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Cellular Targets (cellular + target)
Selected AbstractsT-cell activation: a cellular target for antiretroviral therapyEUROPEAN JOURNAL OF CLINICAL INVESTIGATION, Issue 2 2000Fidler First page of article [source] The MUC1 oncoprotein as a functional target: Immunotoxin binding to ,/, junction mediates cell killingINTERNATIONAL JOURNAL OF CANCER, Issue 1 2009Daniel B. Rubinstein Abstract MUC1, a heavily glycosylated mucin, has generated considerable interest as a target for tumor killing because of its overexpression in malignancies. Full-length MUC1 (MUC1/TM) is proteolytically cleaved after synthesis generating , and , subunits, which specifically bind in a noncovalent interaction. Although the , chain remains on the cell surface, the , chain binds in an on-and-off interaction. Most anti-MUC1 antibodies (Abs) described to date recognize epitopes within the highly immunogenic ,-chain tandem repeat. Because the ,-chain is shed, such Abs are sequestered and fail to reach MUC1-expressing cells. Immunizing with cDNA encoding MUC1/TM and the spliced MUC1/X isoform from which the tandem repeat has been deleted yielded antibodies to the MUC1 ,/, junction. Pseudomonas toxin PE38 linked to polyclonal anti-MUC1 ,/, junction Abs both bound and killed MUC1-positive malignant cells. Monoclonal DMC209 binds the MUC1 ,/, junction in both MUC1/X and MUC1/TM. When injected into SCID mice xenotransplanted with human breast cancer MDA-MB-231, monoclonal DMC209 showed significant in vivo tumor-suppressive activity. The MUC1/X ,/, junction presents a biologically-significant target in MUC1-expressing malignancies because (i) antibodies directed against cell-bound ,/, junction epitopes reach the intended cellular target, (ii) antibodies to junction epitope are internalized into cells, (iii) anti ,/, junction antibodies can effectively kill high MUC1-expressing cancer cells as antibody-toxin conjugates and (iv) antibodies targeting the MUC1 cell-bound ,/, junction results in tumor suppression in vivo. Our results indicate that cell-bound MUC1 ,/, junction, unlike shed alpha chain, represents a highly effective moiety for targeting and killing MUC1-expressing malignancies. © 2008 Wiley-Liss, Inc. [source] In Vitro Toxicity towards Kiwifruit Pollen of the Antimicrobial Peptides Magainins 1 and 2PLANT BIOLOGY, Issue 6 2007A. M. Speranza Abstract: In vitro toxicity of the antimicrobial peptides (AMPs) magainin 1 and 2 to a higher plant organism, i.e., the bicellular male gametophyte of Actinidia deliciosa (kiwifruit), is investigated. Heavy damage to the plasma membrane, the primary cellular target of the peptides, was rapidly induced: in as few as 15 min, from 70 to nearly 100 % of pollen grains were rendered unviable by 20 ,M magainin 1 or 2, respectively. Therefore, kiwifruit pollen sensitivity to natural magainins seemed to be higher if compared to the sensitivity of other pollen species towards magainin 2 amide or synthetic magainin analogues. Strong dose-dependent inhibitory effects on kiwifruit pollen performance were registered: as for magainin 1, the EC50 at 120 min varied from 14.0 (germination) to 15.8 ,M (tube elongation). The inhibitory effect was much greater when administering magainin 1 to elongating tubes rather than to ungerminated pollen grains. The two peptides differentially affected kiwifruit pollen, in line with the previously documented greater activity of magainin 2 in other cell systems. Furthermore, 20 ,M magainin 1-treated pollen grains took on a shrivelled shape within 30 min of incubation, an increasingly widespread effect with higher peptide concentration. At the ultrastructural level, both protoplast shrinkage and striking organelle alterations were evident, including chromatin condensation, swelling and loss of mitochondrial cristae, dilation of rough endoplasmic reticulum cisternae, and vacuolization of cytoplasm. To our knowledge, similar alterations in animal or plant cells treated with AMPs have not been described yet. [source] Synthetic dimer of indole-3-carbinol: Second generation diet derived anti-cancer agent in hormone sensitive prostate cancerTHE PROSTATE, Issue 5 2006Venkata P.S. Garikapaty Abstract BACKGROUND Cruciferous vegetables have been found to have anti-prostate cancer effects. The active compounds mediating these effects include indoles such as indole-3-carbinol (I3C) and isothiocyanates. I3C is unstable having tissue tropic effects and clinical utility has been partly addressed by the synthesis of a more stable dimer diindolylmethane (DIM). METHODS Anti-proliferative activity was measured by XTT assay and cytosolic proteins quantitated by Western blot analysis. RESULTS DIM (IC50 50 µM) is a better anti-proliferative agent than I3C (IC50 150 µM) in androgen dependent LNCaP cells, inhibits DNA synthesis, and growth of R1881 stimulated LNCaP cells. Androgen receptor (AR), cyclin D1, and cdk4, induced by R1881, are downregulated by DIM. DIM downregulates phosphorylated Akt and phosphatidyl inositol 3-kinase and downstream inhibition of cyclin D1 and cdk4. CONCLUSION These studies provide evidence