina; Correa, Merce; Salamone, John D

    2016-02-01

    Tetrabenazine (TBZ) is prescribed for the treatment of chorea associated with Huntington's disease. Via inhibition of the vesicular monoamine transporter (VMAT-2), TBZ blocks dopamine (DA) storage and depletes striatal DA; this drug also has been shown to induce Parkinsonian motor side effects in patients. Recently, TBZ was shown to induce tremulous jaw movements (TJMs) in rats and mice. TJMs are an oral tremor that has many of the characteristics of Parkinsonian tremor in humans. The present study focused upon the ability of the well-estabilished antiparkinsonian agent deprenyl to attenuate the behavioral and neurochemical effects of 2.0mg/kg TBZ. Deprenyl is a selective and irreversible inhibitor of monoamine oxidase-B, and administration of deprenyl produced a dose-related suppression of TBZ-induced TJMs. A second experiment employed in vivo microdialysis to examine extracellular DA levels in the ventrolateral striatum, the neostriatal region most closely associated with the production of TJMs, after administration of TBZ and deprenyl. Consistent with the behavioral data, TBZ alone produced a biphasic effect on extracellular DA, with an initial increases followed by a prolonged decrease during the period in which TJMs are displayed. Co-administration of deprenyl with TBZ increased DA levels compared to rats treated with TBZ alone. These results provide support for use of TBZ as a rodent model of Parkinsonism, and future studies should utilize this model to evaluate putative anti-Parkinsonian agents. PMID:26590367

  6. The Rho kinase inhibitor Y-27632 increases erythrocyte deformability and low oxygen tension-induced ATP release.

    PubMed

    Thuet, K M; Bowles, E A; Ellsworth, M L; Sprague, R S; Stephenson, A H

    2011-11-01

    Low oxygen (O(2)) tension and mechanical deformation are stimuli for ATP release from erythrocytes. It has been shown previously that rabbit erythrocytes made less deformable with diamide, a thiol cross-linking agent, release less ATP in response to low O(2) tension, suggesting a link between these two stimuli. In nonerythroid cells, activation of the Rho/Rho kinase signaling pathway has been reported to decrease cell deformability by altering Rho kinase-dependent cytoskeleton-protein interactions. We investigated the hypothesis that the Rho kinase inhibitor Y-27632 would increase erythrocyte deformability and thereby increase low O(2) tension-induced ATP release from erythrocytes. Here we show that Y-27632 (1 ?M) increases erythrocyte deformability (5%) and increases low O(2) tension-induced ATP release (203%) from healthy human erythrocytes. In addition, we found that, when erythrocytes were made less deformable by incubation with diamide (100 ?M), Y-27632 restored both deformability and low O(2) tension-induced ATP release to levels similar to those measured in the absence of diamide. These findings suggest that the Rho kinase inhibitor Y-27632 is able to reverse the diamide-induced decrease in erythrocyte deformability and rescue low O(2) tension-induced ATP release. These results further support a link between erythrocyte deformability and ATP release in response to low O(2) tension. PMID:21890695

  7. Histone deacetylase inhibitor valproic acid promotes the differentiation of human induced pluripotent stem cells into hepatocyte-like cells.

    PubMed

    Kondo, Yuki; Iwao, Takahiro; Yoshihashi, Sachimi; Mimori, Kayo; Ogihara, Ruri; Nagata, Kiyoshi; Kurose, Kouichi; Saito, Masayoshi; Niwa, Takuro; Suzuki, Takayoshi; Miyata, Naoki; Ohmori, Shigeru; Nakamura, Katsunori; Matsunaga, Tamihide

    2014-01-01

    In this study, we aimed to elucidate the effects and mechanism of action of valproic acid on hepatic differentiation from human induced pluripotent stem cell-derived hepatic progenitor cells. Human induced pluripotent stem cells were differentiated into endodermal cells in the presence of activin A and then into hepatic progenitor cells using dimethyl sulfoxide. Hepatic progenitor cells were matured in the presence of hepatocyte growth factor, oncostatin M, and dexamethasone with valproic acid that was added during the maturation process. After 25 days of differentiation, cells expressed hepatic marker genes and drug-metabolizing enzymes and exhibited drug-metabolizing enzyme activities. These expression levels and activities were increased by treatment with valproic acid, the timing and duration of which were important parameters to promote differentiation from human induced pluripotent stem cell-derived hepatic progenitor cells into hepatocytes. Valproic acid inhibited histone deacetylase activity during differentiation of human induced pluripotent stem cells, and other histone deacetylase inhibitors also enhanced differentiation into hepatocytes. In conclusion, histone deacetylase inhibitors such as valproic acid can be used to promote hepatic differentiation from human induced pluripotent stem cell-derived hepatic progenitor cells. PMID:25084468

  8. Histone Deacetylase Inhibitor Valproic Acid Promotes the Differentiation of Human Induced Pluripotent Stem Cells into Hepatocyte-Like Cells

    PubMed Central

    Kondo, Yuki; Iwao, Takahiro; Yoshihashi, Sachimi; Mimori, Kayo; Ogihara, Ruri; Nagata, Kiyoshi; Kurose, Kouichi; Saito, Masayoshi; Niwa, Takuro; Suzuki, Takayoshi; Miyata, Naoki; Ohmori, Shigeru; Nakamura, Katsunori; Matsunaga, Tamihide

    2014-01-01

    In this study, we aimed to elucidate the effects and mechanism of action of valproic acid on hepatic differentiation from human induced pluripotent stem cell-derived hepatic progenitor cells. Human induced pluripotent stem cells were differentiated into endodermal cells in the presence of activin A and then into hepatic progenitor cells using dimethyl sulfoxide. Hepatic progenitor cells were matured in the presence of hepatocyte growth factor, oncostatin M, and dexamethasone with valproic acid that was added during the maturation process. After 25 days of differentiation, cells expressed hepatic marker genes and drug-metabolizing enzymes and exhibited drug-metabolizing enzyme activities. These expression levels and activities were increased by treatment with valproic acid, the timing and duration of which were important parameters to promote differentiation from human induced pluripotent stem cell-derived hepatic progenitor cells into hepatocytes. Valproic acid inhibited histone deacetylase activity during differentiation of human induced pluripotent stem cells, and other histone deacetylase inhibitors also enhanced differentiation into hepatocytes. In conclusion, histone deacetylase inhibitors such as valproic acid can be used to promote hepatic differentiation from human induced pluripotent stem cell-derived hepatic progenitor cells. PMID:25084468

  9. Bentamapimod (JNK Inhibitor AS602801) Induces Regression of Endometriotic Lesions in Animal Models.

    PubMed

    Palmer, Stephen S; Altan, Melis; Denis, Deborah; Tos, Enrico Gillio; Gotteland, Jean-Pierre; Osteen, Kevin G; Bruner-Tran, Kaylon L; Nataraja, Selvaraj G

    2016-01-01

    Endometriosis is an estrogen (ER)-dependent gynecological disease caused by the growth of endometrial tissue at extrauterine sites. Current endocrine therapies address the estrogenic aspect of disease and offer some relief from pain but are associated with significant side effects. Immune dysfunction is also widely believed to be an underlying contributor to the pathogenesis of this disease. This study evaluated an inhibitor of c-Jun N-terminal kinase, bentamapimod (AS602801), which interrupts immune pathways, in 2 rodent endometriosis models. Treatment of nude mice bearing xenografts biopsied from women with endometriosis (BWE) with 30 mg/kg AS602801 caused 29% regression of lesion. Medroxyprogesterone acetate (MPA) or progesterone (PR) alone did not cause regression of BWE lesions, but combining 10 mg/kg AS602801 with MPA caused 38% lesion regression. In human endometrial organ cultures (from healthy women), treatment with AS602801 or MPA reduced matrix metalloproteinase-3 (MMP-3) release into culture medium. In organ cultures established with BWE, PR or MPA failed to inhibit MMP-3 secretion, whereas AS602801 alone or MPA + AS602801 suppressed MMP-3 production. In an autologous rat endometriosis model, AS602801 caused 48% regression of lesions compared to GnRH antagonist Antide (84%). AS602801 reduced inflammatory cytokines in endometriotic lesions, while levels of cytokines in ipsilateral horns were unaffected. Furthermore, AS602801 enhanced natural killer cell activity, without apparent negative effects on uterus. These results indicate that bentamapimod induced regression of endometriotic lesions in endometriosis rodent animal models without suppressing ER action. c-Jun N-terminal kinase inhibition mediated a comprehensive reduction in cytokine secretion and moreover was able to overcome PR resistance. PMID:26335175

  10. Dihydroorotate dehydrogenase inhibitor A771726 (leflunomide) induces apoptosis and diminishes proliferation of multiple myeloma cells.

    PubMed

    Baumann, Philipp; Mandl-Weber, Sonja; Völkl, Andreas; Adam, Christian; Bumeder, Irmgard; Oduncu, Fuat; Schmidmaier, Ralf

    2009-02-01

    Multiple myeloma is still an incurable disease; therefore, new therapeutics are urgently needed. A771726 is the active metabolite of the immunosuppressive drug leflunomide, which is currently applied in the treatment of rheumatoid arthritis, BK virus nephropathy, and cytomegaly viremia. Here, we show that dihydroorotate dehydrogenase (DHODH) is commonly expressed in multiple myeloma cell lines and primary multiple myeloma cells. The DHODH inhibitor A771726 inhibits cell growth in common myeloma cell lines at clinically achievable concentrations in a time- and dose-dependent manner. Annexin V-FITC/propidium iodide staining revealed induction of apoptosis of multiple myeloma cell lines and primary multiple myeloma cells. The 5-bromo-2'-deoxyuridine cell proliferation assay showed that inhibition of cell growth was partly due to inhibition of multiple myeloma cell proliferation. A771726 induced G(1) cell cycle arrest via modulation of cyclin D2 and pRb expression. A771726 decreased phosphorylation of protein kinase B (Akt), p70S6K, and eukaryotic translation initiation factor 4E-binding protein-1 as shown by Western blotting experiments. Furthermore, we show that the stimulatory effect of conditioned medium of HS-5 bone marrow stromal cells on multiple myeloma cell growth is completely abrogated by A771726. In addition, synergism studies revealed synergistic and additive activity of A771726 together with the genotoxic agents melphalan, treosulfan, and doxorubicin as well as with dexamethasone and bortezomib. Taken together, we show that inhibition of DHODH by A771726/leflunomide is effective in multiple myeloma. Considering the favorable toxicity profile and the great clinical experience with leflunomide in rheumatoid arthritis, this drug represents a potential new candidate for targeted therapy in multiple myeloma. PMID:19174558

  11. Protective effects of amifostine and cyclooxygenase-1 inhibitor against normal human epidermal keratinocyte toxicity induced by methotrexate and 5-fluorouracil.

    PubMed

    Maiguma, Takayoshi; Kaji, Hiroaki; Makino, Kazutaka; Teshima, Daisuke

    2009-07-01

    Our study aimed to find more effective protective agents against mucosa toxicity induced by methotrexate and 5-fluorouracil. We focused on the relationship between oral mucositis and keratinocyte injury and examined methotrexate and 5-fluorouracil-induced cytotoxicity in normal human epidermal keratinocyte cell lines. Cell viability and superoxide radical activity were measured based on converting WST-1 (4-[3-(4-indophenyl)-2-(4-nitrophenyl)-2H-5-tetrazolio]-1,3-benzen disulfonate) to a water-soluble formazan dye. DNA synthesis by 5-bromo-2'-deoxyuridine incorporation was measured as an indirect parameter of cell proliferation. Allopurinol and amifostine were used as the radical scavengers. l-glutamine was used as a mucosa-protective agent. A cyclooxygenase inhibitor interrupting the production of hydroxyl radicals in the arachidonic acid cascade was also examined. 5-fluorouracil and methotrexate caused cytotoxicity due to the activation of intracellular superoxide radicals specifically on normal human epidermal keratinocytes. From the electron spin resonance study, it was found that allopurinol was a superoxide radical scavenger, while amifostine was hydroxyl radical scavenger. Allopurinol showed no effect on the cytotoxicity due to 5-fluorouracil and methotrexate. The cell injury induced by methotrexate was restored by amifostine. However, the cell injury induced by 5-fluorouracil was markedly recovered by a selective cyclooxygenase-1 inhibitor compared to amifostine. It was suggested that amifostine and cyclooxygenase-1 inhibitor could be useful protective agents against methotrexate and 5-fluorouracil chemotherapeutic toxicity. Additionally, this in vitro cell injury model using normal human epidermal keratinocytes may be useful for understanding the pathophysiology of oral mucositis induced by chemotherapeutic agents. PMID:19371262

  12. High-level expression and molecular cloning of genes encoding Candida tropicalis peroxisomal proteins.

    PubMed Central

    Kamiryo, T; Okazaki, K

    1984-01-01

    The development of peroxisomes in the cells of Candida tropicalis grown on oleic acid was accompanied by a markedly high expression of peroxisomal proteins. On the basis of this finding, the nuclear DNA library of this yeast was screened by differential hybridization, and 102 clones of oleic acid-inducible sequences were isolated. Seven coding regions were found to form clusters in three stretches of the genomic DNA. Five of the regions were identified as genes for peroxisomal polypeptides (PXPs). The coding sequence for PXP-2 hybrid selected an additional mRNA for PXP-4, the subunit of long-chain acyl coenzyme A oxidase, which was the most abundant PXP. PXP-2 and PXP-4 were close in apparent molecular weight and generated similar peptides when digested with a protease. The gene for PXP-4 was adjacent to that for PXP-2 on the genome and also hybridized to the mRNA coding for PXP-5. These and other similar results suggest that the genes for the peroxisomal proteins of this organism arose by duplication of a few ancestral genes. Images PMID:6504042

  13. Reduced estradiol synthesis by letrozole, an aromatase inhibitor, is protective against development of pentylenetetrazole-induced kindling in mice.

    PubMed

    Rashid, Davood; Panda, B P; Vohora, Divya

    2015-11-01

    Neurosteroids, such as testosterone and their metabolites, are known to modulate neuronal excitability. The enzymes regulating the metabolism of these neurosteroids, thus, may be targeted as a noval strategy for the development of new antiepileptic drugs. The present work targeted two such enzymes i,e aromatase and 5?-reductase in order to explore the potential of letrozole (an aromatase inhibitor) on pentylenetetrazole (PTZ)-induced kindling in mice and the ability of finasteride (a 5?-reductase inhibitor) to modulate any such effects. PTZ (30 mg/kg, i.p.), when administered once every two days (for a total of 24 doses) induced kindling in Swiss albino mice. Letrozole (1 mg/kg, p.o.), administered prior to PTZ, significantly reduced the % incidence of kindling, delayed mean onset time of seizures and reduced seizure severity score. Letrozole reduced the levels of plasma 17?-estradiol after induction of kindling. The concurrent administration of finasteride and letrozole produced effects similar to letrozole on PTZ-kindling and on estradiol levels. This implies that the ability of letrozole to redirect the synthesis of dihydrotestosterone (DHT) and 5?-androstanediol from testosterone doesn't appear to play a significant role in the protective effects of letrozole against PTZ kindling. Letrozole, however, increased the levels of 5?-DHT in mice plasma. The aromatase inhibitors, thus, may be exploited for inhibiting the synthesis of proconvulsant (17?-estradiol) and/or redirecting the synthesis of anticonvulsant (DHT and 5?-androstanediol) neurosteroids. PMID:26449311

  14. Peroxisomal D-bifunctional protein deficiency

    PubMed Central

    Lines, Matthew A.; Jobling, Rebekah; Brady, Lauren; Marshall, Christian R.; Scherer, Stephen W.; Rodriguez, Amadeo R.; Lee, Liesly; Lang, Anthony E.; Mestre, Tiago A.; Wanders, Ronald J.A.; Ferdinandusse, Sacha

    2014-01-01

    Objective: To determine the causative genetic lesion in 3 adult siblings with a slowly progressive, juvenile-onset phenotype comprising cerebellar atrophy and ataxia, intellectual decline, hearing loss, hypogonadism, hyperreflexia, a demyelinating sensorimotor neuropathy, and (in 2 of 3 probands) supratentorial white matter changes, in whom numerous prior investigations were nondiagnostic. Methods: The patients’ initial clinical assessment included history and physical examination, cranial MRI, and nerve conduction studies. We performed whole-exome sequencing of all 3 probands, followed by variant annotation and selection of rare, shared, recessive coding changes to identify the gene responsible. We next performed a panel of peroxisomal investigations in blood and cultured fibroblasts, including assessment of D-bifunctional protein (DBP) stability and activity by immunoblot and enzymologic methods, respectively. Results: Exome sequencing identified compound heterozygous mutations in HSD17B4, encoding peroxisomal DBP, in all 3 probands. Both identified mutations alter a conserved residue within the active site of DBP’s enoyl-CoA hydratase domain. Routine peroxisomal screening tests, including very long-chain fatty acids and phytanic acid, were normal. DBP enzymatic activity was markedly reduced. Conclusion: Exome sequencing provides a powerful and elegant tool in the specific diagnosis of “mild” or “atypical” neurometabolic disorders. Given the broad differential diagnosis and the absence of detectable biochemical abnormalities in blood, molecular testing of HSD17B4 should be considered as a first-line investigation in patients with compatible features. PMID:24553428

  15. Multikinase Inhibitors Induce Cutaneous Toxicity through OAT6-Mediated Uptake and MAP3K7-Driven Cell Death.

    PubMed

    Zimmerman, Eric I; Gibson, Alice A; Hu, Shuiying; Vasilyeva, Aksana; Orwick, Shelley J; Du, Guoqing; Mascara, Gerard P; Ong, Su Sien; Chen, Taosheng; Vogel, Peter; Inaba, Hiroto; Maitland, Michael L; Sparreboom, Alex; Baker, Sharyn D

    2016-01-01

    The use of multikinase inhibitors (MKI) in oncology, such as sorafenib, is associated with a cutaneous adverse event called hand-foot skin reaction (HFSR), in which sites of pressure or friction become inflamed and painful, thus significantly impacting quality of life. The pathogenesis of MKI-induced HFSR is unknown, and the only available treatment options involve dose reduction or discontinuation of therapy, which have negative effects on primary disease management. To investigate the underlying mechanisms by which sorafenib promotes keratinocyte cytotoxicity and subsequent HFSR induction, we performed a transporter-directed RNAi screen in human epidermal keratinocytes and identified SLC22A20 (OAT6) as an uptake carrier of sorafenib. Further investigations into the intracellular mechanism of sorafenib activity through in situ kinome profiling identified the mitogen-activated protein kinase MAP3K7 (TAK1) as a target of sorafenib that induces cell death. Finally, we demonstrate that sorafenib induced keratinocyte injury in vivo and that this effect could be reversed by cotreatment with the OAT6 inhibitor probenecid. Collectively, our findings reveal a novel pathway that regulates the entry of some MKIs into keratinocytes and explains the basis underlying sorafenib-induced skin toxicity, with important implications for the therapeutic management of HFSR. Cancer Res; 76(1); 117-26. ©2015 AACR. PMID:26677977

  16. Recombinant Buckwheat Trypsin Inhibitor Induces Mitophagy by Directly Targeting Mitochondria and Causes Mitochondrial Dysfunction in Hep G2 Cells.

    PubMed

    Wang, Zhuanhua; Li, Shanshan; Ren, Rong; Li, Jiao; Cui, Xiaodong

    2015-09-01

    Mitochondria are essential targets for cancer chemotherapy and other disease treatments. Recombinant buckwheat trypsin inhibitor (rBTI), a member of the potato type I proteinase inhibitor family, was derived from tartary buckwheat extracts. Our results showed that rBTI directly targeted mitochondria and induced mitochondrial fragmentation and mitophagy. This occurs through enhanced depolarization of the mitochondrial membrane potential, increasing reactive oxygen species (ROS) generation associated with the rise of the superoxide dismutase and catalase activity and glutathione peroxidase (GSH) content, and changes in the GSH/oxidized glutathione ratio. Mild and transient ROS induced by rBTI were shown to be important signaling molecules required to induce Hep G2 mitophagy to remove dysfunctional mitochondria. Furthermore, rBTI could directly induce mitochondrial fragmentation. It was also noted that rBTI highly increased colocalization of mitochondria in treated cells compared to nontreated cells. Tom 20, a subunit of the translocase of the mitochondrial outer membrane complex responsible for recognizing mitochondrial presequences, may be the direct target of rBTI. PMID:26301894

  17. The effect of an specific inducible NO synthase inhibitor, S-methylisothiourea hemisulfate on cisplatin-induced nephrotoxicity; gender-related differences

    PubMed Central

    Ghayyoomi, Mansooreh; Soltani, Nepton; Nematbakhsh, Mehdi; Moslemi, Fatemeh; Talebi, Ardeshir; Shirdavani, Soheila; Razmjoo, Farzaneh

    2015-01-01

    Backgrounds: It has been previously demonstrated that the increase of nitric oxide (NO) level may promote cisplatin (CP)-induced nephrotoxicity. The aim of this study was to investigate the role of inducible NO synthase (iNOS) inhibitor to prevent CP-induced nephrotoxicity. Materials and Methods: Four groups of male and four groups of female rats were treated daily with vehicle, S-methylisothiourea hemisulfate (SMT) as a selective iNOS inhibitor (5 mg/kg/twice a day), CP (2.5 mg/kg/day), and CP + SMT for 6 days. Then, all animals were sacrificed and the serum levels of creatinine (Cr), blood urea nitrogen (BUN), nitrite, and malondialdehyde (MDA) were measured. The kidney was removed immediately for histopathological study. Results: Our results showed that inhibition of iNOS by SMT could make different response in male and female animals. SMT therapy in male animals decreased serum BUN, Cr, nitrite, and MDA levels; and it also protected kidney against CP-induced nephrotoxicity. Conclusion: It is concluded that decrease in NO production by SMT has a beneficial effect on reducing CP-induced nephrotoxicity in male. However, such beneficial effect was not observed in female animals. PMID:26322278

  18. Protective effects of a squalene synthase inhibitor, lapaquistat acetate (TAK-475), on statin-induced myotoxicity in guinea pigs

    SciTech Connect

    Nishimoto, Tomoyuki; Ishikawa, Eiichiro; Anayama, Hisashi; Hamajyo, Hitomi; Nagai, Hirofumi; Hirakata, Masao; Tozawa, Ryuichi

    2007-08-15

    High-dose statin treatment has been recommended as a primary strategy for aggressive reduction of LDL cholesterol levels and protection against coronary artery disease. The effectiveness of high-dose statins may be limited by their potential for myotoxic side effects. There is currently little known about the molecular mechanisms of statin-induced myotoxicity. Previously we showed that T-91485, an active metabolite of the squalene synthase inhibitor lapaquistat acetate (lapaquistat: a previous name is TAK-475), attenuated statin-induced cytotoxicity in human skeletal muscle cells [Nishimoto, T., Tozawa, R., Amano, Y., Wada, T., Imura, Y., Sugiyama, Y., 2003a. Comparing myotoxic effects of squalene synthase inhibitor, T-91485, and 3-hydroxy-3-methylglutaryl coenzyme A. Biochem. Pharmacol. 66, 2133-2139]. In the current study, we investigated the effects of lapaquistat administration on statin-induced myotoxicity in vivo. Guinea pigs were treated with either high-dose cerivastatin (1 mg/kg) or cerivastatin together with lapaquistat (30 mg/kg) for 14 days. Treatment with cerivastatin alone decreased plasma cholesterol levels by 45% and increased creatine kinase (CK) levels by more than 10-fold (a marker of myotoxicity). The plasma CK levels positively correlated with the severity of skeletal muscle lesions as assessed by histopathology. Co-administration of lapaquistat almost completely prevented the cerivastatin-induced myotoxicity. Administration of mevalonolactone (100 mg/kg b.i.d.) prevented the cerivastatin-induced myotoxicity, confirming that this effect is directly related to HMG-CoA reductase inhibition. These results strongly suggest that cerivastatin-induced myotoxicity is due to depletion of mevalonate derived isoprenoids. In addition, squalene synthase inhibition could potentially be used clinically to prevent statin-induced myopathy.

  19. Cox-2 is needed but not sufficient for apoptosis induced by Cox-2 selective inhibitors in colon cancer cells.

    PubMed

    Agarwal, B; Swaroop, P; Protiva, P; Raj, S V; Shirin, H; Holt, P R

    2003-12-01

    The role of Cox-2 in NSAID-induced apoptosis is debated. We studied the role of Cox-2 inhibition in apoptosis induced by a selective Cox-2 inhibitor, SC236 (a structural analogue of celecoxib) in two colon cancer cell lines, HT29 (expressing Cox-2 protein) and HCT116 (not expressing Cox-2 protein). Apoptosis was quantified by flow cytometry. SC236 0-75 microM decreased cell numbers and induced apoptosis to identical levels in HT29 and HCT116 cells. However, SC236, concentrations >75 microM reduced Cox-2 protein expression in HT29 cells and induced greater levels of apoptosis in HT29 than in HCT116 cells. In contrast, sulindac sulfide (SSD) (which inhibits Cox-1 and Cox-2) 0-200 microM or sulindac sulfone (SSN) 0-500 microM (without significant activity against Cox-1 or Cox-2) caused identical decreases in cell number and increases in apoptosis in HT29 and HCT116 cells. Neither SSD nor SSN altered the expression of Cox-2 in HT29 cells. To determine that the higher levels of apoptosis in HT29 cells with SC236 >75 microM were related to decreased Cox-2 protein levels, we decreased Cox-2 protein expression in HT29 cells with curcumin (diferuloylmethane) and studied its effect on SC236-induced apoptosis. Curcumin augmented apoptosis induced by SC236 in HT29 cells but not in Cox-2 lacking HCT116 cells. In conclusion, selective Cox-2 inhibitors can induce apoptosis independent of Cox-2 expression. However they may selectively target cells that express Cox-2 by decreasing their Cox-2 protein expression. PMID:14739610

  20. NOX4 mediates cytoprotective autophagy induced by the EGFR inhibitor erlotinib in head and neck cancer cells

    SciTech Connect

    Sobhakumari, Arya; Schickling, Brandon M.; Love-Homan, Laurie; Raeburn, Ayanna; Fletcher, Elise V.M.; Case, Adam J.; Domann, Frederick E.; Miller, Francis J.; and others

    2013-11-01

    Most head and neck squamous cell carcinomas (HNSCCs) overexpress epidermal growth factor receptor (EGFR) and EGFR inhibitors are routinely used in the treatment of HNSCC. However, many HNSCC tumors do not respond or become refractory to EGFR inhibitors. Autophagy, which is a stress-induced cellular self-degradation process, has been reported to reduce the efficacy of chemotherapy in various disease models. The purpose of this study is to determine if the efficacy of the EGFR inhibitor erlotinib is reduced by activation of autophagy via NOX4-mediated oxidative stress in HNSCC cells. Erlotinib induced the expression of the autophagy marker LC3B-II and autophagosome formation in FaDu and Cal-27 cells. Inhibition of autophagy by chloroquine and knockdown of autophagy pathway genes Beclin-1 and Atg5 sensitized both cell lines to erlotinib-induced cytotoxicity, suggesting that autophagy may serve as a protective mechanism. Treatment with catalase (CAT) and diphenylene iodonium (DPI) in the presence of erlotinib suppressed the increase in LC3B-II expression in FaDu and Cal-27 cells. Erlotinib increased NOX4 mRNA and protein expression by increasing its promoter activity and mRNA stability in FaDu cells. Knockdown of NOX4 using adenoviral siNOX4 partially suppressed erlotinib-induced LC3B-II expression, while overexpression of NOX4 increased expression of LC3B-II. These studies suggest that erlotinib may activate autophagy in HNSCC cells as a pro-survival mechanism, and NOX4 may play a role in mediating this effect. - Highlights: • Erlotinib increased LC3B-II and autophagosome formation in HNSCC cells. • Inhibition of autophagy sensitized HNSCC cells to erlotinib. • Erlotinib increased NOX4 promoter and 3?UTR luciferase activity. • Manipulating NOX4 decreases or increases autophagy.

  1. A signal from inside the peroxisome initiates its division by promoting the remodeling of the peroxisomal membrane

    PubMed Central

    Guo, Tong; Gregg, Christopher; Boukh-Viner, Tatiana; Kyryakov, Pavlo; Goldberg, Alexander; Bourque, Simon; Banu, Farhana; Haile, Sandra; Milijevic, Svetlana; San, Karen Hung Yeung; Solomon, Jonathan; Wong, Vivianne; Titorenko, Vladimir I.

    2007-01-01

    We define the dynamics of spatial and temporal reorganization of the team of proteins and lipids serving peroxisome division. The peroxisome becomes competent for division only after it acquires the complete set of matrix proteins involved in lipid metabolism. Overloading the peroxisome with matrix proteins promotes the relocation of acyl-CoA oxidase (Aox), an enzyme of fatty acid ?-oxidation, from the matrix to the membrane. The binding of Aox to Pex16p, a membrane-associated peroxin required for peroxisome biogenesis, initiates the biosynthesis of phosphatidic acid and diacylglycerol (DAG) in the membrane. The formation of these two lipids and the subsequent transbilayer movement of DAG initiate the assembly of a complex between the peroxins Pex10p and Pex19p, the dynamin-like GTPase Vps1p, and several actin cytoskeletal proteins on the peroxisomal surface. This protein team promotes membrane fission, thereby executing the terminal step of peroxisome division. PMID:17438077

  2. Histone deacetylase inhibitor valproic acid promotes the induction of pluripotency in mouse fibroblasts by suppressing reprogramming-induced senescence stress.

    PubMed

    Zhai, Yingying; Chen, Xi; Yu, Dehai; Li, Tao; Cui, Jiuwei; Wang, Guanjun; Hu, Ji-Fan; Li, Wei

    2015-09-10

    Histone deacetylase inhibitor valproic acid (VPA) has been used to increase the reprogramming efficiency of induced pluripotent stem cell (iPSC) from somatic cells, yet the specific molecular mechanisms underlying this effect is unknown. Here, we demonstrate that reprogramming with lentiviruses carrying the iPSC-inducing factors (Oct4-Sox2-Klf4-cMyc, OSKM) caused senescence in mouse fibroblasts, establishing a stress barrier for cell reprogramming. Administration of VPA protected cells from reprogramming-induced senescent stress. Using an in vitro pre-mature senescence model, we found that VPA treatment increased cell proliferation and inhibited apoptosis through the suppression of the p16/p21 pathway. In addition, VPA also inhibited the G2/M phase blockage derived from the senescence stress. These findings highlight the role of VPA in breaking the cell senescence barrier required for the induction of pluripotency. PMID:26112217

  3. Disruption of Src function potentiates Chk1-inhibitor–induced apoptosis in human multiple myeloma cells in vitro and in vivo

    PubMed Central

    Dai, Yun; Chen, Shuang; Shah, Rena; Pei, Xin-Yan; Wang, Li; Almenara, Jorge A.; Kramer, Lora B.; Dent, Paul

    2011-01-01

    Ras/MEK/ERK pathway activation represents an important compensatory response of human multiple myeloma (MM) cells to checkpoint kinase 1 (Chk1) inhibitors. To investigate the functional roles of Src in this event and potential therapeutic significance, interactions between Src and Chk1 inhibitors (eg, UCN-01 or Chk1i) were examined in vitro and in vivo. The dual Src/Abl inhibitors BMS354825 and SKI-606 blocked Chk1-inhibitor–induced extracellular signal-regulated kinase 1/2 (ERK1/2) activation, markedly increasing apoptosis in association with BimEL up-regulation, p34cdc2 activation, and DNA damage in MM cell lines and primary CD138+ MM samples. Loss-of-function Src mutants (K297R, K296R/Y528F) or shRNA knock-down of Src prevented the ERK1/2 activation induced by Chk1 inhibitors and increased apoptosis. Conversely, constitutively active Ras or mitogen-activated protein kinase/ERK kinase 1 (MEK1) significantly diminished the ability of Src inhibitors to potentiate Chk1-inhibitor lethality. Moreover, Src/Chk1-inhibitor cotreatment attenuated MM-cell production of vascular endothelial growth factor and other angiogenic factors (eg, ANG [angiogenin], TIMP1/2 [tissue inhibitor of metalloproteinases 1/2], and RANTES [regulated on activation normal T-cell expressed and secreted]), and inhibited in vitro angiogenesis. Finally, coadministration of BMS354825 and UCN-01 suppressed human MM tumor growth in a murine xenograft model, increased apoptosis, and diminished angiogenesis. These findings suggest that Src kinase is required for Chk1-inhibitor–mediated Ras ? ERK1/2 signaling activation, and that disruption of this event sharply potentiates the anti-MM activity of Chk1 inhi-bitors in vitro and in vivo. PMID:21148814

  4. The SUV39H1 inhibitor chaetocin induces differentiation and shows synergistic cytotoxicity with other epigenetic drugs in acute myeloid leukemia cells

    PubMed Central

    Lai, Y-S; Chen, J-Y; Tsai, H-J; Chen, T-Y; Hung, W-C

    2015-01-01

    Epigenetic modifying enzymes have a crucial role in the pathogenesis of acute myeloid leukemia (AML). Methylation of lysine 9 on histone H3 by the methyltransferase G9a and SUV39H1 is associated with inhibition of tumor suppressor genes. We studied the effect of G9a and SUV39H1 inhibitors on viability and differentiation of AML cells and tested the cytotoxicity induced by combination of G9a and SUV39H1 inhibitors and various epigenetic drugs. The SUV39H1 inhibitor (chaetocin) and the G9a inhibitor (UNC0638) caused cell death in AML cells at high concentrations. However, only chaetocin-induced CD11b expression and differentiation of AML cells at non-cytotoxic concentration. HL-60 and KG-1a cells were more sensitive to chaetocin than U937 cells. Long-term incubation of chaetocin led to downregulation of SUV39H1 and reduction of H3K9 tri-methylation in HL-60 and KG-1a cells. Combination of chaetocin with suberoylanilide hydroxamic acid (SAHA, a histone deacetylase inhibitor) or JQ (a BET (bromodomain extra terminal) bromodomain inhibitor) showed synergistic cytotoxicity. Conversely, no synergism was found by combining chaetocin and UNC0638. More importantly, chaetocin-induced differentiation and combined cytotoxicity were also found in the primary cells of AML patients. Collectively, the SUV39H1 inhibitor chaetocin alone or in combination with other epigenetic drugs may be effective for the treatment of AML. PMID:25978433

  5. Non-canonical Smads phosphorylation induced by the glutamate release inhibitor, riluzole, through GSK3 activation in melanoma.

    PubMed

    Abushahba, Walid; Olabisi, Oyenike O; Jeong, Byeong-Seon; Boregowda, Rajeev K; Wen, Yu; Liu, Fang; Goydos, James S; Lasfar, Ahmed; Cohen-Solal, Karine A

    2012-01-01

    Riluzole, an inhibitor of glutamate release, has shown the ability to inhibit melanoma cell xenograft growth. A phase 0 clinical trial of riluzole as a single agent in patients with melanoma resulted in involution of tumors associated with inhibition of both the mitogen-activated protein kinase (MAPK) and phophoinositide-3-kinase/AKT (PI3K/AKT) pathways in 34% of patients. In the present study, we demonstrate that riluzole inhibits AKT-mediated glycogen synthase kinase 3 (GSK3) phosphorylation in melanoma cell lines. Because we have demonstrated that GSK3 is involved in the phosphorylation of two downstream effectors of transforming growth factor beta (TGF?), Smad2 and Smad3, at their linker domain, our aim was to determine whether riluzole could induce GSK3?-mediated linker phosphorylation of Smad2 and Smad3. We present evidence that riluzole increases Smad2 and Smad3 linker phosphorylation at the cluster of serines 245/250/255 and serine 204 respectively. Using GSK3 inhibitors and siRNA knock-down, we demonstrate that the mechanism of riluzole-induced Smad phosphorylation involved GSK3?. In addition, GSK3? could phosphorylate the same linker sites in vitro. The riluzole-induced Smad linker phosphorylation is mechanistically different from the Smad linker phosphorylation induced by TGF?. We also demonstrate that riluzole-induced Smad linker phosphorylation is independent of the expression of the metabotropic glutamate receptor 1 (GRM1), which is one of the glutamate receptors whose involvement in human melanoma has been documented. We further show that riluzole upregulates the expression of INHBB and PLAU, two genes associated with the TGF? signaling pathway. The non-canonical increase in Smad linker phosphorylation induced by riluzole could contribute to the modulation of the pro-oncogenic functions of Smads in late stage melanomas. PMID:23077590

  6. Myelin Associated Inhibitors: A Link Between Injury-Induced and Experience-Dependent Plasticity

    PubMed Central

    Akbik, Feras; Cafferty, William B. J.; Strittmatter, Stephen M.

