B Inhibitor (b + inhibitor)

Distribution by Scientific Domains
Distribution within Medical Sciences

Kinds of B Inhibitor

  • cathepsin b inhibitor

  • Selected Abstracts

    NF-,B and apoptosis in colorectal tumourigenesis

    M. M. Aranha
    Abstract Background, Nuclear factor-,B (NF-,B) may play an important role in colorectal tumourigenesis, controlling cell cycle and apoptosis gene expression. In addition, imbalances between cell proliferation and cell death are thought to underlie neoplastic development. The aims of this study were to investigate apoptosis and expression of several apoptosis-related proteins, and to determine correlations with colorectal tumour progression. Materials and methods, Apoptosis was evaluated by the TUNEL assay in 48 patient samples, including adenomas, adenocarcinomas and adjacent normal mucosas. Immunohistochemistry was performed for Bcl-2 and NF-,B. Expression levels of p53, Bax and I,B proteins were determined by immunoblotting. Cultured human colon cancer cells were used to evaluate NF-,B expression and nuclear translocation by immunocytochemistry and immunoblotting. Results, Apoptosis and NF-,B immunoreactivity were significantly higher in tumour tissue compared with normal mucosa (P < 001), increasing in association with histological tumour progression (P < 001). Bcl-2 was consistently higher in normal mucosa (P < 001) and inversely correlated with the percentage of apoptosis (P < 001). Phosphorylated p53 and Bax levels were similar in tumour tissue and normal mucosa; however, the NF-,B inhibitor, I,B, tended to decrease in tumours. In vitro, nuclear translocation of NF-,B was greater in proliferative than in resting phases of colon cancer cells. Conclusions, NF-,B expression and apoptosis are increased from adenoma to poorly differentiated adenocarcinoma tissues. Apoptosis is correlated with suppression of Bcl-2 expression, but appears to proceed through a p53- and Bax-independent pathway. Activation of NF-,B may play an important role in colorectal tumour progression. [source]

    Biotransformation of xenobiotics by amine oxidases

    Margherita Strolin Benedetti
    Although the cytochrome P450 (CYP) system ranks first in terms of catalytic versatility and the wide range of xenobiotics it detoxifies or activates to reactive intermediates, the contribution of amine oxidases and in particular of monoamine oxidases (MAOs) to the metabolism of xenobiotics is far from negligible but has been largely neglected. In this review on the involvement of amine oxidases in the metabolism of xenobiotics, the major characteristics reported for the CYP system (protein, reaction, tissue distribution, subcellular localisation, substrates, inhibitors, inducers, genetic polymorphism, impact of different physiopathological conditions on the activity, turnover) will be compared, whenever possible, with the corresponding characteristics of amine oxidases (MAOs in particular). The knowledge of the involvement of MAO-A, -B or both in the metabolism of a drug allows us to predict interactions with selective or non-selective MAO inhibitors (e.g. the metabolism of a drug deaminated by both forms of MAO is not necessarily inhibited in vivo by a selective MAO-A or -B inhibitor). If a drug is metabolized by MAOs, competitive interactions can occur with other drugs that are MAO substrates, e.g. with ,-adrenoceptor agonists and antagonists, prodrugs of dopamine, serotonin 5-HT1 -receptor agonists as well as with primaquine, flurazepam and citalopram. Moreover, the knowledge of the involvement of MAOs in the metabolism of a drug may suggest possible, although not obligatory, interactions with tyramine-containing food or drink, with over the counter medicines sold to relieve the symptoms of coughs and colds (generally containing the indirectly-acting sympathomimetic amine phenylpropanolamine) or with phenylephrine-containing preparations. Finally, biotransformation by amine oxidases, as by CYP, does not always lead to detoxication but can produce toxic compounds. [source]

    The dietary histone deacetylase inhibitor sulforaphane induces human ,-defensin-2 in intestinal epithelial cells

    IMMUNOLOGY, Issue 2 2008
    Markus Schwab
    Summary Antimicrobial peptides like human ,-defensin-2 (HBD-2) play an important role in the innate immune system protecting the intestinal mucosa against bacterial invasion. The dietary histone deacetylase (HDAC) inhibitors sulforaphane (SFN) and butyrate have received a great deal of attention because of their ability to simultaneously modulate multiple cellular targets involved in cellular protection. In this study the influence of SFN and butyrate on HBD-2 expression as well as the molecular pathways involved in SFN-mediated induction of HBD-2 were scrutinized. Treatment of Caco-2, HT-29 and SW480 cells with SFN led to a time- and dose-dependent upregulation of HBD-2 mRNA expression as determined by semi-quantitative reverse transcription,polymerase chain reaction. Moreover, HBD-2 protein production increased in response to SFN, measured by enzyme-linked immunosorbent assay. Induction of HBD-2 was also observed in response to butyrate. Immunofluorescence analysis revealed that the protein was localized in the cytosol. Coincubation of SFN with a vitamin D receptor (VDR), or an extracellular-regulated kinase 1/2 or a nuclear factor-,B inhibitor all reduced HBD-2 mRNA upregulation. In contrast, transfection of cells with a dominant-negative peroxisome proliferator-activated receptor , (PPAR,) mutant vector to inhibit PPAR, wild-type action and inhibition of p38 mitogen-activated protein kinase (MAPK) signalling did not affect SFN-mediated upregulation of HBD-2 mRNA. Moreover, SFN induced the expression of VDR, PPAR, and phosphorylated ERK1/2 but did not affect p38 MAPK activation. The data clearly demonstrate for the first time that the dietary HDAC inhibitor SFN is able to induce antimicrobial peptides in colonocytes. In this process HBD-2 expression is regulated via VDR, mitogen-activated protein kinase kinase/extracellular-regulated kinase and nuclear factor-,B signalling. [source]