that DIM is a second-generation chemopreventive agent with a viable cellular target and has clinical potential as an anti-prostate cancer chemopreventive. © 2005 Wiley-Liss, Inc. [source] Effect of intermittent and continuous exposure to electromagnetic fields on cultured hippocampal cellsBIOELECTROMAGNETICS, Issue 2 2002A. Boland Abstract This study was designed to assess the effect of 50 Hz electromagnetic fields (EMFs) on hippocampal cell cultures in the presence or absence of either sodium nitroprusside (SNP, a NO donor) or Fe2+ induced oxidative stress. One week old cultured rat hippocampal cells were exposed to either intermittent EMFs (IEMFs, 50 Hz, 0,5 mT, 1 min ON/OFF cycles, repeated 10 times every 2 h, 6 times/day during 48 h) or continuous EMFs (CEMFs, 50 Hz, 0,5 mT for 48 h). In a second set of experiments, the effect on such EMFs applied in combination with oxidative stress induced by 0.5 ,M Fe2+ or SNP was estimated. At the end of both sets of experiments, cell mortality was assessed by lactate dehydrogenase measurements (LDH). Neither type of exposure to EMFs was observed to modify the basal rate of cell mortality. The exposure to CEMFs in presence of either NO or Fe2+ did not induce any significant increase in cell death. However, when cells were exposed to EMFs in the presence of NO, we observed a significant increase in cell death of 11 and 23% (P<0.001) at 2.5 and 5 mT, respectively. This effect had some specificity because IEMFs did not modify the effect of Fe2+ on cell mortality. Although the effects of IEMFs reported in this study were only observed at very high intensities, our model may prove valuable in trying to identify one cellular target of EMFs. Bioelectromagnetics 23:97,105, 2002. © 2002 Wiley-Liss, Inc. [source] A Novel Cyanobacterial Nostocyclopeptide is a Potent Antitoxin against MicrocystinsCHEMBIOCHEM, Issue 11 2010Jouni Jokela Abstract Cyanobacterial hepatotoxins (microcystins and nodularins) cause numerous animal poisonings worldwide each year and are threats to human health. However, we found that extracts from several cyanobacteria isolates failed to induce hepatotoxicity even if they contained high concentrations of the liver toxin microcystin. The antitoxic activity abolishes all morphological hallmarks of microcystin-induced apoptosis, and therefore invalidates cell-based assays of the microcystin content of bloom-forming cyanobacteria. The antitoxin was purified from a cyanobacterial isolate (Nostoc sp. XSPORK 13A) from the Baltic Sea, and the activity was shown to reside in a novel cyclic peptide of the nostocyclopeptide family (nostocyclopeptide M1, Ncp-M1) that consists of seven amino acids (Tyr1 -Tyr2 - D -HSe3 - L -Pro4 - L -Val5 -(2S,4S)-4-MPr6 -Tyr7; MW=881) with an imino linkage between Tyr1 and Tyr7. Ncp-M1 did not compete with labelled microcystin for binding to protein phosphatase 2A; this explains why the antitoxin did not interfere with phosphatase-based microcystin assays. Currently used agents that interfere with microcystin action, such as inhibitors of ROS formation, microcystin uptake and Cam-kinase activity, are themselves inherently toxic. Since Ncp-M1 is potent and nontoxic it promises to become a useful mechanistic tool as soon as its exact cellular target is elucidated. [source] Laminin and fibronectin modulate inner ear spiral ganglion neurite outgrowth in an in vitro alternate choice assayDEVELOPMENTAL NEUROBIOLOGY, Issue 13 2007Amaretta R. Evans Abstract Extracellular matrix (ECM) molecules have been shown to function as cues for neurite guidance in various populations of neurons. Here we show that laminin (LN) and fibronectin (FN) presented in stripe micro-patterns can provide guidance cues to neonatal (P5) inner ear spiral ganglion (SG) neurites. The response to both ECM molecules was dose-dependent. In a LN versus poly- L -lysine (PLL) assay, neurites were more often observed on PLL at low coating concentrations (5 and 10 ,g/mL), while they were more often on LN at a high concentration (80 ,g/mL). In a FN versus PLL assay, neurites were more often on PLL than on FN stripes at high coating concentrations (40 and 80 ,g/mL). In a direct competition between LN and FN, neurites were observed on LN significantly more often than on FN at both 10 and 40 ,g/mL. The data suggest a preference by SG neurites for LN at high concentrations, as well as avoidance of both LN at low and FN at high concentrations. The results also support a potential model for neurite guidance in the developing inner ear in vivo. LN, in the SG and osseus spiral lamina may promote SG dendrite growth toward the organ of Corti. Within the organ of Corti, lower concentrations of LN may slow neurite growth, with FN beneath each row of hair cells providing a stop or avoidance signal. This could allow growth cone filopodia increased time to sample their cellular targets, or direct the fibers upward toward the hair cells. © 2007 Wiley Periodicals, Inc. Develop Neurobiol, 2007 [source] Macrophages and neurons are targets of retinoic acid signaling after spinal cord contusion injuryEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 