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PKC Activity (pkc + activity)
Selected AbstractsMelatonin increases stress fibers and focal adhesions in MDCK cells: participation of Rho-associated kinase and protein kinase CJOURNAL OF PINEAL RESEARCH, Issue 2 2007Gerardo Ramírez-Rodríguez Abstract:, Melatonin cyclically modifies water transport measured as dome formation in MDCK cells. An optimal increase in water transport, concomitant with elevated stress fiber (SF) formation, occurs at nocturnal plasma melatonin concentrations (1 nm) after 6 hr of incubation. Blockage in melatonin-elicited dome formation was observed with protein kinase C (PKC) inhibitors. Despite, this information on the precise mechanism by which melatonin increases SF formation involved in water transport is not known. Focal adhesion contacts (FAC) are cytoskeletal structures, which participate in MDCK membrane polarization. SF organization and vinculin phosphorylation are involved in FAC assembly and both processes are mediated by PKC, an enzyme stimulated by melatonin; in these processes also involved is Rho-associated kinase (ROCK). Thus, we studied FAC formation and the ROCK/PKC pathway as the mechanism by which melatonin increases SF formation and water transport. The results showed that 1 nM melatonin and the PKC agonist phorbol-12-miristate-13-acetate increased FAC. The PKC inhibitor GF109203x, and the ROCK inhibitor Y27632, blocked increased FAC caused by melatonin. ROCK and PKC activities, vinculin phosphorylation and FAC formation were increased with melatonin. The PKC inhibitor, GF109203x, abolished both melatonin stimulated FAC in whole cells and ROCK activity, indicating that ROCK is a downstream kinase in the melatonin-stimulated PKC pathway in MDCK cultured cells that causes an increase in SF and FAC formation. Data also document that melatonin modulates water transport through modifications of the cytoskeletal structure. [source] Protein kinase C modulation of the regulation of sarcoplasmic reticular function by protein kinase A-mediated phospholamban phosphorylation in diabetic ratsBRITISH JOURNAL OF PHARMACOLOGY, Issue 2 2004Satoko Watanuki The goal of this study was to elucidate the possible mechanisms by which protein kinase A (PKA)-mediated regulation of the sarcoplasmic reticulum (SR) via phospholambin protein phosphorylation is functionally impaired in streptozotocin-induced diabetic rats. Phospholamban (PLB) protein and mRNA levels were 1.3-fold higher in diabetic than in control hearts, while protein expression of cardiac SR Ca2+ -ATPase (SERCA2a) was unchanged. Basal and isoprenaline-stimulated phosphorylation of PLB at Ser16 or Thr17 was unchanged in diabetic hearts. However, stronger immunoreactivity was observed at the basal level in diabetic hearts when antiphosphoserine antibody was used. Basal 32P incorporation into PLB was significantly higher in diabetic than in control SR vesicles, but the extent of the PKA-mediated increase in PLB phosphorylation was the same in the two groups of vesicles. Stimulation of Ca2+ uptake by PKA-catalyzed PLB phosphorylation was weaker in diabetic than in control SR vesicles. The PKA-induced increase in Ca2+ uptake was attenuated when control SR vesicles were preincubated with protein kinase C (PKC). PKC activities were increased by more than two-fold in the membranous fractions from diabetic hearts in comparison with control values, regardless of whether Ca2+ was present. This was associated with increases in the protein content of PKC,, PKC,, PKC,, and PKC, in diabetic membranous fractions. The changes observed in diabetic rats were reversed by insulin therapy. These results suggest that PKA-dependent phosphorylation may incompletely counteract the function of PLB as an inhibitor of SERCA2a activity in diabetes in which PKC expression and activity are enhanced. British Journal of Pharmacology (2004) 141, 347,359. doi:10.1038/sj.bjp.0705455 [source] Epidermal Growth Factor Regulates Amino Acid Transport in Chick Embryo Hepatocytes via Protein Kinase CEXPERIMENTAL PHYSIOLOGY, Issue 4 2000Maria Marino System A-mediated amino acid transport, activation