    2011-01-01

    SUMMARY In the adult, both neurologic recovery and anatomical growth after a CNS injury are limited. Two classes of growth inhibitors, myelin associated inhibitors (MAIs) and extracellular matrix associated inhibitors, limit both functional recovery and anatomical rearrangements in animal models of spinal cord injury. Here we focus on how MAIs limit a wide spectrum of growth that includes regeneration, sprouting, and plasticity in both the intact and lesioned CNS. Three classic myelin associated inhibitors, Nogo-A, MAG, and OMgp, signal through their common receptors, Nogo-66 Receptor-1 (NgR1) and Paired-Immunoglobulin-like-Receptor-1 (PirB), to regulate cytoskeletal dynamics and inhibit growth. Initially described as inhibitors of axonal regeneration, subsequent work has demonstrated that MAIs also limit activity and experience-dependent plasticity in the intact, adult CNS. MAIs therefore represent a point of convergence for plasticity that limits anatomical rearrangements regardless of the inciting stimulus, blurring the distinction between injury studies and more “basic” plasticity studies. PMID:21699896

  7. The DNA methyltransferase inhibitor zebularine induces mitochondria-mediated apoptosis in gastric cancer cells in vitro and in vivo

    SciTech Connect

    Tan, Wei; Zhou, Wei; Yu, Hong-gang; Luo, He-Sheng; Shen, Lei

    2013-01-04

    Highlights: Black-Right-Pointing-Pointer Zebularine inhibited cell growth of gastric cancer in a time- and dose-dependent manner. Black-Right-Pointing-Pointer Chromatin condensation and nuclear fragmentation were induced. Black-Right-Pointing-Pointer Zebularine promoted apoptosis via mitochondrial pathways. Black-Right-Pointing-Pointer Tumorigenicity was inhibited by zebularine. -- Abstract: DNA methyltransferase (DNMT) inhibitor zebularine has been reported to potentiate the anti-tumor effect by reactivating the expression of tumor suppressor genes and apoptosis-related genes in various malignant cells. However, the apoptotic signaling pathway in gastric cancer cells induced by zebularine is not well understood. In the study, the effects of zebularine on the growth and apoptosis of gastric cancer cells were investigated by MTT assay, Hoechst assay, Western blot analysis, flow cytometric analysis of annexin V-FITC/PI staining, and TUNEL assay. Zebularine was an effective inhibitor of human gastric cancer cells proliferation in vitro and in vivo. The effects were dose dependent. A zebularine concentration of 50 {mu}M accounted for the inhibition of cell proliferation of 67% at 48 h. The treatment with zebularine upregulated Bax, and decreased Bcl-2 protein. Caspase-3 was activated, suggesting that the apoptosis is mediated by mitochondrial pathways. Moreover, zebularine injection successfully inhibited the tumor growth via apoptosis induction which was demonstrated by TUNEL assay in xenograft tumor mouse model. These results demonstrated that zebularine induced apoptosis in gastric cancer cells via mitochondrial pathways, and zebularine might become a therapeutic approach for the treatment of gastric cancer.

  8. Reversal of High dietary fructose-induced PPAR? suppression by oral administration of lipoxygenase/cyclooxygenase inhibitors

    PubMed Central

    Kelley, Glen L; Azhar, Salman

    2005-01-01

    High fructose feeding causes diet-induced alterations of lipid metabolism and decreased insulin sensitivity, hallmark of which is a rapid and profound hypertriglyceridemia. One of the mechanisms that contribute to serum hypertriglyceridemia in this model is suppression of hepatic PPAR?. HMG-CoA inhibitors, which reduce serum triglycerides in these animals, also elevate/restore hepatic PPAR?. Previously we demonstrated that two known lipoxygenase/cyclooxygenase inhibitors reversed diet-induced hypertriglyceridemia in this model and that reversal of certain inflammatory markers in the liver correlated with the metabolic benefit. In this paper we extended these studies by examining the impact of these compounds on expression of PPAR?, both at the level of transcription and expression. Our data show that diet-induced suppression of hepaic PPAR? is reversed upon treatment with lipoxygenase/cyclooxygenase compounds. We then tested one of these compounds, BW-755c, over a range of doses from 10 mg/kg to 100 mg/kg to establish a dose-response relationship with the reduction of serum hypertriglyceridemia in this model. These experiments support the concept of using anti-inflammatory medications as one method to correct metabolic dysfunction. PMID:16091142

  9. Dual p38/JNK Mitogen Activated Protein Kinase Inhibitors Prevent Ozone-Induced Airway Hyperreactivity in Guinea Pigs

    PubMed Central

    Verhein, Kirsten C.; Salituro, Francesco G.; Ledeboer, Mark W.; Fryer, Allison D.; Jacoby, David B.

    2013-01-01

    Ozone exposure causes airway hyperreactivity and increases hospitalizations resulting from pulmonary complications. Ozone reacts with the epithelial lining fluid and airway epithelium to produce reactive oxygen species and lipid peroxidation products, which then activate cell signaling pathways, including the mitogen activated protein kinase (MAPK) pathway. Both p38 and c-Jun NH2 terminal kinase (JNK) are MAPK family members that are activated by cellular stress and inflammation. To test the contribution of both p38 and JNK MAPK to ozone-induced airway hyperreactivity, guinea pigs were pretreated with dual p38 and JNK MAPK inhibitors (30 mg/kg, ip) 60 minutes before exposure to 2 ppm ozone or filtered air for 4 hours. One day later airway reactivity was measured in anesthetized animals. Ozone caused airway hyperreactivity one day post-exposure, and blocking p38 and JNK MAPK completely prevented ozone-induced airway hyperreactivity. Blocking p38 and JNK MAPK also suppressed parasympathetic nerve activity in air exposed animals, suggesting p38 and JNK MAPK contribute to acetylcholine release by airway parasympathetic nerves. Ozone inhibited neuronal M2 muscarinic receptors and blocking both p38 and JNK prevented M2 receptor dysfunction. Neutrophil influx into bronchoalveolar lavage was not affected by MAPK inhibitors. Thus p38 and JNK MAPK mediate ozone-induced airway hyperreactivity through multiple mechanisms including prevention of neuronal M2 receptor dysfunction. PMID:24058677

  10. Arachidonic acid-induced inflammation: inhibition by dual inhibitor of arachidonic acid metabolism, SK&F 86002.

    PubMed

    Griswold, D E; Webb, E; Schwartz, L; Hanna, N

    1987-06-01

    The antiinflammatory activity of the structurally novel dual inhibitor of arachidonic acid metabolism, SK&F 86002 was evaluated using arachidonic acid-induced edema and inflammatory cell infiltration. Histological examination demonstrated extensive subcutaneous edema and neutrophil (PMN) accumulation in perivascular and interstitial locations one hour after application of arachidonic acid to the ear. SK&F 86002 and, to a lesser extent, phenidone demonstrated potent inhibition of this inflammatory response following oral and topical administration. Nordihydroguaiaretic acid (NDGA) displayed only topical activity. The selective cyclooxygenase inhibitors ibuprofen and naproxen were either inactive or stimulated ear swelling. Histological evaluation of the lesion in drug-treated animals revealed that SK&F 86002 impaired edema formation and caused a significant reduction in numbers of infiltrating neutrophils. Using arachidonic acid-induced peritoneal exudation, a reduction in the cellular infiltrate was observed after oral treatment with SK&F 86002 or phenidone, but not with naproxen. Taken together, these data illustrate the potent antiinflammatory effects of SK&F 86002 and support the suggestion that 5-lipoxygenase products play a significant role in both the edematous and cellular phases of arachidonic acid-induced inflammation. PMID:3108157

  11. JNK inhibitor SP600125 protects against lipopolysaccharide-induced acute lung injury via upregulation of claudin-4

    PubMed Central

    ZHENG, YUELIANG; ZHANG, MEIQI; ZHAO, YIMING; CHEN, JIE; LI, BING; CAI, WENWEI

    2014-01-01

    Although in vitro studies have previously demonstrated that mitogen-activated protein kinases are important for the activation of transcription factors and the regulation of proinflammatory mediators, the function of c-Jun NH2-terminal kinase (JNK) in acute lung injury (ALI) remains to be fully elucidated. The present study aimed to investigate the effect of the JNK selective inhibitor SP600125 on lipopolysaccharide (LPS)-induced ALI. Pulmonary edema, the expression of inflammatory cytokines and pathological alterations were found to be significantly attenuated in LPS-induced ALI following treatment with SP600125 in vivo. In vitro, it was demonstrated that SP600125 administration significantly improved A549 cell viability in a dose-dependent manner using the Cell Counting kit-8 and the 5-ethynyl-2?-deoxyuridine incorporation assay. Furthermore, flow cytometric analysis demonstrated that the apoptotic rate was significantly reduced in a concentration-dependent manner following SP600125 injection. At the molecular level, SP600125 treatment dose-dependently inhibited JNK activation and upregulated claudin-4 expression in vivo and in vitro. In combination, the results from the present study indicated that the JNK inhibitor SP600125 protected against LPS-induced ALI in vivo and in vitro, possibly by upregulating the expression of claudin-4. PMID:24944614

  12. The cathepsin B inhibitor, z-FA-CMK is toxic and readily induced cell death in human T lymphocytes

    SciTech Connect

    Liow, K.Y.; Chow, S.C.

    2013-11-01

    The cathepsin B inhibitor, benzyloxycarbonyl-phenylalanine-alanine-chloromethylketone (z-FA-CMK) was found to be toxic and readily induced cell death in the human T cell line, Jurkat, whereas two other analogs benzyloxycarbonyl-phenylalanine-alanine-fluoromethylketone (z-FA-FMK) and benzyloxycarbonyl-phenylalanine-alanine-diazomethylketone (z-FA-DMK) were not toxic. The toxicity of z-FA-CMK requires not only the CMK group, but also the presence of alanine in the P1 position and the benzyloxycarbonyl group at the N-terminal. Dose–response studies showed that lower concentrations of z-FA-CMK induced apoptosis in Jurkat T cells whereas higher concentrations induced necrosis. In z-FA-CMK-induced apoptosis, both initiator caspases (-8 and -9) and effector caspases (-3, -6 and -7) were processed to their respective subunits in Jurkat T cells. However, only the pro-form of the initiator caspases were reduced in z-FA-CMK-induced necrosis and no respective subunits were apparent. The caspase inihibitor benzyloxycarbonyl-valine-alanine-aspartic acid-(O-methyl)-fluoromehylketone (z-VAD-FMK) inhibits apoptosis and caspase processing in Jurkat T cells treated with low concentration of z-FA-CMK but has no effect on z-FA-CMK-induced necrosis and the loss of initiator caspases. This suggests that the loss of initiator caspases in Jurkat T cells during z-FA-CMK-induced necrosis is not a caspase-dependent process. Taken together, we have demonstrated that z-FA-CMK is toxic to Jurkat T cells and induces apoptosis at low concentrations, while at higher concentrations the cells die of necrosis. - Highlights: • z-FA-CMK is toxic and induce cell death in the human T cells. • z-FA-CMK toxicity requires the CMK group, alanine and the benzyloxycarbonyl group. • z-FA-CMK induced apoptosis at low concentration and necrosis at high concentration.

  13. Peroxisomal localization and activation by bivalent metal ions of ureidoglycolate lyase, the enzyme involved in urate degradation in Candida tropicalis

    SciTech Connect

    Takada, Y.; Tsukiji, N.

    1987-05-01

    Ureidoglycolate lyase was found only in the peroxisomes in urate-induced Candida tropicalis. The enzyme was markedly activated by the bivalent metal ions Mn/sup 2 +/, Fe/sup 2 +/, and Ni/sup 2 +/. The activation by Mn/sup 2 +/ was suggested to be the result of its binding to the apoenzyme.

  14. An Aminopyridazine Inhibitor of Death Associated Protein Kinase Attenuates Hypoxia-Ischemia Induced Brain Damage

    SciTech Connect

    Velentza, A.V.; Wainwright, M.S.; Zasadzki, M.; Mirzoeva, S.; Haiech, J.; Focia, P.J.; Egli, M.; Watterson, D.M.

    2010-03-08

    Death associated protein kinase (DAPK) is a calcium and calmodulin regulated enzyme that functions early in eukaryotic programmed cell death, or apoptosis. To validate DAPK as a potential drug discovery target for acute brain injury, the first small molecule DAPK inhibitor was synthesized and tested in vivo. A single injection of the aminopyridazine-based inhibitor administered 6 h after injury attenuated brain tissue or neuronal biomarker loss measured, respectively, 1 week and 3 days later. Because aminopyridazine is a privileged structure in neuropharmacology, we determined the high-resolution crystal structure of a binary complex between the kinase domain and a molecular fragment of the DAPK inhibitor. The co-crystal structure describes a structural basis for interaction and provides a firm foundation for structure-assisted design of lead compounds with appropriate molecular properties for future drug development.

  15. Design, synthesis and biological evaluation of ?-boswellic acid based HDAC inhibitors as inducers of cancer cell death.

    PubMed

    Sharma, Simmi; Ahmad, Mudassier; Bhat, Javeed Ahmad; Kumar, Arvind; Kumar, Manjeet; Zargar, Mohmmad Afzal; Hamid, Abid; Shah, Bhahwal Ali

    2014-10-01

    The synthesis and bio-evaluation of naturally occurring boswellic acids (BAs) as an alternate CAP for the design of new HDAC inhibitors is described. All the compounds were screened against a panel of human cancer cell lines to identify leads, which were subsequently examined for their potential to inhibit HDACs. The identified lead compound showed IC50 value of 6?m for HDACs, found to induce G1 cell cycle arrest at significantly low concentration (1?M) and caused significant loss in mitochondrial membrane potential at 5 and 10?M. Furthermore, specific interactions of the lead molecule inside the catalytic domain were also studied through in silico molecular modeling. PMID:25176189

  16. Mammalian Target of Rapamycin Inhibitors Induce Tumor Cell Apoptosis In Vivo Primarily by Inhibiting VEGF Expression and Angiogenesis

    PubMed Central

    Berlanger, Eileen; Mysore, Veena; Hoang, Bao; Shi, YiJiang; Gera, Joseph; Lichtenstein, Alan

    2013-01-01

    We found that rapalog mTOR inhibitors induce G1 arrest in the PTEN-null HS Sultan B-cell lymphoma line in vitro, but that administration of rapalogs in a HS Sultan xenograft model resulted in significant apoptosis, and that this correlated with induction of hypoxia and inhibition of neoangiogenesis and VEGF expression. Mechanistically, rapalogs prevent cap-dependent translation, but studies have shown that cap-independent, internal ribosome entry site (IRES)-mediated translation of genes, such as c-myc and cyclin D, can provide a fail-safe mechanism that regulates tumor survival. Therefore, we tested if IRES-dependent expression of VEGF could likewise regulate sensitivity of tumor cells in vivo. To achieve this, we developed isogenic HS Sultan cell lines that ectopically express the VEGF ORF fused to the p27 IRES, an IRES sequence that is insensitive to AKT-mediated inhibition of IRES activity and effective in PTEN-null tumors. Mice challenged with p27-VEGF transfected tumor cells were more resistant to the antiangiogenic and apoptotic effects of the rapalog, temsirolimus, and active site mTOR inhibitor, pp242. Our results confirm the critical role of VEGF expression in tumors during treatment with mTOR inhibitors and underscore the importance of IRES activity as a resistance mechanism to such targeted therapy. PMID:23533410

  17. A trypsin inhibitor from rambutan seeds with antitumor, anti-HIV-1 reverse transcriptase, and nitric oxide-inducing properties.

    PubMed

    Fang, Evandro Fei; Ng, Tzi Bun

    2015-04-01

    Nephelium lappaceum L., commonly known as "rambutan," is a typical tropical tree and is well known for its juicy and sweet fruit which has an exotic flavor. Chemical studies on rambutan have led to the identification of various components such as monoterpene lactones and volatile compounds. Here, a 22.5-kDa trypsin inhibitor (N . lappaceum trypsin inhibitor (NLTI)) was isolated from fresh rambutan seeds using liquid chromatographical techniques. NLTI reduced the proteolytic activities of both trypsin and ?-chymotrypsin. Dithiothreitol reduced the trypsin inhibitory activity of NLTI at a concentration of 1 mM, indicating that an intact disulfide bond is essential to the activity. NLTI inhibited HIV-1 reverse transcriptase with an IC50 of 0.73 ?M. In addition, NLTI manifested a time- and dose-dependent inhibitory effect on growth in many tumor cells. NLTI is one of the few trypsin inhibitors with nitric oxide-inducing activity and may find application in tumor therapy. PMID:25820360

  18. Isolation of hypoxia-inducible factor 1 (HIF-1) inhibitors from frankincense using a molecularly imprinted polymer.

    PubMed

    Lakka, Achillia; Mylonis, Ilias; Bonanou, Sophia; Simos, George; Tsakalof, Andreas

    2011-10-01

    Hypoxia-Inducible Factor 1 (HIF-1), a transcriptional activator, is highly involved in the pathology of cancer. Inhibition of HIF-1 retards tumor growth and enhances treatment efficiency when used in combination with chemo- or radiation therapy. The recent validation of HIF-1 as an important drug target in cancer treatment has stimulated efforts to identify and isolate natural or synthetic HIF-1 inhibitors. In the present study, quercetin, a known inhibitor of HIF-1, was imprinted in a polymer matrix in order to prepare a Molecularly Imprinted Polymer (MIP), which was subsequently used for the selective isolation of new inhibitors from frankincense, a gum resin used as anticancer remedy in traditional medicine. The frankincense components isolated by Solid Phase Extraction on MIP (MIP-SPE), efficiently inhibited the transcriptional activity of HIF-1 and decreased the protein levels of HIF-1?, the regulated subunit of HIF-1. The selective retention of acetyl 11-ketoboswellic acid (AKBA, one of the main bioactive components of frankincense) by MIP led to the revealing of its inhibitory activity on the HIF-1 signaling pathway. AKBA was selectively retained by SPE on the quercetin imprinted polymer, with an imprinting effect of 8.1 ± 4.6. Overall, this study demonstrates the potential of MIP application in the screening, recognition and isolation of new bioactive compounds that aim selected molecular targets, a potential that has been poorly appreciated until. PMID:20437079

  19. Brainstem brain-derived neurotrophic factor signaling is required for histone deacetylase inhibitor-induced pain relief.

    PubMed

    Tao, Wenjuan; Chen, Quan; Wang, Lu; Zhou, Wenjie; Wang, Yunping; Zhang, Zhi

    2015-06-01

    Our previous study demonstrated that persistent pain can epigenetically suppress the transcription of Gad2 [encoding glutamic acid decarboxylase 65 (GAD65)] and consequently impair the inhibitory function of GABAergic synapses in central pain-modulating neurons. This contributes to the development of persistent pain sensitization. Histone deacetylase (HDAC) inhibitors increased GAD65 activity considerably, restored GABA synaptic function, and rendered sensitized pain behavior less pronounced. However, the molecular mechanisms by which HDAC regulates GABAergic transmission through GAD65 under pain conditions are unknown. This work showed that HDAC inhibitor-induced increases in colocalization of GAD65 and synaptic protein synapsin I on the presynaptic axon terminals of the nucleus raphe magnus (NRM) were blocked by a TrkB receptor antagonist K252a [(9S,10R,12R)-2,3,9,10,11,12-hexahydro-10-hydroxy-9-methyl-1-oxo-9,12-epoxy-1H-diindolo[1,2,3-fg:3',2',1'-kl]pyrrolo[3,4-i][1,6]benzodiazocine-10-carboxylic acid methyl ester], indicating that BDNF-TrkB signaling may be required in GAD65 modulation of GABA synaptic function. At the brain-derived neurotrophic factor (BDNF) promoter, HDAC inhibitors induced significant increases in H3 hyperacetylation, consistent with the increase in BDNF mRNA and total proteins. Although exogenous BDNF facilitated GABA miniature inhibitory postsynaptic currents and GAD65 accumulation in NRM neuronal synapses in normal rats, it failed to do so in animals subjected to persistent inflammation. In addition, blockade of the TrkB receptor with K252a has no effect on miniature inhibitory postsynaptic currents and synaptic GAD65 accumulation under normal conditions. In addition, the analgesic effects of HDAC inhibitors on behavior were blocked by NRM infusion of K252a. These findings suggest that BDNF-TrkB signaling is required for drugs that reverse the epigenetic effects of chronic pain at the gene level, such as HDAC inhibitors. PMID:25852071

  20. Peroxisome-proliferator-activated receptors and the control of levels of prostaglandin-endoperoxide synthase 2 by arachidonic acid in the bovine uterus

    PubMed Central

    Sheldrick, E. Linda R.; Derecka, Kamila; Marshall, Elaine; Chin, Evonne C.; Hodges, Louise; Wathes, D. Claire; Abayasekara, D. Robert E.; Flint, Anthony P. F.

    2007-01-01

    Arachidonic acid is a potential paracrine agent released by the uterine endometrial epithelium to induce PTGS2 [PG (prostaglandin)-endoperoxide synthase 2] in the stroma. In the present study, bovine endometrial stromal cells were used to determine whether PTGS2 is induced by arachidonic acid in stromal cells, and to investigate the potential role of PPARs (peroxisome-proliferator-activated receptors) in this effect. Arachidonic acid increased PTGS2 levels up to 7.5-fold within 6 h. The cells expressed PPAR? and PPAR? (also known as PPAR?) (but not PPAR?). PTGS2 protein level was increased by PPAR agonists, including polyunsaturated fatty acids, synthetic PPAR ligands, PGA1 and NSAIDs (non-steroidal anti-inflammatory drugs) with a time course resembling that of arachidonic acid. Use of agonists and antagonists indicated PPAR? (but not PPAR? or PPAR?) was responsible for PTGS2 induction. PTGS2 induction by arachidonic acid did not require PG synthesis. PTGS2 levels were increased by the PKC (protein kinase C) activators 4?-PMA and PGF2?, and the effects of arachidonic acid, NSAIDs, synthetic PPAR ligands and 4?-PMA were blocked by PKC inhibitors. This is consistent with PPAR phosphorylation by PKC. Induction of PTGS2 protein by 4?-PMA in the absence of a PPAR ligand was decreased by the NF-?B (nuclear factor ?B) inhibitors MG132 and parthenolide, suggesting that PKC acted through NF-?B in addition to PPAR phosphorylation. Use of NF-?B inhibitors allowed the action of arachidonic acid as a PPAR agonist to be dissociated from an effect through PKC. The results are consistent with the hypothesis that arachidonic acid acts via PPAR? to increase PTGS2 levels in bovine endometrial stromal cells. PMID:17516915

  1. Use of a Soluble Epoxide Hydrolase Inhibitor in Smoke-Induced Chronic Obstructive Pulmonary Disease

    E-print Network

    Hammock, Bruce D.

    in the spontaneously hypertensive rat. Both t-TUCB and Rolipram in- hibited neutrophil elevation in bronchoalveolar, pulmonary hypertension, and small-airway obstruction. Soluble epoxide hydrolase inhibitors (sEHIs) demonstrated anti-inflammatory properties in a rat model after acute exposure to tobacco smoke. We compared

  2. The effect of marimastat, a metalloprotease inhibitor, on allergen-induced asthmatic hyper-reactivity

    SciTech Connect

    Bruce, Colleen; Thomas, Paul S. . E-mail: paul.thomas@unsw.edu.au

    2005-06-01

    This pilot study was designed to assess whether a synthetic matrix metalloproteinase (MMP) inhibitor has anti-inflammatory properties in mild asthma. Tumor necrosis factor alpha (TNF{alpha}) has been shown to be an important cytokine in the pathogenesis of allergic airway inflammatory responses, and its release can be inhibited by MMP inhibitors. Twelve atopic asthmatic subjects received the MMP inhibitor marimastat (5 mg) or placebo, twice daily for 3 weeks, separated by a 6-week washout period in a randomized, double-blind, cross-over manner. All subjects underwent an allergen inhalation provocation test to Dermatophagoides pteronyssinus before and after each study phase. Spirometry, exhaled NO (eNO) levels, differential sputum cell counts, an asthma symptom questionnaire, peak flow, and {beta}{sub 2}-agonist usage were measured. Nine subjects completed the study, and, when compared with placebo, marimastat reduced bronchial hyper-responsiveness to inhaled allergen in these subjects from an allergen PC{sub 20} of 22.2 AU/ml (95%CI 11.7-32.6) to 17.0 AU/ml (95%CI 7.6-26.4, P = 0.02). The marimastat phase showed a nonsignificant fall in sputum inflammatory cells. Marimastat did not modify eNO, FEV{sub 1}, asthma symptoms, or albuterol usage. In conclusion, airway responsiveness to allergen may be modified by a MMP inhibitor, perhaps via TNF{alpha} playing a role in airway inflammation and remodeling.

  3. Close encounters of the lysosome/peroxisome kind

    PubMed Central

    Jin, Yui; Strunk, Bethany S.; Weisman, Lois S.

    2015-01-01

    Lysosomes provide a major source for cellular cholesterol; however, most of this cholesterol is trafficked to the plasma membrane via unknown mechanisms. In this issue of Cell, Chu et al. identify an unexpected role for peroxisomes in the transport of cholesterol from the lysosome to the plasma membrane via a lysosome-peroxisome membrane contact site. PMID:25860602

  4. In vitro modulation of MMP-2 and MMP-9 in pediatric human sarcoma cell lines by cytokines, inducers and inhibitors

    PubMed Central

    ROOMI, M.W.; KALINOVSKY, T.; RATH, M.; NIEDZWIECKI, A.

    2014-01-01

    The highly aggressive pediatric sarcomas are characterized by high levels of matrix metalloproteinase (MMP)-2 and MMP-9, which play crucial roles in tumor invasion and metastasis by degradation of the extracellular membrane leading to cancer cell spread to distal organs. We examined the effects of cytokines, mitogens, inducers and inhibitors on MMP-2 and -9 expression in osteosarcoma (U2OS) and rhabdomyosarcoma (RD). The selected compounds included natural cytokines and growth factors, as well as chemical compounds applied in therapy of sarcoma and natural compounds that have demonstrated anticancer therapeutic potential. These cell lines were cultured in their respective media to near confluence and the cells were washed with PBS and incubated in serum-free medium with various concentrations of several cytokines, mitogens and inhibitors. After 24 h the media were removed and analyzed for MMP-2 and -9 by gelatinase zymography and quantitated by densitometry. Osteosarcoma and rhabdomyosarcoma showed bands corresponding to MMP-2 and -9 with dose-dependent enhancement of MMP-9 with phorbol 12-myristate 13-acetate (PMA) treatment. Tumor necrosis factor-?, interleukin-1? and LPS enhanced osteosarcoma U2OS MMP-9 secretion but had no effect on MMP-2 secretion. Tumor necrosis factor-? stimulated rhabdomyosarcoma MMP-2 expression, but had no effect on MMP-9 secretion. Doxycycline, epigallocatechin gallate, nutrient mixture (NM), actinomycin-D, cyclohex-amide, retinoic acid and dexamethasone inhibited MMP-2 and -9 in U2OS osteosarcoma cells. PMA-treated RD cells showed dose-response inhibition of MMP-9 by doxycycline and epigallocatechin gallate and both MMPs by NM. Dexamethasone and actinomycin-D showed inhibition of MMP-2 secretion of RD cells. Our results show that cytokines, mitogens and inducers show variable upregulation of U2OS osteosarcoma and RD rhabdomyosarcoma MMP-2 and -9 secretion, and inhibitors demonstrate downregulation under stimulatory conditions, suggesting the application of these agents for the development of effective therapies in pediatric sarcomas. PMID:24190483

  5. Omeprazole increases the efficacy of a soluble epoxide hydrolase inhibitor in a PGE2 induced pain model.

    PubMed

    Goswami, Sumanta Kumar; Inceoglu, Bora; Yang, Jun; Wan, Debin; Kodani, Sean D; da Silva, Carlos Antonio Trindade; Morisseau, Christophe; Hammock, Bruce D

    2015-12-15

    Epoxyeicosatrienoic acids (EETs) are potent endogenous analgesic metabolites produced from arachidonic acid by cytochrome P450s (P450s). Metabolism of EETs by soluble epoxide hydrolase (sEH) reduces their activity, while their stabilization by sEH inhibition decreases both inflammatory and neuropathic pain. Here, we tested the complementary hypothesis that increasing the level of EETs through induction of P450s by omeprazole (OME), can influence pain related signaling by itself, and potentiate the anti-hyperalgesic effect of sEH inhibitor. Rats were treated with OME (100mg/kg/day, p.o., 7days), sEH inhibitor TPPU (3mg/kg/day, p.o.) and OME (100mg/kg/day, p.o., 7days)+TPPU (3mg/kg/day, p.o., last 3days of OME dose) dissolved in vehicle PEG400, and their effect on hyperalgesia (increased sensitivity to pain) induced by PGE2 was monitored. While OME treatment by itself exhibited variable effects on PGE2 induced hyperalgesia, it strongly potentiated the effect of TPPU in the same assay. The significant decrease in pain with OME+TPPU treatment correlated with the increased levels of EETs in plasma and increased activities of P450 1A1 and P450 1A2 in liver microsomes. The results show that reducing catabolism of EETs with a sEH inhibitor yielded a stronger analgesic effect than increasing generation of EETs by OME, and combination of both yielded the strongest pain reducing effect under the condition of this study. PMID:26522832

  6. Isoliquiritigenin is a potent inhibitor of NLRP3 inflammasome activation and diet-induced adipose tissue inflammation.

    PubMed

    Honda, Hiroe; Nagai, Yoshinori; Matsunaga, Takayuki; Okamoto, Naoki; Watanabe, Yasuharu; Tsuneyama, Koichi; Hayashi, Hiroaki; Fujii, Isao; Ikutani, Masashi; Hirai, Yoshikatsu; Muraguchi, Atsushi; Takatsu, Kiyoshi

    2014-12-01

    Inflammasome activation initiates the development of many inflammatory diseases, including obesity and type 2 diabetes. Therefore, agents that target discrete activation steps could represent very important drugs. We reported previously that ILG, a chalcone from Glycyrrhiza uralensis, inhibits LPS-induced NF-?B activation. Here, we show that ILG potently inhibits the activation of NLRP3 inflammasome, and the effect is independent of its inhibitory potency on TLR4. The inhibitory effect of ILG was stronger than that of parthenolide, a known inhibitor of the NLRP3 inflammasome. GL, a triterpenoid from G. uralensis, had similar inhibitory effects on NLRP3 activity, but high concentrations of GL were required. In contrast, activation of the AIM2 inflammasome was inhibited by GL but not by ILG. Moreover, GL inhibited NLRP3- and AIM2-activated ASC oligomerization, whereas ILG inhibited NLRP3-activated ASC oligomerization. Low concentrations of ILG were highly effective in IAPP-induced IL-1? production compared with the sulfonylurea drug glyburide. In vivo analyses revealed that ILG potently attenuated HFD-induced obesity, hypercholesterolemia, and insulin resistance. Furthermore, ILG treatment improved HFD-induced macrovesicular steatosis in the liver. Finally, ILG markedly inhibited diet-induced adipose tissue inflammation and IL-1? and caspase-1 production in white adipose tissue in ex vivo culture. These results suggest that ILG is a potential drug target for treatment of NLRP3 inflammasome-associated inflammatory diseases. PMID:25210146

  7. Tacrine, an oral acetylcholinesterase inhibitor, induced hepatic oxidative damage, which was blocked by liquiritigenin through GSK3-beta inhibition.

    PubMed

    Park, Sang Mi; Ki, Sung Hwan; Han, Nu Ri; Cho, Il Je; Ku, Sae Kwang; Kim, Sang Chan; Zhao, Rong Jie; Kim, Young Woo

    2015-01-01

    Although the cholinesterase inhibitor tacrine has been successfully used for the treatment of Alzheimer's disease, it is known to have hepatotoxic effects. Liquiritigenin (LQ), an active flavonoid in Glycyrrhizae radix, exerts protective effects against liver damage. This study investigated the toxic effect of tacrine on hepatocytes and the beneficial effect of LQ on tacrine intoxication in vivo and in vitro, and the underlying mechanism involved. In hepatocyte cell lines, tacrine induced cell death and oxidative stress, as indicated by decreases in cell viability and glutathione (GSH) contents, which were blocked by pretreatment with LQ. Fluorescent activated cell sorter (FACS) analysis revealed that LQ inhibited cellular H2O2 production and mitochondrial dysfunction induced by tacrine in HepG2 cells. Furthermore, LQ promoted inhibitory phosphorylation of glycogen synthase kinase-3? (GSK3?) and prevented decreases in GSK3? phosphorylation induced by tacrine. In rats treatment with tacrine at 30 mg/kg increased hepatic damage as assessed by blood biochemistry and histopathology. Administration of LQ (10 or 30 mg/kg/d, per os (p.o.)) or the hepatoprotective drug sylimarin (100 mg/kg/d) for 3 d inhibited elevations in alanine aminotransferase, aspartate aminotransferase, and histological changes induced by tacrine. These results show that LQ efficaciously protects the rat liver against tacrine-induced liver damage, and suggest that LQ is a therapeutic candidate for ameliorating the hepatotoxic effects of tacrine. PMID:25747977

  8. Stresgenin B, an inhibitor of heat-induced heat shock protein gene expression, produced by Streptomyces sp. AS-9.

    PubMed

    Akagawa, H; Takano, Y; Ishii, A; Mizuno, S; Izui, R; Sameshima, T; Kawamura, N; Dobashi, K; Yoshioka, T

    1999-11-01

    Stresgenin B was isolated as an inhibitor of heat-induced heat shock protein (HSP) gene expression from a culture broth of Streptomyces sp. AS-9 by silica gel chromatography and HPLC. The molecular formula of the novel compound was determined as C11H13NO5 by high resolution FAB-MS analysis, and the structure was determined by UV, 1H NMR, 13C NMR, HMQC, HMBC, and NOESY spectra. Stresgenin B inhibited heat-induced luciferase reporter-gene expression directed by the human hsp70B promoter in Chinese hamster ovary (CHO) cells at concentrations lower than the concentrations for inhibition of dexamethasone-induced luciferase reporter-gene expression directed by the mouse mammary tumor virus (MMTV)-LTR promoter. The inhibition of heat-induced reporter gene expression was evident even when cells were exposed to stresgenin B only during heat stress treatment. Moreover, the compound inhibited heat-induced syntheses of hsp72/73, hsp90, and hsp110 and thereby suppressed the induction of thermotolerance. Stresgenin B showed moderate cytotoxic activities against several neoplastic cell lines and also showed antibacterial activities against Micrococcus luteus, Bacillus subtilis and Staphylococcus aureus strains. PMID:10656568

  9. Encorafenib (LGX818), a potent BRAF inhibitor, induces senescence accompanied by autophagy in BRAFV600E melanoma cells.

    PubMed

    Li, Zhen; Jiang, Ke; Zhu, Xiaofang; Lin, Guibin; Song, Fei; Zhao, Yongfu; Piao, Yongjun; Liu, Jiwei; Cheng, Wei; Bi, Xiaolin; Gong, Peng; Song, Zhiqi; Meng, Songshu

    2016-01-28

    Encorafenib (LGX818) is a new-generation BRAF inhibitor that is under evaluation in clinical trials. However, the underlying mechanism remains to be elucidated. Here we show that LGX818 potently decreased ERK phosphorylation and inhibited proliferation in BRAFV600E melanoma cell lines. Moreover, LGX818 downregulated CyclinD1 in a glycogen synthase kinase 3?-independent manner and induced cell cycle arrest in the G1 phase, Surprisingly, LGX818 triggered cellular senescence in BRAFV600E melanoma cells, as evidenced by increased ?-galactosidase staining, while no appreciable induction of apoptosis was detected, as determined by Annexin V and propidium iodide staining and immunoblot analysis of caspase-3 processing and poly (ADP-ribose) polymerase cleavage. Increased p27KIP1 expression and retinoblastoma protein activation were detected during LGX818-induced senescence. Additionally, inhibition of dual-specificity tyrosine phosphorylation-regulated kinase 1B by AZ191 reversed LGX818-induced CyclinD1 turnover and senescence. Interestingly, autophagy is triggered through inhibition of the mTOR/70S6K pathway during LGX818-induced senescence. Moreover, autophagy inhibition by pharmacological and genetic regulation attenuates LGX818-induced senescence. Notably, combining LGX818 with autophagy modulators has anti-proliferative effect in LGX818-resistant BRAF mutant melanoma cells. Altogether, we uncovered a mechanism by which LGX818 exerts its anti-tumor activity in BRAFV600E melanoma cells. PMID:26586345

  10. The Aurora-A inhibitor MLN8237 affects multiple mitotic processes and induces dose-dependent mitotic abnormalities and aneuploidy.

    PubMed

    Asteriti, Italia Anna; Di Cesare, Erica; De Mattia, Fabiola; Hilsenstein, Volker; Neumann, Beate; Cundari, Enrico; Lavia, Patrizia; Guarguaglini, Giulia

    2014-08-15

    Inhibition of Aurora kinase activity by small molecules is being actively investigated as a potential anti-cancer strategy. A successful therapeutic use of Aurora inhibitors relies on a comprehensive understanding of the effects of inactivating Aurora kinases on cell division, a challenging aim given the pleiotropic roles of those kinases during mitosis. Here we have used the Aurora-A inhibitor MLN8237, currently under phase-I/III clinical trials, in dose-response assays in U2OS human cancer cells synchronously proceeding towards mitosis. By following the behaviour and fate of single Aurora-inhibited cells in mitosis by live microscopy, we show that MLN8237 treatment affects multiple processes that are differentially sensitive to the loss of Aurora-A function. A role of Aurora-A in controlling the orientation of cell division emerges. MLN8237 treatment, even in high doses, fails to induce efficient elimination of dividing cells, or of their progeny, while inducing significant aneuploidy in daughter cells. The results of single-cell analyses show a complex cellular response to MLN8237 and evidence that its effects are strongly dose-dependent: these issues deserve consideration in the light of the design of strategies to kill cancer cells via inhibition of Aurora kinases. PMID:25153724

  11. Lisofylline suppresses ex vivo release by murine spleen cells of hematopoietic inhibitors induced by cancer chemotherapeutic agents.