2 2006Kirsten Schrage Abstract The physiological reactions after spinal cord injury are accompanied by local synthesis of the transcriptional activator retinoic acid (RA). RA exerts its effects by binding to retinoic acid receptors (RAR) which heterodimerize with retinoid X receptors (RXR) and then act as ligand-activated transcription factors. To identify possible cellular targets of RA we investigated protein levels and cellular distribution of retinoid receptors in the rat spinal cord at 4, 7, 14 and 21 days after a contusion injury. In the nonlesioned spinal cord, immunoreactivity for RAR,, RXR,, RXR, and RXR, was localized in the cytosol of neurons, that of RXR, and RXR, in astrocytes and that of RAR,, RXR, and RXR, in some oligodendrocytes. After contusion injury RAR, and all RXRs appeared in the cell nuclei of reactive microglia and macrophages. This nuclear staining began at 4 days, was most prominent at 7 and 14 days and had decreased at 21 days after injury. A similar nuclear translocation was also observed for the RAR,, RXR, and RXR, staining in neurons situated around the border of the contusion. These observations suggest that RA participates as a signal for the physiological responses of microglia and neurons after CNS injury. [source] A disaccharide derived from chondroitin sulphate proteoglycan promotes central nervous system repair in rats and mice,EUROPEAN JOURNAL OF NEUROSCIENCE, Issue 8 2004Asya Rolls Abstract Chondroitin sulphate proteoglycan (CSPG) inhibits axonal regeneration in the central nervous system (CNS) and its local degradation promotes repair. We postulated that the enzymatic degradation of CSPG generates reparative products. Here we show that an enzymatic degradation product of CSPG, a specific disaccharide (CSPG-DS), promoted CNS recovery by modulating both neuronal and microglial behaviour. In neurons, acting via a mechanism that involves the PKC, and PYK2 intracellular signalling pathways, CSPG-DS induced neurite outgrowth and protected against neuronal toxicity and axonal collapse in vitro. In microglia, via a mechanism that involves ERK1/2 and PYK2, CSPG-DS evoked a response that allowed these cells to manifest a neuroprotective phenotype ex vivo. In vivo, systemically or locally injected CSPG-DS protected neurons in mice subjected to glutamate or aggregated ,-amyloid intoxication. Our results suggest that treatment with CSPG-DS might provide a way to promote post-traumatic recovery, via multiple cellular targets. [source] Resveratrol inhibits proliferation of human epidermoid carcinoma A431 cells by modulating MEK1 and AP-1 signalling pathwaysEXPERIMENTAL DERMATOLOGY, Issue 7 2006Arianna L. Kim Abstract:, Resveratrol (trans -3,4,,5-trihydroxystilbene) is a naturally occurring polyphenolic phytoalexin found in grapes, and has been shown to inhibit the growth of various types of cancer cells. We investigated the mechanism of the antiproliferative effect of resveratrol in A431-transformed keratinocytes harbouring mutant p53, and show that it is accompanied by G1 cell cycle arrest, which coincides with a marked inhibition of G1 cell cycle regulatory proteins, including cyclins A and D1 and cyclin-dependent kinase (CDK)6 and p53-independent induction of p21WAF1. Cell cycle arrest was also associated with the accumulation of hypophosphorylated Rb and p27KIP1. Resveratrol inhibited mitogen-activated protein kinase/extracellular signal-regulated kinase kinase (MEK)1 > extracellular signal-regulated protein kinase (ERK)1/2 signalling, downregulated c-Jun, and suppressed activating protein (AP)-1 DNA-binding and promoter activity. In addition, the inhibition of MEK1 > ERK1/2 signalling appears to be independent of retinoblastoma protein (pRb) hypophosphorylation in A431 cells, as PD098059 did not suppress pRb phosphorylation. Our results demonstrate that resveratrol affects multiple cellular targets in A431 cells, and that the downregulation of both AP-1 and pRb contributes to its antiproliferative activity in these cells. [source] Dothistroma (red-band) needle blight of pines and the dothistromin toxin: a reviewFOREST PATHOLOGY, Issue 3 2004R. E. Bradshaw Summary Dothistroma (red-band) needle blight has been a problem in plantations of exotic pines in the southern hemisphere for many decades. The prevalence of this disease is currently increasing in the northern hemisphere and is now affecting trees in their native ranges. The fungal pathogen Mycosphaerella pini with its anamorph Dothistroma pini, which is responsible for the disease, produces a toxin, dothistromin, that is closely related to the potent carcinogen, aflatoxin. Understandably this has provoked concern about possible effects on the health of forestry workers. This review gives a broad coverage of literature on both disease and toxin. The fungus has a complicated taxonomy with many synonyms and in most countries only the anamorph is found. It is a necrotrophic pathogen that kills needle tissue and completes its life cycle in the lesion thus formed. Dispersal of the disease is normally by rain splash of conidiospores but there is evidence that long range dispersal has occurred