of different steps of signal transduction and involvement of different isoforms of protein kinase C (PKC) have been investigated in chick embryo hepatocytes after epidermal growth factor (EGF) stimulation. EGF rapidly (10 min) increased the rate of aminoisobutyric acid (AIB) uptake in chick embryo hepatocytes freshly isolated on the 19th day of embryonic life, while no change was detectable at other embryonal stages. The growth factor stimulation was abolished by PKC and tyrosine kinase inhibitors and was mimicked by 4-phorbol-12-myristate-13-acetate, dimethyl-2 (PMA). EGF treatment did not modify the phosphorylation of the , isoform of phospholipase C (PLC-,), and inositol trisphosphate (IP3) and intracellular calcium levels, but it induced an increase in PKC activity. Our data show that EGF regulates amino acid uptake, via PKC and without PLC-, activation, only in the last period of chick embryo hepatocyte development. The effects of growth factor on PKC activity suggest the involvement of PKC-, and -, isoforms in EGF modulation of amino acid transport. [source] CREB-dependent cyclooxygenase-2 and microsomal prostaglandin E synthase-1 expression is mediated by protein kinase C and calciumJOURNAL OF CELLULAR BIOCHEMISTRY, Issue 6 2006Hung Pham Abstract Cellular production of prostaglandins (PGs) is controlled by the concerted actions of cyclooxygenases (COX) and terminal PG synthases on arachidonic acid in response to agonist stimulation. Recently, we showed in an ileal epithelial cell line (IEC-18), angiotensin II-induced COX-2-dependent PGI2 production through p38MAPK, and calcium mobilization (J. Biol. Chem. 280: 1582,1593, 2005). Agonist binding to the AT1 receptor results in activation of PKC activity and Ca2+ signaling but it is unclear how each pathway contributes to PG production. IEC-18 cells were stimulated with either phorbol-12,13-dibutyrate (PDB), thapsigargin (TG), or in combination. The PG production and COX-2 and PG synthase expression were measured. Surprisingly, PDB and TG produced PGE2 but not PGI2. This corresponded to induction of COX-2 and mPGES-1 mRNA and protein. PGIS mRNA and protein levels did not change. Activation of PKC by PDB resulted in the activation of ERK1/2, JNK, and CREB whereas activation of Ca2+ signaling by TG resulted in the delayed activation of ERK1/2. The combined effect of PKC and Ca2+ signaling were prolonged COX-2 and mPGES-1 mRNA and protein expression. Inhibition of PKC activity, MEK activity, or Ca2+ signaling blocked agonist induction of COX-2 and mPGES-1. Expression of a dominant negative CREB (S133A) blocked PDB/TG-dependent induction of both COX-2 and mPGES-1 promoters. Decreased CREB expression by siRNA blocked PDB/TG-dependent expression of COX-2 and mPGES-1 mRNA. These findings demonstrate a coordinated induction of COX-2 and mPGES-1 by PDB/TG that proceeds through PKC/ERK and Ca2+ signaling cascades, resulting in increased PGE2 production. J. Cell. Biochem. © 2006 Wiley-Liss, Inc. [source] Possible involvement of protein kinase C in the induction of adipose differentiation-related protein by Sterol ester in RAW 264.7 macrophagesJOURNAL OF CELLULAR BIOCHEMISTRY, Issue 2 2001Jin-Shan Chen Abstract The accumulation of lipid droplets in macrophages contributes to the formation of foam cells, an early event in atherosclerosis. It is, therefore, important to elucidate the mechanisms by which lipid droplets accumulate and are utilized. Sterol ester (SE)-laden RAW 264.7 macrophages accumulated lipid droplets in a time-dependent manner up to 16 h, which was enhanced by cotreatment with 0.1 ,M phorbol 12-myristate 13-acetate (PMA). Inhibition of protein kinase C (PKC) activity by cotreatment with 0.3 ,M calphostin C CAL for 16 h resulted in coalescence of small lipid droplets into large ones and increased accumulation of lipid droplets, although to a lesser extent than after PMA cotreatment. Immunostaining for adipose differentiation-related protein (ADRP) revealed a fluorescent rim at the surface of each medium to large lipid droplet. ADRP appearance correlated with lipid droplet accumulation and was regulated by PMA in a time-dependent manner. Induction of ADRP expression by PMA or CAL required SE, since ADRP levels in PMA- or CAL-treated non-SE-laden macrophages were comparable to those in untreated cells. Removal of SE from the incubation medium resulted in the concomitant dissolution of lipid droplets and down-regulation of ADRP. In conclusion, the above results suggest that ADRP may be an important protein in the regulation of lipid droplet metabolism in lipid-laden macrophages and that this regulation may be mediated by PKC activity. J. Cell. Biochem. 