PKC

Cellular responses to changes in the extracellular environment are mediated by intracellular signaling systems. One of the most extensively studied systems is adenylate cyclase which generates the second messenger molecule cAMP. Another one is the phosphatidylinositol (PI) second messenger system giving rise to IP3 and diacylglycerol, the latter stimulating protein kinase C. Recently, a third potential signaling system has attracted increased scientific attention: the phospholipase A2 system which generates arachidonic acid. This substance may be used for eicosanoid synthesis or serve as a second messenger molecule. The present report gives more evidence about mechanisms how these signaling pathways interact in cultured astrocytes. Substances commonly used for stimulation of arachidonic acid release and prostaglandin synthesis in these cultures (A23187, TPA) had no influence on intracellular cAMP levels. Pertussis toxin that had previously been shown to inhibit prostaglandin synthes...

Purpose: The second generation of tyrosine kinase inhibitors is a group of compounds that inhibit c-kit receptor activity and therefore widely used in the treatment of mastocytosis. In this research, the relationship between the mechanism of action of tyrosine kinase inhibitors and protein kinase C is investigated in HMC-1 560 or HMC-1 560,816 cell lines. Results: From all the tyrosine kinase inhibitors tested, nilotinib is the compound that has the highest cytotoxic effect against HMC-1 560 mast cell line, while midostaurin is the most potent in HMC-1 560,816. Moreover, an increase on histamine release is observed after protein kinase C activation either in HMC-1 560 or HMC-1 560,816 cells. Furthermore, dasatinib increases histamine release in both mast cell lines, which could be related with the secondary reactions previously described in dasatinib-treated patients. Dasatinib also induces Ca 2+-dependent protein kinase C isoforms translocation from the cytosol to the membrane, whereas protein kinase Cd is translocated from the cytosol to the nucleus in the HMC-1 560,816 cell line, but not in HMC-1 560 cells. Conclusion: Results obtained demonstrate that dasatinib induces an important cytotoxic effect in both HMC-1 cell lines and differently regulates protein kinase Cd in HMC-1 560 and HMC-1 560,816 cells. Finally, our results confirm that PKCd is an essential target for dasatinib.

The innate immune response constitutes the first line of defense against invading pathogens and consists of a variety of immune defense mechanisms including the respiratory burst of phagocytes. Respiratory burst can be used as a reliable measure of the immune response of a host, and numerous assays have been developed to measure this response in a variety of mammal and fish species. Phagocytes, like granulocytes and macrophages, that are derived from different tissues, or grown in cell culture, have been employed in a range of assay formats employing a variety of detection methods. The small size of the zebrafish has prevented the large-scale extraction of these cells for respiratory burst assays in the zebrafish. In this work, we describe a respiratory burst assay developed for the zebrafish using intact kidneys and embryos as sources of phagocytes. Phorbol myristate acetate (PMA)-inducible reactive oxygen species (ROS) were detected following the oxidation of a non-fluorescent dye 2′,7′-dihydrodichlorofluorescein diacetate (H2DCFDA) to dichlorofluorescein (DCF), a fluorescent product. Embryos from 1 day post-fertilization until 5 days post-fertilization (dpf) were employed in this assay. Abrogation of H2DCFDA oxidation by the protein kinase C (PKC) inhibitor bisindolylmaleimide I (BisI) indicated a reduction in the respiratory burst. Fluorescence from the PMA-induced respiratory burst in kidneys and embryos was significantly elevated above DMSO-treated controls, while preincubation with BisI inhibited the increase in fluorescence. Colocalization of cell-associated chloromethyl-dihydrodichlorofluorescein diacetate (CM-H2DCFDA) with the phagocyte-selective dye neutral red is consistent with the observation that macrophages and granulocytes are the ROS-producing cells in the zebrafish.