    PubMed

    de Vries, P; Singer, J W

    2000-08-01

    Many cytotoxic cancer therapeutic drugs activate stress response signaling pathways that transcriptionally activate a variety of genes. We decided to determine if cytotoxic therapies induce inflammatory cytokines with inhibitory effects on hematopoiesis and if lisofylline (LSF), a novel antiinflammatory compound, suppresses this induction. Mice were treated with cytosine beta-d-arabinofuranoside (AraC), cis-platinum(II)diammine-dichloride (CisP), etoposide (VP-16), or melphalan at clinically relevant doses, with or without LSF. Spleen cell conditioned media (CM) derived from mice treated with cytotoxic agents, but not from control or LSF treated mice, reduced colony formation by murine bone marrow progenitors belonging to the myeloid, erythroid, megakaryocytic, and B-lymphoid lineages. LSF (100 mg/kg), administered either simultaneously with or up to 48 hours before the cytotoxic agents, suppressed the release of this inhibitory activity. Treatment of inhibitory CM with neutralizing antibodies against known growth inhibitory cytokines, including tumor necrosis factor alpha, transforming growth factor beta, and macrophage inflammatory protein-1alpha, resulted in enhanced colony growth. We conclude that treatment of mice with chemotherapeutic drugs induces the ex vivo production of multilineage hematopoietic inhibitors and that induction of these inhibitors could be abrogated by treatment with LSF. These findings suggest a mechanism whereby LSF can accelerate recovery of hematopoiesis following cytotoxic therapies. PMID:10989192

  12. The RIP1-kinase inhibitor necrostatin-1 prevents osmotic nephrosis and contrast-induced AKI in mice.

    PubMed

    Linkermann, Andreas; Heller, Jan-Ole; Prókai, Agnes; Weinberg, Joel M; De Zen, Federica; Himmerkus, Nina; Szabó, Attila J; Bräsen, Jan H; Kunzendorf, Ulrich; Krautwald, Stefan

    2013-10-01

    The pathophysiology of contrast-induced AKI (CIAKI) is incompletely understood due to the lack of an appropriate in vivo model that demonstrates reduced kidney function before administration of radiocontrast media (RCM). Here, we examine the effects of CIAKI in vitro and introduce a murine ischemia/reperfusion injury (IRI)-based approach that allows induction of CIAKI by a single intravenous application of standard RCM after injury for in vivo studies. Whereas murine renal tubular cells and freshly isolated renal tubules rapidly absorbed RCM, plasma membrane integrity and cell viability remained preserved in vitro and ex vivo, indicating that RCM do not induce apoptosis or regulated necrosis of renal tubular cells. In vivo, the IRI-based CIAKI model exhibited typical features of clinical CIAKI, including RCM-induced osmotic nephrosis and increased serum levels of urea and creatinine that were not altered by inhibition of apoptosis. Direct evaluation of renal morphology by intravital microscopy revealed dilation of renal tubules and peritubular capillaries within 20 minutes of RCM application in uninjured mice and similar, but less dramatic, responses after IRI pretreatment. Necrostatin-1 (Nec-1), a specific inhibitor of the receptor-interacting protein 1 (RIP1) kinase domain, prevented osmotic nephrosis and CIAKI, whereas an inactive Nec-1 derivate (Nec-1i) or the pan-caspase inhibitor zVAD did not. In addition, Nec-1 prevented RCM-induced dilation of peritubular capillaries, suggesting a novel role unrelated to cell death for the RIP1 kinase domain in the regulation of microvascular hemodynamics and pathophysiology of CIAKI. PMID:23833261

  13. The RIP1-Kinase Inhibitor Necrostatin-1 Prevents Osmotic Nephrosis and Contrast-Induced AKI in Mice

    PubMed Central

    Linkermann, Andreas; Heller, Jan-Ole; Prókai, Ágnes; Weinberg, Joel M.; De Zen, Federica; Himmerkus, Nina; Szabó, Attila J.; Bräsen, Jan H.; Krautwald, Stefan

    2013-01-01

    The pathophysiology of contrast-induced AKI (CIAKI) is incompletely understood due to the lack of an appropriate in vivo model that demonstrates reduced kidney function before administration of radiocontrast media (RCM). Here, we examine the effects of CIAKI in vitro and introduce a murine ischemia/reperfusion injury (IRI)–based approach that allows induction of CIAKI by a single intravenous application of standard RCM after injury for in vivo studies. Whereas murine renal tubular cells and freshly isolated renal tubules rapidly absorbed RCM, plasma membrane integrity and cell viability remained preserved in vitro and ex vivo, indicating that RCM do not induce apoptosis or regulated necrosis of renal tubular cells. In vivo, the IRI-based CIAKI model exhibited typical features of clinical CIAKI, including RCM-induced osmotic nephrosis and increased serum levels of urea and creatinine that were not altered by inhibition of apoptosis. Direct evaluation of renal morphology by intravital microscopy revealed dilation of renal tubules and peritubular capillaries within 20 minutes of RCM application in uninjured mice and similar, but less dramatic, responses after IRI pretreatment. Necrostatin-1 (Nec-1), a specific inhibitor of the receptor-interacting protein 1 (RIP1) kinase domain, prevented osmotic nephrosis and CIAKI, whereas an inactive Nec-1 derivate (Nec-1i) or the pan-caspase inhibitor zVAD did not. In addition, Nec-1 prevented RCM-induced dilation of peritubular capillaries, suggesting a novel role unrelated to cell death for the RIP1 kinase domain in the regulation of microvascular hemodynamics and pathophysiology of CIAKI. PMID:23833261

  14. The tyrosine kinase inhibitor genistein induces the detachment of rotavirus particles from the cell surface.

    PubMed

    López, Tomás; López, Susana; Arias, Carlos F

    2015-12-01

    Group A rotaviruses are a major cause of severe gastroenteritis in young infants. In this work we evaluated the potential role of protein tyrosine kinases on rotavirus infectivity and viral progeny production. From the broad-spectrum inhibitors tested, only genistein, a flavonoid, inhibited rotavirus infectivity. The inhibition observed was dose and strain dependent, with more than 10-fold IC50 differences for some rotavirus strains, and the effect of the drug was shown to be dependent of their activity as a protein tyrosine kinase inhibitor, since the inactive analogue of genistein, daidzein, had no effect on virus infection. Investigation of the stage of virus replication blocked by the drug showed that it interferes with the early interactions of the virus with receptors and/or co-receptors, since treatment of the cells with genistein promoted the detachment of the virus from the cell surface. PMID:26216271

  15. Proteome Analysis of Peroxisomes from Etiolated Arabidopsis Seedlings Identifies a Peroxisomal Protease Involved in ?-Oxidation and Development1[C][W][OPEN

    PubMed Central

    Quan, Sheng; Yang, Pingfang; Cassin-Ross, Gaëlle; Kaur, Navneet; Switzenberg, Robert; Aung, Kyaw; Li, Jiying; Hu, Jianping

    2013-01-01

    Plant peroxisomes are highly dynamic organelles that mediate a suite of metabolic processes crucial to development. Peroxisomes in seeds/dark-grown seedlings and in photosynthetic tissues constitute two major subtypes of plant peroxisomes, which had been postulated to contain distinct primary biochemical properties. Multiple in-depth proteomic analyses had been performed on leaf peroxisomes, yet the major makeup of peroxisomes in seeds or dark-grown seedlings remained unclear. To compare the metabolic pathways of the two dominant plant peroxisomal subtypes and discover new peroxisomal proteins that function specifically during seed germination, we performed proteomic analysis of peroxisomes from etiolated Arabidopsis (Arabidopsis thaliana) seedlings. The detection of 77 peroxisomal proteins allowed us to perform comparative analysis with the peroxisomal proteome of green leaves, which revealed a large overlap between these two primary peroxisomal variants. Subcellular targeting analysis by fluorescence microscopy validated around 10 new peroxisomal proteins in Arabidopsis. Mutant analysis suggested the role of the cysteine protease RESPONSE TO DROUGHT21A-LIKE1 in ?-oxidation, seed germination, and growth. This work provides a much-needed road map of a major type of plant peroxisome and has established a basis for future investigations of peroxisomal proteolytic processes to understand their roles in development and in plant interaction with the environment. PMID:24130194

  16. The histone deacetylase inhibitor trichostatin A induces neurite outgrowth in PC12 cells via the epigenetically regulated expression of the nur77 gene.

    PubMed

    Tomioka, Takuma; Maruoka, Hiroki; Kawa, Hiromichi; Yamazoe, Ryosuke; Fujiki, Daichi; Shimoke, Koji; Ikeuchi, Toshihiko

    2014-11-01

    Histone deacetylase (HDAC) inhibitors induce histone acetylation and gene expression by changing local chromatin structures. They can thereby influence various cells to proliferate or differentiate. It has been reported that trichostatin A (TSA) or valproic acid (VPA) can induce the neuronal differentiation of mouse embryonic neural stem cells and rat cerebellar granule cells. It is unclear however which gene is responsible for the neuronal differentiation induced by HDAC inhibitors. In this study, we investigated the contribution of immediate early gene (IEG) nur77 to the neuronal differentiation induced by TSA. We report that TSA induces neurite outgrowth in PC12 cells, and C646, an inhibitor of HAT (histone acetyl transferase) (p300), prevents TSA-induced neurite formation. The acetylation of the Lys14 residue of histone H3, and mRNA and protein expression of nur77 gene were found to be stimulated after treatment with TSA, but not in the presence of C646. A knock-down of nur77 inhibits the neurite outgrowth induced by TSA. Furthermore, the ectopic expression of nur77 significantly elicits neurite formation in PC12 cells. These results suggest that the expression of nur77, which is up-regulated via the TSA-induced acetylation of Lys14 on histone H3, is essential for the neuronal differentiation in TSA-induced PC12 cells. PMID:25128386

  17. Azanitrile Cathepsin K Inhibitors: Effects on Cell Toxicity, Osteoblast-Induced Mineralization and Osteoclast-Mediated Bone Resorption

    PubMed Central

    Ren, Zhong-Yuan; Machuca-Gayet, Irma; Domenget, Chantal; Buchet, Rene; Wu, Yuqing; Jurdic, Pierre; Mebarek, Saida

    2015-01-01

    Aim The cysteine protease cathepsin K (CatK), abundantly expressed in osteoclasts, is responsible for the degradation of bone matrix proteins, including collagen type 1. Thus, CatK is an attractive target for new anti-resorptive osteoporosis therapies, but the wider effects of CatK inhibitors on bone cells also need to be evaluated to assess their effects on bone. Therefore, we selected, among a series of synthetized isothiosemicarbazides, two molecules which are highly selective CatK inhibitors (CKIs) to test their effects on osteoblasts and osteoclasts. Research Design and Methods Cell viability upon treatment of CKIs were was assayed on human osteoblast-like Saos-2, mouse monocyte cell line RAW 264.7 and mature mouse osteoclasts differentiated from bone marrow. Osteoblast-induced mineralization in Saos-2 cells and in mouse primary osteoblasts from calvaria, with or without CKIs,; were was monitored by Alizarin Red staining and alkaline phosphatase activity, while osteoclast-induced bone resorption was performed on bovine slices. Results Treatments with two CKIs, CKI-8 and CKI-13 in human osteoblast-like Saos-2, murine RAW 264.7 macrophages stimulated with RANKL and mouse osteoclasts differentiated from bone marrow stimulated with RANKL and MCSF were found not to be toxic at doses of up to 100 nM. As probed by Alizarin Red staining, CKI-8 did not inhibit osteoblast-induced mineralization in mouse primary osteoblasts as well as in osteoblast-like Saos-2 cells. However, CKI-13 led to a reduction in mineralization of around 40% at 10–100 nM concentrations in osteoblast-like Saos-2 cells while it did not in primary cells. After a 48-hour incubation, both CKI-8 and CKI-13 decreased bone resorption on bovine bone slices. CKI-13 was more efficient than the commercial inhibitor E-64 in inhibiting bone resorption induced by osteoclasts on bovine bone slices. Both CKI-8 and CKI-13 created smaller bone resorption pits on bovine bone slices, suggesting that the mobility of osteoclasts was slowed down by the addition of CKI-8 and CKI-13. Conclusion CKI-8 and CKI-13 screened here show promise as antiresorptive osteoporosis therapeutics but some off target effects on osteoblasts were found with CKI-13. PMID:26168340

  18. ?-Tocotrienol protects against ovariectomy-induced bone loss via mevalonate pathway as HMG-CoA reductase inhibitor.

    PubMed

    Deng, Lili; Ding, Yuedi; Peng, Ying; Wu, Yu; Fan, Jun; Li, Wenxin; Yang, Runlin; Yang, Meiling; Fu, Qiang

    2014-10-01

    ?-Tocotrienol (GT3), an analogue of vitamin E, has gained increasing scientific interest recently as it provides significant health benefits. GT3 exerts its biological effects not only by virtue of antioxidant properties but also by inhibiting hydroxy-methyl-glutaryl-coenzyme A (HMG-CoA) reductase. Studies have reported that the mevalonate pathway is relevant for bone metabolism and HMG-CoA reductase inhibitors can increase bone mass and are useful in osteoporosis therapy. However, whether it is involved in the bone anabolic activity of GT3 is not clear. This study was conducted to investigate the ability of GT3 to protect against ovariectomy-induced bone loss, as well as the correlation between the protections and mevalonate pathway. Results showed that mice supplemented with 100mg/kg emulsified GT3 via subcutaneous injection once per month for three months were significantly protected from ovariectomy-induced bone loss as evaluated by various bone structural parameters, bone metabolic gene expression levels and serum levels of biochemical markers for bone resorption and bone formation. Importantly, the effect of GT3 on preventing against ovariectomy-induced bone loss could be reversed by daily supplementation with mevalonate, indicating that GT3 may via an HMG-CoA reductase-dependent mechanism to protect against ovariectomy-induced bone loss. Our results suggest that GT3 is suitable as dietary supplement and has potential as an alternative drug to treat or prevent osteoporosis. PMID:25019595

  19. Neuronal NOS inhibitor and conventional antidepressant drugs attenuate stress-induced fos expression in overlapping brain regions.

    PubMed

    Silva, Michelle; Aguiar, Daniele C; Diniz, Cassiano R A; Guimarães, Francisco Silveira; Joca, Sâmia R L

    2012-04-01

    Recent evidence indicates that the administration of inhibitors of neuronal nitric oxide synthase (nNOS) induces antidepressant-like effects in animal models such as the forced swimming test (FST). However, the neural circuits involved in these effects are not yet known. Therefore, this study investigated the expression of Fos protein, a marker of neuronal activity, in the brain of rats submitted to FST and treated with the preferential nNOS inhibitor, 7-nitroindazole (7-NI), or with classical antidepressant drugs (Venlafaxine and Fluoxetine). Male Wistar rats were submitted to a forced swimming pretest (PT) and, immediately after, started receiving a sequence of three ip injections (0, 5, and 23 h after PT) of Fluoxetine (10 mg/kg), Venlafaxine (10 mg/kg), 7-NI (30 mg/kg) or respective vehicles. One hour after the last drug injection the animals were submitted to the test session, when immobility time was recorded. After the FST they were sacrificed and had their brains removed and processed for Fos immunohistochemistry. Independent group of non-stressed animals received the same drug treatments, or no treatment (naïve). 7-NI, Venlafaxine or Fluoxetine reduced immobility time in the FST, an antidepressant-like effect. None of the treatments induce significant changes in Fos expression per se. However, swimming stress induced significant increases in Fos expression in the following brain regions: medial prefrontal cortex, nucleus accumbens, locus coeruleus, raphe nuclei, striatum, hypothalamic nucleus, periaqueductal grey, amygdala, habenula, paraventricular nucleus of hypothalamus, and bed nucleus of stria terminalis. This effect was attenuated by 7-NI, Venlafaxine or Fluoxetine. These results show that 7-NI produces similar behavioral and neuronal activation effects to those of typical antidepressants, suggesting that these drugs share common neurobiological substrates. PMID:22120186

  20. Aggression behaviour induced by oral administration of the Janus-kinase inhibitor tofacitinib, but not oclacitinib, under stressful conditions.

    PubMed

    Fukuyama, Tomoki; Tschernig, Thomas; Qi, Yulin; Volmer, Dietrich A; Bäumer, Wolfgang

    2015-10-01

    Janus kinase (JAK) inhibitors have recently been developed for allergic diseases. We focused on the 2 different JAK inhibitors, tofacitinib (selective for JAK3) and oclacitinib (selective for JAK1 and 2), to clarify the mechanism of anti-inflammatory and anti-itching potency of these drugs. In the process of detecting anti-itching potency, we observed that tofacitinib treated mice showed aggression behaviour. The objective of the study reported here was to investigate the aggressive behaviour induced by tofacitinib by using a mouse model of allergic dermatitis and the resident-intruder test. For the allergic dermatitis model, female BALB/c mice were sensitised and challenged topically with toluene-2,4-diisocyanate (TDI). Vehicle, tofacitinib or oclacitinib, was administered orally 30min before TDI challenge. Scratching, aggression and standing behaviours were monitored in the 60min period immediately following challenge of TDI. Another group of male BALB/c mice treated with vehicle, tofacitinib or oclacitinib was evaluated in the resident-intruder test and brains were obtained to determine blood brain barrier penetration. In the allergic dermatitis model, a significant increase in aggression and standing behaviour was only obvious in the tofacitinib treatment group. There was no effect in non-sensitised mice, but similar aggression was also induced by tofacitinib in male resident-intruder test. Penetration of blood-brain barrier was observed both in tofacitinib and oclacitinib treated mice. These results suggest that aggression was induced by tofacitinib under some kind of stressful environment. This study indicates a possible role of the JAK-STAT pathway in modulation of aggression behaviour. PMID:26164790

  1. Mitochondrial complex I inhibitor rotenone induces apoptosis through enhancing mitochondrial reactive oxygen species production.

    PubMed

    Li, Nianyu; Ragheb, Kathy; Lawler, Gretchen; Sturgis, Jennie; Rajwa, Bartek; Melendez, J Andres; Robinson, J Paul

    2003-03-01

    Inhibition of mitochondrial respiratory chain complex I by rotenone had been found to induce cell death in a variety of cells. However, the mechanism is still elusive. Because reactive oxygen species (ROS) play an important role in apoptosis and inhibition of mitochondrial respiratory chain complex I by rotenone was thought to be able to elevate mitochondrial ROS production, we investigated the relationship between rotenone-induced apoptosis and mitochondrial reactive oxygen species. Rotenone was able to induce mitochondrial complex I substrate-supported mitochondrial ROS production both in isolated mitochondria from HL-60 cells as well as in cultured cells. Rotenone-induced apoptosis was confirmed by DNA fragmentation, cytochrome c release, and caspase 3 activity. A quantitative correlation between rotenone-induced apoptosis and rotenone-induced mitochondrial ROS production was identified. Rotenone-induced apoptosis was inhibited by treatment with antioxidants (glutathione, N-acetylcysteine, and vitamin C). The role of rotenone-induced mitochondrial ROS in apoptosis was also confirmed by the finding that HT1080 cells overexpressing magnesium superoxide dismutase were more resistant to rotenone-induced apoptosis than control cells. These results suggest that rotenone is able to induce apoptosis via enhancing the amount of mitochondrial reactive oxygen species production. PMID:12496265

  2. Peroxisome proliferator-activated receptor alpha, PPAR?, directly regulates transcription of cytochrome P450 CYP2C8

    PubMed Central

    Thomas, Maria; Winter, Stefan; Klumpp, Britta; Turpeinen, Miia; Klein, Kathrin; Schwab, Matthias; Zanger, Ulrich M.

    2015-01-01

    The cytochrome P450, CYP2C8, metabolizes more than 60 clinically used drugs as well as endogenous substances including retinoic acid and arachidonic acid. However, predictive factors for interindividual variability in the efficacy and toxicity of CYP2C8 drug substrates are essentially lacking. Recently we demonstrated that peroxisome proliferator-activated receptor alpha (PPAR?), a nuclear receptor primarily involved in control of lipid and energy homeostasis directly regulates the transcription of CYP3A4. Here we investigated the potential regulation of CYP2C8 by PPAR?. Two linked intronic SNPs in PPAR? (rs4253728, rs4823613) previously associated with hepatic CYP3A4 status showed significant association with CYP2C8 protein level in human liver samples (N = 150). Furthermore, siRNA-mediated knock-down of PPAR? in HepaRG human hepatocyte cells resulted in up to ?60 and ?50% downregulation of CYP2C8 mRNA and activity, while treatment with the PPAR? agonist WY14,643 lead to an induction by >150 and >100%, respectively. Using chromatin immunoprecipitation scanning assay we identified a specific upstream gene region that is occupied in vivo by PPAR?. Electromobility shift assay demonstrated direct binding of PPAR? to a DR-1 motif located at positions –2762/–2775 bp upstream of the CYP2C8 transcription start site. We further validated the functional activity of this element using luciferase reporter gene assays in HuH7 cells. Moreover, based on our previous studies we demonstrated that WNT/?-catenin acts as a functional inhibitor of PPAR?-mediated inducibility of CYP2C8 expression. In conclusion, our data suggest direct involvement of PPAR? in both constitutive and inducible regulation of CYP2C8 expression in human liver, which is further modulated by WNT/?-catenin pathway. PPARA gene polymorphism could have a modest influence on CYP2C8 phenotype. PMID:26582990

  3. Peroxisomal organization in normal and cerebrohepatorenal (Zellweger) syndrome fibroblasts.

    PubMed Central

    Santos, M J; Ojeda, J M; Garrido, J; Leighton, F

    1985-01-01

    The reported absence of morphologically detectable peroxisomes in liver and kidney tissue cells from patients affected by the autosomic recessive, inherited metabolic disease known as cerebrohepatorenal, or Zellweger, syndrome was studied in fibroblasts, assuming it to be a generalized defect. Normal cultured fibroblasts were shown to contain peroxisomes according to morphological, biochemical, and subcellular fractionation criteria: particle-bound catalase and fatty acyl-CoA oxidase copurify in subcellular fractionation by differential centrifugation or isopycnic equilibrium in continuous density gradients and peroxidase-positive organelles of approximately equal to 0.1 micron in diameter are detected in the cytoplasm. In Zellweger cultured fibroblasts, these peroxisomal enzymes are present; however, they behave as cytosolic enzymes in the different subcellular fractionation procedures employed and peroxisomes are not detected cytochemically. These findings support the hypothesis that the lack of peroxisomes in this genetic disease is the consequence of a defect in the assembly of the peroxisomal constituents. Furthermore, the value of fibroblasts for subcellular analysis of peroxisomal defects is illustrated. Images PMID:2995971

  4. Expression cloning of genes encoding human peroxisomal proteins

    SciTech Connect

    Spathaky, J.M.; Tate, A.W.; Cox, T.M.

    1994-09-01

    Numerous metabolic disorders associated with diverse peroxisomal defects have been identified but their molecular characterization has been hampered by difficulties associated with the purification of proteins from this fragile organelle. We have utilized antibodies directed against the C-terminal tripeptide peroxisomal targeting signal to detect hitherto unknown peroxisomal proteins in tissue fractions and to isolate genes encoding peroxisonal proteins from human expression libraries. We immunized rabbits with a peptide conjugate encompassing the C-terminal nine amino acids of rat peroxisomal acyl CoA oxidase. Immunoprecipitation assays using radio-labelled peptide showed that the antibody specifically recognizes the terminal SKL motif as well as C-terminal SHL and SRL but not SHL at an internal position. Affinity-purified antibody was used to probe Western blots of crude and peroxisome-enriched monkey liver preparations and detected 8-10 proteins specifically in the peroxisome fractions. 100 positive clones were identified on screening a human liver cDNA expression library in {lambda}-gt11. Sequence analysis has confirmed the identity of cDNA clones for human acyl CoA oxidase and epoxide hydrolase. Four clones show no sequence identity and their putative role in the human peroxisome is being explored.

  5. Characterization of peroxisome-deficient mutants of Hansenula polymorpha.

    PubMed

    Tan, X; Titorenko, V I; van der Klei, I J; Sulter, G J; Haima, P; Waterham, H R; Eyers, M; Harder, W; Veenhuis, M; Cregg, J M

    1995-08-01

    In the methylotrophic yeast Hansenula polymorpha, approximately 25% of all methanol-utilization-defective (Mut-) mutants are affected in genes required for peroxisome biogenesis (PER genes). Previously, we reported that one group of per mutants, termed Pim-, are characterized by the presence of a few small peroxisomes with the bulk of peroxisomal enzymes located in the cytosol. Here, we describe a second major group of per mutants that were observed to be devoid of any peroxisome-like structure (Per-). In each Per- mutant, the peroxisomal methanol-pathway enzymes alcohol oxidase, catalase and dihydroxyacetone synthase were present and active but located in the cytosol. Together, the Pim- and Per- mutant collections involved mutations in 14 different PER genes. Two of the genes, PER5 and PER7, were represented by both dominant-negative and recessive alleles. Diploids resulting from crosses of dominant per strains and wild-type H. polymorpha were Mut- and harbored peroxisomes with abnormal morphology. This is the first report of dominant-negative mutations affecting peroxisome biogenesis. PMID:8529271

  6. EFFECT OF OZONE ON DRUG-INDUCED SLEEPING TIME IN MICE PRETREATED WITH MIXED-FUNCTION OXIDASE INDUCERS AND INHIBITORS

    EPA Science Inventory

    Studies were conducted to investigate the effect of ozone in prolonging pentobarbital (PEN)-induced sleeping time (S.T.). Since ozone is a common air pollutant, an ozone-induced alteration of mechanisms of drug action could have public health implications. It was shown that a 5-h...

  7. Inhibitor of apoptosis signal-regulating kinase 1 protects against acetaminophen-induced liver injury.

    PubMed

    Xie, Yuchao; Ramachandran, Anup; Breckenridge, David G; Liles, John T; Lebofsky, Margitta; Farhood, Anwar; Jaeschke, Hartmut

    2015-07-01

    Metabolic activation and oxidant stress are key events in the pathophysiology of acetaminophen (APAP) hepatotoxicity. The initial mitochondrial oxidative stress triggered by protein adduct formation is amplified by c-jun-N-terminal kinase (JNK), resulting in mitochondrial dysfunction and ultimately cell necrosis. Apoptosis signal-regulating kinase 1 (ASK1) is considered the link between oxidant stress and JNK activation. The objective of the current study was to assess the efficacy and mechanism of action of the small-molecule ASK1 inhibitor GS-459679 in a murine model of APAP hepatotoxicity. APAP (300 mg/kg) caused extensive glutathione depletion, JNK activation and translocation to the mitochondria, oxidant stress and liver injury as indicated by plasma ALT activities and area of necrosis over a 24h observation period. Pretreatment with 30 mg/kg of GS-459679 almost completely prevented JNK activation, oxidant stress and injury without affecting the metabolic activation of APAP. To evaluate the therapeutic potential of GS-459679, mice were treated with APAP and then with the inhibitor. Given 1.5h after APAP, GS-459679 was still protective, which was paralleled by reduced JNK activation and p-JNK translocation to mitochondria. However, GS-459679 treatment was not more effective than N-acetylcysteine, and the combination of GS-459679 and N-acetylcysteine exhibited similar efficacy as N-acetylcysteine monotherapy, suggesting that GS-459769 and N-acetylcysteine affect the same pathway. Importantly, inhibition of ASK1 did not impair liver regeneration as indicated by PCNA staining. In conclusion, the ASK1 inhibitor GS-459679 protected against APAP toxicity by attenuating JNK activation and oxidant stress in mice and may have therapeutic potential for APAP overdose patients. PMID:25818599

  8. Effect of cyclooxygenase inhibitors on gentamicin-induced nephrotoxicity in rats.

    PubMed

    Hosaka, E M; Santos, O F P; Seguro, A C; Vattimo, M F F

    2004-07-01

    The frequent use of nonsteroidal anti-inflammatory drugs (NSAID) in combination with gentamicin poses the additional risk of nephrotoxic renal failure. Cyclooxygenase-1 (COX-1) is the main enzyme responsible for the synthesis of renal vasodilator prostaglandins, while COX-2 participates predominantly in the inflammatory process. Both are inhibited by non-selective NSAID such as indomethacin. Selective COX-2 inhibitors such as rofecoxib seem to have fewer renal side effects than non-selective inhibitors. The objective of the present study was to determine whether the combined use of rofecoxib and gentamicin can prevent the increased renal injury caused by gentamicin and indomethacin. Male Wistar rats (250-300 g) were treated with gentamicin (100 mg/kg body weight, ip, N = 7), indomethacin (5 mg/kg, orally, N = 7), rofecoxib (1.4 mg/kg, orally, N = 7), gentamicin + rofecoxib (100 and 1.4 mg/kg, respectively) or gentamicin + indomethacin (100 and 5 mg/kg, respectively, N = 8) for 5 days. Creatinine clearance and alpha-glutathione-S-transferase concentrations were used as markers of renal injury. Animals were anesthetized with ether and sacrificed for blood collection. The use of gentamicin plus indomethacin led to worsened renal function (0.199 +/- 0.019 ml/min), as opposed to the absence of a nephrotoxic effect of rofecoxib when gentamicin plus rofexicob was used (0.242 +/- 0.011 ml/min). These results indicate that COX-2-selective inhibitors can be used as an alternative treatment to conventional NSAID, especially in situations in which risk factors for nephrotoxicity are present. PMID:15264004

  9. Novel histone deacetylase inhibitors induce growth arrest, apoptosis, and differentiation in sarcoma cancer stem cells.

    PubMed

    Di Pompo, Gemma; Salerno, Manuela; Rotili, Dante; Valente, Sergio; Zwergel, Clemens; Avnet, Sofia; Lattanzi, Giovanna; Baldini, Nicola; Mai, Antonello

    2015-05-14

    Musculoskeletal sarcomas are aggressive malignancies of bone and soft tissues often affecting children and adolescents. Histone deacetylase inhibitors (HDACi) have been proposed to counteract cancer stem cells (CSCs) in solid neoplasms. When tested in human osteosarcoma, rhabdomyosarcoma, and Ewing's sarcoma stem cells, the new HDACi MC1742 (1) and MC2625 (2) increased acetyl-H3 and acetyl-tubulin levels and inhibited CSC growth by apoptosis induction. At nontoxic doses, 1 promoted osteogenic differentiation. Further investigation with 1 will be done in preclinical sarcoma models. PMID:25905694

  10. Peroxisomes: a Nexus for Lipid Metabolism and Cellular Signaling

    PubMed Central

    Lodhi, Irfan J.; Semenkovich, Clay F.

    2014-01-01

    Peroxisomes are often dismissed as the cellular hoi polloi, relegated to cleaning up reactive oxygen chemical debris discarded by other organelles. However, their functions extend far beyond hydrogen peroxide metabolism. Peroxisomes are intimately associated with lipid droplets and mitochondria, and their ability to carry out fatty acid oxidation and lipid synthesis, especially the production of ether lipids, may be critical for generating cellular signals required for normal physiology. Here we review the biology of peroxisomes and their potential relevance to human disorders including cancer, obesity-related diabetes, and degenerative neurologic disease. PMID:24508507

  11. 10058-F4, a c-Myc inhibitor, markedly increases valproic acid-induced cell death in Jurkat and CCRF-CEM T-lymphoblastic leukemia cells

    PubMed Central

    MU, QITIAN; MA, QIULING; LU, SHASHA; ZHANG, TING; YU, MENGXIA; HUANG, XIN; CHEN, JIAN; JIN, JIE

    2014-01-01

    Adult T-cell acute lymphoblastic leukemia (T-ALL) has a poor prognosis. Although it has been found that activation of Notch1 signaling occurs in >50% T-ALL patients, ?-secretase inhibitors that target Notch1 signaling are of limited efficacy. However, c-Myc is an important direct target of Notch1 and, thus, c-Myc is another potential therapeutic target for T-ALL. Valproic acid (VPA), a histone deacetylase inhibitor, has been reported to treat various hematological malignancies. In the present study, we showed that c-Myc expression, at a transcriptional level, was dose-dependently downregulated in VPA-induced growth inhibition in T-ALL cell lines, Jurkat and CCRF-CEM cells. 10058-F4, a small molecule c-Myc inhibitor, could increase the downregulation of c-Myc and markedly increase the growth inhibition and cell death induced by VPA in Jurkat and CCRF-CEM cells, which was accompanied by obvious cleavage of capase-3. Z-VAD-FMK, a caspase inhibitor, partially prevented the anti-leukemic effect. The results of the present study suggest that c-Myc inhibitors increase cell death induced by VPA in a caspase-dependent and -independent manner, and their combination could be a potent therapeutic strategy for adult T-ALL patients. PMID:25120723

  12. Simultaneous Visualization of Peroxisomes and Cytoskeletal Elements Reveals Actin and Not Microtubule-Based Peroxisome Motility in Plants1[w

    PubMed Central

    Mathur, Jaideep; Mathur, Neeta; Hülskamp, Martin

    2002-01-01

    Peroxisomes were visualized in living plant cells using a yellow fluorescent protein tagged with a peroxisomal targeting signal consisting of the SKL motif. Simultaneous visualization of peroxisomes and microfilaments/microtubules was accomplished in onion (Allium cepa) epidermal cells transiently expressing the yellow fluorescent protein-peroxi construct, a green fluorescent protein-mTalin construct that labels filamentous-actin filaments, and a green fluorescent protein-microtubule-binding domain construct that labels microtubules. The covisualization of peroxisomes and cytoskeletal elements revealed that, contrary to the reports from animal cells, peroxisomes in plants appear to associate with actin filaments and not microtubules. That peroxisome movement is actin based was shown by pharmacological studies. For this analysis we used onion epidermal cells and various cell types of Arabidopsis including trichomes, root hairs, and root cortex cells exhibiting different modes of growth. In transient onion epidermis assay and in transgenic Arabidopsis plants, an interference with the actin cytoskeleton resulted in progressive loss of saltatory movement followed by the aggregation and a complete cessation of peroxisome motility within 30 min of drug application. Microtubule depolymerization or stabilization had no effect. PMID:11891258

  13. Fumonisins and fumonisin analogs as inhibitors of ceramide synthase and inducers of apoptosis.

    PubMed

    Desai, Kena; Sullards, M Cameron; Allegood, Jeremy; Wang, Elaine; Schmelz, Eva M; Hartl, Michaela; Humpf, Hans-Ulrich; Liotta, D C; Peng, Qiong; Merrill, Alfred H

    2002-12-30

    Sphingoid bases are growth inhibitory and pro-apoptotic for many types of cells when added to cells exogenously, and can be elevated to toxic amounts endogenously when cells are exposed to inhibitors of ceramide synthase. An important category of naturally occurring inhibitors are the fumonisins, which inhibit ceramide synthase through structural similarities with both the sphingoid base and fatty acyl-CoA co-substrates. Fumonisins cause a wide spectrum of disease (liver and renal toxicity and carcinogenesis, neurotoxicity, induction of pulmonary edema, and others), and most-possibly all-of the pathophysiologic effects of fumonisins are attributable to disruption of the sphingolipid metabolism. The products of alkaline hydrolysis of fumonisins (which occurs during the preparation of masa flour for tortillas) are aminopentols that also inhibit ceramide synthase, but more weakly. Nonetheless, the aminopentols (and other 1-deoxy analogs of sphinganine) are acylated to derivatives that inhibit ceramide synthase, perhaps as product analogs, elevate sphinganine, and kill the cells. Somewhat paradoxically, fumonisins sometimes stimulate growth and inhibit apoptosis, possibly due to elevation of sphinganine 1-phosphate, which is known to have these cellular effects. These findings underscore the complexity of sphingolipid metabolism and the difficulty of identifying the pertinent mediators unless a full profile of the potentially bioactive species is evaluated. PMID:12531553

  14. Anti-Influenza Neuraminidase Inhibitor Oseltamivir Phosphate Induces Canine Mammary Cancer Cell Aggressiveness

    PubMed Central

    de Oliveira, Joana T.; Santos, Ana L.; Gomes, Catarina; Barros, Rita; Ribeiro, Cláudia; Mendes, Nuno; de Matos, Augusto J.; Vasconcelos, M. Helena; Oliveira, Maria José; Reis, Celso A.; Gärtner, Fátima

    2015-01-01

    Oseltamivir phosphate is a widely used anti-influenza sialidase inhibitor. Sialylation, governed by sialyltransferases and sialidases, is strongly implicated in the oncogenesis and progression of breast cancer. In this study we evaluated the biological behavior of canine mammary tumor cells upon oseltamivir phosphate treatment (a sialidase inhibitor) in vitro and in vivo. Our in vitro results showed that oseltamivir phosphate impairs sialidase activity leading to increased sialylation in CMA07 and CMT-U27 canine mammary cancer cells. Surprisingly, oseltamivir phosphate stimulated, CMT-U27 cell migration and invasion capacity in vitro, in a dose-dependent manner. CMT-U27 tumors xenograft of oseltamivir phosphate-treated nude mice showed increased sialylation, namely ?2,6 terminal structures and SLe(x) expression. Remarkably, a trend towards increased lung metastases was observed in oseltamivir phosphate-treated nude mice. Taken together, our findings revealed that oseltamivir impairs canine mammary cancer cell sialidase activity, altering the sialylation pattern of canine mammary tumors, and leading, surprisingly, to in vitro and in vivo increased mammary tumor aggressiveness. PMID:25850034

  15. LW6, a hypoxia-inducible factor 1 inhibitor, selectively induces apoptosis in hypoxic cells through depolarization of mitochondria in A549 human lung cancer cells

    PubMed Central

    SATO, MARIKO; HIROSE, KATSUMI; KASHIWAKURA, IKUO; AOKI, MASAHIKO; KAWAGUCHI, HIDEO; HATAYAMA, YOSHIOMI; AKIMOTO, HIROYOSHI; NARITA, YUICHIRO; TAKAI, YOSHIHIRO

    2015-01-01

    Hypoxia-inducible factor 1 (HIF-1) activates the transcription of genes that act upon the adaptation of cancer cells to hypoxia. LW6, an HIF-1 inhibitor, was hypothesized to improve resistance to cancer therapy in hypoxic tumors by inhibiting the accumulation of HIF-1?. A clear anti-tumor effect under low oxygen conditions would indicate that LW6 may be an improved treatment strategy for cancer in hypoxia. In the present study, the HIF-1 inhibition potential of LW6 on the growth and apoptosis of A549 lung cancer cells in association with oxygen availability was evaluated. LW6 was observed to inhibit the expression of HIF-1? induced by hypoxia in A549 cells at 20 mM, independently of the von Hippel-Lindau protein. In addition, at this concentration, LW6 induced hypoxia-selective apoptosis together with a reduction in the mitochondrial membrane potential. The intracellular reactive oxygen species levels increased in LW6-treated hypoxic A549 cells and LW6 induced a hypoxia-selective increase of mitochondrial O2•?. In conclusion, LW6 inhibited the growth of hypoxic A549 cells by affecting the mitochondria. The inhibition of the mitochondrial respiratory chain is suggested as a potentially effective strategy to target apoptosis in cancer cells. PMID:26017562

  16. New bichalcone analogs as NF-?B inhibitors and as cytotoxic agents inducing Fas/CD95-dependent apoptosis

    PubMed Central

    Vijaya Bhaskar Reddy, M.; Shen, Yuh-Chiang; Hwang, Tsong-Long; Bastow, Kenneth F.; Qian, Keduo; Lee, Kuo-Hsiung; Wu, Tian-Shung

    2011-01-01

    A series of novel bichalcone analogs were synthesized and evaluated in lipopolysaccharide (LPS)-activated microglial cells as inhibitors of nitric oxide (NO) and for in vitro anticancer activity using a limited panel of four human cancer cell lines. All analogs inhibited NO production. Compounds 4 and 11 exhibited optimal activity with IC50 values of 0.3 and 0.5 µM, respectively, and were at least 38-fold better than the positive control. A mechanism of action study showed that both compounds significantly blocked the nuclear translocation of NF-?B p65 and up-regulation of iNOS at 1.0 µM. Compound 4 and three other analogs (3, 20, and 23) exerted significant in vitro anticancer activity GI50 values ranging from 0.70~13.10 µM. A mode of action study using HT-29 colon cancer cells showed that 23 acts by inducing apoptosis signaling. PMID:21377368

  17. Restoration of TRAIL-induced apoptosis in resistant human pancreatic cancer cells by a novel FAK inhibitor, PH11.

    PubMed

    Dao, P; Smith, N; Scott-Algara, D; Garbay, C; Herbeuval, J P; Chen, H

    2015-04-28

    Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) emerges as one of the most-promising experimental cancer therapeutic drugs and is currently being tested in clinical trials. However, both intrinsic and acquired resistance of human cancer cells to TRAIL-induced apoptosis poses a huge problem in establishing clinically efficient TRAIL therapies. To assess the regulation of TRAIL-resistance in human pancreatic cancer cells, we studied the TRAIL resistant pancreatic cell line PANC-1. We show that treatment with PH11, a novel Focal Adhesion Kinase (FAK) inhibitor in association with TRAIL rapidly induces apoptosis in TRAIL-resistant PANC-1 cells, but not in normal human fibroblast cells. To explain sensitization, we showed that PH11 restores TRAIL apoptotic pathway in PANC-1 cells through down-regulation of c-FLIP via inhibition of FAK and the phosphatidylinositol-3 kinase (PI3K)/AKT pathways. These findings suggest that this combined treatment may offer an attractive therapeutic strategy for safely and efficiently treating pancreatic cancer. PMID:25684663

  18. Selective cyclooxygenase-2 inhibitor compound 11b improves haloperidol-induced catatonia by enhancing the striatum dopaminergic neurotransmission.