83: 187,199, 2001. © 2001 Wiley-Liss, Inc. [source] High glucose inhibits fructose uptake in renal proximal tubule cells: Involvement of cAMP, PLC/PKC, p44/42 MAPK, and cPLA2JOURNAL OF CELLULAR PHYSIOLOGY, Issue 3 2004Su Hyung Park The precise signal that regulates fructose transport in renal proximal tubule cells (PTCs) under high glucose conditions is not yet known although fructose has been recommended as a substitute for glucose in the diets of diabetic people. Thus, we investigated that effect of high glucose on fructose uptake and its signaling pathways in primary cultured rabbit renal PTCs. Glucose inhibited the fructose uptake in a time- and dose-dependent manner. A maximal inhibitory effect of glucose on fructose uptake was observed at 25 mM glucose after 48 h, while 25 mM mannitol and l -glucose did not affect fructose uptake. Indeed, 25 mM glucose for 48 h decreased GLUT5 protein level. Thus, the treatment of 25 mM glucose for 48 h was used for this study. Glucose-induced (25 mM) inhibition of fructose uptake was blocked by pertussis toxin (PTX), SQ-22536 (an adenylate cyclase inhibitor), and myristoylated amide 14,22 (a protein kinase A inhibitor). Indeed, 25 mM glucose increased the intracellular cAMP content. Furthermore, 25 mM glucose-induced inhibition of fructose uptake was prevented by neomycin or U-73122 (phospholipase C inhibitors) and staurosporine or bisindolylmaleimide I (protein kinase C inhibitors). In fact, 25 mM glucose increased the total PKC activity and translocation of PKC from the cytosolic to membrane fraction. In addition, PD 98059 (a p44/42 mitogen-activated protein kinase (MAPK) inhibitor) but not SB 203580 (a p38 MAPK inhibitor) and mepacrine or AACOCF3 (phospholipase A2 inhibitors) blocked 25 mM glucose-induced inhibition of fructose uptake. Results of Western blotting using the p44/42 MAPK and GLUT5 antibodies were consistent with the results of uptake experiments. In conclusion, high glucose inhibits the fructose uptake through cAMP, PLC/PKC, p44/42 MAPK, and cytosolic phospholipase A2 (cPLA2) pathways in the PTCs. © 2004 Wiley-Liss, Inc. [source] Characterization of p21Ras -mediated apoptosis induced by protein kinase C inhibition and application to human tumor cell linesJOURNAL OF CELLULAR PHYSIOLOGY, Issue 2 2004James S. Liou Suppression of PKC activity can selectively induce apoptosis in cells expressing a constitutively activated p21Ras protein. We demonstrate that continued expression of p21Ras activity is required in PKC-mediated apoptosis because farnesyltransferase inhibitors abrogated the loss of viability in p21Ras -transformed cells occurring following PKC inhibition. Studies utilizing gene transfer or viral vectors demonstrate that transient expression of oncogenic p21Ras activity is sufficient for induction of apoptosis by PKC inhibition, whereas physiologic activation of p21Ras by growth factor is not sufficient to induce apoptosis. Mechanistically, the p21Ras -mediated apoptosis induced by PKC inhibition is dependent upon mitochondrial dysregulation, with a concurrent loss of mitochondrial membrane potential (,m). Cyclosporine A, which prevented the loss of ,m, also inhibited HMG-induced DNA fragmentation in cells expressing an activated p21Ras. Induction of apoptosis by PKC inhibition in human tumors with oncogenic p21Ras mutations was demonstrated. Inhibition of PKC caused increased apoptosis in MIA-PaCa-2, a human pancreatic tumor line containing a mutated Ki,ras allele, when compared to HS766T, a human pancreatic tumor line with normal Ki,ras alleles. Furthermore, PKC inhibition induced apoptosis in HCT116, a human colorectal tumor line containing an oncogenic Ki,ras allele but not in a subline (Hke3) in which the mutated Ki,ras allele had been disrupted. The PKC inhibitor 1- O -hexadecyl-2- O -methyl-rac-glycerol (HMG), significantly reduced p21Ras -mediated tumor growth in vivo in a nude mouse MIA-PaCa-2 xenograft model. Collectively these studies suggest the therapeutic feasibility of targeting PKC activity in tumors expressing an activated p21Ras oncoprotein. J. Cell. Physiol. 