by transport of contaminated plant tissue and by wind/cloud dispersal of spores in air currents. The severity of disease is affected by humidity, temperature and light. There is variation in susceptibility of different Pinus species and some achieve increased resistance with age. The current method of control in southern hemisphere plantation forests is through spraying with copper fungicides and, with P. radiata, increased disease resistance has been achieved through a breeding programme. The dothistromin toxin is a difuroanthraquinone and is similar in structure to the aflatoxin precursor versicolorin B. Part of a gene cluster encoding dothistromin biosynthetic genes has been cloned and this has confirmed parallels between the dothistromin and aflatoxin biosynthetic pathways. Dothistromin produces damaging oxygen radicals by reductive oxygen activation rather than by photosensitization, but is also thought to exert its toxic effects on specific cellular targets. Studies have shown that dothistromin is a weak mutagen and clastogen and is therefore a potential carcinogen. Although the risks to forest workers are considered very low it is prudent to avoid unnecessary exposure during periods when dothistromin levels are likely to be at their peak. Résumé La maladie des bandes rouges causée par Dothistroma est un problème dans les plantations de pins exotiques de l'hémisphère sud depuis de nombreuses années. La prévalence de cette maladie est en augmentation dans l'hémisphère nord et affecte maintenant les pins dans leurs régions d'origine. Le champignon pathogène Dothistroma pini, responsable de la maladie, produit une toxine, la dothistromine, proche de l'aflatoxine qui est un puissant carcinogène. Ceci pose donc la question des effets possibles sur la santé des travailleurs forestiers. Cette revue repose sur une large couverture de la littérature concernant aussi bien la maladie que la toxine. Le champignon a une taxonomie complexe avec de nombreux synonymes, et seul l'anamorphe se rencontre dans de nombreux pays. C'est un champignon nécrotrophe qui tue les tissus de l'aiguille et réalise son cycle biologique dans la lésion ainsi formée. La dissémination de la maladie s'effectue principalement par éclaboussures de pluie contenant les conidies mais une dissémination à longue distance a été mise en évidence par transport de matériel contaminé ou par dissémination des spores par le vent ou les nuages dans les courants aériens. La sévérité de la maladie est affectée par l'humidité, la température et la lumière. Il existe des différences de sensibilité entre espèces de Pinus, et certaines présentent une résistance accrue avec l'âge. La méthode actuelle de lutte dans les forêts de plantations de l'hémisphère sud consiste à pulvériser des fongicides à base de cuivre ; dans le cas de Pinus radiata, une augmentation de la résistance a été obtenue grâce à un programme d'amélioration génétique. La toxine dothistromine est une difuroanthraquinone, similaire en structure à la versicolorine B, précurseur de l'aflatoxine. Une partie d'une batterie de gènes comprenant des gènes de biosynthèse de la dothistromine a été clonée, confirmant les analogies entre les voies de biosynthèse de la dothistromine et de l'aflatoxine. La dothistromine produit des radicaux oxygène nocifs par activation de la réduction de l'oxygène plutôt que par photosensibilisation, mais ses effets toxiques s'exercent aussi probablement sur des sites cellulaires spécifiques. Des études montrent que la dothistromine est un mutagène et clastogène faible, et donc potentiellement carcinogène. Bien que les risques pour les ouvriers forestiers soient considérés comme très faibles, il est prudent d'éviter dans la mesure du possible de s'exposer dans les périodes où les niveaux de dothistromine sont supposés élevés. Zusammenfassung Die Dothistroma -Nadelbräune ist in der Südhemisphäre in Plantagen mit exotischen Kiefernarten seit vielen Jahren ein Problem. In der Nordhemisphäre nimmt die Bedeutung dieser Krankheit derzeit zu und sie befällt nun Bäume auch in ihren natürlichen Verbreitungsgebieten. Der Erreger ist der Ascomycet Mycosphaerella pini (Anamorphe: Dothistroma pini). Der Pilz bildet das Toxin Dothistromin, das eng mit dem hochtoxischen Karzinogen Aflatoxin verwandt ist. Daraus ergab sich die Frage nach möglichen Nebenwirkungen dieser Baumkrankheit auf die Gesundheit von Waldarbeitern. Dieser Review fasst die Information über die Krankheit und das Toxin zusammen. Der Pilz hat eine komplizierte Taxonomie mit vielen Synonymen und in den meisten Ländern wurde nur die Anamorphe nachgewiesen. Er ist ein nekrotrophes Pathogen, das Blattgewebe abtötet, und in den so gebildeten Läsionen seinen Lebenszyklus abschliesst. Der normale Ausbreitungsweg der Krankheit erfolgt über Konidiosporen mit Regentropfen, aber es gibt auch Hinweise auf einen Ferntransport mit infiziertem Pflanzenmaterial und über die Verbreitung von Sporen mit dem Wind bzw. Wolken in Luftströmungen. Die Krankheitsintensität wird durch Luftfeuchte, Temperatur und Licht beeinflusst. Es gibt Unterschiede in der Anfälligkeit zwischen verschiedenen Kiefernarten