Dissertations / Theses on the topic 'PKC inhibitors'
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Aaltonen, V. (Vesa). "PKC and neurofibromin in the molecular pathology of urinary bladder carcinoma:the effect of PKC inhibitors on carcinoma cell junctions, movement and death." Doctoral thesis, University of Oulu, 2007. http://urn.fi/urn:isbn:9789514285899.
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This study examined the role of tumor suppressor neurofibromin and Protein kinase C (PKC) in urinary bladder cancer, and the effect of PKC inhibitors on cancer cell behaviour.
Tumor suppressor protein neurofibromin is a product of the NF1 gene, a mutation of which causes the most common hereditary tumor syndrome, type 1 neurofibromatosis. NF1 gene mutations and changes in expression have been demonstrated in malignancies, unrelated to type 1 neurofibromatosis. The best known function of neurofibromin is its Ras GTPase accelerating function. Thus, it functions as a Ras inactivator. This study demonstrated for the first time that the NF1 gene is expressed in normal and malignant urinary bladder epithelium and in cultured bladder carcinoma cells in mRNA and at the protein level. Furthermore, neurofibromin expression is decreased during bladder carcinogenesis. It can be speculated that this may lead to increased Ras activity in urinary bladder cancer.
The PKC family is composed of several different isoenzymes which are responsible for a number of important intracellular events and cellular functions. Many of these are also important in cancer development and progression. The results demonstrate changes in expression of PKC α and βI isoenzymes in urinary bladder carcinoma. Furthermore, the results relate the increased expression of isoenzymes to increased PKC enzyme activity and the high proliferation rate of the cancer cells. In addition, this study utilizes small molecular inhibitors of PKC isoenzymes in order to study the effect of the inhibition of these isoenzymes on cancer cell behaviour in vitro and in vivo. The study mainly focuses on the function of PKC α and βI isoenzymes and on the effects of inhibition of these by using Go6976. The results show that Go6976 inhibits cancer cell growth, migration and invasion in vitro, and tumor growth in a mouse model. The use of Go6976 induces changes in desmosomes and adherens junctions, and in focal adhesions and hemidesmosomes. The results also show that Go6976 functions as a cell cycle checkpoint abrogator and increases the cytotoxicity of two classical chemotherapeutic agents, doxorubicin and paclitaxel. In the future, it may be possible that Go6976 or related drugs could be used in clinical cancer treatments.
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Ratnayake, Wishrawana Sarathi Bandara. "Role of Oncogenic Protein Kinase C-iota in Melanoma Progression; A Study Based on Atypical Protein Kinase-C Inhibitors." Scholar Commons, 2019. https://scholarcommons.usf.edu/etd/7895.
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Irrespective of plentiful efforts to enhance primary prevention and early detection, the number of melanoma cases in the United States has increased steadily over the past 30 years, thus greatly affecting public health and the economy. We have investigated the effects of five novel aPKC inhibitors; 2-acetyl-1,3-cyclopentanedione (ACPD), 3,4-Diaminonaphthalene-2,7-disulfonic acid (DNDA), [4-(5-amino-4-carbamoylimidazol-1-yl)-2,3-dihydroxycyclopentyl] methyl dihydrogen phosphate (ICA-1T) along with its nucleoside analog 5-amino-1-((1R,2S,3S,4R)-2,3-dihydroxy-4-methylcyclopentyl)-1H-imidazole-4-carboxamide (ICA-1S) and 8-hydroxy-1,3,6-naphthalenetrisulfonic acid (ζ-Stat) on cell proliferation, apoptosis, migration and invasion of two malignant melanoma cell lines compared to normal melanocyte cell lines. Molecular docking data suggested that both ACPD and DNDA specifically bind to protein kinase C-zeta (PKC-ζ) and PKC-iota (PKC-ι) while both ICA-1 compounds specifically bind to PKC-ι, and ζ-Stat showed a high affinity towards PKC-ζ. Kinase activity assays were carried out to confirm these observations. Results suggest that PKC-ι is involved in melanoma malignancy than PKC-ζ. Both isoforms promote the activation of nuclear factor (NF)-κB and protein kinase B (AKT) thereby supporting survival and progression. In addition, we demonstrated that PKC-ι induced the metastasis of melanoma cells by activating Vimentin, and PKC-ι inhibition downregulated epithilial-mesencymal transition (EMT), while inducing apoptosis. Of note, PKC-ἱ specific inhibitors downregulated the expression of both PKC-ι and phosphorylated PKC-ι, suggesting that PKC-ι plays a role in regulating its own expression in melanoma. We also report the underlaying mechanisms of the transcriptional regulation of PKC-ι (PRKCI gene) expression in melanoma. c-Jun, interferon-stimulated gene factor 3 (ISGF3), paired box gene 3 (PAX3), early growth response protein 1 (EGR1) and forkhead box protein O1 (FOXO1), which bind on or near the promoter sequence of the PRKCI gene, were analyzed for their role in PKC-ι regulation in SK-MEL-2 and MeWo cell lines. We silenced selected transcription factors using siRNA, and the results revealed that the silencing of c-Jun and FOXO1 significantly altered the expression of PRKCI. The levels of both phosphorylated and total PKC-ι increased upon FOXO1 silencing and decreased upon c-Jun silencing, suggesting that c-Jun acts as an upregulator, while FOXO1 acts as a downregulator of PRKCI expression. We also used a multiplex ELISA to analyze multiple pathways other than NF-κB that were affected by treatment with PKC-ι inhibitor. The silencing of NF-κB p65 and PKC-ι by siRNA suggested that the regulation of PKC-ι expression was strongly associated with FOXO1. In addition, we observed a significant decrease in the mRNA levels of both interleukin (IL)-6 and IL-8, with a significant increase in the levels of IL-17E and intercellular adhesion molecule 1 (ICAM-1) upon the knockdown of expression of PKC-ι in both cell lines. This suggested that PKC-ι expression was affected by these cytokines in an autocrine manner. Overall, the findings of this study suggest that PKC-ι inhibition suppresses its own expression, diminishing oncogenic signaling, while upregulating anti-tumor signaling, thus rendering it an effective novel biomarker for use in the design of novel targeted therapeutics for melanoma.
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Panek, Anna [Verfasser]. "Rolle des Connective Tissue Growth Factors (CTGF) und des PKC-enhanced Protein-Phosphatase 1 Inhibitors (KEPI) für die Funktion des adulten Herzen : Studien an transgenen Tiermodellen / Anna Naila Panek." Berlin : Freie Universität Berlin, 2008. http://d-nb.info/1023375095/34.
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Lakshmanan, Aparna. "Modulation of Sodium Iodide Symporter-mediated Thyroidal Radioiodide Uptake by Small Molecule Inhibitors, Natural Plant-based Products and microRNAs." The Ohio State University, 2015. http://rave.ohiolink.edu/etdc/view?acc_num=osu1429407914.
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Esvan, Yannick. "Conception et synthèse de nouveaux composés hétéroaromatiques inhibiteurs potentiels de kinases." Thesis, Clermont-Ferrand 2, 2016. http://www.theses.fr/2016CLF22743.
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Depuis la mise en évidence de l’existence des protéines kinases vers la fin des années 1950 cette famille d’enzymes s’est vu attribuer d’importants rôles dans divers mécanismes pathologiques notamment dans des processus de cancérisations. Plus récemment ces enzymes ont été identifiées comme potentiellement impliquées dans d’autres types de maladies telles que les maladies neurodégénératives.Deux projets de recherche seront présentés. Le premier projet expose la conception et la synthèse de nouveaux composés tricycliques de la famille des pyrido[3,4-g]quinazolines. Les propriétés inhibitrices de kinases des premiers dérivés ont été évaluées sur un panel de cinq kinases (CDK5, CK1, GSK3, CLK1 and DYRK1A) connues pour leurs implications dans la maladie d’Alzheimer. L’intérêt de ces nouveaux squelettes tricycliques comme inhibiteurs de kinases a été validé par des activités inhibitrices nanomolaire à l’encontre des kinases DYRK1A et CLK1. D’autre part l’obtention de structures co-crystallographiques d’interaction de deux dérivés avec le site ATP de la kinase CLK1 a permis de rationnaliser la substitution du motif pyrido[3,4-g]quinazoline. Le second projet présente le développement d’un nouveau dérivé de la staurosporine aglycone (K252c) dans lequel la partie lactame a été remplacée par un noyau pyrazole. Une étude préliminaire des propriétés biologiques de l’indolopyrazolocarbazole obtenu met en avant une cytotoxicité, du même ordre de grandeur que K252c, contre les lignées cellulaires K562 (leucémie humaine) et HCT116 (carcinome du colon). En revanche, le composé chef de file s’est révélé être un faible inhibiteur de cibles connues de K252c, les isoformes α and γ de la protéine kinase C et présente un bon potentiel inhibiteur des kinases Pim 1-3. Ce nouveau chemotype pourrait être un inhibiteur de kinases prometteurIn 1950’s protein kinases were found to play a critical role in cell signaling, rising strong research potential for this enzyme family. Initially investigated for their implications in cancerogenesis they were more recently found to be involved in a wide variety of diseases including neurodegenerative pathologies. Herein will be presented two research projects that offer bright new perspectives for the inhibition of kinases involved whether in neurodegenerative diseases or cancers.First, the design and synthesis of new pyrido[3,4-g]quinazoline derivatives will be described as well as their protein kinase inhibitory potencies toward five CMGC family members (CDK5, CK1, GSK3, CLK1 and DYRK1A) that are known to play a potential role in Alzheimer’s disease. The interest for this original tricyclic heteroaromatic scaffold as modulators of CLK1/ DYRK1A activity was validated by nanomolar potencies. CLK1 co-crystal structures with two inhibitors revealed the binding mode of these compounds within the ATP-binding pocket and led to the synthesis of new diversely substituted pyrido[3,4-g]quinazolines.Then the synthesis of a new derivative of the staurosporine aglycon (K252c), in which the lactam ring was replaced by a pyrazole moiety, will be depicted. The resulting indolopyrazolocarbazole inhibited Pim isoforms 1–3 whereas it did not impair the activity of two known targets of K252c, protein kinase C isoforms α and γ . The lead compound exhibited same cytotoxic activity as K252c toward both human leukemia and colon carcinoma cell lines (K562 and HCT116), strongly suggesting that this new scaffold deserves further investigations for treatment of malignancies associated with kinases activities
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Trachsel, Sébastien Auguste Charles. "Selective responses (Actin polymerization, shape changes, locomotion, pinocytosis) to the PKC-inhibitor Ro 31-8220 suggest thath PKC discriminately regulates functions of human blood lymphocytes /." [S.l : s.n.], 1996. http://www.ub.unibe.ch/content/bibliotheken_sammlungen/sondersammlungen/dissen_bestellformular/index_ger.html.