198: 277,294, 2004. © 2003 Wiley-Liss, Inc. [source] Enhanced generation of Alzheimer's amyloid-, following chronic exposure to phorbol ester correlates with differential effects on alpha and epsilon isozymes of protein kinase CJOURNAL OF NEUROCHEMISTRY, Issue 2 2009Odete A. B. Da Cruz e Silva Abstract Alzheimer's amyloid precursor protein (APP) sorting and processing are modulated through signal transduction mechanisms regulated by protein phosphorylation. Notably, protein kinase C (PKC) appears to be an important component in signaling pathways that control APP metabolism. PKCs exist in at least 11 conventional and unconventional isoforms, and PKC, and PKC, isoforms have been specifically implicated in controlling the generation of soluble APP and amyloid-, (A,) fragments of APP, although identification of the PKC substrate phospho-state-sensitive effector proteins remains challenging. In the current study, we present evidence that chronic application of phorbol esters to cultured cells in serum-free medium is associated with several phenomena, namely: (i) PKC, down-regulation; (ii) PKC, up-regulation; (iii) accumulation of APP and/or APP carboxyl-terminal fragments in the trans Golgi network; (iv) disappearance of fluorescence from cytoplasmic vesicles bearing a green fluorescent protein tagged form of APP; (v) insensitivity of soluble APP release following acute additional phorbol application; and (vi) elevated cellular APP mRNA levels and holoprotein, and secreted A,. These data indicate that, unlike acute phorbol ester application, which is accompanied by lowered A, generation, chronic phorbol ester treatment causes differential regulation of PKC isozymes and increased A, generation. These data have implications for the design of amyloid-lowering strategies based on modulating PKC activity. [source] Ethanol Inhibits Muscarinic Receptor-Induced Axonal Growth in Rat Hippocampal NeuronsALCOHOLISM, Issue 11 2009Kathryn L. VanDeMark Background:, In utero alcohol exposure can lead to fetal alcohol spectrum (FAS) disorders characterized by cognitive and behavioral deficits. In vivo and in vitro studies have shown that ethanol alters neuronal development. One mechanism through which ethanol has been shown to exert its effects is the perturbation of activated signaling cascades. The cholinergic agonist carbachol has been shown to induce axonal outgrowth through intracellular calcium mobilization, protein kinase C (PKC) activation, and ERK1/2 phosphorylation. This study investigated the effect of ethanol on the differentiation of rat hippocampal pyramidal neurons induced by carbachol as a possible mechanism involved in the developmental neurotoxicity of ethanol. Methods:, Prenatal rat hippocampal pyramidal neurons were treated with ethanol (50 to 75 mM) in the presence or absence of carbachol for 24 hours. Neurite outgrowth was assessed spectrophotometrically; axonal length was measured in neurons fixed and immunolabeled with the neuron-specific ,III tubulin antibody; cytotoxicity was analyzed using the thiazolyl blue tetrazolium bromide assay. The effect of ethanol on carbachol-stimulated intracellular calcium mobilization was assessed utilizing the fluorescent calcium probe, Fluo-3AM. The PepTag® assay for nonradioactive detection of PKC from Promega was used to measure PKC activity, and ERK1/2 activation was determined by densitometric analysis of Western blots probed for phospo-ERK1/2. Results:, Ethanol treatment (50 to 75 mM) caused an inhibition of carbachol-induced axonal growth, without affecting neuronal viability. Neuron treatment for 15 minutes with ethanol did not inhibit the carbachol-stimulated