und manche davon werden mit zunehmendem Alter resistenter. Derzeit werden in Plantagen der südlichen Hemisphäre Kupferfungizide zur Kontrolle dieser Krankheit eingesetzt und für Pinus radiata wurde in Züchtungsprogrammen eine erhöhte Resistenz erreicht. Das Toxin Dothistromin ist ein Difuroanthrachinon und ähnelt in seiner Struktur dem Aflatoxin-Präkursor Versicolorin B. Ein Teil des Genclusters, das die Dothistromin-Biosynthese codiert, wurde geklont, und es wurden dabei Parallelen zwischen dem Dothistromin- und dem Aflatoxin-Biosyntheseweg bestätigt. Dothistromin bildet schädliche Sauerstoffradikale (wahrscheinlich eher durch reduktive Sauerstoffaktivierung als durch Photosensibilisierung), es dürfte aber auch auf spezifische Zellkomponenten toxisch wirken. Dothistromin zeigt schwache mutagene und chromosomenschädigende Wirkungen und ist deshalb ein potentielles Karzinogen. Obwohl das Risiko für Waldarbeiter als gering eingeschätzt wird, sollte man in Perioden, in denen der Dothistromingehalt hoch sein dürfte, eine unnötige Exposition vermeiden. [source] The dietary histone deacetylase inhibitor sulforaphane induces human ,-defensin-2 in intestinal epithelial cellsIMMUNOLOGY, Issue 2 2008Markus 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] Mechanisms of regulatory T-cell suppression , a diverse arsenal for a moving targetIMMUNOLOGY, Issue 1 2008Dorothy K. Sojka Summary Naturally-occurring regulatory T cells (Tregs) are emerging as key regulators of immune responses to self-tissues and infectious agents. Insight has been gained into the cell types and the cellular events that are regulated by Tregs. Indeed, Tregs have been implicated in the control of initial activation events, proliferation, differentiation and effector function. However, the mechanisms by which Tregs disable their cellular targets are not well understood. Here we review recent advances in the identification of distinct mechanisms of Treg action and of signals that enable cellular targets to escape regulation. Roles for inhibitory cytokines, cytotoxic molecules, modulators of cAMP and cytokine competition have all been demonstrated. The growing number of inhibitory mechanisms ascribed to Tregs suggests that Tregs take a multi-pronged approach to immune regulation. It is likely that the relative importance of each inhibitory mechanism is context dependent and modulated by the inflammatory milieu and the magnitude of the immune response. In addition, the target cell may be differentially susceptible or resistant to distinct Treg mechanisms depending on their activation or functional status at the time of the Treg encounter. Understanding when and where each suppressive tool is most effective will help to fine tune therapeutic strategies to promote or constrain specific arms of Treg suppression. [source] NF-,B inhibition triggers death of imatinib-sensitive and imatinib-resistant chronic myeloid leukemia cells including T315I Bcr-Abl mutantsINTERNATIONAL JOURNAL OF CANCER, Issue 2 2009Nadia Lounnas Abstract The Bcr-Abl inhibitor imatinib is the current first-line therapy for all newly diagnosed chronic myeloid leukemia (CML). Nevertheless, resistance to imatinib emerges as CML progresses to an acute deadly phase implying that physiopathologically relevant cellular targets should be validated to develop alternative therapeutic strategies. The NF-,B transcription factor that exerts pro-survival actions is found abnormally active in numerous hematologic malignancies. In the present study, using Bcr-Abl-transfected BaF murine cells, LAMA84 human CML cell line and primary CML, we show that NF-,B is active downstream of Bcr-Abl. Pharmacological blockade of NF-,B by the IKK2 inhibitor AS602868 prevented survival of BaF cells expressing either wild-type, M351T or T315I imatinib-resistant mutant forms of Bcr-Abl both in vitro and in vivo using a mouse xenograft model. AS602868 also affected the survival of LAMA84 cells and of an imatinib-resistant variant. Importantly, the IKK2 inhibitor strongly decreased in vitro survival and ability to form hematopoietic colonies of primary imatinib resistant CML cells including T315I cells. Our data strongly support the targeting of NF-,B as a promising new therapeutic opportunity for the treatment of imatinib resistant CML patients in particular in the case of T315I patients. The T315I mutation escapes all currently used Bcr-Abl inhibitors and is likely to become a major clinical problem as it is associated with a poor clinical outcome. © 2009 UICC [source] Calpain-mediated breakdown of cytoskeletal proteins contributes to cholecystokinin-induced damage of rat pancreatic aciniINTERNATIONAL JOURNAL OF EXPERIMENTAL PATHOLOGY, Issue 4 2009Heike Weber Summary The cytosolic cysteine protease calpain is implicated in a multitude of cellular functions but also plays a role in cell damage. Our previous results suggest that an activation of calpain accompanied by a decrease in its endogenous inhibitor calpastatin may contribute to pancreatic damage during cerulein-induced acute pancreatitis. The present study aimed at the time course of secretagogue-induced calpain