    Apoptosis induction by interleukin-2-activated cytotoxic lymphocytes in a squamous cell carcinoma cell line and Daudi cells , involvement of reactive oxygen species-dependent cytochrome c and reactive oxygen species-independent apoptosis-inducing factors

    IMMUNOLOGY, Issue 2 2003
    Tetsuya Yamamoto
    Summary Investigation of the induction of apoptosis by cytotoxic lymphocytes has mainly focused on the signalling associated with Fas and its adaptor proteins. The signal pathway via mitochondria, however, has not been sufficiently elucidated in cytotoxic lymphocyte-induced apoptosis. We examined the release of mitochondrial proapoptotic factors by lymphokine-activated killer (LAK) cells in two cell lines. LAK cell-induced DNA fragmentation of the target cells was suppressed to approximately 50% of control levels by the addition of neutralizing monoclonal antibody to Fas and a granzyme B inhibitor. When intracellular reactive oxygen species (ROS) were scavenged, the LAK cell-induced DNA fragmentation was decreased to approximately 60% of the non-treated cell level. Co-cultivation of Daudi cells with LAK cells increased cytosolic and mitochondrial ROS levels. Activation of procaspase-3 and apoptosis by treatment of oral squamous cell carcinoma cells (OSC) with LAK cells was partially inhibited by pretreatment of OSC cells with ROS scavengers and mitochondrial complex inhibitors. Furthermore, cytochrome c and apoptosis-inducing factor (AIF) were released from mitochondria by OSC cell treatment with supernatants of LAK cells. The supernatant-induced cytochrome c release was suppressed by mitochondrial complex inhibitors, but the inhibitors did not inhibit the release of AIF. These results indicate that LAK cells induce target cell apoptosis via not only the Fas/Fas ligand system and granzyme B, but also ROS-dependent cytochrome c and ROS-independent AIF release. [source]

    Rasagiline: defining the role of a novel therapy in the treatment of Parkinson's disease

    F. Stocchi
    Summary Parkinson's disease (PD) is a therapy area with considerable unmet needs. The current key targets for PD treatment include the slowing of disease progression, improved control of motor fluctuations in advanced disease and the treatment of nonmotor symptoms. In view of such major requirements, it is important to consider how new drug treatments fit into the context of PD therapy, and the practical advantages that they may offer in the management of PD in clinical practice. Rasagiline is a novel, second-generation, irreversible, selective monoamine oxidase type B inhibitor that is indicated for the treatment of idiopathic PD, either as initial monotherapy or as adjunct therapy (with levodopa) for patients experiencing end-of-dose motor fluctuations. This review assesses the outcome from several large-scale clinical studies that have investigated the use of rasagiline in early and advanced PD patient populations and discusses the role of rasagiline within the current scope of PD therapy. [source]

    Activation and induction of cytosolic phospholipase A2 by IL-1, in human tracheal smooth muscle cells: Role of MAPKs/p300 and NF-,B

    Chiang-Wen Lee
    Abstract Cytosolic phospholipase A2 (cPLA2) plays a pivotal role in mediating agonist-induced arachidonic acid (AA) release for prostaglandin (PG) synthesis during inflammation triggered by IL-1,. However, the mechanisms underlying IL-1,-induced cPLA2 expression and PGE2 synthesis in human tracheal smooth muscle cells (HTSMCs) remain unknown. IL-1,-induced cPLA2 protein and mRNA expression, PGE2 production, or phosphorylation of p42/p44 MAPK, p38 MAPK, and JNK1/2, which was attenuated by pretreatment with the inhibitors of MEK1/2 (U0126), p38 MAPK (SB202190), and JNK1/2 (SP600125) or transfection with siRNAs of MEK1, p42, p38, and JNK2. IL-1,-induced cPLA2 expression was also inhibited by pretreatment with a NF-,B inhibitor, helenalin or transfection with siRNA of NIK, IKK,, or IKK,. IL-,-induced NF-,B translocation was blocked by pretreatment with helenalin, but not U0126, SB202190, and SP600125. In addition, transfection with p300 siRNA blocked cPLA2 expression induced by IL-1,. Moreover, p300 was associated with the cPLA2 promoter, which was dynamically linked to histone H4 acetylation stimulated by IL-1,. These results suggest that in HTSMCs, activation of MAPKs, NF-,B, and p300 are essential for IL-1,-induced cPLA2 expression and PGE2 secretion. J. Cell. Biochem. 109: 1045,1056, 2010. 2010 Wiley-Liss, Inc. [source]

    Advanced oxidation protein products inhibit differentiation and activate inflammation in 3T3-L1 preadipocytes,