rise in intracellular calcium, while inhibiting PKC activity at the highest tested concentration and ERK1/2 phosphorylation at both the concentrations used in this study. On the other hand, neuron treatment for 24 hours with ethanol significantly inhibited carbachol-induced increase in intracellular calcium. Conclusions:, Ethanol inhibited carbachol-induced neurite outgrowth by inhibiting PKC and ERK1/2 activation. These effects may be, in part, responsible for some of the cognitive deficits associated with in utero alcohol exposure. [source] Ethanol Uses cAMP-Independent Signal Transduction Mechanisms to Activate Proenkephalin Promoter Activity in Rat C6 Glioma CellsALCOHOLISM, Issue 7 2000Xiaoju Yang Background: Previous in vivo studies show that acute ethanol exposure sequentially increases protein kinase A (PKA) activity, the phosphorylation of the adenosine 3,:5,-cyclic monophosphate (cAMP) dependent transcription factor, CREB, and finally proenkephalin gene expression. The present study was conducted to determine if ethanol could activate directly the adenylyl cyclase pathway and thus enhance proenkephalin promoter activity. Methods: Cultured rat C6 glioma cells stably transfected with a segment of the five prime flanking region of rat proenkephalin promoter (nucleotide -2700 + 53) ligated to the chloramphenicol acetyltransferase (CAT) reporter gene were employed to study the effects of ethanol on proenkephalin promoter activity. This region of proenkephalin promoter contains two cAMP response elements (CRE-1 and CRE-2) and one AP2 site located in the region upstream of the TATA box. Cultures were exposed to ethanol, isoproterenol, and phorbol-12, myristate 13-acetate (PMA) alone and in combination, in the presence and absence of PKA and protein kinase C (PKC) inhibitors. Results: Ethanol and isoproterenol increased proenkephalin promoter activity in a dose-dependent manner. Ethanol had an additive effect on maximal isoproterenol-stimulated proenkephalin promoter activity, which suggested that ethanol used a cAMP-independent signai transduction pathway to increase proenkephalin promoter activation. In contrast with isoproterenol, ethanol exposure did not increase cAMP accumulation, PKA activity, or the phosphorylated form of CREB. However, ethanol exposure modestly increased PKC activity. The PKA-specific inhibitor, Rp-cAMP, dampened isoproterenol-induced activation of CAT activity but did not alter ethanol's ability to increase CAT activity. However, the PKC inhibitors, chelerthyrine and G07874, abrogated ethanol's effect of CAT activity but did not alter isoproterenol's effects. Conclusions: Ethanol enhanced proenkephalin promoter activity and potentiated isoproterenol stimulated promoter activity through a cAMP-independent pathway. [source] Protein kinase C activity in mouse eggs regulates gamete membrane interaction,MOLECULAR REPRODUCTION & DEVELOPMENT, Issue 11 2007Hiroto Akabane Abstract Gamete membrane interaction is critical to initiate the development of a new organism. The signaling pathways governing this event, however, are poorly understood. In this report, we provide the first evidence that protein kinase C activity in mouse eggs plays a crucial role in the regulation of this process. Stimulating PKC activity in mouse eggs by phorbol 12-myristate 13-acetate (PMA) drastically inhibited the egg's membrane ability to bind and fuse with sperm. Surprisingly, this significant reduction of gamete membrane interaction was also observed in eggs treated with the PKC inhibitors staurosporine and calphostin c. In further analysis, we found that while no change of egg actin cytoskeleton was detected after either PMA or calphostin c treatment, the structural morphology of egg surface microvilli was severely altered in the PMA-treated eggs, but not in the calphostin c-treated eggs. Moreover, sperm, which bound but did not fuse with the eggs treated with the anti-CD9 antibody KMC8, were liberated from the egg membrane after PMA, but not calphostin c, treatment. Taken together, these results suggest that egg PKC may be precisely balanced to regulate gamete membrane interaction