activation and cellular substrates of the protease. Isolated rat pancreatic acini were incubated with a supramaximal concentration of cholecystokinin (0.1 ,M CCK) for 30 min in the presence or absence of the calpain inhibitor Z-Val-Phe methyl ester (100 ,M ZVP). The activation of calpain and the expression of calpastatin and the actin cytoskeleton-associated proteins ,II-spectrin, E-cadherin and vinculin were studied by immunoblotting. The cell damage was assessed by lactate dehydrogenase release and ultrastructural analysis including fluorescence-labelled actin filaments. Immediately after administration, CCK led to activation of both calpain isoforms, ,- and m-calpain. The protease activation was accompanied by a decrease in the E-cadherin level and formation of calpain-specific breakdown products of ,II-spectrin. A calpain-specific cleavage product of vinculin appeared concomitantly with changes in the actin filament organization. No effect of CCK on calpastatin was found. Inhibition of calpain by ZVP reduced CCK-induced damage of the actin-associated proteins and the cellular ultrastructure including the actin cytoskeleton. The results suggest that CCK-induced acinar cell damage requires activation of calpain and that the actin cytoskeleton belongs to the cellular targets of the protease. [source] Microtubules: dynamics, drug interaction and drug resistance in LeishmaniaJOURNAL OF CLINICAL PHARMACY & THERAPEUTICS, Issue 5 2002K. G. Jayanarayan MS (Pharm) Summary Microtubules are cytoskeletal polymers essential for the survival of all eukaryotes. These proteins are the proposed cellular targets of many anticancerous, antifungal and antihelminthic drugs. Sufficient differences exist between the microtubules of kinetoplastid parasites like Leishmania and humans to explore the selective targeting of these proteins for therapeutic purposes. This review describes the basic structure of microtubules and its dynamics in general, with specific insights into leishmanial microtubules, the salient features of microtubule,drug interactions including the specificity of certain drugs for parasitic microtubules. Chemotherapy against leishmanial parasites is failing because of the emergence of drug resistant strains. The possible mechanisms of resistance to antimicrotubule agents along with insights into the role of microtubules in mediating drug resistance in Leishmania are discussed. [source] IL-10 and IL-4 regulate type-I and type-II IL-1 receptors expression on IL-1,-activated mouse primary astrocytesJOURNAL OF NEUROCHEMISTRY, Issue 4 2001F. Pousset When activated by its ligand, the interleukin receptor type I (IL-1RI) transduces signals in cooperation with the IL-1 receptor accessory protein (IL-1RacP). In contrast, IL-1RII functions as a decoy receptor without participating in IL-1 signalling. Brain astrocytes are cellular targets of IL-1 and play a pivotal role in brain responses to inflammation. The regulation of IL-1 receptors on astrocytes by anti-inflammatory cytokines such as IL-4 and IL-10 has not been studied, despite its importance for understanding the way these cells respond to IL-1. Using RT-PCR, we first showed that the expression of IL-1RI and IL-1RII, but not IL-1RacP, mRNAs are up-regulated by IL-1, in a time-dependent manner. Using a radioligand binding technique, we then showed that astrocytes display an equivalent number of IL-1RI and IL-1RII. IL-1, decreases the number of IL-1RI binding sites, whereas it increases those of IL-1RII. IL-4 and IL-10 both up-regulate IL-1RII IL-1,-induced, but only IL-4 does so for IL-1RI. At the protein level, IL-4 and IL-10 dramatically reverse the ability of IL-1, to inhibit expression of IL-1RI but neither affects the ability of IL-1, to enhance the number of IL-1RII. Collectively, these results establish the existence of receptor cross-talk between pro- and anti-inflammatory cytokines on a critical type of cell that regulates inflammatory events in the brain. [source] Ethanol Increases Fetal Human Neurosphere Size and Alters Adhesion Molecule Gene ExpressionALCOHOLISM, Issue 2 2008Sharada D. Vangipuram Background:, Ethanol (ETOH) consumption by pregnant women can result in Fetal Alcohol Spectrum Disorder (FASD). To date, the cellular targets and mechanisms responsible for FASD are not fully characterized. Our aim was to determine if ETOH can affect fetal human brain-derived neural progenitor cells (NPC). Methods:, Neural progenitor cells were isolated by positive selection from normal second trimester fetal human brains (n = 4) and cultured, for up to 72 hours, in mitogenic media containing 0, 1, 10, or 100 mM ETOH. From 48 to 72 hours in culture, neurospheres generated in these conditions were filmed using time-lapse video microscopy. At the end of 72 hours, neurosphere diameter and roundness were measured using videographic software. Mitotic phase analysis of cell-cycle activity and apoptotic cell count were also performed at this time, by flow cytometry using propidium iodide (PI) staining. Real-time PCR was used to estimate expression of genes associated with cell adhesion pathways. Results:, Neurosphere diameter correlated