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Makhoul, Stephanie [Verfasser]. "GPIb-V-IX- and GPVI-specific intracellular signaling and their regulation by PKA/PKG-dependent inhibitory pathways in washed human platelets / Stephanie Makhoul." Mainz : Universitätsbibliothek Mainz, 2019. http://d-nb.info/1194096891/34.
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Hofstetter, Barbara. "Klinisch relevante PKC-Inhibitoren als radiosensibilisierende Substanzen für Tumorzellen genaue Untersuchung der Wirkung auf die PI3K/Akt-Signalübermittlungskaskade /." Zürich : ETH, Eidgenössische Technische Hochschule Zürich, Departement Angewandte Biowissenschaften, Institut für Pharmazeutische Wissenschaften, 2001. http://e-collection.ethbib.ethz.ch/show?type=dipl&nr=5.
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Heinzmann, Kathrin. "Investigating the effects or AKT/PKB inhibitors on [18F]-FDG uptake in cancer cells." Thesis, Institute of Cancer Research (University Of London), 2012. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.553702.
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Increased AKT activity due to hyperactivated PI3K or inactivation of PTEN is frequently found in cancer. Thus the development of AKT inhibitors and pharmacodynamic (PO) biomarkers for AKT inhibition is of major interest for cancer therapy. [18F]-FDG PET is a non-invasive imaging technique which allows the isualization of glucose metabolism. As AKT activity has been associated with glucose uptake and phosphorylation, it has been hypothesized that CSF]-FOG PET is a potential PO biomarker for the assessment of AKT inhibitor action in the clinic. The primary aim of the work presented in this thesis was to investigate whether (18F]-FOG uptake can be used as a PO biomarker for AKT inhibition. An in vitro system was established including assembly of a gamma counter which was used to demonstrate that both ATP competitive and allosteric inhibitors of AKT block [18F]-FDG uptake in the PTEN null U87MG human glioblastoma cell line. These results led to a detailed investigation of the mechanism(s) by which the AKT inhibitors affect [18F]-FDG uptake. However, no change in expression or translocation of the glucose transporters GLUT-1 and GLUT-4, or in hexokinase phosphorylation or activity was seen after inhibitor treatment. Reduction of [18F]-FDG uptake was then confirmed with a later generation of AKT inhibitors and in a second cellular model, namely the human breast cancer cell line BT474. These results were also complimented by MRS studies of lactate production. AKT knock down studies using siRNA were also carried out. However, complete functional removal of AKT was not achieved, although AKT protein level was substantially reduced. In addition, in vivo studies demonstrated that [18F]-FDG uptake and AKT signalling were decreased in U87MG xenografts by the AKT inhibitor CCT129254. Hence, it was concluded that [18F]-FDG uptake can be used as a PD biomarker of AKT inhibition for the types of compound studied.
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Gottlieb, Rachel. "Use of S. pombe to Characterize Mammalian Adenylyl Cyclases and Their Inhibitors." Thesis, Boston College, 2015. http://hdl.handle.net/2345/bc-ir:104220.
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Thesis advisor: Charles HoffmanThe study of mammalian cAMP signaling has often been confounded by the fact that ten different genes encode adenylyl cyclases (ACs) that produce cAMP from ATP and 16 different genes encode phosphodiesterases (PDEs) that hydrolyze cAMP to AMP. In this study, mammalian AC cDNAs were cloned and integrated into strains of the fission yeast Schizosaccharomyces pombe that lack their endogenous AC to determine the basal activity of all ten AC isoforms. In addition, response to the stimulatory mammalian Gsα was determined by co-expression of a mutationally-activated form of the human GNAS1 gene. AC activity was assessed using an fbp1-GFP reporter that is repressed by cAMP production and PKA activity. Results confirm that all ten isoforms have detectable basal activity, and AC1-9 definitively respond to Gsα stimulation. When matched with a sufficiently potent mammalian phosphodiesterase (PDE), strains expressing mammalian ACs make good candidates for small molecule high throughput screening (HTS) to detect AC inhibitors. A 100,000 compound screen was recently performed to detect AC and Gsα inhibitors as well as PDE activators. A promising “hit” was progesterone, which has been previously suggested to inhibit ACs in Xenopus. Initial results suggest that progesterone inhibits AC1 and the closely-related AC3. These data demonstrate the utility of using S. pombe as an effective platform for identifying inhibitors of both basal and GNAS1-stimulated AC activity
Thesis (BS) — Boston College, 2015
Submitted to: Boston College. College of Arts and Sciences
Discipline: Departmental Honors
Discipline: Biology
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Lali, Ferdinand Vuciri. "An investigation of the role of p38 MAP kinase and p13-kinase/PKB pathways in IL-2-induced lymphocyte proliferation." Thesis, Imperial College London, 2000. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.342220.
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McDougal, Robert A. "Excitatory-Inhibitory Interactions as the Basis of Working Memory." The Ohio State University, 2011. http://rave.ohiolink.edu/etdc/view?acc_num=osu1313004219.
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Plo, Isabelle. "Activation des voies anti-apoptotiques par les ligands de mort et les médicaments antitumoraux." Toulouse 3, 2002. http://www.theses.fr/2002TOU30048.
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ZOUKHRI, DRISS. "Caracterisation de la proteine kinase c des glandes lacrymales de rat. Effets des esters de phorbol et des inhibiteurs de la pkc sur la secretion." Paris 11, 1992. http://www.theses.fr/1992PA112350.
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Dans les glandes lacrymales extraorbitaires de rat, la secretion des proteines est essentiellement sous le controle des recepteurs cholinergiques, de type muscarinique. Ces recepteurs sont couples a une phospholipase c qui hydrolyse le phosphatidylinositol 4,5 biphosphate, liberant deux messagers intracellulaires: l'inositol 1,4,5-triphosphate (ip3) et le diacylglycerol (dag). L'ip3 mobilise du calcium a partir de pools intracellulaires et le dag active une proteine kinase dependante du calcium et des phospholipides, la proteine kinase c (pkc). Dans la premiere partie de ce travail, nous avons caracterise la pkc a partir d'une fraction cytosolique de glandes lacrymales. Nous avons montre que cette kinase est activable par le dag et les esters de phorbol actifs (pma et pdbu). De plus nous avons mis en evidence l'existence d'une activite histone kinase stimulable par le pma et l'acide arachidonique en absence de calcium et des phospholipides. Cette activite reste confondue dans les memes fractions que la kpc apres trois etapes successives de purification (deae-cellulose, phenyl-sepharose et hydroxyapatite sur hplc). D'un autre cote, nous avons identifie trois isoformes de pkc dans la fraction cytosolique des glandes lacrymales: alpha, delta et epsilon. Dans la deuxieme partie de ce travail, nous avons recherche l'implication potentielle de la pkc dans la regulation de la secretion des proteines. Les resultats obtenus montrent que les esters de phorbol stimulent la decharge des proteines tritiees et que cette reponse est insensible a l'action de trois inhibiteurs de la pkc: le h7, la sphingosine et la chelerytrine. Seules la staurosporine et la tfp sont capables d'inhiber cette reponse mais a des concentrations qui ne sont plus specifiques de la pkc. Nos resultats, pris ensemble suggerent que la pkc peut ne pas etre impliquee dans la secretion
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Lafont, Florian. "Implication des modifications post-traductionnelles de DNA-PKcs dans la régulation de la réponse aux dommages à l'ADN." Thesis, Nantes, 2017. http://www.theses.fr/2017NANT1023/document.