in a biphasic mode, and this biphasic regulation is executed through two different mechanisms. Mol. Reprod. Dev. 74: 1465,1472, 2007. © 2007 Wiley-Liss, Inc. [source] Protein Kinase C Activators as Synaptogenic and Memory TherapeuticsARCHIV DER PHARMAZIE, Issue 12 2009Miao-Kun Sun Abstract The last decade has witnessed a rapid progress in understanding of the molecular cascades that may underlie memory and memory disorders. Among the critical players, activity of protein kinase C (PKC) isoforms is essential for many types of learning and memory and their dysfunction, and is critical in memory disorders. PKC inhibition and functional deficits lead to an impairment of various types of learning and memory, consistent with the observations that neurotoxic amyloid inhibits PKC activity and that transgenic animal models with PKC, deficit exhibit impaired capacity in cognition. In addition, PKC isozymes play a regulatory role in amyloid production and accumulation. Restoration of the impaired PKC signal pathway pharmacologically results in an enhanced memory capacity and synaptic remodeling / repair and synaptogenesis, and, therefore, represents a potentially important strategy for the treatment of memory disorders, including Alzheimer's dementia. The PKC activators, especially those that are isozyme-specific, are a new class of drug candidates that may be developed as future memory therapeutics. [source] Modulation of protein kinase C by curcumin; inhibition and activation switched by calcium ionsBRITISH JOURNAL OF PHARMACOLOGY, Issue 2 2007Y A Mahmmoud Background and purpose: Previous studies have identified the natural polyphenol curcumin as a protein kinase C (PKC) inhibitor. In contrast, we found significant stimulation of PKC activity following curcumin treatment. Thus, the mechanism of curcumin interaction with PKC was investigated. Experimental approach: We employed phosphorylation assays in the presence of soluble or membrane-bound PKC substrates, followed by SDS,PAGE, autoradiography and phosphorylation intensity measurements. Key results: Curcumin inhibited PKC in the absence of membranes whereas stimulation was observed in the presence of membranes. Further analysis indicated that curcumin decreased PKC activity by competition with Ca2+ stimulation of the kinase, resulting in inhibition of activity at lower Ca2+ concentrations and stimulation at higher Ca2+ concentrations. The role of the membrane is likely to be facilitation of Ca2+ -binding to the kinase, thus relieving the curcumin inhibition observed at limited Ca2+ concentrations. Curcumin was found to mildly stimulate the catalytic subunit of PKC, which does not require Ca2+ for activation. In addition, studies on Ca2+ -independent PKC isoforms as well as another curcumin target (the sarcoplasmic reticulum Ca2+ -ATPase) confirmed a correlation between Ca2+ concentration and the curcumin effects. Conclusions and Implications: Curcumin competes with Ca2+ for the regulatory domain of PKC, resulting in a Ca2+ -dependent dual effect on the kinase. We propose that curcumin interacts with the Ca2+ -binding domains in target proteins. To our knowledge, this is the first study that defines an interaction domain for curcumin, and provides a rationale for the broad specificity of this polyphenol as a chemopreventive drug. British Journal of Pharmacology (2007) 150, 200,208. doi:10.1038/sj.bjp.0706970 [source] YC-1 increases cyclo-oxygenase-2 expression through protein kinase G- and p44/42 mitogen-activated protein kinase-dependent pathways in A549 cellsBRITISH JOURNAL OF PHARMACOLOGY, Issue 4 2002Ming-Shyan Chang YC-1, an activator of soluble guanylate cyclase (sGC), has been shown to increase the intracellular cGMP concentration. This study was designed to investigate the signaling pathway involved in the YC-1-induced COX-2 expression in A549 cells. YC-1 caused a concentration- and time-dependent increase in COX activity and COX-2 expression in A549 cells. Pretreatment of the cells with the sGC inhibitor (ODQ), the protein kinase G (PKG) inhibitor (KT-5823), and the PKC inhibitors (Go 6976 and GF10923X), attenuated the YC-1-induced