positively (r = 0.87) with increasing ETOH concentrations. There was no significant difference in cell-cycle activity and no significant increase in apoptosis with increasing ETOH concentrations. Time-lapse video microscopy showed that ETOH (100 mM) reduced the time for neurosphere coalescence. Real-time PCR analysis showed that ETOH significantly altered the expression of genes involved in cell adhesion. There was an increase in the expression of , and , Laminins 1, , Integrins 3 and 5, Secreted phosphoprotein1 and Sarcoglycan ,. No change in the expression of , Actin was observed while the expression of , Integrin 2 was significantly suppressed. Conclusions:, ETOH had no effect on NPC apoptosis but, resulted in more rapid coalescence and increased volume of neurospheres. Additionally, the expression of genes associated with cell adhesion was significantly altered. ETOH induced changes in NPC surface adhesion interactions may underlie aspects of neurodevelopmental abnormalities in FASD. [source] Terminal nerve and visionMICROSCOPY RESEARCH AND TECHNIQUE, Issue 1-2 2004U. Behrens Abstract The vertebrate retina receives efferent input from different parts of the central nervous system. Efferent fibers are thought to influence retinal information processing but their functional role is not well understood. One of the best-described retinopetal fiber systems in teleost retinae belongs to the terminal nerve complex. Gonadotropin-releasing hormone (GnRH) and molluscan cardioexcitatory tetrapeptide (FMRFamide)-containing fibers from the ganglion of the terminal nerve form a dense fiber plexus in the retina at the border of the inner nuclear and inner plexiform layer. Peptide-containing fibers surround and contact perikarya of dopaminergic interplexiform cells in teleost retina. In vitro experiments demonstrated that exogenously supplied GnRH mediates dopaminergic effects on the membrane potential and on the morphology of dendritic tips (spinules) of cone horizontal cells. These effects can be specifically blocked by GnRH-antagonists, indicating that the release of dopamine and dopamine-dependent effects on light adaptation of retinal neurons are affected by the terminal nerve complex. Recent data have shown that olfactory information has an impact on retinal physiology, but its precise role is not clear. The efferent fiber of the terminal nerve complex is one of the first retinopetal fiber systems for which the sources of the fibers, their cellular targets, and several physiological, morphological, and behavioral effects are known. The terminal nerve complex is therefore a model system for the analysis of local information processing which is influenced by a distinct fiber projection. Microsc. Res. Tech. 65:25,32, 2004. © 2004 Wiley-Liss, Inc. [source] Fluorescence Study on the Interaction Between Hypericin and Lens Protein ",-Crystallin"PHOTOCHEMISTRY & PHOTOBIOLOGY, Issue 4 2009Tareq Youssef Hypericin has been reported as a potent photosensitizing agent exhibiting antiviral, antibacterial, antineoplastic activities. Although its photophysics and mode of action are strongly modulated by the binding protein, detailed information about its mechanism of interaction with possible cellular targets, including proteins, is still lacking. Previous in vitro studies demonstrated that hypericin can be uptaken by intact lens and is able to bind to the major lens protein ",-crystallin." In this study, the mechanism of interaction of this potent drug with ,-crystallin was studied using the chemical denaturant guanidine hydrochloride (GdnHCl) and the hydrophobic surface probe, 8-anilino-1-naphthalenesulfonic acid (ANS). Fluorescence measurements showed that the increased exposure of tryptophan resulting from partial unfolding of ,-crystallin incubated with 1.0 mol L,1 of GdnHCl corresponds to the maximum accessibility of hydrophobic sites to ANS at the same GdnHCl concentration. Interestingly at this additional hydrophobicity of the protein, hypericin exhibited its maximum fluorescence intensity. This in vitro study implied that hydrophobic sites of ,-crystallin play a significant role in its interaction with hypericin. The binding between ,-crystallin and hypericin was found to be enhanced by partial perturbation of the protein. [source] Residual tumor cells are unique cellular targets in glioblastoma,ANNALS OF NEUROLOGY, Issue 2 2010Martin Glas MD Residual tumor cells remain beyond the margins of every glioblastoma (GBM) resection. Their resistance to postsurgical therapy is considered a major driving force of mortality, but their biology remains largely uncharacterized. In this study, residual tumor cells were derived via experimental biopsy of the resection margin after standard neurosurgery for direct comparison with samples from the routinely resected tumor tissue. In vitro analysis of proliferation, invasion, stem cell qualities, GBM-typical antigens, genotypes, and in vitro drug and irradiation challenge studies revealed these cells as unique entities. Our findings suggest a need for characterization of residual tumor cells to optimize diagnosis and treatment of GBM. ANN