    PubMed

    Aghasadeghi, Mohammad Reza; Siadat, Seyyed Davar; Shafiee Ardestani, Mehdi; Jabbari Arabzadeh, Ali; Elmi, Mitra; Fathi Moghaddam, Hadi

    2012-01-01

    The aim of this research was to investigate the Cyclooxygenase-2 (COX-2) selective inhibition effect on haloperidol-induced catatonia. In this study, the effect of orally, acutely and Sub-chronically administrations of compound 11b [1-(phenyl)-5-(4-methylsulfonylphenyl)-2-ethylthioimidazole] (2, 4 and 8 mg/kg), a newly selective COX-2 inhibitor, was investigated against the haloperidol-induced catatonia phenomenon comparing to the standard drug scopolamine (1 mg/Kg) followed by microdialysis analysis of Striatum dopaminergic neurotransmission. The results showed a great potency for compound 11b in improvement of catalepsy followed by enhancing the dopaminergic neurotransmission p < 0.05. In addition, our statistical analysis showed that the protective effect of compound 11b against haloperidol-induced catatonia was both dose- and time-dependent. These findings are additional pharmacological data that suggest the effectiveness of compound 11b in treatment of schizophrenic drug overdoses and also Parkinson's disease (PD) affiliated rigidity. PMID:24250457

  19. The selective cyclooxygenase-2 inhibitor, the compound 11b improves haloperidol induced catatonia by enhancing the striatum dopaminergic neurotransmission.

    PubMed

    Fathi-Moghaddam, Hadi; Shafiee Ardestani, Mehdi; Saffari, Mostafa; Jabbari Arabzadeh, Ali; Elmi, Mitra

    2010-01-01

    A substantial amount of evidence has proposed an important role for Cyclooxygenase-2 (COX-2) enzyme in brain diseases and affiliate disorders. The purpose of this research was studying the effects of COX-2 selective inhibition on haloperidol-induced catatonia in an animal model of drug overdose and Parkinson's disease (PD). In this study, the effect of acute and Sub-chronic oral administration of a new selective COX-2 inhibitor, i.e. the compound 11b or 1-(Phenyl)-5-(4-methylsulfonylphenyl)-2-ethylthioimidazole, in a dosage of 2, 4 and 8 mg/kg on haloperidol-induced catatonia was evaluated and compared to the standard drug scopolamine (1 mg/kg) by microanalysis of Striatum dopaminergic neurotransmission. The results showed a very high potency for 11b in improving the catalepsy by enhancing the dopaminergic neurotranmission (p < 0.05). In addition, statistical analysis showed the dose- and time-dependent behavior of the observed protective effect of 11b against the haloperidol-induced catatonia and enhancement of the dopaminergic neurotransmission. These findings are additional pharmacological data that suggest the effectiveness of COX-2 inhibition in treatment of schizophreny-associated rigidity. PMID:24381603

  20. Effect of inhibitors of polyamine biosynthesis on gibberellin-induced internode growth in light-grown dwarf peas

    NASA Technical Reports Server (NTRS)

    Kaur-Sawhney, R.; Dai, Y. R.; Galston, A. W.

    1986-01-01

    When gibberellic acid (GA3) is sprayed on 9-day-old light-brown dwarf Progress pea (Pisum sativum) seedlings, arginine decarboxylase (ADC; EC 4.1.1.9) activity increases within 3 h and peaks at about 9 h after GA3 application. This is followed by a second lower peak at about 30 h; both peaks were higher than the corresponding peaks in the controls. In contrast, no appreciable effect of GA3 on internode length was observed until about 12 h, after which time a dramatic increase in growth rate occurred and persisted for about 12 h. Specific (DL-alpha-difluoromethylarginine) and non-specific (D-arginine and L-canavanine) inhibitors of ADC strongly inhibited ADC activity and to a lesser extent internode growth. The inhibition was reversed only slightly by the addition of polyamines. Actinomycin D and cycloheximide inhibited the rise in ADC activity induced by GA3. The half-life of the enzyme was increased by GA3 treatment. The results suggest that part of the GA3-induced increase in internode growth may result from enhanced polyamine biosynthesis through the ADC pathway. Furthermore, the GA3 induced increase in ADC activity probably requires de novo synthesis of both RNA and protein.

  1. The LSD1 inhibitor RN-1 induces fetal hemoglobin synthesis and reduces disease pathology in sickle cell mice.

    PubMed

    Cui, Shuaiying; Lim, Kim-Chew; Shi, Lihong; Lee, Mary; Jearawiriyapaisarn, Natee; Myers, Greggory; Campbell, Andrew; Harro, David; Iwase, Shigeki; Trievel, Raymond C; Rivers, Angela; DeSimone, Joseph; Lavelle, Donald; Saunthararajah, Yogen; Engel, James Douglas

    2015-07-16

    Inhibition of lysine-specific demethylase 1 (LSD1) has been shown to induce fetal hemoglobin (HbF) levels in cultured human erythroid cells in vitro. Here we report the in vivo effects of LSD1 inactivation by a selective and more potent inhibitor, RN-1, in a sickle cell disease (SCD) mouse model. Compared with untreated animals, RN-1 administration leads to induced HbF synthesis and to increased frequencies of HbF-positive cells and mature erythrocytes, as well as fewer reticulocytes and sickle cells, in the peripheral blood of treated SCD mice. In keeping with these observations, histologic analyses of the liver and spleen of treated SCD mice verified that they do not exhibit the necrotic lesions that are usually associated with SCD. These data indicate that RN-1 can effectively induce HbF levels in red blood cells and reduce disease pathology in SCD mice, and may therefore offer new therapeutic possibilities for treating SCD. PMID:26031919

  2. The endoplasmic reticulum stress inhibitor salubrinal inhibits the activation of autophagy and neuroprotection induced by brain ischemic preconditioning

    PubMed Central

    Gao, Bo; Zhang, Xiang-yang; Han, Rong; Zhang, Tong-tong; Chen, Cheng; Qin, Zheng-hong; Sheng, Rui

    2013-01-01

    Aim: To investigate whether endoplasmic reticulum (ER) stress participates in the neuroprotective effects of ischemic preconditioning (IPC)-induced neuroprotection and autophagy activation in rat brains. Methods: The right middle cerebral artery in SD rats was occluded for 10 min to induce focal cerebral IPC, and was occluded permanently 24 h later to induce permanent focal ischemia (PFI). ER stress inhibitor salubrinal (SAL) was injected via intracerebral ventricle infusion 10 min before the onset of IPC. Infarct volume and motor behavior deficits were examined after the ischemic insult. The protein levels of LC3, p62, HSP70, glucose-regulated protein 78 (GRP 78), p-eIF2? and caspase-12 in the ipsilateral cortex were analyzed using immunoblotting. LC3 expression pattern in the sections of ipsilateral cortex was observed with immunofluorescence. Results: Pretreatment with SAL (150 pmol) abolished the neuroprotective effects of IPC, as evidenced by the significant increases in mortality, infarct volume and motor deficits after PFI. At the molecular levels, pretreatment with SAL (150 pmol) significantly increased p-eIF2? level, and decreased GRP78 level after PFI, suggesting that SAL effectively inhibited ER stress in the cortex. Furthermore, the pretreatment with SAL blocked the IPC-induced upregulation of LC3-II and downregulation of p62 in the cortex, thus inhibiting the activation of autophagy. Moreover,SAL blocked the upregulation of HSP70, but significantly increased the cleaved caspase-12 level, thus promoting ER stress-dependent apoptotic signaling in the cortex. Conclusion: ER stress-induced autophagy might contribute to the neuroprotective effect of brain ischemic preconditioning. PMID:23603983

  3. Matrix metalloproteinase inhibitors prevent sepsis-induced refractoriness to vasoconstrictors in the cecal ligation and puncture model in rats.

    PubMed

    de Souza, Priscila; Schulz, Richard; da Silva-Santos, José Eduardo

    2015-10-15

    Previous studies have shown that the loss of contractility in aortas from lipopolysaccharide (LPS)-treated rats is related to intracellular activation of matrix metalloproteinase (MMPs). However, the role of MMPs in the vascular refractoriness to vasoconstrictors has not been investigated in a model of polymicrobial sepsis. We evaluated the effects of the oral administration of the MMP inhibitors doxycycline or ONO-4817 in the in vitro vascular reactivity of aortic rings from rats subjected to the cecal ligation and puncture (CLP) model of sepsis. Both doxycycline and ONO-4817 did not change vascular responses in sham-operated rats, but fully prevented hyporeactivity to KCl, phenylephrine and angiotensin II in vessels from CLP rats. This protective effect was not associated with changes in hematological parameters or blood nitrate and nitrite. The refractoriness to contractile agents was accompanied by enhanced activity of MMP-2 in aorta from CLP rats, which was abrogated by MMP inhibitors. CLP-induced sepsis did not impair the levels of MMP-2 in aorta, but significantly reduced calponin-1, a regulatory protein of vascular contraction. In addition, augmented levels of TIMP-1 were found in vessels from CLP rats. All these differences were prevented by either doxycycline or ONO-4817. Our study shows, for the first time in the CLP rat model of sepsis, that the vascular refractoriness to different contractile agents induced by polymicrobial sepsis is associated with increased activity of MMP-2 and reduced amounts of calponin-1 in the aorta. These findings reinforce the importance of the enhanced activity of MMPs for vascular failure in septic shock. PMID:26297976

  4. CXC Receptor 1 and 2 and Neutrophil Elastase Inhibitors Alter Radiation-induced Lung Disease in the Mouse

    SciTech Connect

    Fox, Jessica; Haston, Christina K.

    2013-01-01

    Purpose: We previously reported increased numbers of neutrophils to be associated with the development of the radiation-induced lung responses of alveolitis (pneumonitis) and fibrosis in mice. In the present study we investigated whether CXC receptor 1 and 2 antagonism with DF2156A, a small molecule inhibitor of neutrophil chemotaxis, or the neutrophil elastase inhibitor sivelestat decreases the lung response to irradiation. Methods and Materials: KK/HIJ mice received 14 Gy whole-thorax irradiation, and a subset of them received drug treatment 3 times per week from the day of irradiation until they were killed because of respiratory distress symptoms. Results: Irradiated mice receiving sivelestat survived 18% longer than did mice receiving radiation alone (73 vs 60 days for female mice, 91 vs 79 days for male mice), whereas postirradiation survival times did not differ between the group of mice receiving DF2156A and the radiation-only group. The numbers of neutrophils in lung tissue and in bronchoalveolar lavage fluid did not differ among groups of irradiated mice, but they significantly exceeded the levels in unirradiated control mice. The extent of alveolitis, assessed histologically, did not differ between irradiated mice treated with either drug and those receiving radiation alone, when assessed at the end of the experiment, but it was significantly reduced, as were the neutrophil measures, in sivelestat-treated mice at the common kill time of 60 days after irradiation. Mice treated with radiation and DF2156A developed significantly less fibrosis than did mice receiving radiation alone, and this difference was associated with decreased expression of interleukin-13 in lung tissue. Conclusions: We conclude that neutrophil elastase inhibition affects alveolitis and prolongs survival, whereas CXCR1/2 antagonism reduces radiation-induced fibrotic lung disease in mice without affecting the onset of distress.

  5. Inducible Nitric Oxide Inhibitors Block NMDA Antagonist-Stimulated Motoric Behaviors and Medial Prefrontal Cortical Glutamate Efflux

    PubMed Central

    Bergstrom, Hadley C.; Darvesh, Altaf S.; Berger, S. P.

    2015-01-01

    Nitric oxide (NO) plays a critical role in the motoric and glutamate releasing action of N-methyl-D-aspartate (NMDA)-antagonist stimulants. Earlier studies utilized neuronal nitric oxide synthase inhibitors (nNOS) for studying the neurobehavioral effects of non-competitive NMDA-antagonist stimulants such as dizocilpine (MK-801) and phencyclidine (PCP). This study explores the role of the inducible nitric oxide synthase inhibitors (iNOS) aminoguanidine (AG) and (-)-epigallocatechin-3-gallate (EGCG) in NMDA-antagonist induced motoric behavior and prefrontal cortical glutamate e?ux. Adult male rats were administered a dose range of AG, EGCG, or vehicle prior to receiving NMDA antagonists MK-801, PCP, or a conventional psychostimulant (cocaine) and tested for motoric behavior in an open arena. Glutamate in the medial prefrontal cortex (mPFC) was measured using in vivo microdialysis after a combination of AG or EGCG prior to MK-801. Acute administration of AG or EGCG dose-dependently attenuated the locomotor and ataxic properties of MK-801 and PCP. Both AG and EGCG were unable to block the motoric effects of cocaine, indicating the acute pharmacologic action of AG and EGCG is specific to NMDA antagonism and not generalizable to all stimulant class drugs. AG and EGCG normalized MK-801-stimulated mPFC glutamate e?ux. These data demonstrate that AG and EGCG attenuates NMDA antagonist-stimulated motoric behavior and cortical glutamate e?ux. Our results suggest that EGCG-like polyphenol nutraceuticals (contained in “green tea” and chocolate) may be clinically useful in protecting against the adverse behavioral dissociative and cortical glutamate stimulating effects of NMDA antagonists. Medications that interfere with NMDA antagonists such as MK-801 and PCP have been proposed as treatments for schizophrenia. PMID:26696891

  6. Induction of peroxisomal enzymes in livers of neonatal rats exposed to lactating mothers treated with hypolipidaemic drugs. Role of drug metabolite transfer in milk.

    PubMed Central

    Fahl, W E; Lalwani, N D; Reddy, M K; Reddy, J K

    1983-01-01

    Lactating rats were administered by gavage 100 mg/kg body wt. twice a day of either nafenopin or Wy-14,643, two hypolipidaemic drugs with hepatic peroxisome proliferative property. Neonatal rats, after feeding from the drug-treated mothers for 8-14 days, showed sustained increases in both the proliferation of hepatic peroxisomes, as well as in levels of the peroxisome-associated enzymes catalase (3-fold), carnitine acetyltransferase (15-35-fold), peroxisomal enoyl-CoA hydratase (29-46-fold), and palmitoyl-CoA oxidation (12-14-fold). These increases in enzyme activities in suckling rats were similar to those seen in the livers of the drug-treated, lactating mothers after 14 days of treatment. After administering [3H]nafenopin or [3H]Wy-14,643 to lactating rats, significant levels of drug-derived radioactivity were observed in suckling rat gastric milk curds by 2-4 h with significant radioactivity seen in suckling rat livers by 4-6 h. T.l.c. analysis of organic extracts of milk samples from [3H]Wy-14,643 treated animals indicated no detectable levels of the parent drug, only more-polar metabolites. Wy-14,643 metabolites preparatively purified from a rat liver microsomal fraction incubation induced peroxisome proliferation when injected into a neonatal rat. Preparative high pressure liquid chromatography purification and mass spectral analysis has allowed preliminary assessment of the structures of the Wy-14,643 microsomal metabolites. It is concluded that one or more of the metabolite fractions of Wy-14,643 transferred in milk exert the biological ability to induce peroxisome proliferation and peroxisomal enzymes in neonatal livers. Images Fig. 2. PLATE 1 PLATE 2 PMID:6683505

  7. SENESCENCE-ASSOCIATED DECLINE IN HEPATIC PEROXISOMAL ENZYME ACTIVITIES CORRESPONDS WITH DIMINISHED LEVELS OF RETINOID X RECEPTOR ALPHA, BUT NOT PEROXISOME PROLIFERATOR-ACTIVATED RECEPTOR ALPHA1

    EPA Science Inventory

    Abstract

    Aging is associated with alterations in hepatic peroxisomal metabolism and susceptibility to hepatocarcinogenecity produced by agonists of peroxisome proliferator-activated receptor alpha (PPARa). Mechanisms involved in these effects are not well understood. Howev...

  8. Bindarit, an Inhibitor of Monocyte Chemotactic Protein Synthesis, Protects against Bone Loss Induced by Chikungunya Virus Infection

    PubMed Central

    Chen, Weiqiang; Foo, Suan-Sin; Taylor, Adam; Lulla, Aleksei; Merits, Andres; Hueston, Linda; Forwood, Mark R.; Walsh, Nicole C.; Sims, Natalie A.; Herrero, Lara J.

    2014-01-01

    ABSTRACT The recent global resurgence of arthritogenic alphaviruses, in particular chikungunya virus (CHIKV), highlights an urgent need for the development of therapeutic intervention strategies. While there has been significant progress in defining the pathophysiology of alphaviral disease, relatively little is known about the mechanisms involved in CHIKV-induced arthritis or potential therapeutic options to treat the severe arthritic symptoms associated with infection. Here, we used microcomputed tomographic (?CT) and histomorphometric analyses to provide previously undescribed evidence of reduced bone volume in the proximal tibial epiphysis of CHIKV-infected mice compared to the results for mock controls. This was associated with a significant increase in the receptor activator of nuclear factor-?B ligand/osteoprotegerin (RANKL/OPG) ratio in infected murine joints and in the serum of CHIKV patients. The expression levels of the monocyte chemoattractant proteins (MCPs), including MCP-1/CCL2, MCP-2/CCL8, and MCP-3/CCL7, were also highly elevated in joints of CHIKV-infected mice, accompanied by increased cellularity within the bone marrow in tibial epiphysis and ankle joints. Both this effect and CHIKV-induced bone loss were significantly reduced by treatment with the MCP inhibitor bindarit. Collectively, these findings demonstrate a unique role for MCPs in promoting CHIKV-induced osteoclastogenesis and bone loss during disease and suggest that inhibition of MCPs with bindarit may be an effective therapy for patients affected with alphavirus-induced bone loss. IMPORTANCE Arthritogenic alphaviruses, including chikungunya virus (CHIKV) and Ross River virus (RRV), cause worldwide outbreaks of polyarthritis, which can persist in patients for months following infection. Previous studies have shown that host proinflammatory soluble factors are associated with CHIKV disease severity. Furthermore, it is established that chemokine (C-C motif) ligand 2 (CCL2/MCP-1) is important in cellular recruitment and inducing bone-resorbing osteoclast (OC) formation. Here, we show that CHIKV replicates in bone and triggers bone loss by increasing the RANKL/OPG ratio. CHIKV infection results in MCP-induced cellular infiltration in the inflamed joints, and bone loss can be ameliorated by treatment with an MCP-inhibiting drug, bindarit. Taken together, our data reveal a previously undescribed role for MCPs in CHIKV-induced bone loss: one of recruiting monocytes/OC precursors to joint sites and thereby favoring a pro-osteoclastic microenvironment. This suggests that bindarit may be an effective treatment for alphavirus-induced bone loss and arthritis in humans. PMID:25339772

  9. Peroxisomal membrane monocarboxylate transporters: evidence for a redox shuttle system?

    E-print Network

    McClelland, Grant B.

    -oxidation was greatly stimulated by the addition of pyruvate and this increase was partially inhibited by the addition; Catalase Liver peroxisomes are cellular organelles important not only for the detoxification process

  10. Genetics Home Reference: Peroxisomal acyl-CoA oxidase deficiency

    MedlinePLUS

    ... body. It is unclear exactly how VLCFA accumulation leads to the specific features of peroxisomal acyl-CoA oxidase deficiency. However, researchers suggest that the abnormal fatty acid accumulation triggers inflammation in the nervous system that ...

  11. Evaluation of Whether Gemfibrozil is a Peroxisome Proliferator in Fish

    EPA Science Inventory

    Gemfibrozil is a pharmaceutical that indirectly modulates cholesterol biosynthesis through effects on peroxisome proliferator-activated receptors (PPAR), which are transcriptional cofactors that regulate expression of genes related to lipid metabolism. An enzyme found in the pero...

  12. What is the role of hydrogen peroxide in plant peroxisomes?

    PubMed

    Corpas, Francisco J

    2015-11-01

    Plant peroxisomes are unusual subcellular compartments with an apparent simple morphology but with complex metabolic activity. The presence of signal molecules, such as hydrogen peroxide (H2 O2 ) and nitric oxide inside plant peroxisomes have added new functions in the cross-talk events among organelles and cells under physiological and stress conditions. Moreover, recent advances in proteomic analyses of plant peroxisomes have identified new protein candidates involved in several novel metabolic pathways. With all these new data, the present concise manuscript will focus on the relevance of the peroxisomal H2 O2 and its two main antioxidant enzymes, catalase and membrane-bound ascorbate peroxidase, which regulate its level and consequently its potential functions. PMID:26242708

  13. [Possibilities for inhibiting tumor-induced angiogenesis: results with multi-target tyrosine kinase inhibitors].

    PubMed

    Török, Szilvia; Döme, Balázs

    2012-03-01

    Functional blood vasculature is essential for tumor progression. The main signalization pathways that play a key role in the survival and growth of tumor vessels originate from the VEGF-, PDGF- and FGF tyrosine kinase receptors. In the past decade, significant results have been published on receptor tyrosine kinase inhibitors (RTKIs). In this paper, the mechanisms of action and the results so far available of experimental and clinical studies on multi-target antiangiogenic TKIs are discussed. On the one hand, notable achievements have been made recently and these drugs are already used in clinical practice in some patient populations. On the other hand, the optimal combination and dosage of these drugs, selection of the apropriate biomarker and better understanding of the conflicting role of PDGFR and FGFR signaling in angiogenesis remain future challenges. PMID:22403757

  14. Ring-truncated deguelin derivatives as potent Hypoxia Inducible Factor-1? (HIF-1?) inhibitors.

    PubMed

    Kim, Ho Shin; Hong, Mannkyu; Lee, Su-Chan; Lee, Ho-Young; Suh, Young-Ger; Oh, Dong-Chan; Seo, Ji Hae; Choi, Hoon; Kim, Jun Yong; Kim, Kyu-Won; Kim, Jeong Hun; Kim, Joohwan; Kim, Young-Myeong; Park, So-Jung; Park, Hyun-Ju; Lee, Jeewoo

    2015-11-01

    A series of fluorophenyl and pyridine analogues of 1 and 2 were synthesized as ring-truncated deguelin surrogates and evaluated for their HIF-1? inhibition. Their structure-activity relationship was systematically investigated based on the variation of the linker B-region moiety. Among the inhibitors, compound 25 exhibited potent HIF-1? inhibition in a dose-dependent manner and significant antitumor activity in H1299 with less toxicity than deguelin. It also inhibited in vitro hypoxia-mediated angiogenic processes in HRMECs. The docking study indicates that 25 occupied the C-terminal ATP-binding pocket of HSP90 in a similar mode as 1, which implies that the anticancer and antiangiogenic activities of 25 are derived from HIF-1? destabilization by binding to the C-terminal ATP-binding site of hHSP90. PMID:26457742

  15. Enhanced susceptibility of cyclin kinase inhibitor p21 knockout mice to high fat diet induced atherosclerosis

    PubMed Central

    Khanna, Ashwani K

    2009-01-01

    Cyclin kinase inhibitor p21 is one of the most potent inhibitors of aortic smooth muscle cell proliferation, a key mediator of atherosclerosis. This study tests if p2l deficiency will result in severe atherosclerosis in a mouse model. p21-/- and strain matched wild type mice were fed with high fat diet for 21 weeks. Analysis for biochemical parameters (cholesterol, triglycerides) in serum and mRNA expression of CD36, HO-1, TGF-?, IFN-?, TNF-?, PPAR-? and NADPH oxidase components (p22phox, NOX-1 and Rac-1) was performed in aortic tissues by Real Time PCR. p21-/- mice gained significantly (p < 0.01) more weight than wild type mice, triglycerides (p < 0.05) and cholesterol levels (p < 0.01) were more pronounced in the sera of p21-/- compared to wild type mice fed with high fat diet. High fat diet resulted in significantly decreased TGF-? (p < 0.02), HO-l (p < 0.02) and increased CD36 (p < 0.03) mRNA expression in aortic tissues of p21-/- mice compared to animal fed with regular diet. IFN-? mRNA expression (235 ± 11 folds) increased significantly in high fat diet fed p21-/- mice and a multifold modulation of PPAR-?(136 ± 7), p22phox, NOX-1 and Rac-1 (15–35-folds) mRNA in aortic tissues from p21-/- mice compared to the wild type mice. Severity of atherosclerotic lesions was significantly higher in p21-/- compared to wild type mice. The results demonstrate that the deficiency of p21 leads to altered expression of pro-atherogenic genes, and severe atherosclerosis in mice fed with high fat diet. This opens the possibility of p21 protein as a therapeutic tool to control progression of atherosclerosis. PMID:19604372

  16. Largazole, a class I histone deacetylase inhibitor, enhances TNF-?-induced ICAM-1 and VCAM-1 expression in rheumatoid arthritis synovial fibroblasts

    SciTech Connect

    Ahmed, Salahuddin; Riegsecker, Sharayah; Beamer, Maria; Rahman, Ayesha; Bellini, Joseph V.; Bhansali, Pravin; Tillekeratne, L.M. Viranga

    2013-07-15

    In the present study, we evaluated the effect of largazole (LAR), a marine-derived class I HDAC inhibitor, on tumor necrosis factor-? (TNF-?)-induced expression of intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1), and matrix metalloproteinase-2 (MMP-2) activity. LAR (1–5 ?M) had no adverse effect on the viability of RA synovial fibroblasts. Among the different class I HDACs screened, LAR (0.5–5 ?M) inhibited the constitutive expression of HDAC1 (0–30%). Surprisingly, LAR increased class II HDAC [HDAC6] by ? 220% with a concomitant decrease in HDAC5 [30–58%] expression in RA synovial fibroblasts. SAHA (5 ?M), a pan-HDAC inhibitor, also induced HDAC6 expression in RA synovial fibroblasts. Pretreatment of RA synovial fibroblasts with LAR further enhanced TNF-?-induced ICAM-1 and VCAM-1 expression. However, LAR inhibited TNF-?-induced MMP-2 activity in RA synovial fibroblasts by 35% when compared to the TNF-?-treated group. Further, the addition of HDAC6 specific inhibitor Tubastatin A with LAR suppressed TNF-? + LAR-induced ICAM-1 and VCAM-1 expression and completely blocked MMP-2 activity, suggesting a role of HDAC6 in LAR-induced ICAM-1 and VCAM-1 expression. LAR also enhanced TNF-?-induced phospho-p38 and phospho-AKT expression, but inhibited the expression of phospho-JNK and nuclear translocation of NF-?Bp65 in RA synovial fibroblasts. These results suggest that LAR activates p38 and Akt pathways and influences class II HDACs, in particular HDAC6, to enhance some of the detrimental effects of TNF-? in RA synovial fibroblasts. Understanding the exact role of different HDAC isoenzymes in RA pathogenesis is extremely important in order to develop highly effective HDAC inhibitors for the treatment of RA. - Highlights: • Largazole enhances TNF-?-induced ICAM-1 and VCAM-1. • Largazole upregulates class II HDAC (HDAC6) in RA synovial fibroblasts. • Largazole also induces the expression of phospho-p38 and phospho-Akt. • A selective HDAC isoform inhibitor may be more effective than a class inhibitor. • Further studies are required to understand the role of class II HDACs in RA.

  17. Ectodomain shedding of TNF receptor 1 induced by protein synthesis inhibitors regulates TNF-{alpha}-mediated activation of NF-{kappa}B and caspase-8

    SciTech Connect

    Ogura, Hirotsugu; Tsukumo, Yoshinori; Sugimoto, Hikaru; Igarashi, Masayuki; Nagai, Kazuo; Kataoka, Takao

    2008-04-01

    The transcription factor nuclear factor {kappa}B (NF-{kappa}B) plays a major role in the inducible resistance to death receptor-mediated apoptosis. It has been established that the protein synthesis inhibitor cycloheximide (CHX) sensitizes many types of cells to tumor necrosis factor (TNF)-{alpha}-induced apoptosis, mainly due to its ability to block de novo synthesis of cellular FLICE-inhibitory protein (c-FLIP). Nevertheless, we have surprisingly found that CHX, as well as its structural analogue acetoxycycloheximide (Ac-CHX), prevents TNF-{alpha}-mediated activation of NF-{kappa}B and caspase-8 in human lung carcinoma A549 cells. Both CHX and Ac-CHX reduced the expression of cell surface TNF receptor 1 (TNF-R1) in a dose-dependent manner, while Ac-CHX was approximately 100-fold more effective than CHX. Consistent with this observation, Ac-CHX induced the proteolytic cleavage of TNF-R1 and its release into the culture medium. CHX and Ac-CHX profoundly decreased constitutive and inducible expression of c-FLIP, whereas these compounds potentiated TNF-{alpha}-induced caspase-8 activation only when metalloprotease inhibitors were present. Thus, our results indicate that ectodomain shedding of TNF-R1 induced by protein synthesis inhibitors regulates TNF-{alpha}-mediated activation of NF-{kappa}B and caspase-8.

  18. Proton pump inhibitor-induced Sweet’s syndrome: report of acute febrile neutrophilic dermatosis in a woman with recurrent breast cancer

    PubMed Central

    Cohen, Philip R.

    2015-01-01

    Background: Sweet’s syndrome, also referred to as acute febrile neutrophilic dermatosis, can either occur as an idiopathic disorder or associated with another condition, including cancer, or induced by exposure to a drug. Proton pump inhibitors selectively inhibit gastric parietal cell H+-K+-adenosine triphosphatase and are most commonly used for the treatment of gastroesophageal reflux disease. Purpose: Proton pump inhibitor-associated Sweet’s syndrome is described in a woman with recurrent breast cancer. Methods: PubMed was used to search the following terms, separately and in combination: acute febrile neutrophilic dermatosis, breast cancer, malignancy, paraneoplastic, proton pump inhibitor, and Sweet’s syndrome. All papers were reviewed and relevant manuscripts, along with their reference citations, were evaluated. Results: Proton pump inhibitors have previously been associated with cutaneous adverse reactions including maculopapular rash, subacute cutaneous lupus erythematosus and toxic epidermal necrolysis. However, drug-induced Sweet’s syndrome has not been observed in patients receiving proton pump inhibitors. The reported woman developed Sweet’s syndrome after initial exposure and subsequent repeat challenge to proton pump inhibitors; subsequent studies also observed recurrence of her breast cancer presenting as metastases to her stomach and bone. Conclusions: Drug-induced Sweet’s syndrome has most commonly been associated with granulocyte colony stimulating factor in oncology patients. Malignancy-associated Sweet’s syndrome has been observed in patients with solid tumors, including breast cancer. Confirmation of proton pump inhibitor-induced Sweet’s syndrome, by repeat challenge with another medication in the same class of drug, was observed in a woman with breast cancer; although the subsequent discovery of recurrent breast cancer presenting as gastric mucosa and vertebral metastases also raises the possibility of concurrent paraneoplastic Sweet’s syndrome, her Sweet’s syndrome symptoms and lesions resolved without recurrence while her recurrent metastatic visceral malignancy persisted. In summary, medication-associated Sweet’s syndrome can occur in oncology patients and proton pump inhibitors should be added to the list of medications associated with the potential to cause drug-induced Sweet’s syndrome. PMID:26114067

  19. Peroxisomal amine oxidase of Hansenula polymorpha does not require its SRL-containing C-terminal sequence for targeting.

    PubMed

    Faber, K N; Haima, P; de Hoop, M J; Harder, W; Veenhuis, M; Ab, G

    1993-04-01

    Amine oxidase (AMO) is a peroxisomal matrix protein of Hansenula polymorpha, which is induced during growth of the yeast in media containing primary amines as a sole nitrogen source. The deduced amino acid sequence of the protein contains an SRL sequence at nine amino acids from the C-terminus. In this study, we have examined the possible role of the SRL motif in sorting of AMO to peroxisomes by mutating the corresponding gene sequence. For this purpose, we have developed a DNA construct that is specifically integrated into the AMO locus of the H. polymorpha genome, placing the mutant gene under the control of the endogenous AMO promoter and eliminating expression of the wild-type gene. Analysis of a stable transformant, containing the desired gene configuration, showed that mutation of the C-terminal sequence neither interfered with correct targeting of the protein into the peroxisome nor displayed significant effects on its activity. From this, it was concluded that the SRL-containing C-terminus is not essential for peroxisomal targeting of AMO in H. polymorpha. PMID:8511963

  20. Peroxisome biogenesis disorders: Biological, clinical and pathophysiological perspectives.

    PubMed

    Braverman, Nancy E; D'Agostino, Maria Daniela; Maclean, Gillian E

    2013-01-01

    The peroxisome biogenesis disorders (PBD) are a heterogeneous group of autosomal recessive disorders in which peroxisome assembly is impaired, leading to multiple peroxisome enzyme deficiencies, complex developmental sequelae and progressive disabilities. Mammalian peroxisome assembly involves the protein products of 16 PEX genes; defects in 14 of these have been shown to cause PBD. Three broad phenotypic groups are described on a spectrum of severity: Zellweger syndrome is the most severe, neonatal adrenoleukodystrophy is intermediate and infantile Refsum disease is less severe. Another group is Rhizomelic chondrodysplasia punctata spectrum. Recently, atypical phenotypes have been described, indicating that the full spectrum of these disorders remains to be identified. For most patients, there is a correlation between clinical severity and effect of the mutation on PEX protein function. Diagnosis relies on biochemical measurements of peroxisome functions and PEX gene sequencing. There are no targeted therapies, although management protocols have been suggested and research endeavors continue. In this review we will discuss peroxisome biology and PBD, and research contributions to pathophysiology and treatment. PMID:23798008

  1. Gamma Secretase Inhibitors enhance Vincristine-induced apoptosis in T-ALL in a NOTCH-independent manner

    PubMed Central

    Yoon, Sun-Ok; Zapata, Mariana C.; Singh, Akannsha; Jo, Wol Soon; Spencer, Nakia; Choi, Yong Sung

    2014-01-01

    Background Activating mutations in the NOTCH1 gene are found in over 50% of T-ALL cases. Since Notch signaling contributes to the leukemia cell survival and growth, targeting Notch signaling using ?-secretase inhibitors (GSI) has been proposed as a molecularly targeted therapy for the treatment of T-ALL. However, not all T-ALL with NOTCH1 activating mutations respond to GSI treatment. Methods We examined whether GSI could enhance the cytotoxic effect of anti-leukemic agents in the GSI-resistant T-ALL cells although GSI does not have anti-tumor effect as a single agent. The cytotoxic effect of the combination treatment was evaluated by measuring viable cells. To determine if Notch signaling is involved in the synergism between GSI and Vincristine (VCR), loss- and gain-of-function assays were performed. To further dissect the synergistic GSI effect in combination with VCR, cell cycle progression was analyzed and apoptosis was measured by various methods. Results We discovered that GSI synergized with VCR in both GSI-resistant and GSI-sensitive T-ALL in a Notch-independent manner. GSI augmented VCR-induced mitotic arrest, followed by apoptosis. GSI accelerated VCR-triggered loss of mitochondrial membrane potential and caspase-mediated apoptosis. Conclusion GSI promoted VCR-induced apoptosis in T-ALL. Incorporating GSI into VCR- containing therapeutic regimen may be beneficial in treating T-ALL. PMID:25156146

  2. The Pan-Aurora Kinase Inhibitor, PHA-739358, Induces Apoptosis and Inhibits Migration in Melanoma Cell Lines

    PubMed Central

    Xie, Lifang; Meyskens, Frank L

    2014-01-01

    Treatment of metastatic melanoma has long been a challenge due to its resistance to traditional chemotherapeutics leading to the search for alternative strategies. Aurora kinases are key mitotic regulators that are frequently overexpressed in various cancers including melanoma, making them ideal targets for anticancer therapeutics. Several Aurora kinase inhibitors have been developed and tested pre-clinically and clinically. PHA-739358 is currently the most advanced clinical compound; however its antitumor effect has not been tested in melanoma. In this study, the anti-proliferative and anti-invasive effects of PHA-739358 were investigated in melanoma cell lines. The results demonstrated that PHA-739358 produces a time and dose dependent inhibition of cell proliferation, induction of apoptosis, and inhibition of cell migration. Downregulation of MMP-2 via inhibition of NF?B signaling pathway may contribute to PHA-739358-induced migration inhibition. Furthermore, PHA-739358 enhanced temozolomide-induced caspase activation. This study provides a promising new strategy for the treatment of advanced melanoma. PMID:23344158

  3. A comparative study of neuroprotective effect of angiotensin converting enzyme inhibitors against scopolamine-induced memory impairments in rats

    PubMed Central

    Jawaid, Talha; Jahan, Shah; Kamal, Mehnaz

    2015-01-01

    The comparative study of neuroprotective effect of angiotensin converting enzyme inhibitors against scopolamine-induced neuroinflammation in albino Wistar rats was studied. Male albino rats were administered with scopolamine to induce memory impairment. The standard nootropic agent, piracetam (200 mg/kg b.w., [i.p.]), perindopril (0.1 mg/kg b.w., [i.p.]), enalapril (0.1 mg/kg b.w., [i.p.]), and ramipril (0.1 mg/kg b.w., [i.p.]) were administered in different group of animals for 5 days. On 5th day, scopolamine (1 mg/kg b.w., i.p.) was administered after 60 min of the last dose of test drug. Memory function was evaluated in Morris water maze (MWM) test and pole climbing test (PCT). Biochemical estimations like glutathione (GSH), malondialdehyde (MDA), and acetylcholinesterase activity in the brain were estimated after completion of behavior study. All three test groups shows improvement in learning and memory in comparison to control group. Perindopril treated group showed a more effective significant decrease in escape latency time and transfer latency time compared to enalapril and ramipril treated group on day 4 in MWM test and PCT, respectively. Perindopril shows a significant reduction in MDA level and acetylcholinesterase activity and a significant rise in GSH level compared to enalapril and ramipril. The finding of this study indicates that Perindopril is more effective in memory retention compared to enalapril and ramipril. PMID:26317078

  4. Tyrosine kinase inhibitor, methyl 2,5-dihydromethylcinnimate, induces PML nuclear body formation and apoptosis in tumor cells

    SciTech Connect

    Komura, Naoyuki; Asakawa, Mayako; Umezawa, Kazuo . E-mail: umezawa@applc.keio.ac.jp; Segawa, Kaoru

    2007-08-01

    Promyelocytic leukemia (PML) nuclear bodies (PML-NBs) are the nuclear structure consisting of various proteins such as PML, SUMO-1, and p53. PML-NBs are implicated in the regulation of tumor suppression, antiviral responses, and apoptosis. In this study, we searched for bioactive metabolites that would promote the formation of PML-NBs in tumor cells. As a result, methyl 2,5-dihydromethylcinnimate (2,5-MeC), a tyrosine kinase inhibitor, enhanced expression and/or stability of PML proteins and induced PML-NB formation in p53 null H1299 cells established from non-small cell lung cancer (NSCLC) and wild-type p53-expressing U2OS cells derived from osteosarcoma. Furthermore, it enhanced apoptosis by exogenously expressed wild type p53 and the expression of p53-responsive genes, such as PUMA and p21, in H1299 cells. 2,5-MeC also activated endogenous p53 and induced apoptosis in U2OS cells. The results suggest that 2,5-MeC is likely to be a promising candidate drug for the clinical treatment of terminal cancer-expressing wild-type p53.