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Les cellules humaines sont soumises à des stress induisant des cassures double-brin de l’ADN principalement réparées par la voie NHEJ, où la kinase DNA-PKcs joue un rôle central. L’activité de DNA-PKcs, régulée par de nombreuses phosphorylations, est cruciale pour le maintien de l’intégrité génomique. Plus récemment, il a été montré que cette protéine était également modifiée par l’O-GlcNAcylation dans la lignée COS7. Sachant l’équilibre existant entre phosphorylation et O-GlcNAcylation, nous avons étudié le rôle de cette nouvelle MPT dans la régulation de l’activité de DNA- PKcs. Nous avons montré que DNA-PKcs est O-GlcNAcylée dans les cellules HeLa. Puis nous avons montré que la modulation de l’O-GlcNAcylation de DNA-PKcs impacte son autophosphorylation en Ser2056, suggérant l’existence d’une balance O-GlcNAcylation /phosphorylation, ainsi que la capacité des cellules à réparer les DSBs par la voie NHEJ. De plus, nos résultats nous laissent envisager que cette modification puisse jouer un rôle dans la stabilité de la protéine. DNA-PKcs est une cible potentielle dans les stratégies de lutte contre le cancer. Nous avons étudié l’impact d’un composé sur DNA-PKcs. Cette molécule provoque une réduction de la quantité et de l’activité de DNA-PKcs, impliquant son ubiquitinylation et sa dégradation par le protéasome et menant à une sensibilisation des cellules à un traitement génotoxique. Dans ce contexte, nous avons développé une puce à anticorps pour évaluer le profil phosphoprotéique des voies de réparation de l’ADN et ainsi évaluer l’effet d’inhibiteurs de DNA-PKcs. L’ensemble de ces résultats contribuent à une meilleure compréhension de la régulation de DNA-PKcsHuman cells are subjected to stresses inducing DNA double-strand breaks mainly repaired by the NHEJ pathway, where the kinase DNA-PKcs plays a central role. The activity of DNA-PKcs, regulated by numerous phosphorylations, is crucial for the maintenance of genomic integrity. More recently, it has been shown that this protein is also modified by O-GlcNAcylation in the COS7 cell line. Knowing the balance between phosphorylation and O-GlcNAcylation, we studied the role of this new PTM in the regulation of DNA-PKcs activity. We have shown that DNA-PKcs is O-GlcNAcylated in HeLa cells. We then showed that the modulation of DNA-PKcs O-GlcNAcylation affects its autophosphorylation on Ser2056, suggesting an O-GlcNAcylation/phosphorylation balance, as well as the ability of cells to repair DSBs by NHEJ pathway. Moreover, our results allow us to consider that this modification may play a role in protein stability. DNA-PKcs is a potential target in anticancer strategies. We studied the impact of a chemical compound on DNA-PKcs activity. This molecule causes a reduction in the amount and activity of DNA-PKcs, through its ubiquitinylation and its degradation by the proteasome and leading to sensitization of the cells to genotoxic treatment. In this context, we have developed an antibody microarray to evaluate the phosphoprotein level of DNA repair pathways and thus estimate the effect of DNA-PKcs inhibitors. All these results contribute to a better understanding of the regulation of DNA-PKcs
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Tautu, Michel Théodore. "Etude de nouveaux rôles de la DNA-PKcs, indépendants de NHEJ, dans la sensibilité aux inhibiteurs de Topoisomérase I de la famille des camptothécines." Bordeaux 2, 2008. http://www.theses.fr/2008BOR21576.
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La Topoisomérase 1 (Top 1) est une enzyme nucléaire essentielle à la plupart des transactions de l'ADN. Elle est la cible privilégiée des camptothécines (CPT) qui sont très utilisées en chimiothérapie. Ces poisons stabilisent les complexes covalents ADN-Top 1 qui sont transformés en cassures cytotoxiques au cours de la phase S dy cycle cellulaire. Plusieurs études ont montré que des cellules sans DNA-PKcs, une kinase essentielle à la réparation de l'ADN par recombinaison non-homologue (NHEJ), étaient hypersensibles à la CPT et suggéraient que cela était dû à une déficience en réparation. Dans ce travail, nous démontrons que c'est l'absence physique de la DNA-PKcs et non la perte de son activité kinase essentielle au NHEJ qui est à l'origine de la plus grande sensibilité des cellules DNA-PKcs-/-à la CPT, par un mécanisme impliquant l'absence d'interaction entre la DNA-PKcs et la Top1. La DNA-PKcs, sans les protéines Ku70 et Ku80 est en effet capable de le lier au domaine N-terminal de la Topo1 et inhibe son activité de coupure et de relaxation de l'ADN in vitro. Nous montrons également que dans les cellules DNA-PKcs-/-où cette interaction n'e'st pas possible, il existe un niveau basal plus élevé de complexes de clivage Top1-ADN que dans les lignées DNA PKcs+/+, qui est restauré lorsque le niveau de Topo 1 est régulé à la baisse ou en présence de substrats spécifiques de DNA-PKcs. Nos données mettent en évidence un nouveau rôle de "scaffold protein" de la DNA-PKcs et supportent un modèle dans lequel l'interaction entre DNA-PKcs et Topo 1 permettrait de réguler la disponibilité de Topo 1 libre susceptible de former des complexes avec l'ADNDNA topoisomerase 1 (Top 1) is a conserved nuclear enzyme that constitutes the cellular target of the camptothecin (CPT) class of chemotherapeutics. These drugs reversibly stabilize covalent Top1-DNA intermediates, which are converted into potentially lethal DNA lesions during S-phase. The enhanced CPT sensitivity of mammalian cell lines that are deficient for DNA-PKcs, the catalytic subunit of DNAdependent protein kinase, suggests that error-prone nonhomologous end joining (NHEJ) is a critical determinant of tumor cell sensitivity to drugs that poison Top1. However, contrary to this view, we report that the loss of the DNA-PKcs protein scaffold, rather than DNA-PK kinase activity, enhances mammalian cell sensivity to CPT. DNA-PKcs associated with the N-terminal domain of Top1, in the absence of Ku70 and Ku80, to suppress DNA cleavage by Top1. Interestingly, DNA-PKcs (-/-) cells exhibited increased basal levels of covalent Top1-DNA complexes, which were abolished by siRNAtargeted downregulation of Top1. Our data support a model whereby DNA-PKcs binding to Top1 limits DNA binding and consequently, the levels of covalent Top1-DNA complexes in vivo. This NHEJ-independent regulation of Top1 catalysis directly impacts cell sensitivity to CPT by regulating the steady state levels of the cellular target of this important class of chemotherapeutics. Moreover, we posit that this mechanism impacts cellular responses to other genotoxic stress, such as ionizing radiation and oxidative damage, that can also induce endogenous Top1-DNA adducts
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Roche, Emmanuel. "Etude des mécanismes de résistance des cancers de prostate aux inhibiteurs de topoisomérases I de la famille des camptothécines." Thesis, Bordeaux, 2014. http://www.theses.fr/2014BORD0423/document.
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Les ADN-Topoisomérases (Topo) de type I et II sont des enzymes essentielles à la suppression des surenroulements de engendrés par la plupart des transactions de l’ADN. Elles sont des cibles de médicaments anticancéreux très utilisés en clinique. Parmi eux, les inhibiteurs de Topo1 de la famille des camptothécines (CPT) exercent leur cytotoxicité en produisant des cassures double-brin de l’ADN provenant de la collision des fourches de réplications avec les complexes ADN-Topo1 stabilisés par ces inhibiteurs. Les dérivés de CPT sont approuvés pour le traitement des cancers coliques, de l’ovaire et du poumon, mais il existe de multiples mécanismes de résistance à ces agents qui sont à l’origine de l’échec du traitement. Les cancers de la prostate sont réfractaires aux CPT, mais très peu d’études ont été réalisées pour expliquer cette résistance « intrinsèque ». Ce travail de thèse visait à identifier les mécanismes de cette résistance en nous appuyant (1) sur des résultats antérieurs de l’équipe montrant que l’interaction entre la DNA-PKcs, une kinase impliquée dans la réparation de l’ADN par recombinaison non-homologue, et la Topo1 pouvait réguler la sensibilité aux CPT de manière indépendante de la réparation de l’ADN et (2) sur une étude ayant mis en évidence une interaction entre DNA-PKcs et le facteur de transcription ERG dont le gène est remanié dans plus de 50% des tumeurs de prostate. Nos résultats montrent pour la première fois que ERG est effectivement impliqué dans la régulation de la réponse aux CPT dans la lignée VCaP présentant un gène de fusion TMPRSS2-ERG. La répression de ERG dans la lignée VCaP induit une sensibilisation à la CPT mais pas à l’étoposide (un inhibiteur de Topo2) et est accompagnée d’une augmentation du nombre de complexes ADN/Topo1. Ce mécanisme peut-être soit lié à un effet de ERG sur l’interaction DNA-PKcs/Topo1 ou à une régulation transcriptionnelle de gènes impliqués dans la réponse aux CPT incluant la Topo1 elle-même. Nous avons confirmé cette deuxième hypothèse, en démontrant que ERG régule la transcription du micro ARN miR-24 et que l’expression de Topo1 est également sous contrôle de miR-24 dans la lignée VCaP. Des résultats similaires ont été obtenus dans la lignée LNCaP (présentant le gène de fusion TMPRSS2-ETV1) dans laquelle la répression de ETV1 confère aussi une sensibilisation à la CPT. Au cours de notre travail nous avons également recherché des inhibiteurs de l’interaction entre la DNA-PKcs et Topo1 afin de pouvoir utiliser ces composés comme agents de potentialisation des dérivés de CPT en clinique. Les résultats du criblage d’une banque de 320 composés naturels réalisé par la technologie AlphaScreen n’ont malheureusement pu identifier que des inhibiteurs catalytiques de Topo1. Nous avons néanmoins pu montrer que l’un d’entre eux, la mahanimbine, présentait une forte activité cytotoxique vis-à-vis de lignées résistantes aux dérivés de CPT et vis-à-vis de la lignée VCaP ce qui permet d’envisager le développement de nouvelles classes d’inhibiteurs catalytiques Topo1 pouvant contourner la résistance des dérivés de CPT en cliniqueType I and II DNA Topoisomerases (Top) are essential enzymes involved in the removal of DNA torsional constraints induced by most DNA transactions. They are the targets of various anticancer agents used in the clinic. Among them, Top1 inhibitors from the camptothecins (CPT) family exert their cytotoxicity by producing DNA double-strand breaks that are generated by the collision of advancing replication forks with DNA-Top1 complexes that are stabilized by these inhibitors. CPT derivatives are approved for the treatment of colon, ovary and lung cancers but resistance mechanisms are developed and lead to treatment failure. Prostate cancers are refractory to CPT, but few studies have addressed the mechanisms of such “intrinsic” resistance. This work was aimed at identifying such mechanisms based on (1) previous results from the laboratory showing that interaction of Top1 with DNA-PKcs, a kinase that is essential for non-homologous end-joining, could regulate cell sensitivity to CPT independently of DNA repair and (2) a study that showed an interaction of DNA-PKcs with ERG, a transcription factor from the ETS family which is rearranged in more than 50% of prostate tumors. Our results show for the first time that ERG is indeed involved in the regulation of prostate cancer cell response to CPT as its repression sensitized VCaP cells displaying the TMPRSS2-ERG gene fusion to CPT but not to the Top2 inhibitor etoposide. This effect is accompanied with an increase in Top1-DNA complexes. This could be due to either an effect of ERG on DNA-PKcs/Top1 interaction, or to the transcriptional regulation of genes involved in cell response to CPT, including Top1 itself. We confirmed the latter hypothesis by showing that ERG can regulate the transcription levels of the microRNA miR-24 and that Top1 expression relies, at least in part, on miR24 levels in VCaP cells. We obtained similar results in LNCaP cells (characterized by a TMPRSS2-ETV1 gene fusion), in which ETV1 repression also sensitizes cells to CPT. In parallel, we also searched for inhibitors of DNA-PKcs/Top1 interaction in order to use these compounds to potentiate CPT derivatives in the clinic. We screened a chemical library of 320 natural compounds using the AlphaScreen technology. The results were disappointing as we only identified compounds that are catalytic inhibitors of Top1. Nevertheless, we could show that among them, mahanimbine displayed a potent cytotoxic activity towards CPT-resistant colon cancer cell lines and could efficiently inhibit the growth of VCaP cells that are highly resistant to CPT. This opens new avenues for the development of new classes of Top1 catalytic inhibitors that could be used to circumvent the clinical resistance to CPT derivatives
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Blakeney, Bryan Adam. "Branched Short Chain Fatty Acid Isovaleric Acid Causes Smooth Muscle Relaxation via cAMP/PKA Pathway, Inhibits Gastrointestinal Motility, and Disrupts Peristaltic Movement." VCU Scholars Compass, 2018. https://scholarscompass.vcu.edu/etd/5548.