We present two new schemes for efficient certificate revocation. Our first scheme is a direct improvement on a well-known tree-based variant of the NOVOMODO system of Micali [11]. Our second scheme is a direct improvement on a tree-based variant of a multi-certificate revocation system by Aiello, Lodha, and Ostrovsky [1]. At the core of our schemes is a novel construct termed a QuasiModo tree, which is like a Merkle tree but contains a length-2 chain at the leaves and also directly utilizes interior nodes. This concept is of independent interest, and we believe such trees will have numerous other applications. The idea, while simple, immediately provides a strict improvement in the relevant time and communication complexities over previously published schemes.

We present new and efficient key-recovery chosen-ciphertext attacks on NTRUencrypt. Our attacks are somewhat intermediate between chosen-ciphertext attacks on NTRUencrypt previously published at CRYPTO ’00 and CRYPTO ’03. Namely, the attacks only work in the presence of decryption failures; we only submit valid ciphertexts to the decryption oracle, where the plaintexts are chosen uniformly at random; and the number of oracle queries is small. Interestingly, our attacks can also be interpreted from a provable security point of view: in practice, if one had access to a NTRUencrypt decryption oracle such that the parameter set allows decryption failures, then one could recover the secret key. For instance, for the initial NTRU-1998 parameter sets, the output of the decryption oracle on a single decryption failure is enough to recover the secret key.

Dysfunction of GABAA receptor-mediated inhibition is implicated in a number of neurological and psychiatric conditions including epilepsy and affective disorders. Some of these conditions have been associated with abnormal levels of certain endogenously occurring neurosteroids, which potently and selectively enhance the function of the brain’s major inhibitory receptor, the GABAA receptor. Consistent with their ability to enhance neuronal inhibition, such steroids exhibit in animals and humans anxiolytic, anticonvulsant and anesthetic actions. Neurosteroids, exemplified by the potent progesterone metabolite, 5α-pregnan-3α-ol-20-one can be synthesized de novo in the CNS both in neurones and glia in levels sufficient to modulate GABAA receptor function. Neurosteroid levels are not static, but are subject to dynamic fluctuations, for example during stress, or the later stages of pregnancy. These observations suggest that these endogenous modulators may refine the function of the brain’s major inhibitory receptor and thus, play an important physiological and pathophysiological role. However, given the ubiquitous expression of GABAA receptors throughout the mammalian CNS, changes in neurosteroid levels should be widely experienced, causing a generalized enhancement of neuronal inhibition. Such a non-specific action would seem incompatible with a physiological role. However, neurosteroid action is both brain region and neurone selective. This specificity results from a variety of molecular mechanisms including receptor subunit composition, local steroid metabolism and phosphorylation. This paper will evaluate the relative contribution these mechanisms play in defining the interaction of neurosteroids with synaptic and extra-synaptic GABAA receptors.

Protein kinase C (PKC) plays a major role in regulation of both pre and postsynaptic neurotransmission. Excessive activation of PKC results in symptoms related to bipolar disorder. Tamoxifen, a widely used breast cancer drug is known to inhibit PKC and demonstrate antimanic properties in human. We describe herein the synthesis of endoxifen, a tamoxifen active metabolite and compared its PKC inhibitory activity with that of tamoxifen. Endoxifen exhibited fourfold higher potency compared to tamoxifen.A novel synthesis of endoxifen and its PKC inhibitory activity is reported.

Agmatine is an endogenous neuromodulator which, based on animal and human studies, is a putative novel antidepressant drug. In this study, we investigated the ability of sub-chronic (21 days) p.o. agmatine administration to produce an antidepressant-like effect in the tail suspension test and examined the hippocampal cell signaling pathways implicated in such an effect. Agmatine at doses of 0.01 and 0.1 mg/kg (p.o.) produced a significant antidepressant-like effect in the tail suspension test and no effect in the open-field test. Additionally, agmatine (0.001-0.1 mg/kg, p.o.) increased the phosphorylation of protein kinase A substrates (237-258% of control), protein kinase B/Akt (Ser(473)) (116-127% of control), glycogen synthase kinase-3β (Ser(9)) (110-113% of control), extracellular signal-regulated kinases 1/2 (119-137% and 121-138% of control, respectively) and cAMP response elements (Ser(133)) (127-152% of control), and brain-derived-neurotrophic factor (137-175% of control) im...