increase in COX activity and COX-2 expression. Exposure of A549 cells to YC-1 caused an increase in PKC activity; this effect was inhibited by ODQ, KT-5823 or Go 6976. Western blot analyses showed that PKC-,, -,, -,, -, and -, isoforms were detected in A549 cells. Treatment of A549 cells with YC-1 or PMA caused a translocation of PKC-,, but not other isoforms, from the cytosol to the membrane fraction. Long-term (24 h) treatment of A549 cells with PMA down-regulated the PKC-,. The MEK inhibitor, PD 98059 (10,50 ,M), concentration-dependently attenuated the YC-1-induced increases in COX activity and COX-2 expression. Treatment of A549 cells with YC-1 caused an activation of p44/42 MAPK; this effect was inhibited by KT-5823, Go 6976, long-term (24 h) PMA treatment or PD98059, but not the p38 MAPK inhibitor, SB 203580. These results indicate that in human pulmonary epithelial cells, YC-1 might activate PKG through an upstream sGC/cGMP pathway to elicit PKC-, activation, which in turn, initiates p44/42 MAPK activation, and finally induces COX-2 expression. British Journal of Pharmacology (2002) 136, 558,567; doi:10.1038/sj.bjp.0704777 [source] Effects of PKC , on early genome transcription activation in mouse 1-cell stage fertilized eggsCELL BIOCHEMISTRY AND FUNCTION, Issue 6 2007Bing-zhi Yu Abstract Effects of PKC , on the activation of embryonic transcription in 1-cell stage fertilized mouse eggs were explored. The effects of PKC antagonist calphostin C and PKC , specific inhibitor on the activation of embryonic early transcription were observed by Western blotting and cell immunofluorescence. PKC activity increased gradually from G1 phase to late G2 phase in mouse 1-cell stage fertilized eggs, and reached a maximum in G2 stage. Calphostin C inhibited PKC activity by about 47% in 1-cell stage fertilized eggs. Calphostin C inhibited early transcription in 1-cell stage fertilized eggs (p,<,0.01). PKC ,-Thr410 in G2 were about 27% and 110% higher than those in G1 phase of 1-cell stage fertilized eggs and MII oocytes, respectively. PKC , specific inhibitor can also inhibit early transcription in 1-cell stage fertilized eggs (p,<,0.05). The results suggest that PKC , participates in early transcription activation in mouse 1-cell stage fertilized eggs. Copyright © 2006 John Wiley & Sons, Ltd. [source] Effects of the PKC inhibitor PD 406976 on cell cycle progression, proliferation, PKC isozymes and apoptosis in glioma and SVG-transformed glial cellsCELL PROLIFERATION, Issue 2 2005C. Russell However, reports differ on which PKC isozymes are responsible for glioma proliferation. As a means to further elucidate this, the objectives of our research were to determine how inhibition of PKC-,, PKC-, and PKCµ with PD 406976 regulates the cell cycle, cell proliferation and PKC during glioma growth and development. To establish the cell cycle effects of PD 406976 on brain cells (SVG, U-138MG and U-373MG glioma cells), specimens were treated with either dimethylsulfoxide (DMSO; control) or PD 406976 (2 µm). Results from flow cytometry demonstrated that PD 406976 delayed the entry DNA synthesis phase in SVG cells and delayed the number of cells entering and exiting the DNA synthesis phase in both U-138MG and U-373MG cells, indicating that PD 406976 may inhibit G1/S and S phase progression. Assessment of cell viability demonstrated a cytostatic effect of PD 406976 on SVG, U-138MG and U-373MG glioma cell proliferation. The PD 406976-induced decreased proliferation was sustained at 48,96 h. A PKC activity assay was quantified and demonstrated that exposure of SVG and U-373MG glioma cells to PD 406976 suppressed PKC activity. Western blotting demonstrated reduced PKC-,1, PKC-, and PKC-, protein content in cells treated with PD 406976. We determined that the growth inhibitory effect of PD 406976 was not as a result of apoptosis. [source] DOPAMINE D2 RECEPTOR STIMULATION INHIBITS ANGIOTENSIN II-INDUCED HYPERTROPHY IN CULTURED NEONATAL RAT VENTRICULAR MYOCYTESCLINICAL AND EXPERIMENTAL PHARMACOLOGY AND PHYSIOLOGY, Issue 3 2009Hong Li SUMMARY 1Myocardial hypertrophy is a common pathological change that accompanies cardiovascular disease. Dopamine D2 receptors have been demonstrated in cardiovascular tissues. However, the pathophysiological involvement of D2 receptors in myocardial hypertrophy is unclear. Therefore, the effects of the D2 receptor agonist bromocriptine and the D2 receptor antagonist haloperidol on angiotensin (Ang) II- or endothelin (ET)-1-induced hypertrophy of cultured neonatal rat ventricular myocytes were investigated in the present study. 