    Qiu Gen Zhou
    Accumulation of advanced oxidation protein products (AOPPs) is prevalent in metabolic syndromes, a condition with impaired preadipocytes differentiation. In the present study, we tested the hypothesis that AOPPs disturb preadipocyte differentiation. Exposure of 3T3-L1 preadipocytes to increased levels of AOPPs inhibited accumulation of intracellular triglyceride and decreased the expression of the essential markers of matured adipocytes, such as adipocyte fatty-acid-binding protein (aP2), CAAT/enhancer-binding protein (C/EBP)-,, and peroxisome proliferator-activated receptor (PPAR)-,, in response to standard adipogenic induction. Inhibitory effects of AOPPs on preadipocytes differentiation was time sensitive, which occurred at the early stage of differentiation. In the presence of AOPPs, induction of preadipocytes differentiation resulted in upregulated expression of C/EBP homologous protein (CHOP) and CUG-Triplet repeat-binding protein (CUGBP), two important inhibitors of preadipocytes differentiation. In addition, treatment with AOPPs increased abundance of C/EBP-,-liver enriched inhibitory protein (C/EBP-,-LIP), a truncated C/EBP-, isoform without adipogenic activity. Moreover, AOPPs-treated preadipocytes expressed a macrophage marker F4/80 and overexpressed tumor necrosis factor-, and interleukin-6 via nuclear factor-,B (NF-,B)-dependent pathway. However, blocking inflammation with NF-,B inhibitor failed to improve AOPPs-induced inhibition of preadipocytes differentiation. These data suggest that accumulation of AOPPs may inhibit differentiation of preadipocytes and activate inflammation in these cells. This information might have implication for understanding the impairment of preadipocytes differentiation and fat inflammation seen in metabolic syndrome. J. Cell. Physiol. 225: 42,51, 2010. 2010 Wiley-Liss, Inc. [source]

    c-DNA Microarray to determine molecular events in neurodegeneration and neuroprotection

    M. B. H. Youdim
    Cell death in CNS involves complex processes, many of which have not been identified biochemically. At the present biochemical techniques cannot adequately establish these. However, the advent of cDNA microarray or microchips, in which the expression of thousands of genes can be measured at once to give a global assessment in disease pathology, its progress or animal models, has simplified this. We have employed this technique to study the mechanism of neurotoxicity of MPTP and 6-hydroxydoapmine induced in neuronally derived cells in culture, in the animal models of Parkinson's disease and neuroprotection initiated by monoamine oxidase B inhibitor, rasagiline; iron chelators, R-apomorphine and EGCG and other neuroprotective drugs. Our studies have clearly indicated that MPTP induced early gene expression, prior to cell death (first 24 h), are prerequirement for 51 late gene expression changes implicated at the time of neuronal death. The latter genes include those involved in iron metabolism, oxidative stress, inflammatory processes, glutaminergic excitotoxicity, nitric oxide, growth factors, transcription factors, cell cycle, intermediatory metabolism and other gene previously not identified. The expressions of many of the latter genes, also identified by in situ hybridization, are prevented when the animals are pretreated with the above neuroprotective drugs. These studies have clearly shown that neurodegeneratrion is a complex cascades of ,domino' effect. Thus a single neuroprotective drug treatment may not be adequate to prevent it, but, that a cocktail of drugs might. [source]

    Neurovascular and neuronal protection by E64d after focal cerebral ischemia in rats

    Tamiji Tsubokawa
    Abstract Calpains and cathepsins are two families of proteases that play an important role in ischemic cell death. In this study, we investigated the effect of E64d, a ,-calpain and cathepsin B inhibitor, in the prevention of neuronal and endothelial apoptotic cell death after focal cerebral ischemia in rats. Rats underwent 2 hr of transient focal ischemia from middle cerebral artery occlusion (MCAO) and were sacrificed 24 hr later. E64d (5 mg/ kg intraperitoneally) was administered 30 min before MCAO. Assessment included neurological function, infarction volume, brain water content, blood,brain barrier permeability, histology, and immunohistochemistry. The E64d-treated rats had significant brain protection against ischemic damage. We observed a reduction of infarction volume, brain edema, and improved neurological scores in E64d-treated rats compared with the nontreated control. Furthermore, there was a remarkable reduction in both proteases and caspase-3 activation and apoptotic changes in both neurons and endothelial cells in E64d-treated rats. These results suggest that E64d protects the brain against ischemic/reperfusion injury by attenuating neuronal and endothelial apoptosis. 2006 Wiley-Liss, Inc. [source]

    A Functional Polymorphism of the NFKB1 Gene Increases the Risk for Alcoholic Liver Cirrhosis in Patients With Alcohol Dependence

    ALCOHOLISM, Issue 11 2009
    Miguel Marcos
    Background:, The genetic basis for the predisposition to alcoholic liver cirrhosis (ALC) remains unknown. Increasing evidence supports a role for the nuclear factor (NF)-,B, the NF-,B inhibitor , (NFKBIA), and the peroxisome proliferator-activated receptor (PPAR)-, in the pathogenesis of alcoholic liver disease, raising the possibility that common polymorphisms in genes encoding these molecules may confer susceptibility to ALC. The objective of this study was to analyze the relationship between common polymorphisms in NFKB1, NFKBIA, and PPARG2 genes and the presence of ALC. Methods:, A total of 258 male alcoholics (161 without liver disease and 97 with ALC) and 101 healthy controls were genotyped for the ,94ins/delATTG NFKB1, 3,-UTR+126G>A NFKBIA, and 34C>G PPARG2 polymorphisms. The association of these genetic variants with ALC was tested in alcoholic patients with alcohol abuse and alcohol dependence. A logistic regression analysis was further performed to analyze the model of inheritance. Results:, We found an association between the presence of the deletion allele in NFKB1 polymorphism and ALC in patients with alcohol dependence. We found no association between NFKBIA and PPARG2 polymorphisms and the presence of ALC. Conclusions:, The deletion allele of the ,94ins/del NFKB1 polymorphism could be associated with a higher risk of developing ALC through an increase in inflammation, as supported by previous data. [source]

    Neopterin induces pro-atherothrombotic phenotype in human coronary endothelial cells