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Isovaleric Acid (IVA) is a 5-carbon branched chain fatty acid present in fermented foods and produced by the fermentation of leucine by colonic bacteria. IVA activates G-protein coupled receptors such as FFAR2, FFAR3, and OR51E1 known to be expressed on enteric neurons and enteroendocrine cells. We previously reported that the shorter, straight chain fatty acids acetate, propionate and butyrate, differentially affect colonic propulsion; however, the effect of branched chain fatty acids on gastrointestinal motility is unknown. We hypothesize that IVA relaxes smooth muscle in a cAMP/PKA dependent manner by direct action on smooth muscle cells. IVA will also decrease peristalsis and encourage retention of luminal contents. This thesis investigates the effect of IVA on smooth muscle tension and peristaltic activity in isolated colon and individual smooth muscle cells.
Colon segments from C57BL/6J mice were placed in a longitudinal orientation in organ baths in Krebs buffer and fastened to force transducers. Segments were contracted with 10 μM acetylcholine (ACh) and the effects of IVA at several concentrations were measured in the absence and presence of Nitric Oxide Synthase inhibitor L-N-nitroarginine (L-NNA), neuronal action potential inhibitor tetrodotoxin (TTX), and adenylate cyclase inhibitor SQ22536. To study individual live cells, mouse smooth muscle was isolated from colon, suspended in smooth muscle buffer, and after contraction with ACh were relaxed with micromolar concentrations of IVA. For peristalsis studies, whole colonic segments isolated from C57BL/6J were catheterized and placed horizontally in organ baths with circulating Krebs buffer. The colon was clamped on the anal end, and a solution (5 μL per mm of colon length) of either Krebs buffer or 50 mM IVA was delivered from the oral end to the lumen. Video of the peristalsis was then analyzed for diameter, changes in diameter, velocity of diameter changes along the length of the colon, normalized to the anatomical changes in the proximal region.
IVA in concentrations of 10 mM to 50 mM relaxed the ACh-induced contraction in a sigmoidal fashion. In separate studies, L-NNA nor TTX affected the ability of IVA to inhibit relaxation. SQ22536 inhibited IVA induced relaxation in longitudinal colon compared to vehicle control. In isolated cells, SQ22536 and PKA inhibitor H-89 inhibited IVA-induced relaxation. In peristalsis studies, 50 mM IVA in Krebs buffer changed the character of the peristaltic action by increasing proximal diameter, inhibiting contractions in the proximal end of the colon, and decreasing overall velocity of peristaltic contractions in the proximal region.
The data indicate that the branched chain fatty acid IVA causes a concentration-dependent relaxation of colonic smooth muscle that is direct to the smooth muscle and independent of neuronal activity. This relaxation is cAMP/PKA dependent. In addition to the direct relaxation of smooth muscle, intraluminal IVA decreased overall colonic propulsive activity and encouraged retention of the luminal contents. We conclude that the ingestion and production of branched chain fatty acids could affect overall GI motility and is an area for study in dietary and therapeutic control of bowel activity.
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Cevirgen, Ceyhun [Verfasser], Ali [Akademischer Betreuer] El-Armouche, Elisabeth [Gutachter] Zeisberg, and Martin [Gutachter] Oppermann. "Analyse des antifibrotischen Potenzials der betaadrenergen Signalkaskade und des PKA-Effektors Protein-Phosphatase-Inhibitor-1 / Ceyhun Cevirgen ; Gutachter: Elisabeth Zeisberg, Martin Oppermann ; Betreuer: Ali El-Armouche." Göttingen : Niedersächsische Staats- und Universitätsbibliothek Göttingen, 2017. http://d-nb.info/1136471324/34.
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Visseq, Alexia. "Conception et synthèse d’inhibiteurs sélectifs de protéines kinases pour le traitement de l’allodynie mécanique." Thesis, Université Clermont Auvergne (2017-2020), 2019. http://www.theses.fr/2019CLFAC048.
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La douleur chronique constitue aujourd’hui un problème majeur de santé publique qui touche près de 20% de la population. L’allodynie mécanique, une douleur provoquée par une stimulation normalement non douloureuse, est le symptôme le plus fréquent chez les patients atteints de douleur chronique. Malheureusement, les traitements actuellement disponibles ne sont pas réellement efficaces ou bien présentent d’importants effets secondaires ou contre-indications. L’objectif de ce projet est de concevoir, synthétiser et évaluer biologiquement des inhibiteurs sélectifs de protéines kinases pour le traitement de l’allodynie mécanique. Au cours de ce travail de thèse, nous avons découvert de nouveaux inhibiteurs sélectifs de p38α, une protéine kinase bien connue pour son implication dans la douleur chronique et l’allodynie mécanique. Une étude des relations structure-activités a été réalisée pour identifier les éléments structuraux importants permettant d’améliorer l’activité des molécules in vitro et in vivo sur un modèle animal. Les meilleures molécules ont présenté des IC50 submicromolaires sur p38α et une forte inhibition de l’allodynie mécanique in vivoChronic pain is a global public health priority which affects more than 20% of Europeans. Mechanical allodynia, a pain in response to normally innocuous stimuli, is one of the most prevalent pain symptoms. Despite intensive research toward the study of pain mechanisms, currently available treatments of pain are not always effective, and can produce side-effects. In this context, the aim of the project is to design, synthesize and evaluate selective inhibitors of protein kinases for the treatment of mechanical allodynia. During this PhD work, new selective inhibitors of the protein kinase p38α has been discovered. This protein kinase is known for its implication in chronic pain and mechanical allodynia. A structure-activity relationship study was performed to identify the important structural features allowing a gain of activity of molecules in vitro and in vivo using an animal model. The best compounds showed submicromolar IC50 values toward p38α and a strong inhibition of mechanical allodynia in vivo
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Rancillac, Armelle. "Propriétés fonctionnelles et plasticité des synapses glutamatergiques afférentes aux cellules de Purkinje et aux interneurones inhibiteurs de la couche moléculaire du cortex cérébelleux chez le rongeur." Paris 6, 2003. http://www.theses.fr/2003PA066278.
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Tronel, Claire. "Evaluation des effets de molécules à visée neuroprotectrice dans un modèle in vivo de neuroinflammation chez le rat : étude mécanistique et caractérisation du modèle au cours du temps." Thesis, Tours, 2013. http://www.theses.fr/2013TOUR3806/document.
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La mise au point de médicaments ciblant la neuroinflammation, une composante importante de la physiopathologie des maladies neurodégénératives, fait l’objet de nombreuses recherches. Dans ce travail de thèse, nous avons étudié les effets de deux molécules potentiellement anti-inflammatoires et neuroprotectrices : l’hémine, un inducteur de l’hème oxygénase 1(HO-1) et ; le C16, un inhibiteur de la protéine kinase activée par l’ARN (PKR) dans un modèle de neuroinflammation in vivo par injection intrastriatale d’acide quinolinique (AQ) chez le rat. Nos résultats ont montré que l’induction de la HO-1 produit des effets délétères tandis que l’inhibition de la PKR induit des effets neuroprotecteurs et antiapoptotiques. Ce travail a par ailleurs permis de décrire l’évolution cinétique de la neuroinflammation sur 90 jours dans le modèle AQ, la capacité du tissu cérébral à se régénérer après la lésion et l’intérêt de ce modèle dans l’étude des effets d’agents neuroprotecteurs administrés au long coursNeuroinflammation is a key part of the physiopathology of neurodegenerative diseases and is an interesting target in their treatment. In this PhD work, we studied the effects of two potentially anti-inflammatory and neuroprotective molecules, hemin and C16, in an in vivo rat model of neuroinflammation by intrastriatal injection of quinolinic acid (QA). We showed that heme oxygenase 1 (HO-1) induction by hemin has deleterious effects whereas inhibition of the protein kinase RNA activated (PKR) by C16 treatment induced neuroprotective and anti-inflammatory effects. Concurrently, we evaluated longitudinal evolution of neuroinflammation in our model. Results showed the kinetic of the inflammatory phenomena; the ability of cerebral tissue to recover integrity and the capability of this model to evaluate potential neuroprotective and anti-inflammatory drugs in a long-time study
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Humphrey, Peter Saah. "Signal transduction mechanisms for stem cell differentation into cardiomyocytes." Thesis, University of Hertfordshire, 2009. http://hdl.handle.net/2299/3760.