NEUROL 2010;68:264,269 [source] Human Immunodeficiency Virus Infection of the Brain: Pitfalls in Evaluating Infected/Affected Cell PopulationsBRAIN PATHOLOGY, Issue 1 2004Stephanie J. Bissel Monocyte/macrophages and CD4 T-cells are the primary hematopoietic targets of productive HIV infection. In the brain, potential cellular targets for HIV infection include perivascular and parenchymal macrophages/microglia, oligodendrocytes, endothelia, neurons, and astrocytes. We examine evidence of productive and non-productive infection for each cell type in the brains of HIV-infected patients with and without HIV encephalitis. Despite the voluminous literature and substantial experimental effort over the past two decades, evidence for productive infection of any brain cell other than macrophages is left wanting. [source] Mechanisms of Salmonella entry into host cellsCELLULAR MICROBIOLOGY, Issue 9 2007Kim Thien Ly Summary Salmonella enterica is an enteric bacterial pathogen that causes a variety of food and water-borne diseases ranging from gastroenteritis to typhoid fever. Ingested bacteria colonize the intestinal epithelium by triggering their own phagocytosis, using a sophisticated array of effector proteins that are injected into the host cell cytoplasm through a type III secretion apparatus. The synergistic action of these secreted effectors leads to a dramatic reorganization of the host actin cytoskeleton, resulting in vigorous membrane protrusion and the engulfment of attached bacteria. Analysis of these effector proteins and identification of their cellular targets has provided insight into the molecular mechanisms by which bacteria can subvert the host signalling and cytoskeletal machinery for their own purposes. This review is intended to summarize our current understanding of the tools used by Salmonella to enter host cells, with a focus on effectors that modulate the actin cytoskeleton. [source] Show me the substrates: modulation of host cell function by type IV secretion systemsCELLULAR MICROBIOLOGY, Issue 6 2003Hiroki Nagai Summary Evidence for the involvement of type IV protein secretion systems in bacterial virulence is accumulating. Many of the substrate proteins secreted by type IV systems either hijack or interfere with specific host cell pathways. These substrates can be injected directly into host cells via the type IV apparatus or are secreted by the type IV machinery in a state that allows them to gain access to cellular targets without the further assistance of the type IV system. Arguably, the protein substrates of most type IV secretion systems remain undiscovered. Here, we review the activities of known type IV substrates and discuss the putative roles of unidentified substrates. [source] Chemically Induced Cardiomyogenesis of Mouse Embryonic Stem CellsCHEMBIOCHEM, Issue 2 2010Albrecht Berkessel Prof. Dr. Abstract A transgenic murine embryonic stem (ES) cell lineage expressing enhanced green fluorescent protein (EGFP) under the control of ,-myosine heavy chain (,-MHC) promoter (p,-MHC-EGFP) was used to investigate the effects of (thio)urea and cinchona alkaloid derivatives on cardiomyogenesis. The screening of the compounds yielded cardiomyogenesis inducing substances with good (IV-5, V-4) to very good activities (II-16, IV-8), as determined by a 50 to 80,% increase in the EGFP fluorescence compared to untreated cells. Time-dependent screening approaches in which compounds were added at different developmental stages of the ES cells appeared to be of limited suitability for the identification of potential cellular targets. [source] Paramagnetic Liposomes as Innovative Contrast Agents for Magnetic Resonance (MR) Molecular Imaging ApplicationsCHEMISTRY & BIODIVERSITY, Issue 10 2008Enzo Terreno Abstract This article illustrates some innovative applications of liposomes loaded with paramagnetic lanthanide-based complexes in MR molecular imaging field. When a relatively high amount of a GdIII chelate is encapsulated in the vesicle, the nanosystem can simultaneously affect both the longitudinal (R1) and the transverse (R2) relaxation rate of the bulk H2O H-atoms, and this finding can be exploited to design improved thermosensitive liposomes whose MRI response is not longer dependent on the concentration of the probe. The observation that the liposome compartmentalization of a paramagnetic LnIII complex induce a significant R2 enhancement, primarily caused by magnetic susceptibility effects, prompted us to test the potential of such agents in cell-targeting MR experiments. The results obtained indicated that these nanoprobes may have a great potential for the MR visualization of cellular targets (like the glutamine membrane transporters) overexpressing in tumor cells. Liposomes loaded with paramagnetic complexes acting as NMR shift reagents have been recently proposed as highly sensitive CEST MRI agents. The main peculiarity of CEST probes is to allow the MR visualization of different agents present in the same region of interest, and this article provides an illustrative example of the in vivo potential of liposome-based CEST agents. [source] | ||||||||||||||||