    Summary.,Background: Inflammation plays a pivotal role in atherothrombosis. Recent data indicate that serum levels of neopterin, a marker of inflammation and immune modulator secreted by monocytes/macrophages, are elevated in patients with acute coronary syndromes and seem to be a prognostic marker for major cardiovascular events. The aim of the present study was to determine whether neopterin might affect the thrombotic and atherosclerotic characteristics of human coronary artery endothelial cells (HCAECs). Methods and results: In HCAECs, neopterin induced TF-mRNA transcription as demonstrated by real time polymerase chain reaction and expression of functionally active tissue factor (TF) as demonstrated by procoagulant activity assay, and of cellular adhesion molecules (CAMs) as demonstrated by FACS analysis, in a dose-dependent fashion. These neopterin effects were prevented by lovastatin, a HMG-CoA reductase inhibitor. Neopterin-induced TF and CAMs expression was mediated by oxygen free radicals through the activation of the transcription factor, nuclear factor-kappa B (NF- ,B), as demonstrated by electrophoretic mobility shift assay and by suppression of CAMs and TF expression by superoxide dismutase and by NF- ,B inhibitor, pyrrolidine-dithio-carbamate ammonium. Conclusions: These data indicate that neopterin exerts direct effects on HCAECs by promoting CAMs and TF expression and support the hypothesis that neopterin, besides representing a marker of inflammation, might be an effector molecule able to induce a pro-atherothrombotic phenotype in cells of the coronary circulation. [source]

    Signaling pathways in osteoblast proinflammatory responses to infection by Porphyromonas gingivalis

    T. Ohno
    Introduction:, We recently investigated global gene expression in ST2 mouse stromal cells infected by the periodontal pathogen Porphyromonas gingivalis using microarray technology, and found that the bacterium induces a wide range of proinflammatory gene expression. Here, we reported the signaling pathways involved in those proinflammatory responses. Methods:, ST2 cells and primary calvarial osteoblasts from C3H/HeN, C57BL/6, and MyD88-deficient (MyD88,/,) mice were infected with P. gingivalis ATCC33277 and its gingipain-deficient mutant KDP136. Expression of the chemokines CCL5 and CXCL10, and matrix metalloproteinase-9 (MMP9) were quantified by real-time polymerase chain reaction, while phosphorylation of protein kinases and degradation of an inhibitor of nuclear factor-,B, I,B-,, were detected by Western blotting, and activation of transcriptional factors was determined by a luciferase reporter assay. The effects of inhibitors of transcriptional factors and protein kinases were also investigated. Results:, Infection by P. gingivalis elicited gene expression of CCL5, CXCL10, and MMP9 in both ST2 cells and osteoblasts. Western blot and reporter assay results revealed activation of nuclear factor-,B (NF-,B) and activator protein-1 transcription factors. The NF-,B inhibitor suppressed the expression of CCL5 and MMP9, but not that of CXCL10, whereas P. gingivalis infection induced significant CCL5 expression in MyD88,/, osteoblasts. In addition, activation of protease-activated receptors by trypsin elicited significant induction of CXCL10. Conclusion:, Our results suggest that various proinflammatory responses in P. gingivalis -infected stromal/osteoblast cells are NF-,B-dependent, but not always dependent on the Toll-like receptor/MyD88 pathway, while some responses are related to the activation of protease-activated receptors. Thus, P. gingivalis does not fully utilize well-established pathogen recognition molecules such as Toll-like receptors. [source]

    ORIGINAL ARTICLE: Regulation of Nod1 and Nod2 in First Trimester Trophoblast Cells

    Melissa J. Mulla
    Problem:, The cytoplasmic pattern recognition receptors, Nod1 and Nod2, are thought to be important for detecting intracellular bacteria. We have previously reported that first trimester trophoblast cells express Nod1 and Nod2, and that trophoblast Nod2 activation triggers an inflammatory response. The objectives of this study were to characterize the effects of Nod1 stimulation, and to determine the regulation of Nod1 and Nod2, in the trophoblast. Method of Study:, The effect of Nod1 activation on trophoblast cells was determined by analyzing the cytokine response following treatment with ,-D-glutamyl- meso -diaminopimelic acid (iE-DAP). The regulation of Nod1 and Nod2 expression by trophoblast cells was evaluated by RT-PCR. Results:, Treatment of trophoblast cells with iE-DAP significantly increased their production of cytokines and chemokines. In addition, Nod1 and Nod2 mRNA expression was upregulated following treatment of trophoblast cells with lipopolysaccharide (LPS), and this was significantly reduced by the presence of a NF,B inhibitor and a TLR4-dominant negative (DN). Conclusion:, This study demonstrates that LPS, through TLR4, increases trophoblast expression of Nod1 and Nod2 via the NF,B pathway; and that Nod1 is functional in the trophoblast. These findings suggest that extracellular recognition of bacterial LPS by TLR4 may prime the trophoblast in preparation for its cytoplasmic recognition of, and response to, bacterial peptides through the Nod proteins. [source]

    Inflammatory stimuli from macrophages and cancer cells synergistically promote tumor growth and angiogenesis

    CANCER SCIENCE, Issue 12 2007
    Yusuke N. Kimura
    The focus of the present study was whether and how infiltrating macrophages play a role in angiogenesis and the growth of cancer cells in response to the inflammatory cytokine interleukin (IL)-1,. Lewis lung carcinoma cells overexpressing IL-1, grew faster and induced greater neovascularization than a low IL-1,-expressing counterpart in vivo. When macrophages were depleted using clodronate liposomes, both neovascularization and tumor growth were reduced in the IL-1,-expressing tumors. Co-cultivation of IL-1,-expressing cancer cells with macrophages synergistically augmented neovascularization and the migration of vascular endothelial cells. In these co-cultures, production of the angiogenic factors vascular endothelial growth factor-A and IL-8, monocyte chemoattractant protein-1, and matrix metalloproteinase-9 were increased markedly. The production of these factors, induced by IL-1,-stimulated lung cancer cells, was blocked by a nuclear factor (NF)-,B inhibitor, and also by the knockdown of p65 (NF-,B) and c-Jun using small interference RNA, suggesting involvement of the transcription factors NF-,B and AP-1. These results demonstrated that macrophages recruited into tumors by monocyte chemoattractant protein-1 and other chemokines could play a critical role in promoting tumor growth and angiogenesis, through interactions with cancer cells mediated by inflammatory stimuli. (Cancer Sci 2007; 98: 2009,2018) [source]