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Cardiovascular diseases are among the leading causes of death worldwide and particularly in the developed World. The search for new therapeutic approaches for improving the functions of the damaged heart is therefore a critical endeavour. Myocardial infarction, which can lead to heart failure, is associated with irreversible loss of functional cardiomyocytes. The loss of cardiomyocytes poses a major difficulty for treating the damaged heart since terminally differentiated cardiomyocytes have very limited regeneration potential. Currently, the only effective treatment for severe heart failure is heart transplantation but this option is limited by the acute shortage of donor hearts. The high incidence of heart diseases and the scarcity donor hearts underline the urgent need to find alternative therapeutic approaches for treating cardiovascular diseases. Pluripotent embryonic stem (ES) cells can differentiate into functional cardiomyocytes. Therefore the engraftment of ES cell-derived functional cardiomyocytes or cardiac progenitor cells into the damaged heart to regenerate healthy myocardial tissues may be used to treat damaged hearts. Stem cell-based therapy therefore holds a great potential as a very attractive alternative to heart transplant for treating heart failure and other cardiovascular diseases. A major obstacle to the realisation of stem cell-based therapy is the lack of donor cells and this in turn is due to the fact that, currently, the molecular mechanisms or the regulatory signal transduction mechanisms that are responsible for mediating ES cell differentiation into cardiomyocytes are not well understood. Overcoming this huge scientific challenge is absolutely necessary before the use of stem cell-derived cardiomyocytes to treat the damaged heart can become a reality. Therefore the aim of this thesis was to investigate the signal transduction pathways that are involved in the differentiation of stem cells into cardiomyocytes. The first objective was the establishment and use of cardiomyocyte differentiation models using H9c2 cells and P19 stem cells to accomplish the specific objectives of the thesis. The specific objectives of the thesis were, the investigation of the roles of (i) nitric oxide (ii) protein kinase C (PKC), (iii) p38 mitogen-activated protein kinase (p38 MAPK) (vi) phosphoinositide 3-kinase (PI3K) and (vi) nuclear factor-kappa B (NF-kB) signalling pathways in the differentiation of stem cells to cardiomyocytes and, more importantly, to identify where possible any points of convergence and potential cross-talk between pathways that may be critical for differentiation to occur. P19 cells were routinely cultured in alpha minimal essential medium (α-MEM) supplemented with 100 units/ml penicillin /100 μg/ml streptomycin and 10% foetal bovine serum (FBS). P19 cell differentiation was initiated by culturing the cells in microbiological plates in medium containing 0.8 % DMSO to form embryoid bodies (EB). This was followed by transfer of EBs to cell culture grade dishes after four days. H9c2 cells were cultured in Dulbecco’s Modified Eagle’s medium (DMEM) supplemented with 10% FBS. Differentiation was initiated by incubating the cells in medium containing 1% FBS. In both models, when drugs were employed, they were added to cells for one hour prior to initiating differentiation. Cell monolayers were monitored daily over a period of 12 or 14 days. H9c2 cells were monitored for morphological changes and P19 cells were monitored for beating cardiomyocytes. Lysates were generated in parallel for western blot analysis of changes in cardiac myosin heavy chain (MHC), ventricular myosin chain light chain 1(MLC-1v) or troponin I (cTnI) using specific monoclonal antibodies. H9c2 cells cultured in 1% serum underwent differentiation as shown by the timedependent formation of myotubes, accompanied by a parallel increase in expression of both MHC and MLC-1v. These changes were however not apparent until 4 to 6 days after growth arrest and increased with time, reaching a peak at day 12 to 14. P19 stem cells cultured in DMSO containing medium differentiated as shown by the timedependent appearance of beating cardiomyocytes and this was accompanied by the expression of cTnI. The differentiation of both P19 stem cells and H9c2 into cardiomyocytes was blocked by the PI3K inhibitor LY294002, PKC inhibitor BIM-I and the p38 MAPK inhibitor SB2035800. However when LY294002, BIM-I or SB2035800 were added after the initiation of DMSO-induced P19 stem cell differentiation, each inhibitor failed to block the cell differentiation into beating cardiomyocytes. The NF-kB activation inhibitor, CAPE, blocked H9c2 cell differentiation into cardiomyocytes. Fast nitric oxide releasing donors (SIN-1 and NOC-5) markedly delayed the onset of differentiation of H9c2 cells into cardiomyocytes while slow nitric oxide releasing donors (SNAP and NOC-18) were less effective in delaying the onset of differentiation or long term differentiation of H9c2 cells into cardiomyocytes. Akt (protein kinase B) is the key downstream target of PI3K. Our cross-talk data also showed that PKC inhibition and p38 MAPK inhibition respectively enhanced and reduced the activation of Akt, as determined by the phosphorylation of Akt at serine residue 473. In conclusion, PKC, PI3K, p38 MAPK and NF-kB are relevant for the differentiation of stem cells into cardiomyocytes. Our data also show that the PKC, PI3K and p38 MAPK signalling pathways are activated as very early events during the differentiation of stem cells into cardiomyocytes. Our data also suggest that PKC may negatively regulate Akt activation while p38 MAPK inhibition inhibits Akt activation. Our fast NO releasing donor data suggest that nitric oxide may negatively regulate H9c2 cell differentiation.
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Wang, Jen-Jui, and 王仁瑞. "Inhibitory Mechanism of Midazolam in PKC-induced MMPs Expression in Chondrocytes." Thesis, 2010. http://ndltd.ncl.edu.tw/handle/42636205848281078456.
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碩士臺北醫學大學
醫學科學研究所
98
Midazolam, a benzodiazepine, has a hypnotic effect and is widely used as a sedative. The role of midazolam in activation of chondrocytes during inflammation is not known. The aim of this study was to evaluate the anti-inflammatory actions of midazolam in cultured chondrocytes. Using a chondrosarcoma cell line, SW1353 cells, the inhibitory effect of midazolam on protein kinase C (PKC) activator phorbol-12-myristate-13-acetate (PMA) induced matrix metalloproteinases (MMPs) was assessed by Western blot and gelatin zymography. Our results show that PMA-induced up-regulation of MMPs 1, 9 and 13 expressions was significantly inhibited by midazolam in a concentration-dependent manner (5-20 μM). Midazolam also inhibited PMA-mediated activation of extracellular signal-regulated kinase 1/2, p38 mitogen-activated protein kinases but had no effect on c-Jun N-terminal kinase. Treatment with midazolam enhanced re-synthesis of a concentration dependant PMA mediated IκB-α degradation. Thus, overall our results revealed that the inhibitory mechanism of midazolam on MMPs involves depressed expression of p38 and ERK 1/2 and facilitated recovery of IκB-α degradation in an induced chondrosarcoma SW1353 cell line. Midazolam has an anti-inflammatory action by inhibiting activity, synthesis and the expression of inducible MMPs through MAPKs and NF- κB/IκB pathways. These findings may considerably be provided the novel molecular basis of midazolam.
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Alam, Mohammad Intakhab [Verfasser]. "Investigation of the role of PKCα [PKC-alpha] for influenza A virus-induced signalling and of the inhibitory effect of verapamil on virus replication / vorgelegt von Mohammad Intakhab Alam." 2008. http://d-nb.info/989046486/34.
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Syu, Ya-Ting, and 徐雅婷. "PMC Inhibits PDGF-BB-Induced Proliferation of Vascular Smooth Muscle Cells through PKC-delta." Thesis, 2005. http://ndltd.ncl.edu.tw/handle/43780639479372376569.
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碩士臺北醫學大學
藥理學研究所
93
Abnormal proliferation of vascular smooth muscle cells (VSMCs) results in neointima plays an important role in many coronary diseases including atherosclerosis and restenosis. This research was shown the inhibitory mechanisms of PMC (2, 2, 5, 7, 8 -pentamethyl-6-hydroxychromane), as a derivative of -tocopherol, on VSMCs proliferation. By MTT assay and microscopy, we found that PMC (20, 50 and 100 M) inhibited PDGF-BB-induced vascular smooth muscle cells proliferation in a concentration dependent manner. Rottlerin (PKC--specific inhibitor) and Gö6976 (PKC--specific inhibitor) inhibited PDGF-BB-induced vascular smooth muscle cells proliferation. Evidence suggests that PMC may inhibit PDGF-BB-induced vascular smooth muscle cells proliferation through PKC- or PKC- By Western blot analysis and confocal microscopy, we found that PMC inhibited the translocation of PKC- from the cytosolic to membrane fraction stimulated by PDGF-BB in a concentration dependent manner. After stimulating with PDGF-BB, PKC-would not be translocate from the cytosolic to membrane fraction. Upon treating cells PMC, the distruction of PKC-would not be affect. Evidence suggests that PMC may inhibit PDGF-BB-induced vascular smooth muscle cells proliferation through inhibiting the specific translocation of PKC- PMC inhibited the translocation of PKC- and IB degradation, while rottlerin (PKC--specific inhibitor) didn’t inhibit PDGF-BB-induced vascular smooth muscle cells IB degradation. On the other hand, PMC inhibited Akt phosphorylation and IB degradation, while LY294002 (PI3K inhibitor) didn’t inhibit PDGF-BB-induced vascular smooth muscle cells IB degradation. In order to clarify the cross-talk between PKC- and Akt, we found that rottlerin (PKC--specific inhibitor) didn’t inhibit PDGF-BB-induced vascular smooth muscle cells Akt phosphorylation. By Western blot analysis, PMC (100 M) may induce apoptosis by increasing the expression of active caspase-3 stimulated by PDGF-BB. In the Western blot analysis of protein levels of cyclin D1, Cdk4 and 21cip1 in rat vascular smooth muscle cells treated with PMC (100 M) stimulated with PDGF-BB (10 ng/ml), protein levels of cyclin D1, Cdk4 and 21cip1 would not be affected. In conclusion, PMC inhibited vascular smooth muscle cells proliferation through inhibiting the translocation of PKC-Akt phosphorylation, and IBdegradation. In the privous studies, PMC inhibited cell cycle progression by arresting in the G0/G1 phase to S phase and inhibited cell proliferation. PMC (100 M) may induce apoptosis by increasing the expression of active caspase-3 although protein levels of cyclin D1, Cdk4 and 21cip1 would not be affected. Further efforts would be focused on investigating the effects of PMC on other molecular regulators of cell cycle and apoptosis. Effects of anti-aggregation and anti-oxidation of PMC were proven. This research showed PMC inhibited VSMCs proliferation. Because of the anti-proliferation effect of PMC, the further efforts to evaluate the anti-angiogenesis and anti-cancer effects of PMC might be worthwhile.