  5. Naringin Reverses Hepatocyte Apoptosis and Oxidative Stress Associated with HIV-1 Nucleotide Reverse Transcriptase Inhibitors-Induced Metabolic Complications

    PubMed Central

    Adebiyi, Oluwafeyisetan O.; Adebiyi, Olubunmi A.; Owira, Peter M. O.

    2015-01-01

    Nucleoside Reverse Transcriptase Inhibitors (NRTIs) have not only improved therapeutic outcomes in the treatment of HIV infection but have also led to an increase in associated metabolic complications of NRTIs. Naringin’s effects in mitigating NRTI-induced complications were investigated in this study. Wistar rats, randomly allotted into seven groups (n = 7) were orally treated daily for 56 days with 100 mg/kg zidovudine (AZT) (groups I, II III), 50 mg/kg stavudine (d4T) (groups IV, V, VI) and 3 mL/kg of distilled water (group VII). Additionally, rats in groups II and V were similarly treated with 50 mg/kg naringin, while groups III and VI were treated with 45 mg/kg vitamin E. AZT or d4T treatment significantly reduced body weight and plasma high density lipoprotein concentrations but increased liver weights, plasma triglycerides and total cholesterol compared to controls, respectively. Furthermore, AZT or d4T treatment significantly increased oxidative stress, adiposity index and expression of Bax protein, but reduced Bcl-2 protein expression compared to controls, respectively. However, either naringin or vitamin E significantly mitigated AZT- or d4T-induced weight loss, dyslipidemia, oxidative stress and hepatocyte apoptosis compared to AZT- or d4T-only treated rats. Our results suggest that naringin reverses metabolic complications associated with NRTIs by ameliorating oxidative stress and apoptosis. This implies that naringin supplements could mitigate lipodystrophy and dyslipidemia associated with NRTI therapy. PMID:26690471

  6. Chloroplastic oxidative burst induced by tenuazonic acid, a natural photosynthesis inhibitor, triggers cell necrosis in Eupatorium adenophorum Spreng.

    PubMed

    Chen, Shiguo; Yin, Chunyan; Qiang, Sheng; Zhou, Fenyan; Dai, Xinbin

    2010-03-01

    Tenuazonic acid (TeA), a nonhost-specific phytotoxin produced by Alternaria alternata, was determined to be a novel natural photosynthesis inhibitor owning several action sites in chloroplasts. To further elucidate the mode of its action, studies were conducted to assess the production and involvement of reactive oxygen species (ROS) in the toxic activity of TeA. A series of experiments indicated that TeA treatment can induce chloroplast-derived ROS generation including not only (1)O(2) but also superoxide radical, H(2)O(2) and hydroxyl radicals in Eupatorium adenophorum mesophyll cells, resulting from electron leakage and charge recombination in PSII as well as thylakoid overenergization due to inhibition of the PSII electron transport beyond Q(A) and the reduction of end acceptors on the PSI acceptor side and chloroplast ATPase activity. The initial production of TeA-induced ROS was restricted to chloroplasts and accompanied with a certain degree of chloroplast damage. Subsequently, abundant ROS were quickly dispersed throughout whole cell and cellular compartments, causing a series of irreversible cellular harm such as chlorophyll breakdown, lipid peroxidation, plasma membrane rupture, chromatin condensation, DNA cleavage, and organelle disintegration, and finally resulting in rapid cell destruction and leaf necrosis. These results show that TeA causing cell necrosis of host-plants is a result of direct oxidative damage from chloroplast-mediated ROS eruption. PMID:20026008

  7. Mechanisms and biomarkers of cardiovascular injury induced by phosphodiesterase inhibitor III SK&F 95654 in the spontaneously hypertensive rat.

    PubMed

    Zhang, Jun; Herman, Eugene H; Robertson, Donald G; Reily, Michael D; Knapton, Alan; Ratajczak, Helen V; Rifai, Nader; Honchel, Ronald; Blanchard, Kerry T; Stoll, Raymond E; Sistare, Frank D

    2006-01-01

    The cardiovascular injury of the type III selective PDE inhibitor SK&F 95654 was investigated in SHR. Twenty-four hours after a single sc injection of 100 or 200 mg/kg of the drug, rats exhibited cardiomyocyte necrosis and apoptosis, interstitial inflammation, hemorrhage and edema, as well as mesenteric arterial hemorrhage and necrosis, periarteritis, EC and VSMC apoptosis, EC activation, and MC activation and degranulation. Elevated serum levels of cTnT and decreased cTnT immunoperoxidase staining on cardiomyocytes were detected in the drug-treated rats. Serum levels of alpha2-macroglobulin and IL-6 were significantly elevated following drug treatment. NMR spectral patterns of urine samples are significantly different between the drug-treated and control rats. These results indicate that measurement of serum cTnT, acute phase proteins, and cytokines as well as metabonomic urine profiles may serve as potential biomarkers for drug-induced cardiovascular injury in rats. Increased expression of CD63 on MC (tissue biomarker of MC), of nitrotyrosine on MC and EC (an indirect indicator of NO in vivo), and of iNOS on MC and EC (source of NO) suggest that NO produced by activated and degranulated MC as well as activated EC play an important role in SK&F 95654-induced mesenteric vascular injury. PMID:16537294

  8. The DNA methylation inhibitor induces telomere dysfunction and apoptosis of leukemia cells that is attenuated by telomerase over-expression

    PubMed Central

    de Jonge, Nick; Björkholm, Magnus; Xu, Dawei

    2015-01-01

    DNA methyltransferase inhibitors (DNMTIs) such as 5-azacytidine (5-AZA) have been used for treatment of acute myeloid leukemia (AML) and other malignancies. Although inhibiting global/gene-specific DNA methylation is widely accepted as a key mechanism behind DNMTI anti-tumor activity, other mechanisms are likely involved in DNMTI's action. Because telomerase reverse transcriptase (TERT) plays key roles in cancer through telomere elongation and telomere lengthening-independent activities, and TERT has been shown to confer chemo- or radio-resistance to cancer cells, we determine whether DNMTIs affect telomere function and whether TERT/telomerase interferes with their anti-cancer efficacy. We showed that 5-AZA induced DNA damage and telomere dysfunction in AML cell lines by demonstrating the presence of 53-BP1 foci and the co-localization of 53-BP1 foci with telomere signals, respectively. Telomere dysfunction was coupled with diminished TERT expression, shorter telomere and apoptosis in 5-AZA-treated cells. However, 5-AZA treatment did not lead to changes in the methylation status of subtelomere regions. Down-regulation of TERT expression similarly occurred in primary leukemic cells derived from AML patients exposed to 5-AZA. TERT over-expression significantly attenuated 5-AZA-mediated DNA damage, telomere dysfunction and apoptosis of AML cells. Collectively, 5-AZA mediates the down-regulation of TERT expression, and induces telomere dysfunction, which consequently exerts an anti-tumor activity. PMID:25682873

  9. Fasudil, an inhibitor of Rho-associated coiled-coil kinase, attenuates hyperoxia-induced pulmonary fibrosis in neonatal rats

    PubMed Central

    Qi, Xiu-Jie; Ning, Wei; Xu, Feng; Dang, Hong-Xing; Fang, Fang; Li, Jing

    2015-01-01

    Background: Oxygen therapy is important during the management of high-risk neonatal infants, such as those with preterm birth, low birth weight, and asphyxia. However, prolonged exposure to high oxygen concentrations can readily lead to diffuse nonspecific inflammation, which promotes airway remodeling and pulmonary fibrosis. The Rho/Rho-associated coiled-coil kinase (Rho/ROCK) signaling pathway plays an important role in numerous developmental and proliferative diseases. This study was performed to determine the efficacy of ROCK inhibitor fasudil in blocking the development of hyperoxia-induced lung injury and fibrosis in neonatal rats. Methods: Neonatal rats were randomly divided into four groups: air + saline group, air + fasudil group, hyperoxia + saline group, and hyperoxia + fasudil group. The hyperoxia + saline and Hyp + fasudil groups were exposed to 95% oxygen for 21 days and administered intraperitoneal saline or fasudil once daily. The air + saline and air + fasudil group were exposed to 21% oxygen (room air) and administered the same volume of intraperitoneal saline or fasudil. Results: Fasudil-treated rats exhibited improved histopathological changes and decreased lung hydroxyproline content. Fasudil attenuated the protein level of alpha-smooth muscle actin, transforming growth factor-?1, and connective tissue growth factor. Additionally, fasudil reduced the activation of ROCK1 and myosin phosphatase targeting subunit 1 protein in the Rho/ROCK signaling pathway. Conclusions: Fasudil may be a potentially effective therapeutic drug for hyperoxia-induced pulmonary fibrosis. PMID:26722398

  10. Comparing the Efficacy of Bupropion and Amantadine on Sexual Dysfunction Induced by a Selective Serotonin Reuptake Inhibitor

    PubMed Central

    Zahiroddin, Alireza; Faridhosseini, Farhad; Zamani, Azar; Shahini, Najmeh

    2015-01-01

    Background: Antidepressant-induced sexual dysfunction (SD) is a common problem, associated with a significant risk of non-adherence. Selective Serotonin Reuptake Inhibitors (SSRIs) are associated with a substantial risk of SD. Only 10 % of patients show spontaneous improvement during follow up period. Objectives: This study aimed to compare two proposed medication (bupropion vs. amantadine) in alleviating SD in patients treated with SSRIs. Patients and Methods: In a randomized, single-blinded, clinical trial in Iran, 46 patients were recruited based on DSM-IV-TR criteria and semi-structured interview. Then, they were randomized into two treatment groups using table of random numbers. Eight patients were excluded and finally 38 patients completed the study which lasted for 4 weeks. Twenty patients were given bupropion, 18 patients were randomly assigned to another group, and given amantadine. Patients were assessed with the Arizona sexual experience scale (ASEX) at baseline and 4 weeks after the treatment. Results: A total of 38 patients completed the study (18 patients in amantadine vs. 20 patients in bupropion).The mean ASEX scores gradually declined in both study groups during the trial. The reduction of ASEX score in bupropion group was more than that of amantadine group that was statistically significant. So, the addition of bupropion at higher doses appears to be more effective approach in comparison with amantadine. Conclusions: These results provide empirical support for conducting a further study on comparing different add-on strategies for treating drug-induced SD.

  11. A comparative study of neuroprotective effect of angiotensin converting enzyme inhibitors against scopolamine-induced memory impairments in rats.

    PubMed

    Jawaid, Talha; Jahan, Shah; Kamal, Mehnaz

    2015-01-01

    The comparative study of neuroprotective effect of angiotensin converting enzyme inhibitors against scopolamine-induced neuroinflammation in albino Wistar rats was studied. Male albino rats were administered with scopolamine to induce memory impairment. The standard nootropic agent, piracetam (200 mg/kg b.w., [i.p.]), perindopril (0.1 mg/kg b.w., [i.p.]), enalapril (0.1 mg/kg b.w., [i.p.]), and ramipril (0.1 mg/kg b.w., [i.p.]) were administered in different group of animals for 5 days. On 5(th) day, scopolamine (1 mg/kg b.w., i.p.) was administered after 60 min of the last dose of test drug. Memory function was evaluated in Morris water maze (MWM) test and pole climbing test (PCT). Biochemical estimations like glutathione (GSH), malondialdehyde (MDA), and acetylcholinesterase activity in the brain were estimated after completion of behavior study. All three test groups shows improvement in learning and memory in comparison to control group. Perindopril treated group showed a more effective significant decrease in escape latency time and transfer latency time compared to enalapril and ramipril treated group on day 4 in MWM test and PCT, respectively. Perindopril shows a significant reduction in MDA level and acetylcholinesterase activity and a significant rise in GSH level compared to enalapril and ramipril. The finding of this study indicates that Perindopril is more effective in memory retention compared to enalapril and ramipril. PMID:26317078

  12. In vivo anti-myeloma activity and modulation of gene expression profile induced by valproic acid, a histone deacetylase inhibitor.

    PubMed

    Neri, Paola; Tagliaferri, Pierosandro; Di Martino, Maria Teresa; Calimeri, Teresa; Amodio, Nicola; Bulotta, Alessandra; Ventura, Monica; Eramo, Pasqua Orietta; Viscomi, Caterina; Arbitrio, Mariamena; Rossi, Marco; Caraglia, Michele; Munshi, Nikhil C; Anderson, Kenneth C; Tassone, Pierfrancesco

    2008-11-01

    Valproic acid (VPA) is a well-tolerated anticonvulsant that exerts anti-tumour activity as a histone deacetylase inhibitor. This study investigated the in vitro and in vivo activity of VPA against multiple myeloma (MM) cells. In vitro exposure of interleukin-6-dependent or -independent MM cells to VPA inhibited cell proliferation in a time- and dose-dependent manner and induced apoptosis. In a cohort of severe combined immunodeficiency mice bearing human MM xenografts, VPA induced tumour growth inhibition and survival advantage in treated animals versus controls. Flow cytometric analysis performed on MM cells from excised tumours showed increase of G(0)-G(1) and a decreased G(2)/M- and S-phase following VPA treatment, indicating in vivo effects of VPA on cell cycle regulation. Gene expression profiling of MM cells exposed to VPA showed downregulation of genes involved in cell cycle progression, DNA replication and transcription, as well as upregulation of genes implicated in apoptosis and chemokine pathways. Pathfinder analysis of gene array data identified cell growth, cell cycle, cell death, as well as DNA replication and repair as the most important signalling networks modulated by VPA. Taken together, our data provide the preclinical rationale for VPA clinical evaluation as a single agent or in combination, to improve patient outcome in MM. PMID:18986388

  13. Yeast pex1 cells contain peroxisomal ghosts that import matrix proteins upon reintroduction of Pex1.

    PubMed

    Knoops, Kèvin; de Boer, Rinse; Kram, Anita; van der Klei, Ida J

    2015-12-01

    Pex1 and Pex6 are two AAA-ATPases that play a crucial role in peroxisome biogenesis. We have characterized the ultrastructure of the Saccharomyces cerevisiae peroxisome-deficient mutants pex1 and pex6 by various high-resolution electron microscopy techniques. We observed that the cells contained peroxisomal membrane remnants, which in ultrathin cross sections generally appeared as double membrane rings. Electron tomography revealed that these structures consisted of one continuous membrane, representing an empty, flattened vesicle, which folds into a cup shape. Immunocytochemistry revealed that these structures lack peroxisomal matrix proteins but are the sole sites of the major peroxisomal membrane proteins Pex2, Pex10, Pex11, Pex13, and Pex14. Upon reintroduction of Pex1 in Pex1-deficient cells, these peroxisomal membrane remnants (ghosts) rapidly incorporated peroxisomal matrix proteins and developed into peroxisomes. Our data support earlier views that Pex1 and Pex6 play a role in peroxisomal matrix protein import. PMID:26644511

  14. Interruption of the Ras/MEK/ERK signaling cascade enhances Chk1 inhibitor–induced DNA damage in vitro and in vivo in human multiple myeloma cells

    PubMed Central

    Dai, Yun; Chen, Shuang; Pei, Xin-Yan; Almenara, Jorge A.; Kramer, Lora B.; Venditti, Charis A.; Dent, Paul

    2008-01-01

    The role of the Ras/MEK/ERK pathway was examined in relation to DNA damage in human multiple myeloma (MM) cells exposed to Chk1 inhibitors in vitro and in vivo. Exposure of various MM cells to marginally toxic concentrations of the Chk1 inhibitors UCN-01 or Chk1i modestly induced DNA damage, accompanied by Ras and ERK1/2 activation. Interruption of these events by pharmacologic (eg, the farnesyltransferase inhibitor R115777 or the MEK1/2 inhibitor PD184352) or genetic (eg, transfection with dominant-negative Ras or MEK1 shRNA) means induced pronounced DNA damage, reflected by increased ?H2A.X expression/foci formation and by comet assay. Increased DNA damage preceded extensive apoptosis. Notably, similar phenomena were observed in primary CD138+ MM cells. Enforced MEK1/2 activation by B-Raf transfection prevented R115777 but not PD184352 from inactivating ERK1/2 and promoting Chk1 inhibitor–induced ?H2A.X expression. Finally, coadministration of R115777 diminished UCN-01–mediated ERK1/2 activation and markedly potentiated ?H2A.X expression in a MM xenograft model, associated with a striking increase in tumor cell apoptosis and growth suppression. Such findings suggest that Ras/MEK/ERK activation opposes whereas its inhibition dramatically promotes Chk1 antagonist–mediated DNA damage. Together, these findings identify a novel mechanism by which agents targeting the Ras/MEK/ERK pathway potentiate Chk1 inhibitor lethality in MM. PMID:18614762

  15. Proteasome inhibitor MG-132 enhances histone deacetylase inhibitor SAHA-induced cell death of chronic myeloid leukemia cells by an ROS-mediated mechanism and downregulation of the Bcr-Abl fusion protein

    PubMed Central

    ZHOU, WENJING; ZHU, WEIWEI; MA, LIYA; XIAO, FENG; QIAN, WENBIN

    2015-01-01

    Recently, there has been progress in the treatment of chronic myeloid leukemia (CML). However, novel therapeutic strategies are required in order to address the emerging problem of imatinib resistance. Histone deacetylase inhibitors (HDACi) and proteasome inhibitors are promising alternatives, and may be amenable to integration with current therapeutic approaches. However, the mechanisms underlying the interaction between these two agents remain unclear. The present study assessed the cytotoxic effect of the HDACi, suberoylanilide hydroxamic acid (SAHA), in combination with the proteasome inhibitor, MG-132, in imatinib-sensitive K562 and imatinib-resistant K562G cells, and investigated the mechanism underlying this effect. Cell viability was measured using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide method and protein expression levels were determined by western blotting. Reactive oxygen species (ROS) generation levels were observed under a fluorescence microscope The results indicated that SAHA and MG-132 act in a synergistic manner to induce cell death in K562 and K562G cells. This effect was associated with Bcr-Abl downregulation and the production of ROS. Notably, the ROS scavenger, N-acetyl-L-cysteine, almost fully reversed the cell death and Bcr-Abl downregulation that was induced by the combination of SAHA and MG-132. By contrast, the pan-caspase inhibitor, z-VAD-fmk, only partially reversed the cell death induced by these two drugs in CML cells. These results indicated that increased intracellular ROS levels are important in the induction of cell death and the downregulation of Bcr-Abl. In conclusion, the present results suggested that combined SAHA and MG-132 may be a promising treatment for CML. PMID:26722260

  16. RhoA Regulates Peroxisome Association to Microtubules and the Actin Cytoskeleton

    PubMed Central

    Lay, Dorothee; Wiese, Sebastian; Meyer, Helmut E.; Warscheid, Bettina; Saffrich, Rainer; Peränen, Johan; Gorgas, Karin; Just, Wilhelm W.

    2010-01-01

    The current view of peroxisome inheritance provides for the formation of new peroxisomes by both budding from the endoplasmic reticulum and autonomous division. Here we investigate peroxisome-cytoskeleton interactions and show by proteomics, biochemical and immunofluorescence analyses that actin, non-muscle myosin IIA (NMM IIA), RhoA, Rho kinase II (ROCKII) and Rab8 associate with peroxisomes. Our data provide evidence that (i) RhoA in its inactive state, maintained for example by C. botulinum toxin exoenzyme C3, dissociates from peroxisomes enabling microtubule-based peroxisomal movements and (ii) dominant-active RhoA targets to peroxisomes, uncouples the organelles from microtubules and favors Rho kinase recruitment to peroxisomes. We suggest that ROCKII activates NMM IIA mediating local peroxisomal constrictions. Although our understanding of peroxisome-cytoskeleton interactions is still incomplete, a picture is emerging demonstrating alternate RhoA-dependent association of peroxisomes to the microtubular and actin cytoskeleton. Whereas association of peroxisomes to microtubules clearly serves bidirectional, long-range saltatory movements, peroxisome-acto-myosin interactions may support biogenetic functions balancing peroxisome size, shape, number, and clustering. PMID:21079737

  17. An Organometallic Protein Kinase Inhibitor Pharmacologically Activates p53 and Induces Apoptosis in

    E-print Network

    Meggers, Eric

    intervention. The majority of cancers harbor point mutations in p53, most of which are in the central region/2 compound was barely able to induce cell death in melanoma cells with p53 mutations, further confirming of the protein responsible for DNA binding (5). Unlike other solid tumors, melanomas typically lack p53 mutations

  18. TOPICAL CORTICOSTEROIDS BUT NOT CALCINEURIN INHIBITORS INDUCED ATROPHY AFTER FOUR WEEKS.

    PubMed

    Jiráková, A; Rob, F; Se?níková, Z; Koblová, K; Džambová, M; Rajská, L; Göpfertová, D; Pellacani, G; Malý, M; Lotti, T; Hercogová, J

    2015-01-01

    Reflectance confocal microscopy (RCM) is a non-invasive, in vivo technique for real-time imaging of the epidermis and superficial dermis at the cellular resolution. We performed a pilot study focusing on the evaluation of the effect of topical corticosteroids and calcineurin inhibitors on the epidermis of patients with atopic dermatitis (AD). The effect was assessed by RCM. A total of 45 patients with AD took part in the study. Patients were selected according to the standardized protocol and divided into two groups. Twenty-three patients used methylprednisolone aceponat topically on the skin with lesions of AD once a day for three months (group A). Twenty-one patients applied topical tacrolimus on the skin with lesions of AD twice a day for three months (B). RCM imaging was performed on the day of intiating the study (T0), then after one (T1), two (T2) and three months (T3). In group A, there was a visible decrease of the stratum corneum and the epidermis thickness which was statistically significant. In comparison, in group B, such changes were not noted and the differences between the groups in time course were statistically significant. In group A, an increase in the percentage of blurred keratinocytes in the stratum spinosum was also recorded, especially between the first (T0) and the second visit (T1). RCM is a useful method for evaluating the changes in epidermis due to the different topical treatment in patients with AD. PMID:26403410

  19. Retinoic acid induces CDK inhibitors and growth arrest specific (Gas) genes in neural crest cells.

    PubMed

    Wang, Linping; Mear, John P; Kuan, Chia-Yi; Colbert, Melissa C

    2005-04-01

    Retinoic acid (RA), the active metabolite of vitamin A, regulates cellular growth and differentiation during embryonic development. In excess, this vitamin is also highly teratogenic to animals and humans. The neural crest is particularly sensitive to RA, and high levels adversely affect migration, proliferation and cell death. We investigated potential gene targets of RA associated with neural crest proliferation by determining RA-mediated changes in gene expression over time, using microarrays. Statistical analysis of the top ranked RA-regulated genes identified modest changes in multiple genes previously associated with cell cycle control and proliferation including the cyclin-dependent kinase inhibitors Cdkn1a (p21), Cdkn2b (p15(INK4b)), and Gas3/PMP22. The expression of p21 and p15(INK4b) contribute to decreased proliferation by blocking cell cycle progression at G1-S. This checkpoint is pivotal to decisions regulating proliferation, apoptosis, or differentiation. We have also confirmed the overexpression of Gas3/PMP22 in RA-treated neural crests, which is associated with cytoskeletal changes and increased apoptosis. Our results suggest that increases in multiple components of diverse regulatory pathways have an overall cumulative effect on cellular decisions. This heterogeneity contributes to the pleiotropic effects of RA, specifically those affecting proliferation and cell death. PMID:15839997

  20. Effect of c-Met Inhibitor on HGF-induced Ovarian Carcinoma Cell Migration

    NASA Astrophysics Data System (ADS)

    Lo, Chun-Min; Lo, Jun-Chih; Yip, Kay-Pong

    2010-03-01

    The dysregulation of hepatocyte growth factor (HGF) and its receptor, c-Met, in cell migration contributes to tumor invasion and metastasis in numerous cancers including ovarian cancer. Specific inhibitors against HGF/c-Met signaling like SU11274, therefore, may have important therapeutic potential for the treatment of cancers. Here, we applied electric cell-substrate impedance sensing (ECIS) and traction force microscopy to evaluate the effect of SU11274 on HGF-treated SKOV-3 ovarian cancer cells. Our results showed that, compared with control cells, HGF-treated cell monolayer displayed lower junctional resistance between cells, larger cell-substrate separation, and higher cell micromotion. In addition, individual HGF-treated SKOV-3 cells demonstrated weaker traction forces on the collagen-coated polyacrylamide substrate than did control cells. These changes lead to faster directional movement of HGF-treated cells, as demonstrated with wound healing assay. Treatment of SKOV-3 cells with SU11274 indicated significant inhibition of HGF stimulation on all assays tested.

  1. BRAF Inhibitor-Induced Antitumoral Granulomatous Dermatitis Eruption in Advanced Melanoma.

    PubMed

    Garrido, Maria C; Gutierrez, Carlota; Riveiro-Falkenbach, Erica; Ortiz, Pablo; Rodriguez-Peralto, Jose L

    2015-10-01

    Recent advances in targeting BRAF mutations, which occur in roughly 50% of the melanomas, have improved response rates and overall survival in patients with advanced disease. With the increasingly extensive use of the drug, new, nonpreventable, cutaneous and noncutaneous toxicities keep arising as infrequent adverse effects. We report a 55-year-old man with a history of metastatic melanoma treated with the dabrafenib who presented, 10 months after the initiation of the treatment, with erythematous, slightly squamous, round plaques on his upper trunk and on his left upper arm. Two skin biopsies from the lesions revealed a granulomatous dermatitis in the superficial reticular dermis. One of them showed admixed abundant melanophages from tumoral melanosis. No melanoma cells were seen in any of the specimens. No interruption of the treatment was necessary. Our observation indicates that such a response may represent a positive immune activation triggered by BRAF inhibitors. The erythematous rash was initially concerning for progression of metastatic disease, which suggests that a close monitoring of the patients with advanced melanomas treated with vemurafenib is advisable to prevent unnecessary discontinuation of the therapy. PMID:26381028

  2. An EGF receptor inhibitor induces RAR-{beta} expression in breast and ovarian cancer cells

    SciTech Connect

    Grunt, Thomas W. . E-mail: thomas.grunt@meduniwien.ac.at; Puckmair, Klaudia; Tomek, Katharina; Kainz, Birgit; Gaiger, Alexander

    2005-04-22

    Inhibition of the epidermal growth factor (EGF)-receptor (EGFR) has become a promising anticancer treatment strategy. In addition, application of retinoids yields encouraging results for cancer prevention and therapy. Many tumors express no or low amounts of retinoic acid receptor-{beta}2 (RAR-{beta}2) due to epigenetic silencing via DNA hypermethylation. RAR-{beta}2 is the main mediator of the antiproliferative effect of retinoids. RAR-{beta}2 re-expression causes reversal of transformation, cell cycle arrest, and restoration of retinoid sensitivity. RAR-{beta}2 is thus a tumor suppressor. Western blotting, colorimetric in vitro cell proliferation assays, and reverse transcription-polymerase chain reaction demonstrated that the EGFR inhibitor PD153035 not only blocked activation of EGFR and inhibited cell growth, but also stimulated RAR-{beta} expression in MDA-MB-468 breast and OVCAR-3 ovarian carcinoma cells. Upregulation of RAR-{beta} by PD153035 was confirmed by real-time reverse transcription-polymerase chain reaction. In contrast, expression of other retinoid receptors and of estrogen receptor-{alpha} was not affected. PD153035-mediated re-induction of RAR-{beta} was associated with demethylation of the RAR-{beta}2 gene promoter P2 as demonstrated by methylation-specific polymerase chain reaction. These novel results on the ErbB/retinoid receptor cross-talk may be useful for designing future anticancer combination regimens.

  3. Lisofylline, an inhibitor of unsaturated phosphatidic acid generation, ameliorates interleukin-1 beta-induced dysfunction in cultured rat islets.

    PubMed

    Bleich, D; Chen, S; Bursten, S L; Nadler, J L

    1996-11-01

    Interleukin-1 beta (IL-1 beta) causes rat islet cell dysfunction through mechanisms that involve inducible nitric oxide synthase (iNOS). However, IL-1 beta also activates several lipid pathways, including those generating phosphatidic acid (PA). Lisofylline (LSF), a water-soluble, nontoxic, selective inhibitor of the PA-1 alpha subspecies, which is stimulated by IL-1 beta and tumor necrosis factor-alpha, has been shown to prevent cytokine-induced cytotoxicity in in vivo animal models. To evaluate the effect of LSF on acute IL-1 beta-induced islet dysfunction, rat islets were exposed to IL-1 beta (0.1 ng/ml) with or without LSF (100 microM) for 24 h, followed by 25 mM glucose (G) stimulation, measurement of rat insulin by RIA, and calculation of the insulin secretion rate. In other experiments, rat islets were precultured for 48 h, then treated for 48 h in 25 mM G with or without IL-1 beta (0.1 ng/ml) and LSF (400 microM), and aliquots of medium were removed at 0, 24, and 48 h for measurement of rat insulin. In addition, islets were exposed to 25 mM G with or without IL-1 beta and LSF, lipids were then extracted, and PA subspecies were identified by TLC and mass spectroscopy, and quantitated using normal phase HPLC. Islets were also exposed to IL-1 beta with or without LSF, and Western immunoblots were performed to evaluate the effect of LSF on iNOS protein expression. IL-1 beta caused a 44% decrease in islet G-stimulated insulin secretion compared to that in untreated islets (P < 0.0005), which was totally reversed by LSF. In addition, IL-1 beta decreased the G-stimulated medium insulin content by 75% at 24 h (P = 0.0004) and 86% at 48 h compared to that in control islets (P < 0.0001). LSF-treated islets maintained 70% of medium insulin content at 24 h (P = 0.11) and 50% at 48 h (P < 0.0001) compared to control islets. HPLC quantitation of PA-1 alpha extracted from islets treated with IL-1 beta alone showed an approximately 15-fold increase over the PA-1 alpha content of islets treated with IL-1 beta and LSF. IL-1 beta-induced expression of iNOS was unchanged with the addition of LSF. These results suggest that LSF is effective in reducing IL-1 beta-induced islet dysfunction, thus supporting the role of lipid mediators such as PA in cytokine-induced islet toxicity. PMID:8895359

  4. Non-Catalytic Site HIV-1 Integrase Inhibitors Disrupt Core Maturation and Induce a Reverse Transcription Block in Target Cells

    PubMed Central

    Tsai, Luong; O’Sullivan, Christopher; Bam, Rujuta A.; Tsai, Angela; Niedziela-Majka, Anita; Stray, Kirsten M.; Sakowicz, Roman; Cihlar, Tomas

    2013-01-01

    HIV-1 integrase (IN) is the target for two classes of antiretrovirals: i) the integrase strand-transfer inhibitors (INSTIs) and ii) the non-catalytic site integrase inhibitors (NCINIs). NCINIs bind at the IN dimer interface and are thought to interfere primarily with viral DNA (vDNA) integration in the target cell by blocking IN-vDNA assembly as well as the IN-LEDGF/p75 interaction. Herein we show that treatment of virus-producing cells, but not of mature virions or target cells, drives NCINI antiviral potency. NCINIs target an essential late-stage event in HIV replication that is insensitive to LEDGF levels in the producer cells. Virus particles produced in the presence of NCINIs displayed normal Gag-Pol processing and endogenous reverse transcriptase activity, but were defective at initiating vDNA synthesis following entry into the target cell. NCINI-resistant virus carrying a T174I mutation in the IN dimer interface was less sensitive to the compound-induced late-stage effects, including the reverse transcription block. Wild-type, but not T174I virus, produced in the presence of NCINIs exhibited striking defects in core morphology and an increased level of IN oligomers that was not observed upon treatment of mature cell-free particles. Collectively, these results reveal that NCINIs act through a novel mechanism that is unrelated to the previously observed inhibition of IN activity or IN-LEDGF interaction, and instead involves the disruption of an IN function during HIV-1 core maturation and assembly. PMID:24040198

  5. Short-term treatment with VEGF receptor inhibitors induces retinopathy of prematurity-like abnormal vascular growth in neonatal rats.

    PubMed

    Nakano, Ayuki; Nakahara, Tsutomu; Mori, Asami; Ushikubo, Hiroko; Sakamoto, Kenji; Ishii, Kunio

    2016-02-01

    Retinal arterial tortuosity and venous dilation are hallmarks of plus disease, which is a severe form of retinopathy of prematurity (ROP). In this study, we examined whether short-term interruption of vascular endothelial growth factor (VEGF) signals leads to the formation of severe ROP-like abnormal retinal blood vessels. Neonatal rats were treated subcutaneously with the VEGF receptor (VEGFR) tyrosine kinase inhibitors, KRN633 (1, 5, or 10 mg/kg) or axitinib (10 mg/kg), on postnatal day (P) 7 and P8. The retinal vasculatures were examined on P9, P14, or P21 in retinal whole-mounts stained with an endothelial cell marker. Prevention of vascular growth and regression of some preformed capillaries were observed on P9 in retinas of rats treated with KRN633. However, on P14 and P21, density of capillaries, tortuosity index of arterioles, and diameter of veins significantly increased in KRN633-treated rats, compared to vehicle (0.5% methylcellulose)-treated animals. Similar observations were made with axitinib-treated rats. Expressions of VEGF and VEGFR-2 were enhanced on P14 in KRN633-treated rat retinas. The second round of KRN633 treatment on P11 and P12 completely blocked abnormal retinal vascular growth on P14, but thereafter induced ROP-like abnormal retinal blood vessels by P21. These results suggest that an interruption of normal retinal vascular development in neonatal rats as a result of short-term VEGFR inhibition causes severe ROP-like abnormal retinal vascular growth in a VEGF-dependent manner. Rats treated postnatally with VEGFR inhibitors could serve as an animal model for studying the mechanisms underlying the development of plus disease. PMID:26500193

  6. The HDAC inhibitor, MPT0E028, enhances erlotinib-induced cell death in EGFR-TKI-resistant NSCLC cells.

    PubMed

    Chen, M-C; Chen, C-H; Wang, J-C; Tsai, A-C; Liou, J-P; Pan, S-L; Teng, C-M

    2013-01-01

    Epidermal growth factor receptor (EGFR), which promotes cell survival and division, is found at abnormally high levels on the surface of many cancer cell types, including many cases of non-small cell lung cancer. Erlotinib (Tarceva), an oral small-molecule tyrosine kinase inhibitor, is a so-called targeted drug that inhibits the tyrosine kinase domain of EGFR, and thus targets cancer cells with some specificity while doing less damage to normal cells. However, erlotinib resistance can occur, reducing the efficacy of this treatment. To develop more effective therapeutic interventions by overcoming this resistance problem, we combined the histone deacetylase inhibitor, MPT0E028, with erlotinib in an effort to increase their antitumor effects in erlotinib-resistant lung adenocarcinoma cells. This combined treatment yielded significant growth inhibition, induced the expression of apoptotic proteins (PARP, ?H2AX, and caspase-3), increased the levels of acetylated histone H3, and showed synergistic effects in vitro and in vivo. These effects were independent of the mutation status of the genes encoding EGFR or K-Ras. MPT0E028 synergistically blocked key regulators of the EGFR/HER2 signaling pathways, attenuating multiple compensatory pathways (e.g., AKT, extracellular signal-regulated kinase, and c-MET). Our results indicate that this combination therapy might be a promising strategy for facilitating the effects of erlotinib monotherapy by activating various networks. Taken together, our data provide compelling evidence that MPT0E028 has the potential to improve the treatment of heterogeneous and drug-resistant tumors that cannot be controlled with single-target agents. PMID:24052078

  7. Propranolol as an inhibitor of some cytochrome P450-dependent monooxygenase activities in native and induced rat liver microsomes.