    Zaire Ebola virus entry into human dendritic cells is insensitive to cathepsin L inhibition

    Osvaldo Martinez
    Summary Cathepsins B and L contribute to Ebola virus (EBOV) entry into Vero cells and mouse embryonic fibroblasts. However, the role of cathepsins in EBOV-infection of human dendritic cells (DCs), important targets of infection in vivo, remains undefined. Here, EBOV-like particles containing a ,-lactamase,VP40 fusion reporter and Ebola virus were used to demonstrate the cathepsin dependence of EBOV entry into human monocyte-derived DCs. However, while DC infection is blocked by cathepsin B inhibitor, it is insensitive to cathepsin L inhibitor. Furthermore, DCs pre-treated for 48 h with TNF, were generally less susceptible to entry and infection by EBOV. This decrease in infection was associated with a decrease in cathepsin B activity. Thus, cathepsin L plays a minimal, if any, role in EBOV infection in human DCs. The inflammatory cytokine TNF, modulates cathepsin B activity and affects EBOV entry into and infection of human DCs. [source]

    Montelukast inhibits tumour necrosis factor-,-mediated interleukin-8 expression through inhibition of nuclear factor-,B p65-associated histone acetyltransferase activity

    F. Tahan
    Summary Background Montelukast is a potent cysteinyl leukotriene-1 receptor antagonist possessing some anti-inflammatory effects although the molecular mechanism of these anti-inflammatory effects is unknown. In this study, we aimed to investigate the effect of montelukast on nuclear factor (NF)-,B-associated histone acetylation activity in phorbol myristate acetate (PMA)-differentiated U937 cells. Methods We examined the inhibitory effects of montelukast on TNF-,-induced IL-8 production in PMA-differentiated U-937 cells. U-937 cells were exposed to PMA (50 ng/mL) for 48 h to allow differentiation to macrophages. Macrophages were then exposed to TNF-, (10 ng/mL) in the presence or absence of montelukast (0.01,10 ,m) for 24 h. After this time, the concentration of IL-8 in the culture supernatant was measured by sandwich-type ELISA kit. The effect of signalling pathways on TNF-,-induced IL-8 release was examined pharmacologically using selective NF-,B/IKK2 (AS602868, 3 ,m), (PD98059, 10 ,m) and p38 mitogen activated protein kinase (MAPK) (SB203580, 1 ,m) inhibitors. NF-,B DNA binding activity was measured by a DNA-binding ELISA-based assay. NF-,B-p65-associated histone acetyltransferase (HAT) activity was measured by immunoprecipitation linked to commercial flourescent HAT. Results TNF-,-induced IL-8 release was suppressed by an NF-,B inhibitor but not by MEK or p38 MAPK inhibitors. Montelukast induced a concentration-dependent inhibition of TNF-,-induced IL-8 release and mRNA expression that reached a plateau at 0.1 ,m without affecting cell viability. Montelukast did not affect NF-,B p65 activation as measured by DNA binding but suppressed NF-,B p65-associated HAT activity. Conclusion Montelukast inhibits TNF-,-stimulated IL-8 expression through changes in NF-,B p65-associated HAT activity. Drugs targeting these enzymes may enhance the anti-inflammatory actions of montelukast. [source]

    TREM-1 expression in macrophages is regulated at transcriptional level by NF-,B and PU.1

    Heng Zeng
    Abstract Triggering receptor expressed on myeloid cells (TREM)-1 is a recently identified immunoglobulin receptor that is expressed on neutrophils and monocytes where it amplifies the acute inflammatory response to bacteria. We examined the transcriptional regulation of TREM-1 in macrophages. Treatment of RAW cells with Escherichia coli LPS or Pseudomonas aeruginosa led to the induction of TREM-1 within 1,h with an expression lasting up to at least 24,h in vitro as detected by RT-PCR. Since the promoter of TREM-1 has multiple binding sites for NF-,B and PU.1 (one of the members of the ets family of transcription factors), we investigated the role of these transcription factors in the induction of TREM-1. Treatment of cells with NF-,B inhibitors abolished the expression of message of TREM-1 induced by LPS and P.,aeruginosa. In contrast, the expression of TREM-1 was increased after stimulation with LPS or P.,aeruginosa in cells that had gene of PU.1 silenced. Additionally, over-expression of PU.1 led to inhibition of TREM-1 induction in response to LPS and P.,aeruginosa. These data suggest that both these transcription factors are involved in the expression of TREM-1. NF-,B functions as a positive regulator whereas PU.1 is a negative regulator of the TREM-1 gene. [source]

    Monoamine oxidase A rather than monoamine oxidase B inhibition increases nicotine reinforcement in rats