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萩原, 和美. "Prox 1 overexpression of Hela cells inhibits PKC beta II transcription through promoter DNA methylation." Thesis, 2012. http://hdl.handle.net/2237/16914.
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chi, Wang szu, and 王思琪. "Virtual Screening of PKA Inhibitors Using Docking Computation：Effect of Flexible Side Chain." Thesis, 2010. http://ndltd.ncl.edu.tw/handle/44642027663578608742.
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碩士國立臺灣師範大學
化學系
98
Protein kinase A (PKA) is a kinase protein that has several functions in cell, including regulation of glycogen, sugar, and lipid metabolism. It also plays significant role in a number of biochemical reaction networks associated with diseases, including lung cancer and colorectal cancers. In the present study, we used docking computation to aid in design and discovery of PKA inhibitors. First, we carried out docking computations for 20 PKA-inhibitor complexes from protein data bank to examine their reproducibility. The results showed that the computed fitnesses values of ligands are in good accord with the experimental IC50 values. Second, crossing docking of selected 5 complexes was carried out to investigate if and how ligand conformations can be regained when a protein structure from different complexes were used. In addition, thirdly, the protein structures from these 5 complexes were used to undergo a virtual screening to see if 10 active compounds can be screened out of 1000 compounds selected from a database. In these computations, the several side chains at active site were allowed to move to examine how this effect affects the docking results. The results showed that better results were obtained in the case of allowing 4 residues to move. Finally, a virtual screening for 24535 compounds was carried out. The interactions between top-ranked compounds and PKA were analyzed and discussed. These computed results and analysis should be of aid in design and discovery of PKA inhibitors.
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Shan-Meei, Tang, and 唐善美. "Involvement of PKC in the inhibitory effects of 18 beta-GA on gap junction in the cardiac myoblast." Thesis, 2001. http://ndltd.ncl.edu.tw/handle/66373002464052138250.
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碩士國立臺灣大學
解剖學暨細胞生物學研究所
89
Abstract Gap junctions are intercellular channels ensuring electrical and metabolic coupling and are responsible for the synchronous contraction of cardiomyocytes. The major gap junctional protein in working cardiomyocytes is connexin43(Cx43). In this study, immunofluorescence microscopy and Lucifer yellow dye coupling assays were used to investigate the role of protein kinase C (PKC) in the inhibitory effects of 18b-glycyrrhetinic(18b-GA) acid on gap junctions in H9c2(2-1) rat cardiac myoblast cells. The inhibitory effect of 18b-GA on gap junction of H9c2(2-1) cells is dose-dependent. There was no significant change in the immunofluorescence staining of Cx43 when H9c2(2-1) cells were treated with 60 mM of 18b-GA for two hours. When the concentration of 18b-GA was raised to 100 mM, a decrease of Cx43 immunostaining intensity was found on H9c2(2-1) cell membranes. The dye coupling of gap junctions was reduced from 62% in controls to 5% in 100 mM 18b-GA treated H9c2(2-1) cells. Treatment of H9c2(2-1) cells with 100 nM PMA or 100 mM 18b-GA plus 100 nM PMA led to disappearing of Cx43 immunostaining spots from cell membranes and the dye coupling of gap junctions were reduced to 8% and 1.6%, respectively. When H9c2(2-1) cells were treated with 2.5 mM chelerythrine, large Cx43 immunostaining spots were detected on the cell membranes and dye coupling was increased to 85%. Likewise, treatment of H9c2(2-1) cells with 0.5 mM calphostin C also led to accumulation of Cx43 immunostaining spots on cell membrane and a slightly increase of dye coupling to 68%. Co-treatment with 100 mM 18b-GA plus 2.5 mM chelerythrine or 100 mM 18b-GA plus 0.5 mM calphostin C both led to a weaker immunostaining intensity of Cx43 in drug-treated groups than in controls and the dye coupling of gap junctions was restored to 30% and 31%, respectively. These results suggest that the inhibitory effects of 18b-GA on gap junctions were attenuated by co-treatment of PKC inhibitors, which can restore partial immunostaining intensity of Cx43 on cell membrane and dye coupling between H9c2(2-1) cells.
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Chang, Pei-Yun, and 張沛昀. "Study on how DAPK inhibits the activation of PKCθ in T cells." Thesis, 2013. http://ndltd.ncl.edu.tw/handle/87897947530085202221.
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碩士國立臺灣大學
免疫學研究所
101
Death-associated protein kinase (DAPK) is well known as a tumor suppressor. Previous studies from our lab have shown that DAPK is activated after TCR stimulation. DAPK inhibits T cell activation through suppression of NF-κB signaling. Furthermore, DAPK specifically inhibits the TCR-induced NF-κB activation but not TNFα- or IL-1β-induced NF-κB activation. Results from our lab also demonstrated that DAPK inhibits TCR-induced PKCθ phosphorylation, but the exact biochemical process targeted by DAPK to inhibit PKCθ activation remains unclear. The specific aim of this study is to delineate the molecular mechanism on how DAPK inhibits TCR-induced PKCθ phosphorylation. MAP4K3/GLK is recently identified as the kinase that phosphorylates and activates PKCθ after TCR stimulation. We found that DAPK interacted with both PKCθ and MAP4K3/GLK in HEK293T cells and Jurkat JE6.1 T cells. We also identified several domains of DAPK that mediate the binding to PKCθ and MAP4K3/GLK. Using in vitro kinase assay, we found that DAPK did not phosphorylate PKCθ. DAPK did not phosphorylate MAP4K3/GLK, but DAPK inhibited MAP4K3/GLK auto-phosphorylation and kinase activity. We also found that DAPK bound SLP-76, the adaptor protein which binds MAP4K3/GLK and is required for MAP4K3/GLK activation. In vitro binding analysis demonstrated that the presence of DAPK decreased the binding between SLP-76 and MAP4K3/GLK. DAPK also inhibited the association between SLP-76 and MAP4K3/GLK in Jurkat JE6.1 T cells. Therefore, DAPK uses at least two different mechanisms to inhibit TCR-induced PKCθ activation: one by suppressing MAP4K3/GLK kinase activity, the other by reducing the binding of MAP4K3/GLK to SLP-76. Further characterization on the processes underlying the inactivation of MAP4K3/GLK and the reduced association of MAP4K3/GLK-SLP-76 by DAPK may help to understand the exact inhibitory mechanism of DAPK in TCR-induced NF-κB signaling.
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Cevirgen, Ceyhun. "Analyse des antifibrotischen Potenzials der betaadrenergen Signalkaskade und des PKA-Effektors Protein-Phosphatase-Inhibitor-1." Doctoral thesis, 2017. http://hdl.handle.net/11858/00-1735-0000-0023-3E46-4.
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Chiu, Po-Hsuan, and 裘博媗. "Activation of P2X7 Receptors Inhibits Proliferation by Arresting Cell Cycle Progression via Ca2+/PKC/ERK1/2 Signal Pathways of Neural Progenitor Cells." Thesis, 2012. http://ndltd.ncl.edu.tw/handle/19397030143482074823.
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碩士國立陽明大學
神經科學研究所
101
ABSTRACT ATP is an extracellular signaling molecule that regulates multiple physiological functions during CNS development. Recent studies implicated that purinergic signaling is involved in neural development, including proliferation. Previous studies also demonstrated the P2X7 purinergic receptor emerged at the stage of neurogenesis. However, the exact role of P2X7 receptors during neural development has not been thoroughly examined. The aim of this study is to investigate the effect of P2X7 receptors on proliferation of neural progenitor cells (NPCs) and elucidate the underlying molecular mechanisms involved. In this study, we found that NPCs expressed functional P2X7 purinergic receptors. Treatment of NPCs with P2X7 receptors selective agonist BzATP decreased cell number, cell viability, and the number of bromouridine (BrdU)-incorporated cells, but did not affect cell survival in trypan blue exclusion test. The effect was inhibited by the action of a P2X7 receptor selective antagonist A 438079 and by using P2X7 shRNA to knockdown the receptor expression. Using flow cytometry, BzATP treatment arrested cell cycle at the S phase. Thus, activation P2X7 receptor decreased proliferation and altered cell cycle progression of NPCs. To further examine the molecular mechanism involved, we also found that BzATP stimulated ERK1/2 activation in a time-dependent manner. By using EGTA to chelate extracellular Ca2+, or by using GF 109203X to inhibit PKC or PD98059 to inhibit MEK, significantly attenuated the BzATP-induced ERK1/2 activation. In contrast the ERK1/2 activation was not affected by using CaMKII inhibitor KN 93, or by using PI3K inhibitor LY294002. In addition, BzATP induced cytosol-to-membrane translocation of PKC α, γ, ε isozymes. Moreover, either PD98059 or GF 109203X blocked the BzATP-decreased proliferation of these cells. The physiological ligand of P2X7 receptors, ATP also inhibited proliferation of NPCs and that was suppressed by P2X7 receptors antagonist but not by P2X1-6 receptor antagonist, TNP-ATP. ATP also induced transient ERK1/2 activation. Together, these results indicated that activation of P2X7 receptors inhibits proliferation by arresting cell cycle progression and that is via Ca2+-PKC-ERK1/2 signal pathway in NPCs.