    PubMed

    Kastelova, A; Yanev, S

    2002-05-01

    The in vitro effect of propranolol (10(-3) M and 10(-4) M), a nonselective and extensively metabolized beta-adrenergic blocking agent, on rat liver drug metabolism in native and induced (with phenobarbital and beta-naphthoflavone [beta-NF]) microsomes was studied. The type of inhibition and the inhibitory constants of some cytochrome P450-dependent microsomal enzyme reactions (hexobarbital oxidation [HBO], ethylmorphine-N-demethylation [EMND], aniline hydroxylation [AH], ethoxycoumarin-O-deethylation [ECOD], ethoxyresorufin-O-dealkylation [EROD] and penthoxyresorufin-O-dealkylation [PROD]) were estimated. The results showed that propranolol competitively inhibited AH activity in native microsomes. The type of inhibition was changed from competitive to noncompetitive in all other enzyme activities studied. This inhibition was more pronounced after phenobarbital induction in PROD (Ki = 0.11 +/- 0.01 mM), ECOD (Ki = 0.40 +/- 0.09 mM) and EMND (Ki = 0.59 +/- 0.1 mM), and after beta-NF induction in AH (Ki = 0.28 +/- 0.05 mM) and in HBO (Ki = 0.35 +/- 0.1 mM) in native microsomes. It was assumed that the noncompetitive type of inhibition is due to the covalent binding of reactive metabolites derived from propranolol to hepatic microsomal proteins. The competitive type of inhibition of AH suggested a common P450 isoenzyme in the metabolism of propranolol and aniline. Thus, in this study, propranolol has been found to be not only a selective inhibitor of CYP2D6 isoenzyme-dependent reactions, but also a nonspecific inhibitor of other cytochrome P450 isoenzymes. PMID:12092004

  8. Accelerated generation of human induced pluripotent stem cells with retroviral transduction and chemical inhibitors under physiological hypoxia

    SciTech Connect

    Shimada, Hidenori; Hashimoto, Yoshiya; Nakada, Akira; Shigeno, Keiji; Nakamura, Tatsuo

    2012-01-13

    Highlights: Black-Right-Pointing-Pointer Very rapid generation of human iPS cells under optimized conditions. Black-Right-Pointing-Pointer Five chemical inhibitors under hypoxia boosted reprogramming. Black-Right-Pointing-Pointer We performed genome-wide DNA methylation analysis. -- Abstract: Induced pluripotent stem (iPS) cells are generated from somatic cells by the forced expression of a defined set of pluripotency-associated transcription factors. Human iPS cells can be propagated indefinitely, while maintaining the capacity to differentiate into all cell types in the body except for extra-embryonic tissues. This technology not only represents a new way to use individual-specific stem cells for regenerative medicine but also constitutes a novel method to obtain large amounts of disease-specific cells for biomedical research. Despite their great potential, the long reprogramming process (up to 1 month) remains one of the most significant challenges facing standard virus-mediated methodology. In this study, we report the accelerated generation of human iPS cells from adipose-derived stem (ADS) cells, using a new combination of chemical inhibitors under a setting of physiological hypoxia in conjunction with retroviral transduction of Oct4, Sox2, Klf4, and L-Myc. Under optimized conditions, we observed human embryonic stem (ES)-like cells as early as 6 days after the initial retroviral transduction. This was followed by the emergence of fully reprogrammed cells bearing Tra-1-81-positive and DsRed transgene-silencing properties on day 10. The resulting cell lines resembled human ES cells in many respects including proliferation rate, morphology, pluripotency-associated markers, global gene expression patterns, genome-wide DNA methylation states, and the ability to differentiate into all three of the germ layers, both in vitro and in vivo. Our method, when combined with chemical inhibitors under conditions of physiological hypoxia, offers a powerful tool for rapidly generating bona fide human iPS cells and facilitates the application of iPS cell technology to biomedical research.

  9. The FGFR1 inhibitor PD173074 induces mesenchymal–epithelial transition through the transcription factor AP-1

    PubMed Central

    Nguyen, P T; Tsunematsu, T; Yanagisawa, S; Kudo, Y; Miyauchi, M; Kamata, N; Takata, T

    2013-01-01

    Background: Epithelial–mesenchymal transition (EMT) is a crucial process in cancer progression that provides cancer cells with the ability to escape from the primary focus, invade stromal tissues and migrate to distant regions. Cell lines that lack E-cadherin show increased tumorigenesis and metastasis, and the expression levels of E-cadherin and Snail correlate inversely with the prognosis of patients suffering from breast cancer or oral squamous cell carcinoma (OSCC). Moreover, recent studies have shown that most EMT cases are regulated by soluble growth factors or cytokines. Among these factors, fibroblast growth factors (FGFs) execute diverse functions by binding to and activating members of the FGF receptor (FGFR) family, including FGFR1–4. Fibroblast growth factor receptor 1 is an oncoprotein that is involved in tumorigenesis, and PD173074 is known to be a selective inhibitor of FGFR1. However, the roles of FGFR1 and FGFR1 inhibitors have not yet been examined in detail. Methods: Here, we investigated the expression of FGFR1 in head and neck squamous cell carcinoma (HNSCC) and the role of the FGFR1 inhibitor PD173074 in carcinogenesis and the EMT process. Results: Fibroblast growth factor receptor 1 was highly expressed in 54% of HNSCC cases and was significantly correlated with malignant behaviours. Nuclear FGFR1 expression was also observed and correlated well with histological differentiation, the pattern of invasion and abundant nuclear polymorphism. Fibroblast growth factor receptor 1 was also overexpressed in EMT cell lines compared with non-EMT cell lines. Furthermore, treatment of HOC313 cells with PD173074 suppressed cellular proliferation and invasion and reduced ERK1/2 and p38 activation. These cells also demonstrated morphological changes, transforming from spindle- to cobble stone-like in shape. In addition, the expression levels of certain matrix metalloproteinases (MMPs), whose genes contain activator protein-1 (AP-1) promoter sites, as well as Snail1 and Snail2 were reduced following PD173074 treatment. Conclusion: Taken together, these data suggest that PD173074 inhibits the MAPK pathway, which regulates the activity of AP-1 and induces MET. Furthermore, this induction of MET likely suppresses cancer cell growth and invasion. PMID:24045665

  10. Identification of Novel Compounds Inhibiting Chikungunya Virus-Induced Cell Death by High Throughput Screening of a Kinase Inhibitor Library

    PubMed Central

    Gomes, Rafael G. B.; da Silva, Camila T.; Taniguchi, Juliana B.; No, Joo Hwan; Lombardot, Benoit; Schwartz, Olivier; Hansen, Michael A. E.; Freitas-Junior, Lucio H.

    2013-01-01

    Chikungunya virus (CHIKV) is a mosquito-borne arthrogenic alphavirus that causes acute febrile illness in humans accompanied by joint pains and in many cases, persistent arthralgia lasting weeks to years. The re-emergence of CHIKV has resulted in numerous outbreaks in the eastern hemisphere, and threatens to expand in the foreseeable future. Unfortunately, no effective treatment is currently available. The present study reports the use of resazurin in a cell-based high-throughput assay, and an image-based high-content assay to identify and characterize inhibitors of CHIKV-infection in vitro. CHIKV is a highly cytopathic virus that rapidly kills infected cells. Thus, cell viability of HuH-7 cells infected with CHIKV in the presence of compounds was determined by measuring metabolic reduction of resazurin to identify inhibitors of CHIKV-associated cell death. A kinase inhibitor library of 4,000 compounds was screened against CHIKV infection of HuH-7 cells using the resazurin reduction assay, and the cell toxicity was also measured in non-infected cells. Seventy-two compounds showing ?50% inhibition property against CHIKV at 10 µM were selected as primary hits. Four compounds having a benzofuran core scaffold (CND0335, CND0364, CND0366 and CND0415), one pyrrolopyridine (CND0545) and one thiazol-carboxamide (CND3514) inhibited CHIKV-associated cell death in a dose-dependent manner, with EC50 values between 2.2 µM and 7.1 µM. Based on image analysis, these 6 hit compounds did not inhibit CHIKV replication in the host cell. However, CHIKV-infected cells manifested less prominent apoptotic blebs typical of CHIKV cytopathic effect compared with the control infection. Moreover, treatment with these compounds reduced viral titers in the medium of CHIKV-infected cells by up to 100-fold. In conclusion, this cell-based high-throughput screening assay using resazurin, combined with the image-based high content assay approach identified compounds against CHIKV having a novel antiviral activity - inhibition of virus-induced CPE - likely by targeting kinases involved in apoptosis. PMID:24205414

  11. Computational prediction and experimental validation of a novel synthesized pan-PIM inhibitor PI003 and its apoptosis-inducing mechanisms in cervical cancer

    PubMed Central

    Gao, Jianglin; Luo, Junhua; Huang, Xian; Gao, Chunfang

    2015-01-01

    PIM protein family, short-lived serine/threonine kinases (PIM1, PIM2 and PIM3), are weak oncogenes but contribute to tumorigenesis as cancer targets. Thus, design of a novel pan-PIM inhibitor is still a challenge for current cancer drug discovery. Herein, we used a Naïve Bayesian model to construct the PIM network and identified Bad and Hsp90 to interact with PIMs. Then, we screened a series of candidate small-molecule compounds targeting PIMs, and subsequently synthesized a novel small-molecule compound PI003 with remarkable anti-proliferative activities in cervical cancer cells. Moreover, we found that PI003 induced apoptosis via the death-receptor and mitochondrial pathways by targeting PIMs and affecting Bad and Hsp90. Combined with microRNA microarray analyses, we demonstrated that some microRNAs such as miR-1296 and miR-1299 could affect PIM1-STAT3 pathway in PI003-induced apoptosis. Finally, we reported that PI003 had remarkable anti-tumor activity and apoptosis-inducing effect in in vivo mouse model. In conclusion, these results demonstrate that PI003, as a novel synthesized pan-PIM inhibitor, induces the death-receptor and mitochondrial apoptosis involved in microRNA regulation, and also possessed remarkable anti-tumor activity and apoptosis-inducing effect in vivo. Thus, these findings would shed light on discovering more potential new small-molecule pan-PIM inhibitors in future cervical cancer therapy. PMID:25749522

  12. Constitutive Smad signaling and Smad-dependent collagen gene expression in mouse embryonic fibroblasts lacking peroxisome proliferator-activated receptor-{gamma}

    SciTech Connect

    Ghosh, Asish K Wei, Jun; Wu, Minghua; Varga, John

    2008-09-19

    Transforming growth factor-{beta} (TGF-{beta}), a potent inducer of collagen synthesis, is implicated in pathological fibrosis. Peroxisome proliferator-activated receptor-{gamma} (PPAR-{gamma}) is a nuclear hormone receptor that regulates adipogenesis and numerous other biological processes. Here, we demonstrate that collagen gene expression was markedly elevated in mouse embryonic fibroblasts (MEFs) lacking PPAR-{gamma} compared to heterozygous control MEFs. Treatment with the PPAR-{gamma} ligand 15d-PGJ{sub 2} failed to down-regulate collagen gene expression in PPAR-{gamma} null MEFs, whereas reconstitution of these cells with ectopic PPAR-{gamma} resulted in their normalization. Compared to control MEFs, PPAR-{gamma} null MEFs displayed elevated levels of the Type I TGF-{beta} receptor (T{beta}RI), and secreted more TGF-{beta}1 into the media. Furthermore, PPAR-{gamma} null MEFs showed constitutive phosphorylation of cellular Smad2 and Smad3, even in the absence of exogenous TGF-{beta}, which was abrogated by the ALK5 inhibitor SB431542. Constitutive Smad2/3 phosphorylation in PPAR-{gamma} null MEFs was associated with Smad3 binding to its cognate DNA recognition sequences, and interaction with coactivator p300 previously implicated in TGF-{beta} responses. Taken together, these results indicate that loss of PPAR-{gamma} in MEFs is associated with upregulation of collagen synthesis, and activation of intracellular Smad signal transduction, due, at least in part, to autocrine TGF-{beta} stimulation.

  13. Dihydroartemisinin Inhibits Glucose Uptake and Cooperates with Glycolysis Inhibitor to Induce Apoptosis in Non-Small Cell Lung Carcinoma Cells

    PubMed Central

    Gao, Jing; Luo, Xian-yang; Liu, Yu; Li, Ning; Li, Chun-lei; Chen, Yu-qiang; Yu, Xiu-yi; Jiang, Jie

    2015-01-01

    Despite recent advances in the therapy of non-small cell lung cancer (NSCLC), the chemotherapy efficacy against NSCLC is still unsatisfactory. Previous studies show the herbal antimalarial drug dihydroartemisinin (DHA) displays cytotoxic to multiple human tumors. Here, we showed that DHA decreased cell viability and colony formation, induced apoptosis in A549 and PC-9 cells. Additionally, we first revealed DHA inhibited glucose uptake in NSCLC cells. Moreover, glycolytic metabolism was attenuated by DHA, including inhibition of ATP and lactate production. Consequently, we demonstrated that the phosphorylated forms of both S6 ribosomal protein and mechanistic target of rapamycin (mTOR), and GLUT1 levels were abrogated by DHA treatment in NSCLC cells. Furthermore, the upregulation of mTOR activation by high expressed Rheb increased the level of glycolytic metabolism and cell viability inhibited by DHA. These results suggested that DHA-suppressed glycolytic metabolism might be associated with mTOR activation and GLUT1 expression. Besides, we showed GLUT1 overexpression significantly attenuated DHA-triggered NSCLC cells apoptosis. Notably, DHA synergized with 2-Deoxy-D-glucose (2DG, a glycolysis inhibitor) to reduce cell viability and increase cell apoptosis in A549 and PC-9 cells. However, the combination of the two compounds displayed minimal toxicity to WI-38 cells, a normal lung fibroblast cell line. More importantly, 2DG synergistically potentiated DHA-induced activation of caspase-9, -8 and -3, as well as the levels of both cytochrome c and AIF of cytoplasm. However, 2DG failed to increase the reactive oxygen species (ROS) levels elicited by DHA. Overall, the data shown above indicated DHA plus 2DG induced apoptosis was involved in both extrinsic and intrinsic apoptosis pathways in NSCLC cells. PMID:25799586

  14. Administration of I?B-kinase inhibitor PS1145 enhances apoptosis in DMBA-induced tumor in male Wistar rats.

    PubMed

    Rajmani, R S; Gandham, Ravi Kumar; Gupta, Shishir Kumar; Sahoo, Aditya P; Singh, Prafull Kumar; Saxena, Shikha; Kumar, Rajiv; Chaturvedi, Uttara; Tiwari, Ashok K

    2015-11-01

    Nuclear factor kappa-B (NF-?B), a key anti-apoptotic factor, plays a critical role in tumor cell growth, metastasis, and angiogenesis. The transcriptional activity of NF-?B is normally suppressed in the cytoplasm due to its association with a natural inhibitor molecule I?B. Phosphorylation of the I?B at Ser 32 and Ser 36 by the I?B kinase complex (IKK) marks the degradation of the molecule by 26S proteasome. As NF-?B is constitutively activated in most of the tumor cells, inhibition of the activities of IKK may significantly sensitize the tumor cells to apoptosis. In the present study, we investigated the effect of I?B kinase-specific blocker PS1145 on DMBA-induced skin tumor of male Wistar rats. We examined the apoptotic effect of PS1145 on DMBA-induced tumor by various histopathological and molecular techniques. Our results demonstrate the significant expression of major pro-apoptotic genes like caspases 2, 3, 8, 9, and p53 in PS1145-treated tumor bearing group at mRNA levels as well as significant (P?induced skin tumor in male Wistar rats by blocking NF-?B and VEGF activities. PMID:26148342

  15. G2019S LRRK2 and aging confer susceptibility to proteasome inhibitor-induced neurotoxicity in nigrostriatal dopaminergic system.

    PubMed

    Xiao, Qian; Yang, Suosuo; Le, Weidong

    2015-12-01

    The leucine-rich repeat kinase 2 (LRRK2) mutation G2019S is one of the most common genetic causes in Parkinson's disease (PD). The penetrance of G2019S LRRK2 is incomplete and is age-dependent, therefore, it has been speculated that environmental toxins and aging could contribute to G2019S LRRK2-related PD pathogenesis. To prove this speculation, we performed a longitudinal investigation in mice bearing G2019S LRRK2 mutation. BAC G2019S LRRK2 transgenic (Tg) mice and their wildtype (Wt) littermates were treated with lactacystin, a specific proteasome inhibitor. The susceptibilities of mice to lactacystin-induced nigrostriatal dopaminergic (DAergic) degeneration were evaluated, at 5 and 12 months of age. We found that lactacystin treatment caused a greater decline of striatal DA content in the Tg mice at either 5 or 12 months of age than their age-matched Wt littermates. Moreover, the lactacystin-treated Tg or Wt mice at 12 months of age lose much more nigral tyrosine hydroxylase (TH)-positive neurons than the mice at 5 months of age, indicating an age-associated DAergic neurotoxicity. Additionally, stereotactic injection of lactacystin induced a dramatic increase of activated microglia in substantia nigra of mice at 12 months of age, compared with mice at 5 months of age. In summary, our study suggests that expression of the G2019S mutation in the mouse LRRK2 gene confers an age-associated high susceptibility to proteasome inhibition-induced nigrostriatal DAergic degeneration. PMID:26253900

  16. Immunomodulatory drugs act as inhibitors of DNA methyltransferases and induce PU.1 up-regulation in myeloma cells.

    PubMed

    Endo, Shinya; Amano, Masayuki; Nishimura, Nao; Ueno, Niina; Ueno, Shikiko; Yuki, Hiromichi; Fujiwara, Shiho; Wada, Naoko; Hirata, Shinya; Hata, Hiroyuki; Mitsuya, Hiroaki; Okuno, Yutaka

    2016-01-01

    Immunomodulatory drugs (IMiDs) such as thalidomide, lenalidomide, and pomalidomide are efficacious in the treatment of multiple myeloma and significantly prolong their survival. However, the mechanisms of such effects of IMiDs have not been fully elucidated. Recently, cereblon has been identified as a target binding protein of thalidomide. Lenalidomide-resistant myeloma cell lines often lose the expression of cereblon, suggesting that IMiDs act as an anti-myeloma agent through interacting with cereblon. Cereblon binds to damaged DNA-binding protein and functions as a ubiquitin ligase, inducing degradation of IKZF1 and IKZF3 that are essential transcription factors for B and T cell development. Degradation of both IKZF1 and IKZF3 reportedly suppresses myeloma cell growth. Here, we found that IMiDs act as inhibitors of DNA methyltransferases (DMNTs). We previously reported that PU.1, which is an ETS family transcription factor and essential for myeloid and lymphoid development, functions as a tumor suppressor in myeloma cells. PU.1 induces growth arrest and apoptosis of myeloma cell lines. In this study, we found that low-dose lenalidomide and pomalidomide up-regulate PU.1 expression through inducing demethylation of the PU.1 promoter. In addition, IMiDs inhibited DNMT1, DNMT3a, and DNMT3b activities in vitro. Furthermore, lenalidomide and pomalidomide decreased the methylation status of the whole genome in myeloma cells. Collectively, IMiDs exert demethylation activity through inhibiting DNMT1, 3a, and 3b, and up-regulating PU.1 expression, which may be one of the mechanisms of the anti-myeloma activity of IMiDs. PMID:26657848

  17. Regulation of the human SLC25A20 expression by peroxisome proliferator-activated receptor alpha in human hepatoblastoma cells

    SciTech Connect

    Tachibana, Keisuke; Takeuchi, Kentaro; Inada, Hirohiko; Yamasaki, Daisuke; The Center for Advanced Medical Engineering and Informatics, Osaka University, 2-2 Yamadaoka, Suita, Osaka 565-0871 ; Ishimoto, Kenji; Graduate School of Medicine, Osaka University, 2-2 Yamadaoka, Suita, Osaka 565-0871 ; Tanaka, Toshiya; Hamakubo, Takao; Sakai, Juro; Kodama, Tatsuhiko; Doi, Takefumi; The Center for Advanced Medical Engineering and Informatics, Osaka University, 2-2 Yamadaoka, Suita, Osaka 565-0871; Graduate School of Medicine, Osaka University, 2-2 Yamadaoka, Suita, Osaka 565-0871

    2009-11-20

    Solute carrier family 25, member 20 (SLC25A20) is a key molecule that transfers acylcarnitine esters in exchange for free carnitine across the mitochondrial membrane in the mitochondrial {beta}-oxidation. The peroxisome proliferator-activated receptor alpha (PPAR{alpha}) is a ligand-activated transcription factor that plays an important role in the regulation of {beta}-oxidation. We previously established tetracycline-regulated human cell line that can be induced to express PPAR{alpha} and found that PPAR{alpha} induces the SLC25A20 expression. In this study, we analyzed the promoter region of the human slc25a20 gene and showed that PPAR{alpha} regulates the expression of human SLC25A20 via the peroxisome proliferator responsive element.

  18. Translation inhibitors sensitize prostate cancer cells to apoptosis induced by tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) by activating c-Jun N-terminal kinase.

    PubMed

    Sah, Nand K; Munshi, Anupama; Kurland, John F; McDonnell, Timothy J; Su, Bing; Meyn, Raymond E

    2003-06-01

    Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) induces apoptosis in several human tumors both in vitro and in vivo, however, some tumors remain resistant for poorly understood reasons. Using a quantitative DNA fragmentation assay for apoptosis, we have shown that human prostate cancer cells are resistant to a wide range of TRAIL doses up to 500 ng/ml. However, translation inhibitors, such as anisomycin, cycloheximide, emetine, harringtonine, and puromycin, unlike several transcription inhibitors, significantly sensitized PC3-neomycin (PC3-neo) cells to TRAIL-induced apoptosis. These effects were inhibited in PC3 cells engineered to express bcl2 (PC3-bcl2). Translation inhibitors led to activation of c-Jun N-terminal kinase (JNK), which plays a role in this sensitization process because inhibition of JNK activation resulted in protection against TRAIL plus translation inhibitor-induced apoptosis. JNK activation may be required for this process, but it is not sufficient because activation of JNK using an MEKK2 expression vector did not mimic the sensitizing effect of translation inhibitors. Other stress-activated protein kinases, such as ERK and p38, play an insignificant role in determining the apoptotic sensitivity. We conclude that activation of JNK is required for sensitization of PC3 cells to TRAIL-induced apoptosis by translation inhibitors in cells that are otherwise TRAIL-resistant. However, in addition to JNK activation, other aspects of translation inhibition such as the suppressed activity of apoptosis-inhibitory proteins or activation of other signal transduction pathways must also be involved. PMID:12663665

  19. Identification of a kinase profile that predicts chromosome damage induced by small molecule kinase inhibitors.

    PubMed

    Olaharski, Andrew J; Gonzaludo, Nina; Bitter, Hans; Goldstein, David; Kirchner, Stephan; Uppal, Hirdesh; Kolaja, Kyle

    2009-07-01

    Kinases are heavily pursued pharmaceutical targets because of their mechanistic role in many diseases. Small molecule kinase inhibitors (SMKIs) are a compound class that includes marketed drugs and compounds in various stages of drug development. While effective, many SMKIs have been associated with toxicity including chromosomal damage. Screening for kinase-mediated toxicity as early as possible is crucial, as is a better understanding of how off-target kinase inhibition may give rise to chromosomal damage. To that end, we employed a competitive binding assay and an analytical method to predict the toxicity of SMKIs. Specifically, we developed a model based on the binding affinity of SMKIs to a panel of kinases to predict whether a compound tests positive for chromosome damage. As training data, we used the binding affinity of 113 SMKIs against a representative subset of all kinases (290 kinases), yielding a 113x290 data matrix. Additionally, these 113 SMKIs were tested for genotoxicity in an in vitro micronucleus test (MNT). Among a variety of models from our analytical toolbox, we selected using cross-validation a combination of feature selection and pattern recognition techniques: Kolmogorov-Smirnov/T-test hybrid as a univariate filter, followed by Random Forests for feature selection and Support Vector Machines (SVM) for pattern recognition. Feature selection identified 21 kinases predictive of MNT. Using the corresponding binding affinities, the SVM could accurately predict MNT results with 85% accuracy (68% sensitivity, 91% specificity). This indicates that kinase inhibition profiles are predictive of SMKI genotoxicity. While in vitro testing is required for regulatory review, our analysis identified a fast and cost-efficient method for screening out compounds earlier in drug development. Equally important, by identifying a panel of kinases predictive of genotoxicity, we provide medicinal chemists a set of kinases to avoid when designing compounds, thereby providing a basis for rational drug design away from genotoxicity. PMID:19629159

  20. Dehydrogenases of the pentose phosphate pathway in rat liver peroxisomes.

    PubMed

    Antonenkov, V D

    1989-07-15

    Subcellular distribution of pentose-phosphate cycle enzymes in rat liver was investigated, using differential and isopycnic centrifugation. The activities of the NADP+-dependent dehydrogenases of the pentose-phosphate pathway (glucose-6-phosphate dehydrogenase and phosphogluconate dehydrogenase) were detected in the purified peroxisomal fraction as well as in the cytosol. Both dehydrogenases were localized in the peroxisomal matrix. Chronic administration of the hypolipidemic drug clofibrate (ethyl-alpha-p-chlorophenoxyisobutyrate) caused a 1.5-2.5-fold increase in the amount of glucose-6-phosphate and phosphogluconate dehydrogenases in the purified peroxisomes. Clofibrate decreased the phosphogluconate dehydrogenase, but did not alter glucose-6-phosphate dehydrogenase activity in the cytosolic fraction. The results obtained indicate that the enzymes of the non-oxidative segment of the pentose cycle (transketolase, transaldolase, triosephosphate isomerase and glucose-phosphate isomerase) are present only in a soluble form in the cytosol, but not in the peroxisomes or other particles, and that ionogenic interaction of the enzymes with the mitochondrial and other membranes takes place during homogenization of the tissue in 0.25 M sucrose. Similar to catalase, glucose-6-phosphate dehydrogenase and phosphogluconate dehydrogenase are present in the intact peroxisomes in a latent form. The enzymes have Km values for their substrates in the millimolar range (0.2 mM for glucose-6-phosphate and 0.10-0.12 mM for 6-phosphogluconate). NADP+, but not NAD+, serves as a coenzyme for both enzymes. Glucose-6-phosphate dehydrogenase was inhibited by palmitoyl-CoA, and to a lesser extent by NADPH. Peroxisomal glucose-6-phosphate and phosphogluconate dehydrogenases have molecular mass of 280 kDa and 96 kDa, respectively. The putative functional role of pentose-phosphate cycle dehydrogenases in rat liver peroxisomes is discussed. PMID:2753047

  1. Tetrahydroindazoles as Interleukin-2 Inducible T-Cell Kinase Inhibitors. Part II. Second-Generation Analogues with Enhanced Potency, Selectivity, and Pharmacodynamic Modulation in Vivo.

    PubMed

    Burch, Jason D; Barrett, Kathy; Chen, Yuan; DeVoss, Jason; Eigenbrot, Charles; Goldsmith, Richard; Ismaili, M Hicham A; Lau, Kevin; Lin, Zhonghua; Ortwine, Daniel F; Zarrin, Ali A; McEwan, Paul A; Barker, John J; Ellebrandt, Claire; Kordt, Daniel; Stein, Daniel B; Wang, Xiaolu; Chen, Yong; Hu, Baihua; Xu, Xiaofeng; Yuen, Po-Wai; Zhang, Yamin; Pei, Zhonghua

    2015-05-14

    The medicinal chemistry community has directed considerable efforts toward the discovery of selective inhibitors of interleukin-2 inducible T-cell kinase (ITK), given its role in T-cell signaling downstream of the T-cell receptor (TCR) and the implications of this target for inflammatory disorders such as asthma. We have previously disclosed a structure- and property-guided lead optimization effort which resulted in the discovery of a new series of tetrahydroindazole-containing selective ITK inhibitors. Herein we disclose further optimization of this series that resulted in further potency improvements, reduced off-target receptor binding liabilities, and reduced cytotoxicity. Specifically, we have identified a correlation between the basicity of solubilizing elements in the ITK inhibitors and off-target antiproliferative effects, which was exploited to reduce cytotoxicity while maintaining kinase selectivity. Optimized analogues were shown to reduce IL-2 and IL-13 production in vivo following oral or intraperitoneal dosing in mice. PMID:25844760

  2. In vitro modulation of MMP-2 and MMP-9 in adult human sarcoma cell lines by cytokines, inducers and inhibitors

    PubMed Central

    ROOMI, M.W.; KALINOVSKY, T.; MONTERREY, J.; RATH, M.; NIEDZWIECKI, A.

    2013-01-01

    The highly aggressive adult sarcomas are characterized by high levels of matrix metalloproteinase (MMP)-2 and -9, which play crucial roles in tumor invasion and metastasis by degradation of the extracellular membrane leading to cancer cell spread to distal organs. We examined the effect of cytokines, mitogens, inducers and inhibitors on MMP-2 and MMP-9 secretion in chondrosarcoma (SW-1353), fibrosarcoma (HT-1080), liposarcoma (SW-872) and synovial sarcoma (SW-982) cell lines. The selected compounds included natural cytokines and growth factors, as well as chemical compounds applied in therapy of sarcoma and natural compounds that have demonstrated anticancer therapeutic potential. MMP-2 and MMP-9 secretions were analyzed by gelatinase zymography following 24-h exposure to the tested agents and quantitated by densitometry. Fibrosarcoma, chondrosarcoma, liposarcoma and synovial sarcoma showed bands corresponding to MMP-2 and MMP-9 with dose-dependent enhancement of MMP-9 with phorbol 12-myristate 13-acetate (PMA) treatment. In chondrosarcoma cells, tumor necrosis factor (TNF)-? had a stimulatory effect on MMP-9 and insignificant effect on MMP-2 and interleukin (IL)-1? stimulated MMP-9 and MMP-2. In fibrosarcoma and liposarcoma cells, TNF-? had a profound stimulatory effect on MMP-9, but no effect on MMP-2 and in synovial sarcoma an inhibitory effect on MMP-2 and no effect on MMP-9. IL-1? had a slight inhibitory effect on fibrosarcoma, liposarcoma and synovial sarcoma MMP-2 and MMP-9 except for MMP-9 in synovial sarcoma which showed slight stimulation. Lipopolysaccharide (LPS) stimulated expression of MMP-2 in fibrosarcoma and chondrosarcoma while inhibited it in liposarcoma. Doxycycline, epigallocatechin gallate and the nutrient mixture inhibited MMP-2 and MMP-9 in all cell lines. Actinomycin-D, cyclohexamide, retinoic acid, and dexamethasone inhibited MMP-2 and -9 in chondrosarcoma and fibrosarcoma cells. Our results show that cytokines, mitogens, inducers and inhibitors have an up or down regulatory effect on MMP-2 and MMP-9 expression in adult sarcoma cell lines, suggesting these agents may be effective strategies to treat these cancers. PMID:24085323

  3. Positive selection of novel peroxisome biogenesis-defective mutants of the yeast Pichia pastoris.

    PubMed Central

    Johnson, M A; Waterham, H R; Ksheminska, G P; Fayura, L R; Cereghino, J L; Stasyk, O V; Veenhuis, M; Kulachkovsky, A R; Sibirny, A A; Cregg, J M

    1999-01-01

    We have developed two novel schemes for the direct selection of peroxisome-biogenesis-defective (pex) mutants of the methylotrophic yeast Pichia pastoris. Both schemes take advantage of our observation that methanol-induced pex mutants contain little or no alcohol oxidase (AOX) activity. AOX is a peroxisomal matrix enzyme that catalyzes the first step in the methanol-utilization pathway. One scheme utilizes allyl alcohol, a compound that is not toxic to cells but is oxidized by AOX to acrolein, a compound that is toxic. Exposure of mutagenized populations of AOX-induced cells to allyl alcohol selectively kills AOX-containing cells. However, pex mutants without AOX are able to grow. The second scheme utilizes a P. pastoris strain that is defective in formaldehyde dehydrogenase (FLD), a methanol pathway enzyme required to metabolize formaldehyde, the product of AOX. AOX-induced cells of fld1 strains are sensitive to methanol because of the accumulation of formaldehyde. However, fld1 pex mutants, with little active AOX, do not efficiently oxidize methanol to formaldehyde and therefore are not sensitive to methanol. Using these selections, new pex mutant alleles in previously identified PEX genes have been isolated along with mutants in three previously unidentified PEX groups. PMID:10101164

  4. The effects of an inhibitor of diglyceride lipase on collagen-induced platelet activation.

    PubMed

    Jackson, Elke C G; Ortar, Giorgio; McNicol, Archie

    2013-12-01

    Human platelet activation by collagen occurs in a dose-dependent manner. High concentrations of collagen bind to a pair of receptors, the ?2?1 integrin and glycoprotein (GP)VI/Fc-receptor ?-chain (FcR?), which stimulate a cascade of events including Syk, LAT, Btk, Gads, and phospholipase C?2, leading to calcium release and protein kinase C (PKC) activation. Calcium and PKC are responsible for a range of platelet responses including exocytosis and aggregation, as well as the cytosolic phospholipase A2 (cPLA2)-mediated release of arachidonic acid, which is converted to thromboxane (Tx)A2. In contrast, low concentrations of collagen are acutely aspirin-sensitive, and calcium release and aggregation are TxA2-dependent. Under these conditions, cPLA2 is not involved and it has been suggested that phospholipase C generates 1,2-diacylglycerol (DG) from which arachidonic acid is liberated by diglyceride lipase (DGL). Here a novel DGL blocker (OMDM-188) inhibited collagen-, but not arachidonic acid-induced aggregation and TxA2 synthesis. Furthermore, OMDM-188 inhibited collagen-induced arachidonic acid release. Finally OMDM-188 inhibited collagen-induced p38(MAPK) phosphorylation, but not extracellular signal-regulated kinase (ERK) phosphorylation, with no effect on the phosphorylation of either enzyme in response to arachidonic acid. Taken together, these data suggest a role for a pathway involving phospholipase C liberating DG from membrane phospholipids in response to minimally activating concentrations of collagen. The DG serves as a substrate for DGL, potentially under the regulations of p38(MAPK), to release arachidonic acid, which is subsequently converted to TxA2, which mediates the final platelet response. PMID:24042163

  5. TNF-inhibitor induced lupus in a patient treated with adalimumab for rheumatoid arthritis.

    PubMed

    Lieberman, Miriam R; Liebman, Tracey N; Alapati, Usha; Khachemoune, Amor

    2015-02-01

    Anti-tumor necrosis factor induced lupus (ATIL) is a rare side effect reported in patients treated with anti-tumor necrosis factor medications such as infliximab, etanercept and adalimumab. Of the three, this condition has been least commonly reported secondary to adalimumab. In this report, we present a case of ATIL in a patient treated for rheumatoid arthritis (RA) with adalimumab. This report will increase physician awareness of the warning signs, diagnostic options and potential complications of ATIL. In this patient, adalimumab was discontinued and treatment was started, leading to improvement in the patient's status. PMID:25756476

  6. The indirect NMDAR inhibitor flupirtine induces sustained post-ischemic recovery, neuroprotection and angioneurogenesis.

    PubMed

    Jaeger, Hanna M; Pehlke, Jens R; Kaltwasser, Britta; Kilic, Ertugrul; Bähr, Mathias; Hermann, Dirk M; Doeppner, Thorsten R

    2015-06-10

    N-methyl-D-aspartate receptor (NMDAR) activation induces excitotoxicity, contributing to post-stroke brain injury. Hitherto, NMDAR deactivation failed in clinical trials due to insufficient pre-clinical study designs and drug toxicity. Flupirtine is an indirect NMDAR antagonist being used as analgesic in patients. Taking into account its tolerability profile, we evaluated effects of flupirtine on post-stroke tissue survival, neurological recovery and brain remodeling.Mice were exposed to stroke and intraperitoneally treated with saline (control) or flupirtine at various doses (1-10 mg/kg) and time-points (0-12 hours). Tissue survival and cell signaling were studied on day 2, whereas neurological recovery and tissue remodeling were analyzed until day 84.Flupirtine induced sustained neuroprotection, when delivered up to 9 hours. The latter yielded enhanced neurological recovery that persisted over three months and which was accompanied by enhanced angioneurogenesis. On the molecular level, inhibition of calpain activation was noted, which was associated with increased signal-transducer-and-activator-of-transcription-6 (STAT6) abundance, reduced N-terminal-Jun-kinase and NF-?B activation, as well as reduced proteasomal activity. Consequently, blood-brain-barrier integrity was stabilized, oxidative stress was reduced and brain leukocyte infiltration was diminished.In view of its excellent tolerability, considering its sustained effects on neurological recovery, brain tissue survival and remodeling, flupirtine is an attractive candidate for stroke therapy. PMID:26050199

  7. Stress exposure results in increased peroxisomal levels of yeast Pnc1 and Gpd1, which are imported via a piggy-backing mechanism.