    Karine Guillem
    Abstract Although nicotine is considered to be responsible for the addictive properties of tobacco, growing evidence underlines the importance of non-nicotine components in smoking reinforcement. It has been shown that tobacco smoke contains monoamine oxidase (MAO) A and B inhibitors and decreases MAO-A and MAO-B activity in smokers. Here, we investigated the effects of clorgyline hydrochloride (irreversible MAO-A inhibitor; 2 mg/kg/day), selegiline (irreversible MAO-B inhibitor; 4 mg/kg) and the beta-carboline norharmane hydrochloride (reversible MAO-B inhibitor; 5 mg/kg/day) treatments on nicotine self-administration (30 g/kg/infusion, free base) in rats. Independent of the responsiveness to novelty and locomotor activity stimulation, only clorgyline hydrochloride treatment increased the intake of nicotine in a fixed-ratio schedule (FR5) of reinforcement. When a progressive-ratio schedule was implemented, both clorgyline hydrochloride and norharmane hydrochloride treatments potentiated the reinforcing effects of nicotine, whereas selegiline had no effect. Taken together, these results indicate that MAO-A inhibition interacts with nicotine to enhance its rewarding effects in rats and suggest that other compounds present in tobacco, such as beta-carboline, may also play an important role in sustaining smoking behavior in humans. [source]

    Biochemical characterization of human cathepsin X revealed that the enzyme is an exopeptidase, acting as carboxymonopeptidase or carboxydipeptidase

    FEBS JOURNAL, Issue 17 2000
    Ivica Klemen
    Cathepsin X, purified to homogeneity from human liver, is a single chain glycoprotein with a molecular mass of ,,33 kDa and pI 5.1,5.3. Cathepsin X was inhibited by stefin A, cystatin C and chicken cystatin (Ki = 1.7,15.0 nm), but poorly or not at all by stefin B (Ki > 250 nm) and l -kininogen, respectively. The enzyme was also inhibited by two specific synthetic cathepsin B inhibitors, CA-074 and GFG-semicarbazone. Cathepsin X was similar to cathepsin B and found to be a carboxypeptidase with preference for a positively charged Arg in P1 position. Contrary to the preference of cathepsin B, cathepsin X normally acts as a carboxymonopeptidase. However, the preference for Arg in the P1 position is so strong that cathepsin X cleaves substrates with Arg in antepenultimate position, acting also as a carboxydipeptidase. A large hydrophobic residue such as Trp is preferred in the P1, position, although the enzyme cleaved all P1, residues investigated (Trp, Phe, Ala, Arg, Pro). Cathepsin X also cleaved substrates with amide-blocked C-terminal carboxyl group with rates similar to those of the unblocked substrates. In contrast, no endopeptidase activity of cathepsin X could be detected on a series of o -aminobenzoic acid-peptidyl- N -[2,-dinitrophenyl]ethylenediamine substrates. Furthermore, the standard cysteine protease methylcoumarine amide substrates (kcat/Km,,5.0 103 m,1s,1) were degraded ,,25-fold less efficiently than the carboxypeptidase substrates (kcat/Km , 120.0 103 m,1s,1). [source]

    Cathepsin B Inhibitory Tetraene Lactones from the Fungus Talaromyces wortmannii

    Yuesheng Dong
    Abstract Wortmannilactones E,H (1,4), four new cathepsin B inhibitors, were produced and isolated from the culture of the soil filamentous fungus Talaromyces wortmannii. Their structures and relative configurations were elucidated on the basis of 1D- and 2D-NMR techniques, three of them (1, 2, and 4) posses an oxabicyclo[2.2.1]heptane moiety. Compounds 1,4 showed inhibitory activities against cathepsin B with IC50 values of 4.3, 6.5, 13.0, and 6.0,,M, respectively. [source]

    Nuclear factor-,B contributes to interleukin-4- and interferon-dependent polymeric immunoglobulin receptor expression in human intestinal epithelial cells

    IMMUNOLOGY, Issue 1 2004
    Laynez W. Ackermann
    Summary Polymeric immunoglobulins (pIgs) that are present at mucosal surfaces play key roles in both the innate and adaptive immune responses. These pIgs are delivered to the mucosal surface via transcytosis across the epithelium, a process mediated by the polymeric immunoglobulin receptor (pIgR). Previous studies demonstrate that expression of the pIgR is regulated by multiple immunomodulatory factors including interleukin-4 (IL-4) and interferon-, (IFN-,). In studies using human intestinal epithelial cells (HT29), multiple inhibitors of the transcription factor nuclear factor-,B (NF-,B), including a dominant negative I,B,-serine mutant, inhibited both IL-4- and IFN-dependent increases in pIgR expression. Under identical conditions, NF-,B inhibitors had no effect on cytokine-dependent increases in expression of the transcription factor interferon regulatory factor-1. Over-expression of the I,B,-serine mutant also inhibited reporter gene expression in response to IL-4, TNF-,, IL-1,, and in some cases IFN-, using constructs with sequences from the pIgR promoter. Reduced levels of pIgR were observed even when inhibitors were added ,24 hr after cytokines suggesting that prolonged activation of NF-,B is required. Finally, reporter gene studies with NF-,B enhancer elements indicated that IFN-, alone and IL-4 in combination with other cytokines activated NF-,B in HT29 cells. Together, these studies provide additional insight into the signalling pathways that contribute to expression of the pIgR, a critical player in mucosal immunity. [source]

    Endogenous TNF, Lowers Maximum Peak Bone Mass and Inhibits Osteoblastic Smad Activation Through NF-,B,,