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Hsu, Ching-Chueh, and 許競爵. "I. The mRNA expressions of PKC isoforms during decidulization at pseudopregnant rats. II. Estimation of the inhibitory effect of Chinese herb extract on liver fibrosis." Thesis, 2002. http://ndltd.ncl.edu.tw/handle/14799440415691413118.
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碩士中山醫學大學
生物化學研究所
90
Part I. Our previous study have demonstrated that the protein levels of PKC isoforms were altered in the proliferation stages of decidualization at pseudopregnant and pregnant rats. In this study, we measured the mRNA expressions of the PKC isoforms and their down-stream related genes by using RT-PCR. The product of RT-PCR was also checked by DNA sequence analysis. The results showed that the levels of PKCα mRNA was significantly increased from day 2 to day 9 in the decidualization. The mRNA expression of the transcription factors (c-jun and c-fos) were also significantly increased during the same times. These specified that the alterations in the PKCα may signify their activation in decidual tissues. In addition, the mRNA levels of the other PKC isoforms (PKCλ and PKCδ were also significantly increased during the decidualization. The increase in the PKCλ mRNA was paralleled with the increase in the mRNA levels of the proliferation markers (cdc2 and cyclin D1). Thus, These data confirmed that the expression of PKC isoforms may be involved in the development of the decidualization. Part II. Hepatitis is one of the most common diseases in Taiwan. Patients with hepatitis may lead to liver fibrosis and cirrhosis. Liver fibrogenesis, which is a dynamic, complex, and progressive process, may be reversible at initial stages. Recently, many labs are extremely focus on discovering new drugs from Chinese herb for anti-fibrosis of liver. According to previous research, to activate hepatic stellate cells have been clearly identified as the major cause of liver fibrogenesis. The present studies aim to set up a fast selective cell model to estimate the anti-fibrotic effects by Chinese herbs. LD50 was determined by the actived/inactived HSCs when treated with water extracts of 36 kinds of Chinese herb. Our results showed that totally 31 kinds of Chinese herbs (86.1%) were detected with the antifibrotic effects. Within these 31 herbs, there are 22 kinds (61.1%) have less damage on inactived HSCs compared with actived. Taken together, our results indicated that the HSCs model may be more suitable to estimate the anti-fibrotic effects of Chinese herbs than animal models.
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Maiuri, Tamara Lise. "Specificity in PI3K-PKB/AKT-PTEN Signaling: Subcellular Locus-specific Functions of Pathway Targets." Thesis, 2010. http://hdl.handle.net/1807/26370.
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The PI3K-PKB/Akt-PTEN signal transduction pathway orchestrates a variety of fundamental cell processes and its deregulation is implicated in several human diseases, including cancer. While the importance of this pathway to many cellular functions is well established, the mechanisms leading to context-specific physiological outcomes in response to a variety of stimuli remain largely unknown.
Spatial restriction of signaling events is one of the means to coordinate specific cellular responses. To investigate the subcellular locus-specific roles of the major PI3K effector PKB/Akt in various cell processes, I have devised a novel experimental system employing cellular compartment-directed PKB/Akt pseudosubstrate inhibitors. The work herein describes the development and characterization of the localized PKB/Akt pseudosubstrate inhibitor system and its application to investigate potential locus-specific functions in established PKB/Akt-regulated cellular processes. Subcellular compartment-restricted PKB/Akt inhibition in the 3T3L1 adipocyte differentiation model revealed that nuclear and plasma membrane, but not cytoplasmic, PKB/Akt activity is required for terminal adipocyte differentiation. Nuclear and plasma membrane pools of PKB/Akt were found to contribute to distinct stages of adipocyte differentiation, revealing that PKB/Akt activity impacts multiple points of this program.
The localized PKB/Akt pseudosubstrate inhibitor system was also utilized to investigate the importance of distinct subcellular pools of PKB/Akt in breast epithelial cells. MCF-10A human breast epithelial cells can be grown in three-dimensional culture to form acinar structures that recapitulate in vivo mammary glandular architecture. Expression of the plasma membrane PKB/Akt inhibitor during cell growth in three-dimensional culture severely impaired acinar formation. On the other hand, expression of the nuclear PKB/Akt inhibitor during acinar development resulted in the formation of large, misshapen, multi-acinar structures. Assessment of the migratory capacity of MCF-10A cells upon localized PKB/Akt inhibition revealed that nuclear PKB/Akt inhibition promoted, while plasma membrane PKB/Akt inhibition impaired, MCF-10A cell migration.
The development of locus-specific PKB/Akt inhibitors represents the first attempt to prioritize the targets of this kinase based on their subcellular localization. This work and its immediate extensions will further our understanding of the biology of PKB/Akt, a multi-tasking kinase with profound roles in development, cellular and organismal homeostasis and disease.
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Montrobert, Ariane [Verfasser]. "Vergleich der Apoptoseaktivierung anhand der Caspase-3-Verteilung in einer Gliomzelllinie und Astrozytenkultur nach Inkubation mit TNF-α [TNF-alpha] und PKC-Inhibitoren / vorgelegt von Ariane Montrobert." 2010. http://d-nb.info/1007453540/34.
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Hussain, Munir, N. Bracken, W. Kent, and C. Pearman. "H-89 inhibits transient outward (Ito) and inward rectifier (IK1) potassium currents independently of pka-mediated phosphorylation in isolated rat ventricular myocytes." 2006. http://hdl.handle.net/10454/3310.
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NoVoltage clamp was used to investigate the effects of N-[2-p-bromo-cinnamylamino)ethyl]-5-isoquinolinesulfonamide (H-89), a potent inhibitor of PKA, on transient outward K+ current (Ito) and inward rectifying K+ current (IK1) in rat cardiac muscle. Initial experiments, performed using descending voltage ramps, showed that H-89 inhibited both the outward and inward ramp currents in a concentration-dependent manner at concentrations between 5 and 60 ¿mol l¿1. A similar degree of inhibition was observed when Ito and IK1 were recorded using square wave depolarising and hyperpolarising voltage steps, respectively. The IC50 was 35.8 ¿mol l¿1 for Ito and 27.8 ¿mol l¿1 for IK1 compared to 5.4 ¿mol l¿1 for L-type Ca2+ current (ICa). The Hill coefficients for Ito, IK1 and ICa were ¿1.97, ¿1.60 and ¿1.21, respectively. In addition to inhibiting Ito amplitude, H-89 also accelerated the time to peak and the rate of voltage-dependent inactivation so that the time course of Ito was abbreviated. Paired-pulse protocols were performed to study the effects of H-89 on steady-state activation and inactivation as well as recovery from voltage-dependent inactivation. H-89 produced a concentration-dependent rightward shift in voltage-dependent activation but had no significant effect on steady-state inactivation. Recovery from voltage-dependent inactivation was delayed, although this was only visible at the highest concentration (60 ¿mol l¿1) used. In experiments investigating the effects of elevated cyclic AMP, the ß-adrenergic agonist isoprenaline and the phosphatase inhibitor calyculin A had no major effects on Ito or IK1. Data suggest that the effects of H-89 on K+ currents are more complex than simple inhibition of PKA-mediated phosphorylation.
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Yun-Wen, Chen, and 陳韻雯. "Effects of ketone body on the expression and ubiquitination of transcription factor Smads and cyclin-dependent kinase inhibitior p21Waf1and p27Kip1 in LLC-PK1." Thesis, 2002. http://ndltd.ncl.edu.tw/handle/87526157800905178631.
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碩士高雄醫學大學
生物化學研究所
90
Diabetic nephropathy (DN) is a major cause of morbidity and mortality in diabetic patients. The hallmark of DN is cellular hyperplasia, hypertrophy, expansion of extracellular matrix and deranged growth factors (e.g. transforming growth factor-β, TGF-β and its downstream smads) and cell cycle regulators (e.g.p21Waf1and p27Kip1) in the gomeruli and renal tubules. Hypertrophy consists of increased synthesis and/or decreased degradation of protein. Ubiquitination and proteasome pathway is one of the important mechanisms for protein degradation (e.g.Smad, p21Waf1and p27Kip1). But the role of protein ubiquitination in DN remains unknown. High glucose and advanced glycation end-product(AGE) have been proposed to be essential factors in DN. For example, we had shown that high glucose decreased cellular mitogenesis while increasing cellular hypertrophy and collage synthesis by inducing TGF-β1 protein expression in the proximal tubule-like LLC-PK1 cells. Apart from patients with diabetic ketoacidosis, serum ketone body (e.g.β- hydroxybutyrate) level is also often elevated in patients with type I or type II diabetes mellitus. However, the effects of ketone body on proximal tubule is still unknown. Therefore, the purpose of this study was to elucidate the effects of -hydroxybutyrate in LLC-PK1 cells in terms of cellular growth. We found that high glucose (500mg/dl) and β-hydroxybutyrate (10mM) decreased cellular proliferation in LLC-PK1 cells at 48 h. Meanwhile, -hydroxybutyrate also increased TGF-β, p21Waf1 and p27Kip1 transcriptional activity at 18-24 h. Interestingly, dominant negative Smad2 and Smad3 plasmids reversed -hydroxy- butyrate—inhibited cellular proliferation. They also reversed β-hydroxy- butyrate—induced p21Waf1 and p27Kip1 protein expression and TGF-β transcriptional activity. Additionally, β-hydroxybutyrate decreased p27Kip1 ubiquitination. In conclusion, β-hydroxybutyrate—inhibited cellular proliferation by inducing TGF-β/Smad2/3, p21Waf1 and p27Kip1 in LLC-PK1 cells. Moreover, β-hydroxybutyrate-induced p27Kip1 protein expression was accompanied by decreased p27Kip1 ubiquitination .