2Protein content and protein synthesis, determined by examining [3H]-leucine uptake, were used as estimates of cardiomyocyte hypertrophy. The expression of D2 receptor protein in neonatal rat ventricular myocytes was determined using western blotting. Changes in [Ca2+]i in cardiomyocytes were observed by laser scanning confocal microscopy. 3Angiotensin II and ET-1, both at 10 nmol/L, induced myocyte hypertrophy, as demonstrated by increased protein content and synthesis, [Ca2+]i levels, protein kinase C (PKC) activity and phosphorylation of extracellular signal-regulated kinase, c-Jun N-terminal kinase and mitogen-activated protein kinase (MAPK) p38 (p38). Concomitant treatment of cells with 10 nmol/L AngII plus 10 µmol/L bromocriptine significantly inhibited cardiomyocyte hypertrophy, MAPK phosphorylation and PKC activity in the membrane, as well as [Ca2+]i signalling pathways, compared with the effects of AngII alone. In addition, 10 µmol/L bromocriptine significantly inhibited cardiomyocyte hypertrophy induced by 10 nmol/L ET-1. However, pretreatment with haloperidol (10 µmol/L) had no significant effects on cardiomyocyte hypertrophy induced by either AngII or ET-1. 4In conclusion, D2 receptor stimulation inhibits AngII-induced hypertrophy of cultured neonatal rat ventricular myocytes via inhibition of MAPK, PKC and [Ca2+]i signalling pathways. [source] HIGH GLUCOSE-INDUCED HUMAN UMBILICAL VEIN ENDOTHELIAL CELL HYPERPERMEABILITY IS DEPENDENT ON PROTEIN KINASE C ACTIVATION AND INDEPENDENT OF THE Ca2+,NITRIC OXIDE SIGNALLING PATHWAYCLINICAL AND EXPERIMENTAL PHARMACOLOGY AND PHYSIOLOGY, Issue 9 2005Lei Dang SUMMARY 1.,Endothelial barrier dysfunction plays a pivotal role in the pathogenesis of diabetic vascular complications. The precise molecular mechanisms by which hyperglycaemia causes the increased permeability in endothelial cells are not yet well understood. In the present study, we investigated whether high concentrations of glucose induce endothelial permeability through the activation of protein kinase C (PKC) and/or the calcium,nitric oxide (NO) signalling pathway in human umbilical vein endothelial cells (HUVEC). 2.,Endothelial permeability was measured by albumin diffusion across endothelial monolayers under the stimuli of high glucose (HG; 20 mmol/L), 100 nmol/L phorbol-myristate-acetate (PMA) or 100 nmol/L histamine. The intracellular calcium concentration ([Ca2+]i) was detected in HUVEC using the fluorescent probe fura-2 AM. The effects of PKC inhibitors (LY379196 and hypocrellin A) and the NO synthase (NOS) inhibitor NG -monomethyl- l -arginine (l -NMMA) on endothelial permeability and [Ca2+]i were determined. 3.,High glucose and PMA increased endothelial permeability associated with decreased [Ca2+]i, whereas histamine triggered significant increases in endothelial permeability, accompanied by increases in [Ca2+]i in HUVEC. Hypocrellin A (HA) and LY379196 reversed both HG- and histamine-induced endothelial permeability. The NOS inhibitor l -NMMA only abolished histamine- and not HG-induced endothelial permeability. Neither LY379196, HA nor l -NMMA had any significant effects on alterations in [Ca2+]i caused by HG and histamine. 4.,These results indicate that increased endothelial permeability in HUVEC induced by HG is dependent on PKC activity and is independent of the [Ca2+]i,NO pathway. Increased endothelial permeability due to other inflammatory factors, such as histamine, may also be mediated by the PKC pathway. Thus, PKC inhibitors would be a potential therapeutic approach to endothelial dysfunction induced by hyperglycaemia, as well as other inflammatory factors, in diabetes. [source] | ||||||||||||||||||||||||||||