    Yan Li
    Abstract Endogenous TNF, prevents the attainment of maximum achievable peak bone mass in vivo. In vitro, TNF, suppresses BMP-2, and TGF,-mediated Smad activation through induction of NF-,B. Consistently, pharmacological suppression of NF-,B augments osteoblast differentiation and mineralization in vitro. Introduction: Osteoporosis is a major health threat. Traditional therapeutic strategies have centered on anti-catabolic drugs that block bone resorption. Recently focus has shifted to anabolic agents that actively rebuild lost bone mass. Future strategies may involve elevating peak bone mass to delay osteoporosis development. Recent in vitro studies show that TNF, represses osteoblast differentiation and mineralization; however, the mechanisms are poorly understood and the impact of basal TNF, concentrations on the acquisition of peak bone mass in vivo is unknown. Materials and Methods: We examined peak BMD, bone volume, and bone turnover makers in mice deficient in TNF, or its receptors. We further examined the effect of TNF, on Smad-induced signaling by TGF, and BMP-2 in vitro using a Smad responsive reporter. The effect of TNF,-induced NF-,B signaling on Smad signaling and on in vitro osteoblast mineralization was examined using specific NF-,B inhibitors and activators, and effects of TNF,-induced NF-,B signaling on BMP-2,induced Runx2 mRNA were examined using RT-PCR. Results: Mice null for TNF, or its p55 receptor had significantly increased peak bone mass, resulting exclusively from elevated bone formation. In vitro, TNF, potently suppressed Smad signaling induced by TGF, and BMP-2, downregulated BMP-2,mediated Runx2 expression, and inhibited mineralization of osteoblasts. These effects were mimicked by overexpression of NF-,B and prevented by pharmacological NF-,B suppression. Conclusions: Our data suggest that TNF, and NF-,B antagonists may represent novel anabolic agents for the maximization of peak basal bone mass and/or the amelioration of pathological bone loss. [source]

    Glioma cells under hypoxic conditions block the brain microvascular endothelial cell death induced by serum starvation

    Yoshifumi Ueda
    Abstract Angiogenesis is one of essential components for the growth of neoplasms, including malignant gliomas. However, tumor vascularization is often poorly organized and marginally functional due to tumor strucutural abnormalities, inducing regional or temporal hypoxic conditions and nutritional shortages in tumor tissues. We investigated how during angiogenesis migrating endothelial cells survive in these hypoxic and reduced nutritional conditions. Human brain microvascular endothelial cells (HBMECs) underwent apoptosis and necrosis after serum withdrawal. This endothelial cell death was blocked by recombinant VEGF protein or the culture medium of U251 glioma cells exposed to hypoxia (H-CM). Hypoxic treatment increased vascular endothelial growth factor (VEGF) and tumor necrosis factor alpha (TNF-,) expression in U251 glioma cells. H-CM activated nuclear factor-,B (NF,B) protein and increased the gene expression of antiapoptotic factors including Bcl-2, Bcl-XL, survivin and X-chromosome-linked inhibitor of apoptosis protein (XIAP) in endothelial cells. The survival activity of H-CM for endothelial cells was abolished by two kinds of VEGF inhibitors {Cyclopeptidic VEGF inhibitor and a VEGF receptor tyrosine kinase inhibitor (4-[(4,-chloro-2,-fluoro) phenylamino]-6, 7-dimethoxyquinazoline)} or NF,B inhibitors (ALLN and BAY 11,7082). These VEGF inhibitors did not block the activation of NF,B induced by H-CM in endothelial cells. On the contrary, TNF-, antagonist WP9QY enhanced the survival activity of H-CM for endothelial cells and blocked NF,B activation induced by H-CM under serum-starved conditions. Taken together, our data suggest that both the secretion of VEGF from glioma cells and activation of NF,B in endothelial cells induced by TNF-, are necessary for endothelial cell survival as they increase the expression of antiapoptotic genes in endothelial cells under conditions of serum starvation. These pathways may be one of the mechanisms by which angiogenesis is maintained in glioma tissues. [source]

    TNF-,,induced NF-,B signaling reverses age-related declines in VEGF induction and angiogenic activity in intervertebral disc tissues

    Tetsuro Ohba
    Abstract We previously demonstrated that VEGF and its receptors were expressed in human herniated discs (HD). TNF-, induced VEGF, resulting in neovascularization of disc tissues in a model of HD. The goal of the current research was to investigate the precise role of TNF-,,induced VEGF and the mechanism of angiogenesis in disc tissues. We performed ELISAs, Western blots, and immunohistological examinations to assess the role of TNF-,,induced VEGF using organ disc cultures with wild type, TNF receptor 1-null (TNF-RInull), or TNF receptor 2-null (TNF-RIInull) mice. VEGF induction was inhibited when we used TNF-RInull -derived disc tissues. NF-,B pathway inhibitors also strongly suppressed VEGF induction. Thus, TNF-, induced VEGF expression in disc cells primarily through the NF-,B pathway. In addition, VEGF immunoreactivity was detected predominantly in annulus fibrosus cells and increased after TNF-, stimulation. TNF-, treatment also resulted in CD31 expression on endothelial cells and formation of an anastomosing network. In contrast, angiogenic activity was strongly inhibited in the presence of NF-,B inhibitors or anti-VEGF antibody. Our data show angiogenesis activity in disc tissues is regulated by VEGF and the NF-,B pathway, both of which are induced by TNF-,. The level of angiogenic activity in disc tissues was closely related to aging. Because neovascularization of HD is indispensable for HD resorption, the prognosis of HD and the rate of the resorption process in patients may vary as a function of the patient's age. 2008 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 27:229,235, 2009 [source]

    Calprotectin release from human neutrophils is induced by Porphyromonas gingivalis lipopolysaccharide via the CD-14,Toll-like receptor,nuclear factor ,B pathway