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Chao, Wen-Yu, and 趙文瑜. "HE-145-111 inhibits hepatitis B virus via down-regulation of the metabolic co-activator PGC-1-α in human hepatoma cells." Thesis, 2013. http://ndltd.ncl.edu.tw/handle/51373510279816831400.
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碩士國立陽明大學
生化暨分子生物研究所
101
Hepatitis B virus (HBV) infection may result in cirrhosis and hepatocellular carcinoma. Although currently there is hepatitis B vaccine can prevent HBV infection, for already infected patients, the clinical therapies for HBV have many side effects. Therefore, development of new anti-hepatitis B drugs is still one of the important research topics. Previous studies from our laboratory, it has showed that natural compound Helioxanthin, HE-145 can suppress both HBV gene expression and secretion of viral particles in human hepatoma HepG2/A2 cells. The role of inhibition by HE-145 is via interfering with the host transcriptional machinery of viral promoter. It also found that HE-145-111, one of derivatives of HE-145 may suppress HBV core promoter activity and gluconeogenesis genes, such as glucose-6-phosphatase (G6Pase) and phosphoenolcarboxykinase (PEPCK) expression in human hepatoma cells. In my study, it is interested to know whether the anti-HBV activity is through regulating of gluconeogenesis gene expression, and further to suppress HBV core promoter activity. It has been demonstrated that the inhibition of HBV by HE-145-111 is through down-regulating of the metabolic co-activator PGC-1α. It was found that HE-145-111 suppressed HBV surface antigen secretion and core protein in Hep3B/T2 cells and 1.3ES2 cells respectively. In order to understand the relationships between suppression of HBV and gluconeogenesis by HE-145-111 in Hep3B/T2 cells, the combination of 8-bromo-cAMP (cAMP) and dexamethasone had a synergistic effect on the induction of HBV core promoter activity and mRNA levels of G6Pase, PEPCK, and PGC-1α; and on the induction of PGC-1α protein levels. Also the identification of CPD2 (nt1656-1675) as a response element in the HBV core promoter using serial deletion as well as point mutation at different regions of the core promoter. The overexpression of PGC-1α construct plasmid can reverse the inhibition of core promoter activity. HE-145-111 suppressed gluconeogenesis, G6Pase, PEPCK and PGC-1αmRNAs and PGC-1α protein and HBV core promoter together identifying CPD2 (nt 1656-1675) as binding sequences of core promoter. In 1.3ES2 cells, it was confirmed that HE-145-111 not only suppressed HBV mRNA and core protein levels but also suppressed the mRNA levels of G6Pase, PEPCK and transcription co-activator PGC-1α. In conclusion, HE-145-111 represses the HBV gene expression by down-regulation PGC-1α protein and through regulating directly the host gluconeogenesis. This mechanism study which HBV is controlled by the hepatic metabolic gluconeogenesis may broaden our understanding of the regulation of HBV expression.
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Hempsch, Birgit [Verfasser]. "Experimentelle Therapie der chronischen lymphatischen Leukämie vom B-Zell-Typ (B-CLL) mit den Small-molecule-Inhibitoren CGP 049090 und PKF 115-584 / vorgelegt von Birgit Hempsch." 2009. http://d-nb.info/994578628/34.
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Park, Sung Woo (Calvin). "Activation of EPAC Inhibits the Aquisition of Nucleus Accumbens Amphetamine Place Preference in a Dose-Dependent Manner in Rats." Thesis, 2008. http://hdl.handle.net/1974/1176.
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Reward-related learning occurs when previously neutral stimuli acquires an enhanced ability to elicit approach and other responses. Studies in the past have shown that dopamine receptor-mediated 3’,5’-cyclic adenosine monophosphate (cAMP)-dependent intracellular signalling is important for reward-related learning. Until recently, cAMP-dependent protein kinase (PKA) was the only known signalling molecule that was activated by cAMP. However, it has been discovered that another enzyme, exchange protein directly activated by cAMP (Epac), is also activated by cAMP. Thus, it is possible that cAMP mediates reward-related learning by an Epac-dependent signalling pathway. The present study used a conditioned place preference (CPP) paradigm to investigate whether Epac is involved in the acquisition of reward-related learning. Bilateral injections of amphetamine (20 µg/0.5μl/side) into the nucleus accumbens (NAc) have been shown in previous studies to reliably produce a CPP. Thus, amphetamine (20 µg) and Sp-adenosine 3’,5’-cyclic monophosphorothioate triethylamanine (Sp-cAMPS) (0.1, 1.0, 10, 15, 20 µg), an agent that activates both PKA and Epac, or amphetamine (20 µg) and 8-(4-chlorophenylthio)-2’-O-methyladenosine-3’,5’-cyclic monophosphate (8-pCPT) (0.73, 1.27, 1.45, 2.89, 5.78, 11.56 µg), an agent that selectively activates Epac, were co-injected into NAc to determine their effects on the acquisition of CPP. Results showed that 8-pCPT (1.45 µg), but not lower or higher doses, inhibited CPP. Sp-cAMPS (0.1, 15, 20 µg) also inhibited CPP, replicating the results of previous studies. The results implicate Epac in the acquisition of reward-related learning.Thesis (Master, Neuroscience Studies) -- Queen's University, 2008-04-25 13:29:37.857
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Lin, Yu-Chen, and 林昱蓁. "Docosahexaenoic Acid Inhibits Tumor Necrosis Factor α-Induced Intercellular Adhesion Molecule-1 Expression through Upregulation of DNA Methyltransferase-3b-Depedent DNA Methylation and Suppression of PKCζ/ERK/Sp1 Pathway in EA.hy926 cells." Thesis, 2014. http://ndltd.ncl.edu.tw/handle/w6m477.
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碩士中國醫藥大學
營養學系碩士班
102
Epigenetic regulation is a key mechanism in either the activation or the silencing of gene transcription. Tumor necrosis factor alpha (TNFα), a potent inflammatory mediator, has multiple effects on the pathogenesis of atherosclerosis. Specificity protein 1 (Sp1), a zinc finger transcription factor, binds to the GC-rich elements in the promoter of inducible genes. Docosahexaenoic acid (DHA), an n-3 fatty acid (n-3 FA), has potent anti-inflammatory effect. A previous study showed that Sp1 is activated by TNFα and induces intercellular adhesion molecule 1 (ICAM-1) expression. In our previous study, we have demonstrated that DHA inhibits TNFα-induced ICAM-1 experssion via enhancing HO-1 expression. In this study, we try to investigate the involvement of Sp1 and epigenetic mechanism in DHA inhibition of TNFα-induced ICAM-1 expression in EA.hy 926 cells. Treatment with the DNA methylation inhibitor reagent 5-aza-dC reversed DHA inhibition of TNFα-induced ICAM-1 expression. Transfection with shDNMT-3b knocks down DNMT-3b expresson and abolished DHA inhibition of TNFα-induced ICAM-1 expression. TNFα induces phosphorylation of Akt, p38 MAPK, ERK1/2 and PKCζ as well as Sp1. Treatment with ERK inhibitor (PD98059), PKCζ pseudo-substrate inhibitor and PKC inhibitor (Rottlerine) blocked TNFα-induced ICAM-1 expression. Transfection with shSp1 knocked down Sp1expresson and blocked TNFα-induced ICAM-1 expression. DHA inhibited TNFα-induced phosphorylation of Sp1.Taken together, the anti-inflammatory effect of DHA is associated with down-regulation of TNFα-induced phosphorylation of ERK1/2 and PKCζ and subsequent Sp1 phosphorylatio which increases ICAM-1 expression in endothelial cells. In addition, an increase in ICAM-1 DNA methylation by DHA may contribute to the down-regulation of TNFα-induced ICAM-1 expression.
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Hofman, Tomáš. "Vliv malých DNA virů na regulaci tvorby interferónu." Master's thesis, 2018. http://www.nusl.cz/ntk/nusl-380361.
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Plasmacytoid dendritic cells (pDC) represent innate immune cells capable to detect viruses in their endosomal environment via Toll-like receptors (TLRs). Viral nuclear acid recognition leads to the massive production of type I interferon (IFN I) and induction of the antiviral state in uninfected cells. Crosslinking of the surface regulatory receptors, such as BDCA-2, with monoclonal antibodies or with some viruses leads to the activation of MEK1/2- ERK signaling pathway and inhibition of IFN I production in pDC. In this study, the role of MEK1/2 kinase has been highlighted. Its inhibition reversed the inhibitory effect of BDCA-2 crosslinking and its direct activation with PMA led to the inhibition of IFN-α production. Yet an unclear role of pDC in sensing of BK polyomavirus virus (BKV) responsible for kidney transplant rejection was investigated as a major topic of this thesis. Experiments with the pDC cell line Gen2.2 and HRPTEC primary cell line showed that pDCs were not able to detect BKV particles, however, exposure of activated Gen2.2 cells to BKV inoculum dramatically upregulated production of IFN-α. Most importantly, coculture of Gen2.2 cells with BKV- infected HRPTEC cells resulted in IFN-α and TNF-α production, which was prevented by Bafilomycin. These results suggest that BKV-infected...
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Surup, Frank. "Metagenom-Technologie zur Wirkstoffsuche sowie Untersuchungen der Iromycine aus Streptomyces sp." Doctoral thesis, 2007. http://hdl.handle.net/11858/00-1735-0000-0006-ACA7-B.
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