    PubMed

    Kumar, Sanjeev; Singh, Ritika; Williams, Chris P; van der Klei, Ida J

    2016-01-01

    Saccharomyces cerevisiae glycerol phosphate dehydrogenase 1 (Gpd1) and nicotinamidase (Pnc1) are two stress-induced enzymes. Both enzymes are predominantly localised to peroxisomes at normal growth conditions, but were reported to localise to the cytosol and nucleus upon exposure of cells to stress. Import of both proteins into peroxisomes depends on the peroxisomal targeting signal 2 (PTS2) receptor Pex7. Gpd1 contains a PTS2, however, Pnc1 lacks this sequence. Here we show that Pnc1 physically interacts with Gpd1, which is required for piggy-back import of Pnc1 into peroxisomes. Quantitative fluorescence microscopy analyses revealed that the levels of both proteins increased in peroxisomes and in the cytosol upon exposure of cells to stress. However, upon exposure of cells to stress we also observed enhanced cytosolic levels of the control PTS2 protein thiolase, when produced under control of the GPD1 promoter. This suggests that these conditions cause a partial defect in PTS2 protein import, probably because the PTS2 import pathway is easily saturated. PMID:26516056

  8. Pharmacological characterization of KLYP961, a dual inhibitor of inducible and neuronal nitric-oxide synthases.

    PubMed

    Symons, Kent T; Nguyen, Phan M; Massari, Mark E; Anzola, John V; Staszewski, Lena M; Wang, Li; Yazdani, Nahid; Dorow, Steven; Muhammad, Jerry; Sablad, Marciano; Rozenkrants, Natasha; Bonefous, Celine; Payne, Joseph E; Rix, Peter J; Shiau, Andrew K; Noble, Stewart A; Smith, Nicholas D; Hassig, Christian A; Zhang, Yan; Rao, Tadimeti S

    2011-02-01

    Nitric oxide (NO) derived from neuronal nitric-oxide synthase (nNOS) and inducible nitric-oxide synthase (iNOS) plays a key role in various pain and inflammatory states. KLYP961 (4-((2-cyclobutyl-1H-imidazo[4,5-b]pyrazin-1-yl)methyl)-7,8-difluoroquinolin-2(1H)-one) inhibits the dimerization, and hence the enzymatic activity of human, primate, and murine iNOS and nNOS (IC(50) values 50-400 nM), with marked selectivity against endothelial nitric-oxide synthase (IC(50) >15,000 nM). It has ideal drug like-properties, including excellent rodent and primate pharmacokinetics coupled with a minimal off-target activity profile. In mice, KLYP961 attenuated endotoxin-evoked increases in plasma nitrates, a surrogate marker of iNOS activity in vivo, in a sustained manner (ED(50) 1 mg/kg p.o.). KLYP961 attenuated pain behaviors in a mouse formalin model (ED(50) 13 mg/kg p.o.), cold allodynia in the chronic constriction injury model (ED(50) 25 mg/kg p.o.), or tactile allodynia in the spinal nerve ligation model (ED(50) 30 mg/kg p.o.) with similar efficacy, but superior potency relative to gabapentin, pregabalin, or duloxetine. Unlike morphine, the antiallodynic activity of KLYP961 did not diminish upon repeated dosing. KLYP961 also attenuated carrageenin-induced edema and inflammatory hyperalgesia and writhing response elicited by phenylbenzoquinone with efficacy and potency similar to those of celecoxib. In contrast to gabapentin, KLYP961 did not impair motor coordination at doses as high as 1000 mg/kg p.o. KLYP961 also attenuated capsaicin-induced thermal allodynia in rhesus primates in a dose-related manner with a minimal effective dose (? 10 mg/kg p.o.) and a greater potency than gabapentin. In summary, KLYP961 represents an ideal tool with which to probe the physiological role of NO derived from iNOS and nNOS in human pain and inflammatory states. PMID:21036913

  9. Evaluation of protease inhibitors and an antioxidant for treatment of sulfur mustard-induced toxic lung injury.

    PubMed

    Anderson, Dana R; Taylor, Stephanie L; Fetterer, David P; Holmes, Wesley W

    2009-09-01

    Sulfur mustard (SM)-induced lung injury has been associated with protease activation, oxidative injury and inflammatory response culminating in tissue necrosis. The protease inhibitors aprotinin and ilomastat and the antioxidant trolox were evaluated for efficacy in ameliorating SM-induced lung injury. Anesthetized spontaneously breathing rats (N=6-8/group) were intratracheally intubated and exposed to 1.4 mg/kg SM (0.35 mg SM in 0.1 ml of ethanol) or ethanol alone by vapor inhalation for 50 min. At 1 min before the exposure rats were treated with one of the following: intravenous aprotinin, 4.4 mg/kg; intraperitoneal (ip) ilomastat, 25mg/kg; or ip trolox, 500 microg/kg. Aprotinin-treated animals received supplemental 2.2mg/kg doses at 1 min and 6h post-exposure (PE). A whole body plethysmograph system was used to monitor pulmonary function (PF) parameters for 1h before exposure (baseline), and from 5-6 and 23-24h post-exposure. SM inhalation caused significant increases in several PF parameters, including tidal volume, peak inspiratory flow, peak expiratory flow, end expiratory pause and enhanced pause. Consistent with the reported development of SM-induced pathology, these changes were minimal at the 5-6-h time and significant at the 23-24-h timepoint. At the later time it is known from previous work that airways are becoming obstructed with loose cellular debris, damaged cells and exudate, which contributed to the changes in PF parameters. Treatment with aprotinin or ilomastat eliminated these PF changes, yielding results comparable with controls for each of these parameters. Lung lavage fluid analysis showed that SM caused a significant increase in total protein (TP) and in the cytokines IL-1alpha and IL-13. Aprotinin treatment prevented the increases in TP and IL-1alpha production, ilomastat prevented the increased production of IL-13, and trolox treatment did not significantly prevent the SM-related increases in TP, IL-1alpha or IL-13. Histopathologic examination of lung tissue 24h post-exposure showed minimal alveolar effects caused by SM, while damage to bronchiolar regions was much more severe due to the highly reactive nature of SM. While aprotinin and ilomastat both alleviated the PF perturbations, surprisingly only aprotinin reduced the observed pathology, both grossly and histologically. These early results indicate that treatment with aprotinin and to a lesser extent ilomastat reduces some of the direct inflammatory response and damage associated with SM-induced lung injury. This research was supported by the Defense Threat Reduction Agency - Joint Science and Technology Office, Medical S&T Division. PMID:18852015

  10. Europium as an inhibitor of Amyloid-?(1-42) induced membrane permeation

    PubMed Central

    Williams, Thomas L.; Urbanc, Brigita; Marshall, Karen E.; Vadukul, Devkee M.; Jenkins, A. Toby A.; Serpell, Louise C.

    2015-01-01

    Soluble Amyloid-beta (A?) oligomers are a source of cytotoxicity in Alzheimer’s disease (AD). The toxicity of A? oligomers may arise from their ability to interact with and disrupt cellular membranes mediated by GM1 ganglioside receptors within these membranes. Therefore, inhibition of A?–membrane interactions could provide a means of preventing the toxicity associated with A?. Here, using Surface Plasmon field-enhanced Fluorescence Spectroscopy, we determine that the lanthanide, Europium III chloride (Eu3+), strongly binds to GM1 ganglioside-containing membranes and prevents the interaction with A?42 leading to a loss of the peptides ability to cause membrane permeation. Here we discuss the molecular mechanism by which Eu3+ inhibits A?42-membrane interactions and this may lead to protection of membrane integrity against A?42 induced toxicity. PMID:26450778

  11. Europium as an inhibitor of Amyloid-?(1-42) induced membrane permeation.

    PubMed

    Williams, Thomas L; Urbanc, Brigita; Marshall, Karen E; Vadukul, Devkee M; Jenkins, A Toby A; Serpell, Louise C

    2015-10-24

    Soluble Amyloid-beta (A?) oligomers are a source of cytotoxicity in Alzheimer's disease (AD). The toxicity of A? oligomers may arise from their ability to interact with and disrupt cellular membranes mediated by GM1 ganglioside receptors within these membranes. Therefore, inhibition of A?-membrane interactions could provide a means of preventing the toxicity associated with A?. Here, using Surface Plasmon field-enhanced Fluorescence Spectroscopy, we determine that the lanthanide, Europium III chloride (Eu(3+)), strongly binds to GM1 ganglioside-containing membranes and prevents the interaction with A?42 leading to a loss of the peptides ability to cause membrane permeation. Here we discuss the molecular mechanism by which Eu(3+) inhibits A?42-membrane interactions and this may lead to protection of membrane integrity against A?42 induced toxicity. PMID:26450778

  12. Multifunctional 8-hydroxyquinoline-appended cyclodextrins as new inhibitors of metal-induced protein aggregation.

    PubMed

    Oliveri, Valentina; Attanasio, Francesco; Puglisi, Antonino; Spencer, John; Sgarlata, Carmelo; Vecchio, Graziella

    2014-07-14

    Mounting evidence suggests a pivotal role of metal imbalances in protein misfolding and amyloid diseases. As such, metal ions represent a promising therapeutic target. In this context, the synthesis of chelators that also contain complementary functionalities to combat the multifactorial nature of neurodegenerative diseases is a highly topical issue. We report two new 8-hydroxyquinoline-appended cyclodextrins and highlight their multifunctional properties, including their Cu(II) and Zn(II) binding abilities, and capacity to act as antioxidants and metal-induced antiaggregants. In particular, the latter property has been applied in the development of an effective assay that exploits the formation of amyloid fibrils when ?-lactoglobulin?A is heated in the presence of metal ions. PMID:24863958

  13. Brief exposure of embryos to steroids or aromatase inhibitor induces sex reversal in Nile tilapia (Oreochromis niloticus).

    PubMed

    Gennotte, Vincent; Mafwila Kinkela, Patrick; Ulysse, Bernard; Akian Djétouan, Dieudonné; Bere Sompagnimdi, Frédéric; Tomson, Thomas; Mélard, Charles; Rougeot, Carole

    2015-01-01

    This study aimed to develop sex reversal procedures targeting the embryonic period as tools to study the early steps of sex differentiation in Nile tilapia with XX, XY, and YY sexual genotypes. XX eggs were exposed to masculinizing treatments with androgens (17?-methyltestosterone, 11-ketotestosterone) or aromatase inhibitor (Fadrozole), whereas XY and YY eggs were subjected to feminizing treatments with estrogen analog (17?-ethynylestradiol). All treatments consisted of a single or double 4-hr immersion applied between 1 and 36 hour post-fertilization (hpf). Concentrations of active substances were 1000 or 2000 ?g l(-1) in XX and XY, and 2000 or 6500 ?g l(-1) in YY. Masculinizing treatments of XX embryos achieved a maximal sex reversal rate of 10% with an exposure at 24 hpf to 1000 ?g l(-1) of 11-ketotestosterone or to 2000 ?g l(-1) of Fadrozole. Feminization of XY embryos was more efficient and induced up to 91% sex reversal with an exposure to 2000 ?g l(-1) of 17?-ethynylestradiol. Interestingly, similar treatments failed to reverse YY fish to females, suggesting either that a sex determinant linked to the Y chromosome prevents the female pathway when present in two copies, or that a gene present on the X chromosome is needed for the development of a female phenotype. PMID:25376842

  14. VEGF receptor inhibitors block the ability of metronomically dosed cyclophosphamide to activate innate immunity-induced tumor regression

    PubMed Central

    Doloff, Joshua C.; Waxman, David J.

    2012-01-01

    In metronomic chemotherapy, frequent drug administration at lower than maximally tolerated doses can improve activity while reducing the dose-limiting toxicity of conventional dosing schedules. Although the anti-tumor activity produced by metronomic chemotherapy is attributed widely to anti-angiogenesis, the significance of this mechanism remains somewhat unclear. In this study, we show that a 6-day repeating metronomic schedule of cyclophosphamide administration activates a potent antitumor immune response associated with brain tumor recruitment of natural killer (NK) cells, macrophages, and dendritic cells that leads to tumor regression. Tumor regression was blocked in NOD-scid-gamma mice, which are deficient or dysfunctional in all these immune cell types. Furthermore, regression was blunted by NK cell depletion in immune competent syngeneic mice or in perforin deficient mice, which are compromised for NK, NKT, and T cell cytolytic functions. Unexpectedly, we found that VEGF receptor inhibitors blocked both innate immune cell recruitment and the associated tumor regression response. Cyclophosphamide administered at a maximum-tolerated dose activated a transient, weak innate immune response, arguing that persistent drug-induced cytotoxic damage or associated cytokine and chemokine responses are required for effective innate immune-based tumor regression. Together, our results reveal an innate immune-based mechanism of tumor regression that can be activated by a traditional cytotoxic chemotherapy administered on a metronomic schedule. These findings suggest the need to carefully evaluate the clinical effects of combination chemotherapies that incorporate anti-angiogenesis drugs targeting VEGF receptor. PMID:22237627

  15. Histone deacetylase inhibitor suberoyl bis-hydroxamic acid suppresses cell proliferation and induces apoptosis in breast cancer cells.

    PubMed

    Yang, Xinmiao; Zhang, Ning; Shi, Zeliang; Yang, Zhangyu; Hu, Xichun

    2015-04-01

    Suberoyl bis?hydroxamic acid (SBHA) is a histone deacetylase inhibitor that has shown anticancer activity against numerous types of human cancer. The aim of the current study was to explore the effects of SBHA on the proliferation and apoptosis of breast cancer cells. MCF?7 breast cancer cells were treated with different concentrations of SBHA and tested for cell viability, apoptosis and gene expression changes. The results showed that SBHA significantly inhibited the proliferation of MCF?7 cells in a concentration?dependent manner, as determined using a Cell Counting kit?8 assay. SBHA?treated MCF?7 cells showed G0/G1 cell?cycle arrest, coupled with elevated expression levels of p21 and p27 proteins. Hoechst 33258 staining revealed cell shrinkage, chromosomal condensation and nuclear fragmentation in MCF?7 cells treated with SBHA. Flow cytometric analysis of Annexin V?stained cells showed that SBHA treatment induced apoptotic cell death in a concentration?dependent manner. Western blot analysis confirmed the upregulation of Bax and the downregulation of Bcl?2 by SBHA. In conclusion, these results indicate that SBHA exerts cytotoxic effects against human breast cancer cells, which involves the modulation of p21, p27 and Bcl?2 family proteins, consequently leading to cell?cycle arrest and apoptosis. PMID:25501628

  16. A Systems Pharmacology Model of Erythropoiesis in Mice Induced by Small Molecule Inhibitor of Prolyl Hydroxylase Enzymes

    PubMed Central

    Singh, I; Nagiec, EE; Thompson, JM; Krzyzanski, W; Singh, P

    2015-01-01

    Mammalian erythropoiesis is a conserved process tightly controlled by the hypoxia-inducible factor (HIF1) pathway. In this study, a small molecule inhibitor (PHI-1) of prolyl-hydroxylase-2 (PHD2) enzyme involved in regulating HIF1? levels was orally administered to male BALB/c mice at 10 and 30 mg/kg. A systems pharmacology model was developed based on the measured PHI-1 plasma exposures, kidney HIF1?, kidney erythropoietin (EPO) mRNA, plasma EPO, reticulocyte counts, red blood cells, and hemoglobin levels. The model fit resulted in the estimation of drug potency (IC50: 1.7?M), and systems parameters such as EPO mRNA turnover (kdeg_EPOmRNA: 0.43 hr-1) and mean lifespan of reticulocytes (Tr: 81 hours). The model correctly described the observed 30–40-fold increase in kidney HIF1? protein, ?1,000 fold increase in EPO mRNA and 2–3-fold increase in the reticulocytes at 30 mg/kg. This study presents the first parsimonious systems model of erythropoiesis to quantitatively describe the in vivo effects of PHD2 inhibition. PMID:26225228

  17. A Systems Pharmacology Model of Erythropoiesis in Mice Induced by Small Molecule Inhibitor of Prolyl Hydroxylase Enzymes.

    PubMed

    Singh, I; Nagiec, E E; Thompson, J M; Krzyzanski, W; Singh, P

    2015-02-01

    Mammalian erythropoiesis is a conserved process tightly controlled by the hypoxia-inducible factor (HIF1) pathway. In this study, a small molecule inhibitor (PHI-1) of prolyl-hydroxylase-2 (PHD2) enzyme involved in regulating HIF1? levels was orally administered to male BALB/c mice at 10 and 30 mg/kg. A systems pharmacology model was developed based on the measured PHI-1 plasma exposures, kidney HIF1?, kidney erythropoietin (EPO) mRNA, plasma EPO, reticulocyte counts, red blood cells, and hemoglobin levels. The model fit resulted in the estimation of drug potency (IC50: 1.7?M), and systems parameters such as EPO mRNA turnover (kdeg_EPOmRNA: 0.43 hr(-1)) and mean lifespan of reticulocytes (Tr : 81 hours). The model correctly described the observed 30-40-fold increase in kidney HIF1? protein, ?1,000 fold increase in EPO mRNA and 2-3-fold increase in the reticulocytes at 30 mg/kg. This study presents the first parsimonious systems model of erythropoiesis to quantitatively describe the in vivo effects of PHD2 inhibition. PMID:26225228

  18. Neuronal developmental gene and miRNA signatures induced by histone deacetylase inhibitors in human embryonic stem cells

    PubMed Central

    Meganathan, K; Jagtap, S; Srinivasan, S P; Wagh, V; Hescheler, J; Hengstler, J; Leist, M; Sachinidis, A

    2015-01-01

    Human embryonic stem cells (hESCs) may be applied to develop human-relevant sensitive in vitro test systems for monitoring developmental toxicants. The aim of this study was to identify potential developmental toxicity mechanisms of the histone deacetylase inhibitors (HDAC) valproic acid (VPA), suberoylanilide hydroxamic acid (SAHA) and trichostatin A (TSA) relevant to the in vivo condition using a hESC model in combination with specific differentiation protocols and genome-wide gene expression and microRNA profiling. Analysis of the gene expression data showed that VPA repressed neural tube and dorsal forebrain (OTX2, ISL1, EMX2 and SOX10)-related transcripts. In addition, VPA upregulates axonogenesis and ventral forebrain-associated genes, such as SLIT1, SEMA3A, DLX2/4 and GAD2. HDACi-induced expression of miR-378 and knockdown of miR-378 increases the expression of OTX2 and EMX2, which supports our hypothesis that HDACi targets forebrain markers through miR-378. In conclusion, multilineage differentiation in vitro test system is very sensitive for monitoring molecular activities relevant to in vivo neuronal developmental toxicity. Moreover, miR-378 seems to repress the expression of the OTX2 and EMX2 and therefore could be a regulator of the development of neural tube and dorsal forebrain neurons. PMID:25950486

  19. The Combined Use of Known Antiviral Reverse Transcriptase Inhibitors AZT and DDI Induce Anticancer Effects at Low Concentrations

    PubMed Central

    Aschacher, Thomas; Sampl, Sandra; Käser, Lisa; Bernhard, David; Spittler, Andreas; Holzmann, Klaus; Bergmann, Michael

    2012-01-01

    A hallmark of tumor cell survival is the maintenance of elongated telomeres. It is known that antiviral reverse transcriptase inhibitors (RTIs) such as azidothymidine (AZT) and didanosine (ddI) lead to telomere shortening at high, potentially toxic concentrations. We hypothesized that those drugs might have synergistic effects enabling successful therapy with low, nontoxic concentrations. Biologic effects of AZT and ddI were analyzed at concentrations that correspond to minimal plasma levels achieved during human immunodeficiency virus therapy. Long-term coapplication of low-dose AZT and ddI induced a significant shortening of telomeres in the tumor cell lines HCT-116, SkMel-28, MelJuso, and Jurkat. Treatment of cells with both RTI, but not with single RTI, led to a significant accumulation of ?H2AX, to p53 phosphorylation, and to cell apoptosis in all cell lines. Oral low-dose dual RTI application but not low-dose single RTI application was associated with a significantly reduced tumor growth of HCT-116 cells in mice. This antiproliferative activity of the combined use of AZT and ddI at low, clinically applicable concentrations warrants clinical testing in human solid cancer. PMID:22355273

  20. BA-j as a novel CDK1 inhibitor selectively induces apoptosis in cancer cells by regulating ROS

    PubMed Central

    Zhang, Shixuan; Bao, Yongming; Ju, Xiulan; Li, Kangjian; Shang, Haiyan; Ha, Lisha; Qian, Yuan; Zou, Liang; Sun, Xiaodan; Li, Jing; Wang, Qianru; Fan, Qingyu

    2015-01-01

    Cyclin-dependent kinase 1 (CDK1) is the only necessary CDK in cell proliferation and a novel target in the development of anticancer drugs. 8-Hydroxypiperidinemethyl-baicalein (BA-j) is a novel selective CDK1 inhibitor with broad spectrum anti-cancer activity (IC50 12.3??M) and 2 tumor xenografts. Because of the differential mechanisms controlling redox-states in normal and cancer cells, BA-j can capture oxygen free radicals (·O2?) and selectively increase the level of H2O2 in cancer cells, thereby specifically oxidize and activate the intrinsic apoptosis pathway bypassing the extrinsic death receptor pathway, thus inducing apoptosis in cancer cells rather than in normal cells. BA-j is different from cytotoxic anticancer drugs which can activate both the intrinsic apoptosis pathway and the extrinsic death receptor pathway, and therefore harm normal cells while killing cancer cells. The molecular and biochemical mechanisms of reactive oxygen species (ROS) regulation suggest that BA-j may be developed into a novel anticancer agent. PMID:26330167

  1. Kinesin spindle protein inhibitor SB743921 induces mitotic arrest and apoptosis and overcomes imatinib resistance of chronic myeloid leukemia cells.

    PubMed

    Yin, Yue; Sun, Huiyan; Xu, Jun; Xiao, Fengjun; Wang, Hua; Yang, Yuefeng; Ren, Hanyun; Wu, Chu-Tse; Gao, Chunji; Wang, Lisheng

    2015-06-01

    Inhibition of the cell mitotic pathway may provide a novel means for therapeutic intervention in chronic myeloid leukemia (CML). Kinesin spindle protein (KSP), a microtubule-associated motor protein which is essential for cell cycle progression, is overexpressed in bcr-abl+ CML cells. Retrovirus mediated bcr-abl transduction increases KSP expression in cord blood CD34 + cells. SB743921 is a selective KSP inhibitor which is being investigated in ongoing clinical trials for treatment of myeloma, leukemia and solid tumors. Treatment of CML cells with SB743921 resulted in reduced proliferation and colony forming cell (CFC) formation ability. SB743921 also actively blocked cell cycle progression, leading to apoptosis in both primary CML cells and cell lines. KSP inhibition sensitized CML cells to imatinib-induced apoptosis. Importantly, SB743921 inhibited the proliferation of various CML cells including T315I mutation-harboring cells. Furthermore, we demonstrated that SB743921 treatment suppressed ERK and AKT activity in CML cells. These data indicate that SB743921 may become a novel treatment agent for patients with CML. PMID:25146433

  2. Pharmacokinetics and pharmacodynamics of the dual FII/FX inhibitor BIBT 986 in endotoxin-induced coagulation.

    PubMed

    Leitner, J M; Jilma, B; Mayr, F B; Cardona, F; Spiel, A O; Firbas, C; Rathgen, K; Stähle, H; Schühly, U; Graefe-Mody, E U

    2007-06-01

    BIBT986 is a dual inhibitor of factors Xa and IIa. The aim of this study was to compare with placebo the effect of three doses of BIBT986 on coagulation, platelet activation, and inflammation. This was a prospective, randomized, double-blind, placebo-controlled, parallel-group dose escalation trial in 48 healthy male volunteers. Participants received one of three doses of BIBT986 or placebo intravenously together with a bolus infusion of 2 ng/kg lipopolysaccharide (LPS). BIBT986 dose-dependently changed global coagulation parameters and in vivo markers of thrombin generation and action: BIBT986 doses, which prolonged activated partial thromboplastin time by 100%, completely suppressed the LPS-induced increases in prothrombin fragment, thrombin-antithrombin complexes, and D-dimer, which were 6.1-, 14.5, and 3.5-fold in the placebo group, respectively. BIBT986 did not influence inflammation, fibrinolysis, or platelet activation. Therefore, BIBT986 is a potent anticoagulant in the human endotoxemia model. PMID:17410124

  3. Effect of the oral thrombin inhibitor dabigatran on allergic lung inflammation induced by repeated house dust mite administration in mice.

    PubMed

    de Boer, Johannes D; Berkhout, Lea C; de Stoppelaar, Sacha F; Yang, Jack; Ottenhoff, Roelof; Meijers, Joost C M; Roelofs, Joris J T H; Van't Veer, Cornelis; van der Poll, Tom

    2015-10-15

    Asthma is a chronic disease of the airways; asthma patients are hampered by recurrent symptoms of dyspnoea and wheezing caused by bronchial obstruction. Most asthma patients suffer from chronic allergic lung inflammation triggered by allergens such as house dust mite (HDM). Coagulation activation in the pulmonary compartment is currently recognized as a feature of allergic lung inflammation, and data suggest that coagulation proteases further drive inflammatory mechanisms. Here, we tested whether treatment with the oral thrombin inhibitor dabigatran attenuates allergic lung inflammation in a recently developed HDM-based murine asthma model. Mice were fed dabigatran (10 mg/g) or placebo chow during a 3-wk HDM airway exposure model. Dabigatran treatment caused systemic thrombin inhibitory activity corresponding with dabigatran levels reported in human trials. Surprisingly, dabigatran did not lead to inhibition of HDM-evoked coagulation activation in the lung as measured by levels of thrombin-antithrombin complexes and D-dimer. Repeated HDM administration caused an influx of eosinophils and neutrophils into the lungs, mucus production in the airways, and a T helper 2 response, as reflected by a rise in bronchoalveolar IL-4 and IL-5 levels and a systemic rise in IgE and HDM-IgG1. Dabigatran modestly improved HDM-induced lung pathology (P < 0.05) and decreased IL-4 levels (P < 0.01), without influencing other HDM-induced responses. Considering the limited effects of dabigatran in spite of adequate plasma levels, these results argue against clinical evaluation of dabigatran in patients with asthma. PMID:26320153

  4. The anti-inflammatory activity of duloxetine, a serotonin/norepinephrine reuptake inhibitor, prevents kainic acid-induced hippocampal neuronal death in mice.

    PubMed

    Choi, Hee-Soo; Park, Joon Ha; Ahn, Ji Hyeon; Hong, Seongkweon; Cho, Jun Hwi; Won, Moo-Ho; Lee, Choong-Hyun

    2015-11-15

    Duloxetine (DXT), a potent serotonin/norepinephrine reuptake inhibitor, is widely used in the treatment of major depressive disorder. In the present study, we examined the effects of DXT treatment on seizure behavior and excitotoxic neuronal damage in the mouse hippocampal CA3 region following intraperitoneal kainic acid (KA) injection. DXT treatment showed no effect on KA-induced behavioral seizure activity. However, treatment with 10mg/kg DXT reduced KA-induced neuronal death in the hippocampal CA3 region at 72h after KA administration, and treatment with 20 and 40mg/kg DXT showed a noticeable neuroprotection in the hippocampal CA3 region after KA injection. In addition, KA-induced activations of microglia and astrocytes as well as KA-induced increases of TNF-? and IL-1? levels were also suppressed by DXT treatment. These results indicate that DXT displays the neuroprotective effect against KA-induced excitotoxic neuronal death through anti-inflammatory action. PMID:26453128

  5. Ligand-dependent interactions of coactivators steroid receptor coactivator-1 and peroxisome proliferator-

    E-print Network

    Tsien, Roger Y.

    Ligand-dependent interactions of coactivators steroid receptor coactivator-1 and peroxisome demonstrate that both peroxisome proliferator-activated receptor binding protein (PBP) and steroid receptor ligand-dependent transcriptional activation. Nu- clear receptors that are regulated by steroid hormones

  6. Myelin peroxisomes - essential organelles for the maintenance of white matter in the nervous system.

    PubMed

    Kassmann, Celia M

    2014-03-01

    Peroxisomes are cellular compartments primarily associated with lipid metabolism. Most cell types, including nervous system cells, harbor several hundred of these organelles. The importance of peroxisomes for central nervous system white matter is evidenced by a variety of human peroxisomal disorders with neurological impairment frequently involving the white matter. Moreover, the most frequent childhood white matter disease, X-linked adrenoleukodystrophy, is a peroxisomal disorder. During the past decade advances in imaging techniques have enabled the identification of peroxisomes within the myelin sheath, especially close to nodes of Ranvier. Although the function of myelin peroxisomes is not solved yet on molecular level, recently acquired knowledge suggests a central role for these organelles in axo-glial metabolism. This review focuses on the biology of myelin peroxisomes as well as on the pathology of myelin and myelinated axons that is observed as a consequence of partial or complete peroxisomal dysfunction in the brain. PMID:24120688

  7. Activation of Peroxisome Proliferator-Activated Receptor (PPAR ) by Rosiglitazone Suppresses

    E-print Network

    Abraham, Nader G.

    Activation of Peroxisome Proliferator-Activated Receptor (PPAR ) by Rosiglitazone Suppresses) that are ligands for peroxisome proliferator-activated receptor (PPAR ). Stimulation of PPAR suppresses bone formation and enhances marrow adipogenesis. We hypothe- sized that activation of PPAR down

  8. Electrophilic peroxisome proliferator-activated receptor-gamma ligands have potent antifibrotic effects in human lung fibroblasts.

    PubMed

    Ferguson, Heather E; Kulkarni, Ajit; Lehmann, Geniece M; Garcia-Bates, Tatiana M; Thatcher, Thomas H; Huxlin, Krystel R; Phipps, Richard P; Sime, Patricia J

    2009-12-01

    Pulmonary fibrosis is a progressive scarring disease with no effective treatment. Transforming growth factor (TGF)-beta is up-regulated in fibrotic diseases, where it stimulates differentiation of fibroblasts to myofibroblasts and production of excess extracellular matrix. Peroxisome proliferator-activated receptor (PPAR) gamma is a transcription factor that regulates adipogenesis, insulin sensitization, and inflammation. We report here that a novel PPARgamma ligand, 2-cyano-3,12-dioxoolean-1,9-dien-28-oic acid (CDDO), is a potent inhibitor of TGF-beta-stimulated differentiation of human lung fibroblasts to myofibroblasts, and suppresses up-regulation of alpha-smooth muscle actin, fibronectin, collagen, and the novel myofibroblast marker, calponin. The inhibitory concentration causing a 50% decrease in aSMA for CDDO was 20-fold lower than the endogenous PPARgamma ligand, 15-deoxy-Delta(12,14)-prostaglandin J(2) (15 d-PGJ(2)), and 400-fold lower than the synthetic ligand, rosiglitazone. Pharmacologic and genetic approaches were used to demonstrate that CDDO mediates its activity via a PPARgamma-independent pathway. CDDO and 15 d-PGJ(2) contain an alpha/beta unsaturated ketone, which acts as an electrophilic center that can form covalent bonds with cellular proteins. Prostaglandin A(1) and diphenyl diselenide, both strong electrophiles, also inhibit myofibroblast differentiation, but a structural analog of 15 d-PGJ(2) lacking the electrophilic center is much less potent. CDDO does not alter TGF-beta-induced Smad or AP-1 signaling, but does inhibit acetylation of CREB binding protein/p300, a critical coactivator in the transcriptional regulation of TGF-beta-responsive genes. Overall, these data indicate that certain PPARgamma ligands, and other small molecules with electrophilic centers, are potent inhibitors of critical TGF-beta-mediated profibrogenic activities through pathways independent of PPARgamma. As the inhibitory concentration causing a 50% decrease in aSMA for CDDO is 400-fold lower than that in rosiglitazone, the translational potential of CDDO for treatment of fibrotic diseases is high. PMID:19286977

  9. Peroxisome proliferator-activated receptors gamma reverses hepatic nutritional fibrosis in mice and suppresses activation of hepatic stellate cells in vitro.

    PubMed

    Yu, Jun; Zhang, Sui; Chu, Eagle S H; Go, Minnie Y Y; Lau, Rebecca H Y; Zhao, Junhong; Wu, Chung-Wah; Tong, Lixin; Zhao, Jingmin; Poon, Terence C W; Sung, Joseph J Y

    2010-06-01

    Nonalcoholic steatohepatitis with fibrosis is a more severe form of nonalcoholic fatty liver disease, one of the most common liver diseases. We have previously shown that peroxisome proliferator-activated receptors gamma (PPARgamma) ligand, rosiglitazone, prevented the development of the methionine choline deficient (MCD) diet-induced fibrosing steatohepatitis. We have now tested whether overexpression of PPARgamma ameliorates established steatohepatitis and fibrosis. Male C57BL6 mice fed with MCD diet for 8 weeks developed hepatic fibrosis with increased hepatic expression of collagen1alpha(I), inhibitors of fibrosis reversal-1, regulator involved in matrix degradation-9 and connective tissue growth factor. After 2 weeks of transduction of PPARgamma through an adenovirus-expressing PPARgamma (Ad-PPARgamma), expression of these genes was reduced in a manner that paralleled the reduction in activated hepatic stellate cells (HSCs) and resolution of liver fibrosis. On the in vitro study, PPARgamma is expressed in primary quiescent HSC, but depleted in culture activated HSC. Conversely, ectopic expression of PPARgamma in activated HSC achieved the phenotypic reversal to the quiescent cell. Such induction markedly suppressed cell viability and cell proliferation, downregulated proliferating cell nuclear antigen, and caused cell cycle arrest at G0/G1 phase. Further, introduction of PPARgamma in HSC increased cell apoptosis, this was confirmed by enhanced expression of FasL, cleaved caspase-3, cleaved caspase-7 and poly ADP-ribose polymerase, indicating an extrinsic apoptosis pathway. In conclusion, the present study shows that MCD diet-induced fibrosing steatohepatitis can be reversed by overexpression of PPARgamma. It is likely that PPARgamma reverses fibrosis by reducing HSCs proliferation, inducing cell cycle arrest and apoptosis. PMID:20156580

  10. GW5074 and PP2 kinase inhibitors implicate nontraditional c-Raf and Lyn function as drivers of retinoic acid-induced maturation?

    PubMed Central

    Jensen, Holly A.; Bunaciu, Rodica P.; Varner, Jeffrey D.; Yen, Andrew

    2015-01-01

    The multivariate nature of cancer necessitates multi-targeted therapy, and kinase inhibitors account for a vast majority of approved cancer therapeutics. While acute promyelocytic leukemia (APL) patients are highly responsive to retinoic acid (RA) therapy, kinase inhibitors have been gaining momentum as co-treatments with RA for non-APL acute myeloid leukemia (AML) differentiation therapies, especially as a means to treat relapsed or refractory AML patients. In this study GW5074 (a c-Raf inhibitor) and PP2 (a Src-family kinase inhibitor) enhanced RA-induced maturation of t(15;17)-negative myeloblastic leukemia cells and rescued response in RA-resistant cells. PD98059 (a MEK inhibitor) and Akti-1/2 (an Akt inhibitor) were less effective, but did tend to promote maturation-uncoupled G1/G0 arrest, while wortmannin (a PI3K inhibitor) did not enhance differentiation surface marker expression or growth arrest. PD98059 and Akti-1/2 did not enhance differentiation markers and have potential, antagonistic off-targets effects on the aryl hydrocarbon receptor (AhR), but neither could the AhR agonist 6-formylindolo(3,2-b)carbazole (FICZ) rescue differentiation events in the RA-resistant cells. GW5074 rescued early CD38 expression in RA-resistant cells exhibiting an early block in differentiation before CD38 expression, while for RA-resistant cells with differentiation blocked later, PP2 rescued the later differentiation marker CD11b; but surprisingly, the combination of the two was not synergistic. Kinases c-Raf, Src-family kinases Lyn and Fgr, and PI3K display highly correlated signaling changes during RA treatment, while activation of traditional downstream targets (Akt, MEK/ERK), and even the surface marker CD38, were poorly correlated with c-Raf or Lyn during differentiation. This suggests that an interrelated kinase module involving c-Raf, PI3K, Lyn and perhaps Fgr functions in a nontraditional way during RA-induced maturation or during rescue of RA induction therapy using inhibitor co-treatment in RA-resistant leukemia cells. PMID:25817574

  11. GW5074 and PP2 kinase inhibitors implicate nontraditional c-Raf and Lyn function as drivers of retinoic acid-induced maturation.