    Jun-ichi Kido
    Objectives:, Calprotectin is a cytosolic protein with antibacterial action in leukocytes and its level increases in some inflammatory diseases, including periodontal diseases, rheumatoid arthritis and ulcerative colitis. Recently, we found that the lipopolysaccharide of Porphyromonas gingivalis (P-LPS) induced calprotectin release from human neutrophils. P-LPS, a major virulence factor of periodontal pathogens, is known to induce the production and release of inflammatory cytokines through CD14, Toll-like receptor (TLR) and nuclear factor ,B (NF-,B). In the present study, we investigated whether calprotectin release by P-LPS is induced via the CD14,TLR,NF-,B pathway and the cellular mechanism of calprotectin release in human neutrophils. Material and methods:, Human neutrophils were isolated from the peripheral blood of healthy donors and pre-incubated in medium containing antibodies against CD14, TLR2 and TLR4, or several inhibitors of NF-,B, microtubules and microfilaments, and then incubated with P-LPS. The calprotectin amount in the culture medium was determined using ELISA, and the nuclear extracts from cells were used for the examination of NF-,B binding activity using electrophoretic mobility shift assays. Results:, P-LPS increased calprotectin release from neutrophils and its induction was inhibited by anti-CD14 and anti-TLR2 antibodies, but not by two anti-TLR4 antibodies. NF-,B inhibitors suppressed P-LPS-induced NF-,B binding activity and calprotectin release. The inhibitors of microtubule and microfilament polymerization significantly decreased P-LPS-induced calprotectin release. Conclusion:, These results suggest that calprotectin release is induced by P-LPS via the CD14,TLR2,NF-,B signal pathway in human neutrophils and may be dependent on microtubule and microfilament systems. [source]

    Nuclear factor-,B inhibition improves myocardial contractility in rats with cirrhotic cardiomyopathy

    Hongqun Liu
    Abstract Background/Aims: Cytokines such as tumour necrosis factor (TNF-,) contribute to the pathogenesis of cirrhotic cardiomyopathy. Nuclear factor-,B (NF-,B) is crucial for cytokine regulation, and induces cardiac dysfunction in several heart disease models. We aimed to elucidate possible NF-,B involvement in cirrhotic cardiomyopathy. Methods: Rats were bile duct ligated (BDL) to produce cirrhosis; controls received sham operation. Animals were studied 4 weeks later. Two NF-,B inhibitors were used: pyrrolidine dithiocarbamate (PDTC) and Bay 11-7082. Four groups were studied in most protocols: sham control, sham+PDTC, BDL and BDL+PDTC. Additional contractility studies were performed with Bay 11-7082. Myocardial NF-,B and TNF-, expression was measured by Western blot and ELISA. The contractility of isolated cardiomyocytes was observed under direct microscopy. Results: Nuclear factor-,B and TNF-, levels were increased in cirrhotic hearts compared with controls. PDTC significantly reduced NF-,B activity and TNF-, expression in cirrhotic hearts; controls were unaffected. Cirrhotic cardiomyocytes showed decreased systolic and diatolic velocity compared with sham controls. Both PDTC and Bay 11-7082 restored contractile function in cirrhotic cardiomyocytes, but did not affect controls. Conclusions: Inhibition of the increased NF-,B activity in cirrhotic hearts was associated with improvement of attenuated cardiomyocyte contractility. NF-,B, via effects on cytokine expression, may contribute to the pathogenesis of cirrhotic cardiomyopathy. [source]

    Down-regulation of myeloid cell leukemia 1 by epigallocatechin-3-gallate sensitizes rheumatoid arthritis synovial fibroblasts to tumor necrosis factor ,,induced apoptosis

    ARTHRITIS & RHEUMATISM, Issue 5 2009
    Salahuddin Ahmed
    Objective Overexpression of the antiapoptotic protein myeloid cell leukemia 1 (Mcl-1) in rheumatoid arthritis (RA) synovial fibroblasts is a major cause of their resistance to tumor necrosis factor , (TNF,),induced apoptosis. This study was undertaken to evaluate the efficacy of epigallocatechin-3-gallate (EGCG) in down-regulating Mcl-1 expression and its mechanism of RA synovial fibroblast sensitization to TNF,-induced apoptosis. Methods EGCG effects on cultured RA synovial fibroblast cell morphology, proliferation, and viability over 72 hours were determined by microscopy and a fluorescent cell enumeration assay. Caspase 3 activity was determined by a colorimetric assay. Western blotting was used to evaluate the apoptosis mediators poly(ADP-ribose) polymerase (PARP), Mcl-1, Bcl-2, Akt, and nuclear translocation of NF-,B. Results In RA synovial fibroblasts, EGCG (5,50 ,M) inhibited constitutive and TNF,-induced Mcl-1 protein expression in a concentration- and time-dependent manner (P < 0.05). Importantly, EGCG specifically abrogated Mcl-1 expression in RA synovial fibroblasts and affected Mcl-1 expression to a lesser extent in osteoarthritis and normal synovial fibroblasts or endothelial cells. Inhibition of Mcl-1 by EGCG triggered caspase 3 activity in RA synovial fibroblasts, which was mediated via down-regulation of the TNF,-induced Akt and NF-,B pathways. Caspase 3 activation by EGCG also suppressed RA synovial fibroblast growth, and this effect was mimicked by Akt and NF-,B inhibitors. Interestingly, Mcl-1 degradation by EGCG sensitized RA synovial fibroblasts to TNF,-induced PARP cleavage and apoptotic cell death. Conclusion Our findings indicate that EGCG itself induces apoptosis and further sensitizes RA synovial fibroblasts to TNF,-induced apoptosis by specifically blocking Mcl-1 expression and, hence, may be of promising adjunct therapeutic value in regulating the invasive growth of synovial fibroblasts in RA. [source]