    PubMed

    Jensen, Holly A; Bunaciu, Rodica P; Varner, Jeffrey D; Yen, Andrew

    2015-08-01

    The multivariate nature of cancer necessitates multi-targeted therapy, and kinase inhibitors account for a vast majority of approved cancer therapeutics. While acute promyelocytic leukemia (APL) patients are highly responsive to retinoic acid (RA) therapy, kinase inhibitors have been gaining momentum as co-treatments with RA for non-APL acute myeloid leukemia (AML) differentiation therapies, especially as a means to treat relapsed or refractory AML patients. In this study GW5074 (a c-Raf inhibitor) and PP2 (a Src-family kinase inhibitor) enhanced RA-induced maturation of t(15;17)-negative myeloblastic leukemia cells and rescued response in RA-resistant cells. PD98059 (a MEK inhibitor) and Akti-1/2 (an Akt inhibitor) were less effective, but did tend to promote maturation-uncoupled G1/G0 arrest, while wortmannin (a PI3K inhibitor) did not enhance differentiation surface marker expression or growth arrest. PD98059 and Akti-1/2 did not enhance differentiation markers and have potential, antagonistic off-targets effects on the aryl hydrocarbon receptor (AhR), but neither could the AhR agonist 6-formylindolo(3,2-b)carbazole (FICZ) rescue differentiation events in the RA-resistant cells. GW5074 rescued early CD38 expression in RA-resistant cells exhibiting an early block in differentiation before CD38 expression, while for RA-resistant cells with differentiation blocked later, PP2 rescued the later differentiation marker CD11b; but surprisingly, the combination of the two was not synergistic. Kinases c-Raf, Src-family kinases Lyn and Fgr, and PI3K display highly correlated signaling changes during RA treatment, while activation of traditional downstream targets (Akt, MEK/ERK), and even the surface marker CD38, were poorly correlated with c-Raf or Lyn during differentiation. This suggests that an interrelated kinase module involving c-Raf, PI3K, Lyn and perhaps Fgr functions in a nontraditional way during RA-induced maturation or during rescue of RA induction therapy using inhibitor co-treatment in RA-resistant leukemia cells. PMID:25817574

  12. Twist1 Is a TNF-Inducible Inhibitor of Clock Mediated Activation of Period Genes

    PubMed Central

    Meier, Daniel; Lopez, Martin; Franken, Paul; Fontana, Adriano

    2015-01-01

    Background Activation of the immune system affects the circadian clock. Tumor necrosis factor (TNF) and Interleukin (IL)-1? inhibit the expression of clock genes including Period (Per) genes and the PAR-bZip clock-controlled gene D-site albumin promoter-binding protein (Dbp). These effects are due to cytokine-induced interference of E-box mediated transcription of clock genes. In the present study we have assessed the two E-box binding transcriptional regulators Twist1 and Twist2 for their role in cytokine induced inhibition of clock genes. Methods The expression of the clock genes Per1, Per2, Per3 and of Dbp was assessed in NIH-3T3 mouse fibroblasts and the mouse hippocampal neuronal cell line HT22. Cells were treated for 4h with TNF and IL-1?. The functional role of Twist1 and Twist2 was assessed by siRNAs against the Twist genes and by overexpression of TWIST proteins. In luciferase (luc) assays NIH-3T3 cells were transfected with reporter gene constructs, which contain a 3xPer1 E-box or a Dbp E-box. Quantitative chromatin immunoprecipitation (ChIP) was performed using antibodies to TWIST1 and CLOCK, and the E-box consensus sequences of Dbp (CATGTG) and Per1 E-box (CACGTG). Results We report here that siRNA against Twist1 protects NIH-3T3 cells and HT22 cells from down-regulation of Period and Dbp by TNF and IL-1?. Overexpression of Twist1, but not of Twist2, mimics the effect of the cytokines. TNF down-regulates the activation of Per1-3xE-box-luc, the effect being prevented by siRNA against Twist1. Overexpression of Twist1, but not of Twist2, inhibits Per1-3xE-box-luc or Dbp-E-Box-luc activity. ChIP experiments show TWIST1 induction by TNF to compete with CLOCK binding to the E-box of Period genes and Dbp. Conclusion Twist1 plays a pivotal role in the TNF mediated suppression of E-box dependent transactivation of Period genes and Dbp. Thereby Twist1 may provide a link between the immune system and the circadian timing system. PMID:26361389

  13. Mitigation of indomethacin-induced gastric mucosal lesions by a potent specific type V phosphodiesterase inhibitor

    PubMed Central

    Karakaya, Kemal; Hanci, Volkan; Bektas, Sibel; Can, Murat; Ucan, Hamdi B; Emre, Ali Ugur; Tasc?lar, Oge; Turan, Is?l Ozkocak; Comert, Mustafa; Irkorucu, Oktay; Cakmak, Guldeniz Karadeniz

    2009-01-01

    AIM: To investigate the gastroprotective effect of vardenafil against indomethacin-induced gastric damage. METHODS: Forty-eight female Wistar albino rats were randomly divided into 6 groups. Group 1 received saline only. Group 2 (indomethacin) received indomethacin. Rats in group 3 and 4 were pretreated with different doses of famotidine. Group 5 and 6 were pretreated with different doses of vardenafil. Rats in groups 3 to 6 received 25 mg/kg indomethacin 30 min after pretreatment. The animals were sacrificed 6 h later and their stomachs were opened. Gastric lesions were counted and measured. The stomach of each animal was divided in two parts for histopathological examinations and nitric oxide (NO) and malondialdehyde (MDA) assays, respectively. RESULTS: There were no gastric mucosal lesion in the saline group but all rats in the indomethacin group had gastric mucosal ulcerations (ulcer count; 6.25 ± 3.49, and mean ulcer area; 21.00 ± 12.35). Ulcer counts were diminished with famotidine 5 mg/kg (4.12 ± 2.47, P > 0.05), 20 mg/kg (2.37 ± 4.43, P < 0.05), vardenafil 2 mg/kg (4.37 ± 3.06), and vardenafil 10 mgkg (1.25 ± 1.38, P < 0.05) compared to the indomethacin group. Gastric mucosal lesion areas were diminished with famotidine 5 mg/kg (8.62 ± 2.97, P < 0.001) , famotidine 20 mg/kg (0.94 ± 2.06, P < 0.001), vardenafil 2 mg/kg (6.62 ± 5.87, P < 0.001), and vardenafil 10 mg/kg (0.75 ± 0.88, P < 0.001) compared to the indomethacin group. MDA levels were significantly higher in indomethacin group (28.48 ± 14.51), compared to the famotidine 5 mg/kg (6,21 ± 1.88, P < 0.05), famotidine 20 mg/kg (5.88 ± 1.60. P < 0.05), vardenafil 2 mg/kg (15.87 ± 3.93, P < 0.05), and vardenafil 10 mg/kg (10.97 ± 4.50, P < 0.05). NO concentration in gastric tissues of the famotidine groups were significantly increased (P < 0.05), but the NO increases in the vardenafil groups were not statistically significant. Histopathology revealed diminished gastric damage for pretreatment groups compared to the indomethacin group (P < 0.05). CONCLUSION: Vardenafil affords a significant dose-dependent protection against indomethacin induced gastric mucosal lesions in rats. PMID:19860004

  14. Cytokine-Induced Loss of Glucocorticoid Function: Effect of Kinase Inhibitors, Long-Acting ?2-Adrenoceptor Agonist and Glucocorticoid Receptor Ligands

    PubMed Central

    Rider, Christopher F.; Shah, Suharsh; Miller-Larsson, Anna; Giembycz, Mark A.; Newton, Robert

    2015-01-01

    Acting on the glucocorticoid receptor (NR3C1), glucocorticoids are widely used to treat inflammatory diseases. However, glucocorticoid resistance often leads to suboptimal asthma control. Since glucocorticoid-induced gene expression contributes to glucocorticoid activity, the aim of this study was to use a 2×glucocorticoid response element (GRE) reporter and glucocorticoid-induced gene expression to investigate approaches to combat cytokine-induced glucocorticoid resistance. Pre-treatment with tumor necrosis factor-? (TNF) or interleukin-1? inhibited dexamethasone-induced mRNA expression of the putative anti-inflammatory genes RGS2 and TSC22D3, or just TSC22D3, in primary human airway epithelial and smooth muscle cells, respectively. Dexamethasone-induced DUSP1 mRNA was unaffected. In human bronchial epithelial BEAS-2B cells, dexamethasone-induced TSC22D3 and CDKN1C expression (at 6 h) was reduced by TNF pre-treatment, whereas DUSP1 and RGS2 mRNAs were unaffected. TNF pre-treatment also reduced dexamethasone-dependent 2×GRE reporter activation. This was partially reversed by PS-1145 and c-jun N-terminal kinase (JNK) inhibitor VIII, inhibitors of IKK2 and JNK, respectively. However, neither inhibitor affected TNF-dependent loss of dexamethasone-induced CDKN1C or TSC22D3 mRNA. Similarly, inhibitors of the extracellular signal-regulated kinase, p38, phosphoinositide 3-kinase or protein kinase C pathways failed to attenuate TNF-dependent repression of the 2×GRE reporter. Fluticasone furoate, fluticasone propionate and budesonide were full agonists relative to dexamethasone, while GSK9027, RU24858, des-ciclesonide and GW870086X were partial agonists on the 2×GRE reporter. TNF reduced reporter activity in proportion with agonist efficacy. Full and partial agonists showed various degrees of agonism on RGS2 and TSC22D3 expression, but were equally effective at inducing CDKN1C and DUSP1, and did not affect the repression of CDKN1C or TSC22D3 expression by TNF. Finally, formoterol-enhanced 2×GRE reporter activity was also proportional to agonist efficacy and functionally reversed repression by TNF. As similar effects were apparent on glucocorticoid-induced gene expression, the most effective strategy to overcome glucocorticoid resistance in this model was addition of formoterol to high efficacy NR3C1 agonists. PMID:25625944

  15. A cytosolic thioredoxin acts as a molecular chaperone for peroxisome matrix proteins as well as antioxidant in peroxisome.

    PubMed

    Du, Hui; Kim, Sunghan; Hur, Yoon-Sun; Lee, Myung-Sok; Lee, Suk-Ha; Cheon, Choong-Ill

    2015-01-01

    Thioredoxin (TRX) is a disulfide reductase present ubiquitously in all taxa and plays an important role as a regulator of cellular redox state. Recently, a redox-independent, chaperone function has also been reported for some thioredoxins. We previously identified nodulin-35, the subunit of soybean uricase, as an interacting target of a cytosolic soybean thioredoxin, GmTRX. Here we report the further characterization of the interaction, which turns out to be independent of the disulfide reductase function and results in the co-localization of GmTRX and nodulin-35 in peroxisomes, suggesting a possible function of GmTRX in peroxisomes. In addition, the chaperone function of GmTRX was demonstrated in in vitro molecular chaperone activity assays including the thermal denaturation assay and malate dehydrogenase aggregation assay. Our results demonstrate that the target of GmTRX is not only confined to the nodulin-35, but many other peroxisomal proteins, including catalase (AtCAT), transthyretin-like protein 1 (AtTTL1), and acyl-coenzyme A oxidase 4 (AtACX4), also interact with the GmTRX. Together with an increased uricase activity of nodulin-35 and reduced ROS accumulation observed in the presence of GmTRX in our results, especially under heat shock and oxidative stress conditions, it appears that GmTRX represents a novel thioredoxin that is co-localized to the peroxisomes, possibly providing functional integrity to peroxisomal proteins. PMID:26013260

  16. Angiogenesis Inhibitors

    MedlinePLUS

    ... inhibitors currently being used to treat cancer in humans? How are angiogenesis inhibitors different from conventional anticancer ... inhibitors currently being used to treat cancer in humans? Yes. The U.S. Food and Drug Administration (FDA) ...

  17. Permeability of the peroxisomal membrane: lessons from the glyoxylate cycle

    PubMed Central

    Kunze, Markus; Hartig, Andreas

    2013-01-01

    Glyoxylate serves as intermediate in various metabolic pathways, although high concentrations of this metabolite are toxic to the cell. In many organisms glyoxylate is fed into the glyoxylate cycle. Enzymes participating in this metabolism are located on both sides of the peroxisomal membrane. The permeability of this membrane for small metabolites paves the way for exchange of intermediates between proteins catalyzing consecutive reactions. A model, in which soluble enzymes accumulate in close proximity to both ends of pore-like structures forming a transmembrane metabolon could explain the rapid and targeted exchange of intermediates. The metabolites passing the membrane differ between the three model organisms Saccharomyces cerevisiae, Arabidopsis thaliana, and Candida albicans, which reflects the ease of evolutionary adaptation processes whenever specific transporter proteins are not involved. The atypical permeability properties of the peroxisomal membrane together with a flexible structural arrangement ensuring the swift and selective transport across the membrane might represent the molecular basis for the functional versatility of peroxisomes. PMID:23966945

  18. [Peroxisomal ABC transporters and X-linked adrenoleukodystrophy].

    PubMed

    Geillon, Flore; Trompier, Doriane; Gondcaille, Catherine; Lizard, Gérard; Savary, Stéphane

    2012-12-01

    X-linked adrenoleukodystrophy (X-ALD) is a complex neurodegenerative disease associated with mutations in the ABCD1 gene, which encodes for a peroxisomal ABC transporter. Thanks to the efforts of the ELA foundation and to the recent successes of gene therapy published in Science in 2009, X-ALD is better known but still remains poorly understood. The exact role of ABCD1 and its homologs, as well as the exact link between the biochemical and metabolic peroxisomal defects and the clinical symptoms of the disease remain to be elucidated. This review summarizes the knowledge concerning the subfamily D of the ABC transporter family and concerning X-ALD, the most frequent peroxisomal disorder. PMID:23290409

  19. Targeting interleukin-2-inducible T-cell kinase (ITK) and resting lymphocyte kinase (RLK) using a novel covalent inhibitor PRN694.

    PubMed

    Zhong, Yiming; Dong, Shuai; Strattan, Ethan; Ren, Li; Butchar, Jonathan P; Thornton, Kelsey; Mishra, Anjali; Porcu, Pierluigi; Bradshaw, J Michael; Bisconte, Angelina; Owens, Timothy D; Verner, Erik; Brameld, Ken A; Funk, Jens Oliver; Hill, Ronald J; Johnson, Amy J; Dubovsky, Jason A

    2015-03-01

    Interleukin-2-inducible T-cell kinase (ITK) and resting lymphocyte kinase (RLK or TXK) are essential mediators of intracellular signaling in both normal and neoplastic T-cells and natural killer (NK) cells. Thus, ITK and RLK inhibitors have therapeutic potential in a number of human autoimmune, inflammatory, and malignant diseases. Here we describe a novel ITK/RLK inhibitor, PRN694, which covalently binds to cysteine residues 442 of ITK and 350 of RLK and blocks kinase activity. Molecular modeling was utilized to design molecules that interact with cysteine while binding to the ATP binding site in the kinase domain. PRN694 exhibits extended target residence time on ITK and RLK and is highly selective for a subset of the TEC kinase family. In vitro cellular assays confirm that PRN694 prevents T-cell receptor- and Fc receptor-induced cellular and molecular activation, inhibits T-cell receptor-induced T-cell proliferation, and blocks proinflammatory cytokine release as well as activation of Th17 cells. Ex vivo assays demonstrate inhibitory activity against T-cell prolymphocytic leukemia cells, and in vivo assays demonstrate durable pharmacodynamic effects on ITK, which reduces an oxazolone-induced delayed type hypersensitivity reaction. These data indicate that PRN694 is a highly selective and potent covalent inhibitor of ITK and RLK, and its extended target residence time enables durable attenuation of effector cells in vitro and in vivo. The results from this study highlight potential applications of this dual inhibitor for the treatment of T-cell- or NK cell-mediated inflammatory, autoimmune, and malignant diseases. PMID:25593320

  20. Human catalase is imported and assembled in peroxisomes of Saccharomyces cerevisiae.

    PubMed

    de Hoop, M J; Holtman, W L; Ab, G

    1993-01-01

    To study the conservation of peroxisomal targeting signals, we have determined the intracellular localization of human peroxisomal catalase when expressed in yeast. Using immunofluorescence, differential centrifugation and immunoelectron microscopy, we show that the protein is targeted to the peroxisomes of the heterologous cell and assembled in its active tetrameric form. These data show the conservation of the catalase targeting signal and import specificity between human and yeast peroxisomes. PMID:8442388

  1. Violent death in a rare peroxisomal disease-Zellweger syndrome.

    PubMed

    Malinescu, Bogdan; Martius, Eliza; Pelin, Ana Maria

    2015-10-01

    Peroxisomal diseases are rare (1:50,000), genetically determined disorders (autosomal recessive), systemic, multiorgan illnesses with prominent involvement of the nervous system, caused either by the failure to form or to maintain the peroxisome, or by a defect in the function of a single or multiple peroxisomal enzymes [2]. Peroxisomes contain approximately 50 enzymes which are responsible for many metabolic reactions, and play an important role in the oxidation of saturated very-long-chain fatty acids (VLCFA) [1]. The authors present the case of a Romanian boy, who died at the age of 1.6 of one of the peroxisomal diseases-Zellweger syndrome. Newborn infants with Zellweger syndrome have a typical dysmorphic facies, neonatal seizures, profound hypotonia, and eye abnormalities. Major abnormalities are present in the liver (fibrotic), kidney (cortical cysts), and brain (lipid-laden macrophages and histiocytes in cortical and periventricular areas, demyelination, centrosylvian polymicrogyria and pachygyria)-cerebro-hepato-renal syndrome (CHRS) (Zellweger) [2]. Infants with Zellweger syndrome rarely live more than a few months [10], but in this case the survival was longer, and the cause of death was not directly the peroxisomal disease but a violent cause of death-mechanical asphyxia with tracheo-bronchial food aspiration. The authors present the results of investigations carried out during the child's life, but also data collected at the autopsy and hystopathological postnecroptic investigations. By presenting this case, the authors wish to bring to your attention a rare pathology in forensic practice by the paradox of finding a common violent cause of death, asphyxia with food aspiration, in a rare metabolic-genetic disease, which is usually fatal by itself. PMID:26235911

  2. The pan erbB inhibitor PD168393 enhances lysosomal dysfunction-induced apoptotic death in malignant peripheral nerve sheath tumor cells

    PubMed Central

    Kohli, Latika; Kaza, Niroop; Lavalley, Nicholas J.; Turner, Kathryn L.; Byer, Stephanie; Carroll, Steven L.; Roth, Kevin A.

    2012-01-01

    Malignant peripheral nerve sheath tumors (MPNSTs) are rapidly progressive Schwann cell neoplasms. The erbB family of membrane tyrosine kinases has been implicated in MPNST mitogenesis and invasion and, thus, is a potential therapeutic target. However, tyrosine kinase inhibitors (TKIs) used alone have limited tumoricidal activity. Manipulating the autophagy lysosomal pathway in cells treated with cytostatic agents can promote apoptotic cell death in some cases. The goal of this study was to establish a mechanistic basis for formulating drug combinations to effectively trigger death in MPNST cells. We assessed the effects of the pan erbB inhibitor PD168393 on MPNST cell survival, caspase activation, and autophagy. PD168393 induced a cytostatic but not a cytotoxic response in MPNST cells that was accompanied by suppression of Akt and mTOR activation and increased autophagic activity. The effects of autophagy modulation on MPNST survival were then assessed following the induction of chloroquine (CQ)–induced lysosomal stress. In CQ-treated cells, suppression of autophagy was accompanied by increased caspase activation. In contrast, increased autophagy induction by inhibition of mTOR did not trigger cytotoxicity, possibly because of Akt activation. We thus hypothesized that dual targeting of mTOR and Akt by PD168393 would significantly increase cytotoxicity in cells exposed to lysosomal stress. We found that PD168393 and CQ in combination significantly increased cytotoxicity. We conclude that combinatorial therapies with erbB inhibitors and agents inducing lysosomal dysfunction may be an effective means of treating MPNSTs. PMID:22259051

  3. Estrogen receptor ? inhibitor activates the unfolded protein response, blocks protein synthesis, and induces tumor regression.

    PubMed

    Andruska, Neal D; Zheng, Xiaobin; Yang, Xujuan; Mao, Chengjian; Cherian, Mathew M; Mahapatra, Lily; Helferich, William G; Shapiro, David J

    2015-04-14

    Recurrent estrogen receptor ? (ER?)-positive breast and ovarian cancers are often therapy resistant. Using screening and functional validation, we identified BHPI, a potent noncompetitive small molecule ER? biomodulator that selectively blocks proliferation of drug-resistant ER?-positive breast and ovarian cancer cells. In a mouse xenograft model of breast cancer, BHPI induced rapid and substantial tumor regression. Whereas BHPI potently inhibits nuclear estrogen-ER?-regulated gene expression, BHPI is effective because it elicits sustained ER?-dependent activation of the endoplasmic reticulum (EnR) stress sensor, the unfolded protein response (UPR), and persistent inhibition of protein synthesis. BHPI distorts a newly described action of estrogen-ER?: mild and transient UPR activation. In contrast, BHPI elicits massive and sustained UPR activation, converting the UPR from protective to toxic. In ER?(+) cancer cells, BHPI rapidly hyperactivates plasma membrane PLC?, generating inositol 1,4,5-triphosphate (IP3), which opens EnR IP3R calcium channels, rapidly depleting EnR Ca(2+) stores. This leads to activation of all three arms of the UPR. Activation of the PERK arm stimulates phosphorylation of eukaryotic initiation factor 2? (eIF2?), resulting in rapid inhibition of protein synthesis. The cell attempts to restore EnR Ca(2+) levels, but the open EnR IP3R calcium channel leads to an ATP-depleting futile cycle, resulting in activation of the energy sensor AMP-activated protein kinase and phosphorylation of eukaryotic elongation factor 2 (eEF2). eEF2 phosphorylation inhibits protein synthesis at a second site. BHPI's novel mode of action, high potency, and effectiveness in therapy-resistant tumor cells make it an exceptional candidate for further mechanistic and therapeutic exploration. PMID:25825714

  4. Rho-Associated Kinase Inhibitor (Y-27632) Attenuates Doxorubicin-Induced Apoptosis of Human Cardiac Stem Cells

    PubMed Central

    Kan, Lijuan; Smith, Aubrie; Chen, Miao; Ledford, Benjamin T.; Fan, Huimin; Liu, Zhongmin; He, Jia-Qiang

    2015-01-01

    Background Recent clinical trials using c-kit+ human cardiac stem cells (CSCs) demonstrated promising results in increasing cardiac function and improving quality of life. However, CSC efficiency is low, likely due to limited cell survival and engraftment after transplantation. The Rho-associated protein kinase (ROCK) inhibitor, Y-27632, significantly increased cell survival rate, adhesion, and migration in numerous types of cells, including stem cells, suggesting a common feature of the ROCK-mediated apoptotic pathway that may also exist in human CSCs. In this study, we examine the hypothesis that pretreatment of human CSCs with Y-27632 protects cells from Doxorubicin (Dox) induced apoptosis. Methods and Results c-kit+ CSCs were cultured in CSC medium for 3–5 days followed by 48hr treatment with 0 to 10?M Y-27632 alone, 0 to 1.0?M Dox alone, or Y-27632 followed by Dox (48hrs). Cell viability, toxicity, proliferation, morphology, migration, Caspase-3 activity, expression levels of apoptotic-related key proteins and c-kit+ were examined. Results showed that 48hr treatment with Y-27632 alone did not result in great changes in c-kit+ expression, proliferation, Caspase-3 activity, or apoptosis; however cell viability was significantly increased and cell migration was promoted. These effects likely involve the ROCK/Actin pathways. In contrast, 48hr treatment with Dox alone dramatically increased Caspase-3 activity, resulting in cell death. Although Y-27632 alone did not affect the expression levels of apoptotic-related key factors (p-Akt, Akt, Bcl-2, Bcl-xl, Bax, cleaved Caspase-3, and Caspase-3) under basal conditions, it significantly inhibited the Dox-induced increase in cleaved Caspase-3 and reduced cell death under Dox treatment. Conclusions We conclude that preconditioning human CSCs with Y-27632 significantly reduces Dox-induced cell death and possibly involves the cleaved Caspase-3 and ROCK/Actin pathways. The beneficial effects of Y-27632 may be applied to stem cell-based therapy to increase cell survival rates after transplantation or to act as a cardiac protective agent for Dox-treated cancer patients. PMID:26645568

  5. SD-208, a Novel Protein Kinase D Inhibitor, Blocks Prostate Cancer Cell Proliferation and Tumor Growth In Vivo by Inducing G2/M Cell Cycle Arrest

    PubMed Central

    Tandon, Manuj; Salamoun, Joseph M.; Carder, Evan J.; Farber, Elisa; Xu, Shuping; Deng, Fan; Tang, Hua; Wipf, Peter; Wang, Q. Jane

    2015-01-01

    Protein kinase D (PKD) has been implicated in many aspects of tumorigenesis and progression, and is an emerging molecular target for the development of anticancer therapy. Despite recent advancement in the development of potent and selective PKD small molecule inhibitors, the availability of in vivo active PKD inhibitors remains sparse. In this study, we describe the discovery of a novel PKD small molecule inhibitor, SD-208, from a targeted kinase inhibitor library screen, and the synthesis of a series of analogs to probe the structure-activity relationship (SAR) vs. PKD1. SD-208 displayed a narrow SAR profile, was an ATP-competitive pan-PKD inhibitor with low nanomolar potency and was cell active. Targeted inhibition of PKD by SD-208 resulted in potent inhibition of cell proliferation, an effect that could be reversed by overexpressed PKD1 or PKD3. SD-208 also blocked prostate cancer cell survival and invasion, and arrested cells in the G2/M phase of the cell cycle. Mechanistically, SD-208-induced G2/M arrest was accompanied by an increase in levels of p21 in DU145 and PC3 cells as well as elevated phosphorylation of Cdc2 and Cdc25C in DU145 cells. Most importantly, SD-208 given orally for 24 days significantly abrogated the growth of PC3 subcutaneous tumor xenografts in nude mice, which was accompanied by reduced proliferation and increased apoptosis and decreased expression of PKD biomarkers including survivin and Bcl-xL. Our study has identified SD-208 as a novel efficacious PKD small molecule inhibitor, demonstrating the therapeutic potential of targeted inhibition of PKD for prostate cancer treatment. PMID:25747583

  6. Combination chemoprevention of HER2/neu-induced breast cancer using a COX-2 inhibitor and an RXR-selective retinoid

    PubMed Central

    Brown, Powel H.; Subbaramaiah, Kotha; Salmon, Amoi P.; Baker, Rebecca; Newman, Robert A.; Yang, Peiying; Zhou, Xi Kathy; Bissonnette, Reid P.; Dannenberg, Andrew J.; Howe, Louise R.

    2008-01-01

    The inducible prostaglandin synthase isoform cyclooxygenase-2 (COX-2) is overexpressed in ~40% of human breast carcinomas, and in precancerous breast lesions, particularly in association with overexpression of human epidermal growth factor receptor 2 (HER2/neu). Experimental breast cancer can be suppressed by pharmacological inhibition or genetic ablation of Cox-2, suggesting potential clinical utility of COX-2 inhibitors with respect to breast cancer. Importantly, several clinical trials have found reduced colorectal adenoma formation in individuals administered selective COX-2 inhibitors. However, such trials also identified increased cardiovascular risk associated with COX-2 inhibitor use. The goal of this research was to test whether improved chemopreventive efficacy could be achieved by combining submaximal doses of a selective COX-2 inhibitor and a retinoid X receptor-selective retinoid (rexinoid). The rate of HER2/neu-induced mammary tumor formation was substantially delayed by coadministration of the COX-2 inhibitor celecoxib (500 ppm in diet) and the rexinoid LGD1069 (10 mg/kg body weight; oral gavage) to MMTV/neu mice. Median time to tumor formation was increased from 304 days to >600 days (P<0.0001). The combination was substantially more effective than either drug individually. Similarly, potent suppression of aromatase activity was observed in mammary tissues from the combination cohort (44% of control; P<0.001). Regulation of aromatase expression and activity by COX-derived prostaglandins is well established. Interestingly however, single agent LGD1069 significantly reduced mammary aromatase activity (71% of control; P<0.001) without modulating eicosanoid levels. Our data demonstrate that simultaneous blockade of COX/prostaglandin signaling and retinoid X receptor-dependent transcription confers potent anticancer efficacy, suggesting a novel avenue for clinical evaluation. PMID:19138958

  7. ortho-Carboranylphenoxyacetanilides as inhibitors of hypoxia-inducible factor (HIF)-1 transcriptional activity and heat shock protein (HSP) 60 chaperon activity.

    PubMed

    Li, Guangzhe; Azuma, Soyoko; Sato, Shinichi; Minegishi, Hidemitsu; Nakamura, Hiroyuki

    2015-07-01

    ortho-Carboranylphenoxy derivatives were synthesized and evaluated for their ability to inhibit hypoxia-induced HIF-1 transcriptional activity using a cell-based reporter gene assay. Among the compounds synthesized, compound 1d showed the most significant inhibition of hypoxia-induced HIF-1 transcriptional activity with the IC50 of 0.53?M. Furthermore, compound 1h was found to possess the most significant inhibition of heat shock protein (HSP) 60 chaperon activity among the reported inhibitors: the IC50 toward the porcine heart malate dehydrogenase (MDH) refolding assay was 0.35?M. PMID:25981686

  8. Placotylene A, an Inhibitor of the Receptor Activator of Nuclear Factor-?B Ligand-Induced Osteoclast Differentiation, from a Korean Sponge Placospongia sp.

    PubMed Central

    Kim, Hiyoung; Kim, Kwang-Jin; Yeon, Jeong-Tae; Kim, Seong Hwan; Won, Dong Hwan; Choi, Hyukjae; Nam, Sang-Jip; Son, Young-Jin; Kang, Heonjoong

    2014-01-01

    A new inhibitor, placotylene A (1), of the receptor activator of nuclear factor-?B ligand (RANKL)-induced osteoclast differentiation, and a regioisomer of placotylene A, placotylene B (2), were isolated from a Korean marine sponge Placospongia sp. The chemical structures of placotylenes A and B were elucidated on the basis of 1D and 2D NMR, along with MS spectral analysis and revealed as an iodinated polyacetylene class of natural products. Placotylene A (1) displayed inhibitory activity against RANKL-induced osteoclast differentiation at 10 ?M while placotylene B (2) did not show any significant activity up to 100 ?M, respectively. PMID:24705502

  9. Placotylene A, an inhibitor of the receptor activator of nuclear factor-?B ligand-induced osteoclast differentiation, from a Korean sponge Placospongia sp.

    PubMed

    Kim, Hiyoung; Kim, Kwang-Jin; Yeon, Jeong-Tae; Kim, Seong Hwan; Won, Dong Hwan; Choi, Hyukjae; Nam, Sang-Jip; Son, Young-Jin; Kang, Heonjoong

    2014-04-01

    A new inhibitor, placotylene A (1), of the receptor activator of nuclear factor-?B ligand (RANKL)-induced osteoclast differentiation, and a regioisomer of placotylene A, placotylene B (2), were isolated from a Korean marine sponge Placospongia sp. The chemical structures of placotylenes A and B were elucidated on the basis of 1D and 2D NMR, along with MS spectral analysis and revealed as an iodinated polyacetylene class of natural products. Placotylene A (1) displayed inhibitory activity against RANKL-induced osteoclast differentiation at 10 ?M while placotylene B (2) did not show any significant activity up to 100 ?M, respectively. PMID:24705502

  10. The peroxisomal enzyme L-PBE is required to prevent the dietary toxicity of medium-chain fatty acids.

    PubMed

    Ding, Jun; Loizides-Mangold, Ursula; Rando, Gianpaolo; Zoete, Vincent; Michielin, Olivier; Reddy, Janardan K; Wahli, Walter; Riezman, Howard; Thorens, Bernard

    2013-10-17

    Specific metabolic pathways are activated by different nutrients to adapt the organism to available resources. Although essential, these mechanisms are incompletely defined. Here, we report that medium-chain fatty acids contained in coconut oil, a major source of dietary fat, induce the liver ?-oxidation genes Cyp4a10 and Cyp4a14 to increase the production of dicarboxylic fatty acids. Furthermore, these activate all ?- and ?-oxidation pathways through peroxisome proliferator activated receptor (PPAR) ? and PPAR?, an activation loop normally kept under control by dicarboxylic fatty acid degradation by the peroxisomal enzyme L-PBE. Indeed, L-pbe(-/-) mice fed coconut oil overaccumulate dicarboxylic fatty acids, which activate all fatty acid oxidation pathways and lead to liver inflammation, fibrosis, and death. Thus, the correct homeostasis of dicarboxylic fatty acids is a means to regulate the efficient utilization of ingested medium-chain fatty acids, and its deregulation exemplifies the intricate relationship between impaired metabolism and inflammation. PMID:24075987

  11. The cyclin-dependent kinase inhibitor SNS-032 induces apoptosis in breast cancer cells via depletion of Mcl-1 and X-linked inhibitor of apoptosis protein and displays antitumor activity in vivo.

    PubMed

    Xie, Gui'e; Tang, Hongping; Wu, Shaoqing; Chen, Jingsong; Liu, Jiangwen; Liao, Can

    2014-08-01

    Inhibitors of cyclin-dependent kinases (Cdks) have been reported to have activities in many types of cancer cells by inhibiting Cdk7 and Cdk9, which control transcription. SNS-032 is a potent and selective inhibitor of Cdk2, Cdk7 and Cdk9 and has emerged in clinical trials. Here, we examined the viability of MCF-7 and MDA-MB-435 breast cancer cells in the presence of SNS-032 and observed a dose-dependent inhibition of cellular proliferation in both cell lines. SNS-032 had a direct apoptosis-inducing effect through both the extrinsic and intrinsic apoptotic pathways in breast cancer cells as shown by a dose-dependent increase in Annexin V-positive cells and terminal deoxynucleotidyl transferase-mediated dUTP nick?end labeling (TUNEL)-positive cells, as well as activation of caspase-8, -9 and poly(ADP-ribose) polymerase (PARP). At the molecular level, SNS-032 induced a marked dephosphorylation of serine 2 and 5 of RNA polymerase (RNA Pol) II and blocked RNA synthesis. Consistent with the inherently rapid turnover rates of their transcripts and proteins, the anti-apoptotic proteins Mcl-1 and X-linked inhibitor of apoptosis protein (XIAP) were rapidly reduced on exposure to SNS-032. Our results also indicated that SNS-032 suppressed the growth of breast cancer xenografts in mice. These data demonstrate that the use of SNS-032 may be a rational and novel therapeutic strategy for human breast cancer and warrants further clinical investigation. PMID:24865236

  12. Mitochondrial division inhibitor 1 protects against mutant huntingtin-induced abnormal mitochondrial dynamics and neuronal damage in Huntington's disease.

    PubMed

    Manczak, Maria; Reddy, P Hemachandra

    2015-12-20

    The objective of this study was to determine the protective effects of the mitochondrial division inhibitor 1 (Mdivi1) in striatal neurons that stably express mutant Htt (STHDhQ111/Q111) and wild-type (WT) Htt (STHDhQ7/Q7). Using gene expression analysis, biochemical methods, transmission electron microscopy (TEM) and confocal microscopy methods, we studied (i) mitochondrial and synaptic activities by measuring mRNA and the protein levels of mitochondrial and synaptic genes, (ii) mitochondrial function and (iii) ultra-structural changes in mutant Htt neurons relative to WT Htt neurons. We also studied these parameters in Mdivil-treated and untreated WT and mutant Htt neurons. Increased expressions of mitochondrial fission genes, decreased expression of fusion genes and synaptic genes were found in the mutant Htt neurons relative to the WT Htt neurons. Electron microscopy of the mutant Htt neurons revealed a significantly increased number of mitochondria, indicating that mutant Htt fragments mitochondria. Biochemical analysis revealed defective mitochondrial functioning. In the Mdivil-treated mutant Htt neurons, fission genes were down-regulated, and fusion genes were up-regulated, suggesting that Mdivil decreases fission activity. Synaptic genes were up-regulated, and mitochondrial function was normal in the Mdivi1-treated mutant Htt neurons. Immunoblotting findings of mitochondrial and synaptic proteins agreed with mRNA findings. The TEM studies revealed that increased numbers of structurally intact mitochondria were present in Mdivi1-treated mutant Htt neurons. Increased synaptic and mitochondrial fusion genes and decreased fission genes were found in the Mdivi1-treated WT Htt neurons, indicating that Mdivi1 beneficially affects healthy neurons. Taken together, these findings suggest that Mdivi1 is protective against mutant Htt-induced mitochondrial and synaptic damage in HD neurons and that Mdivi1 may be a promising molecule for the treatment of HD patients. PMID:26464486

  13. Simvastatin and GGTI-2133, a geranylgeranyl transferase inhibitor, increase erythrocyte deformability but reduce low O(2) tension-induced ATP release.

    PubMed

    Clapp, K M; Ellsworth, M L; Sprague, R S; Stephenson, A H

    2013-03-01

    Statin drugs inhibit 3-hydroxy-3-methylglutaryl CoA reductase, which reduces the synthesis of both cholesterol and isoprenoids (geranylgeranyl pyrophosphate and farnesyl pyrophosphate), with the latter being lipid molecules responsible for the posttranslational modification of small GTP-binding proteins such as Rho. Effects of statins, independent of lowering blood cholesterol levels, are thought to occur by inhibition of Rho/Rho kinase. The Rho kinase inhibitor Y-27632 has been reported to increase both erythrocyte deformability and low O2 tension-induced ATP release. Here, we tested the hypothesis that by inhibiting Rho/Rho kinase, simvastatin would increase both erythrocyte deformability and low O2 tension-induced ATP release. Male Sprague-Dawley rats were divided into two groups, control or simvastatin treated [simvastatin-supplemented chow (0.02%)], for 4 wk. Simvastatin treatment increased rat erythrocyte deformability compared with controls (n = 6, P < 0.05). However, erythrocytes of simvastatin-treated rats (n = 9, P < 0.05) exhibited impaired low O2 tension-induced ATP release. Similarly, the geranylgeranyl transferase inhibitor GGTI-2133 (10 ?M) also increased deformability and impaired low O2 tension-induced ATP release in healthy human erythrocytes (P < 0.05). Interestingly, ATP release in response to mastoparan 7 (n = 7, P < 0.05), which directly activates Gi, and isoproterenol (n = 5, P < 0.05), which signals through Gs, was not altered by incubation with GGTI-2133. These results suggest that although statins increase erythrocyte deformability, likely by inhibiting geranylgeranylation, the finding that both statins and a geranylgeranyl transferase inhibitor attenuated low O2 tension-induced ATP release demonstrates that factors in addition to erythrocyte deformability are critical for ATP release in response to this physiological stimulus. PMID:23335799

  14. Chronic treatment with novel small molecule Hsp90 inhibitors rescues striatal dopamine levels but not ?-synuclein-induced neuronal cell loss.

    TOXLINE Toxicology Bibliographic Information

    McFarland NR; Dimant H; Kibuuka L; Ebrahimi-Fakhari D; Desjardins CA; Danzer KM; Danzer M; Fan Z; Schwarzschild MA; Hirst W; McLean PJ

    2014-01-01

    Hsp90 inhibitors such as geldanamycin potently induce Hsp70 and reduce cytotoxicity due to ?-synuclein expression, although their use has been limited due to toxicity, brain permeability, and drug design. We recently described the effects of a novel class of potent, small molecule Hsp90 inhibitors in cells overexpressing ?-synuclein. Screening yielded several candidate compounds that significantly reduced ?-synuclein oligomer formation and cytotoxicity associated with Hsp70 induction. In this study we examined whether chronic treatment with candidate Hsp90 inhibitors could protect against ?-synuclein toxicity in a rat model of parkinsonism. Rats were injected unilaterally in the substantia nigra with AAV8 expressing human ?-synuclein and then treated with drug for approximately 8 weeks by oral gavage. Chronic treatment with SNX-0723 or the more potent, SNX-9114 failed to reduce dopaminergic toxicity in the substantia nigra compared to vehicle. However, SNX-9114 significantly increased striatal dopamine content suggesting a positive neuromodulatory effect on striatal terminals. Treatment was generally well tolerated, but higher dose SNX-0723 (6-10 mg/kg) resulted in systemic toxicity, weight loss, and early death. Although still limited by potential toxicity, Hsp90 inhibitors tested herein demonstrate oral efficacy and possible beneficial effects on dopamine production in a vertebrate model of parkinsonism that warrant further study.