Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.7.11.13 (protein kinase C)
 49,245 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

BACKGROUND: A number of novel agents that activate or inhibit protein kinase C (PKC) in vitro have been developed to evaluate the physiologic role of PKC in regulation of cellular function. However, most of the PKC inhibitors also affect the protein kinase A, and the effects of these agents in intact myocardium remain still controversial. The present study was carried out to examine the effects of these agents on the positive inotropic effect (PIE) medicated by alpha- and beta-adrenoceptors in isolated rabbit papillary muscle. METHODS AND RESULTS: A potent PKC activator, phorbol 12, 13-dibutyrate (PDBu) at 10 and 30 nM, induced a significant PIE. PDBu at 3 nM and higher inhibited the alpha-mediated PIE and abolished it at 100 nM without affecting the beta-mediated PIE. Phorbol 12-myrisate 13-acetate (PMA) and 1-oleyl-2-acetyl-sn-glycerol (OAG) elicited a similar selective inhibitory action on the alpha-mediated PIE. The PIE of PDBu was abolished by chelerythrine and partially inhibited by staurosporine, but H-7 or calphostin-C did not affect the PIE. These PKC inhibitors consistently inhibited the alpha-mediated PIE by 20-30% at concentrations that they did not affect the beta-mediated PIE. None of the PKC inhibitors influence the PDBu-induced inhibitory action on the alpha-mediated PIE, an indication that they failed to reach the site of the inhibitory action of PDBu. CONCLUSION: Selective modulation by the PKC activators and inhibitors of the alpha-mediated PIE with little effect on the beta-mediated PIE implies that the activation of PKC has a physiological relevance to the alpha-mediated PIE. However, the externally administered PKC activators do not mimic the effect of diacylglycerol that is generated endogenously by alpha-stimulation. By contrast, externally applied PKC inhibitors selectively antagonize the alpha-adrenoreceptor-mediated PIE in rabbit ventricular myocardium.
J Cardiovasc Pharmacol Ther 1997 Jul
PMID:Differential Effects of Protein Kinase C Activators and Inhibitors on alpha- and beta-Adrenoceptor-mediated Positive Inotropic Effect in Isolated Rabbit Papillary Muscle. 1068 55

BACKGROUND: Hydrogen peroxide (H(2)O(2)) in high concentrations has been implicated in heart dysfunction attributable to ischemia-reperfusion. Although H(2)O(2) is also known to increase the intracellular concentration of Ca(2+) ([Ca(2+)](i)) in cardiomyocytes, the mechanisms for such a change are not clear. In this study, the sources and mechanisms of increase in [Ca(2+)](i) caused by high concentrations of H(2)O(2) in cardiomyocytes were explored. METHODS AND RESULTS: Cardiomyocytes were isolated from adult male Sprague-Dawley rats. Cell viability was examined by trypan blue exclusion test. [Ca(2+)](i) was measured by employing cell suspension at room temperature and Fura-2 fluorescence technique. Incubation of cells with 0.25-l mmol/L H(2)O(2) increased [Ca(2+)](i) in a time- and concentration-dependent manner. Catalase attenuated the H(2)O(2)-induced increase in [Ca(2+)](i) significantly, whereas mannitol showed no effect. Neither the presence of verapamil, a sarcolemmal Ca(2+) channel blocker, nor the removal of Ca(2+) from the medium produced any significant reduction in the H(2)O(2)-induced increase in [Ca(2+)](i). Conversely, treatment of cardiomyoctes with staurosporin, a protein kinase C inhibitor, thapsigargin, a sarcoplasmic reticulum Ca(2+)-pump adenosine triphosphatase inhibitor, as well as ryanodine, a sarcoplasmic reticulum Ca(2+)-release channel blocker, markedly prevented the 0.5-mmol/L H(2)O(2)-induced increase in [Ca(2+)](i). The responses of cardiomyoctes to H(2)O(2) and other Ca(2+)-mobilizing agents, such as KCl or adenosine triphosphate, were additive. No changes in cardiomyocyte viability were seen on incubation with 0.5 and 1 mmol/L H(2)O(2). Perfusion of the isolated heart with H(2)O(2) (0.1-0.5 mmol/L) depressed the left ventricular developed pressure, rate of contraction, and rate of relaxation, whereas the left ventricular end-diastolic pressure was increased. CONCLUSIONS: These results indicate that formation of H(2)O(2) under pathophysiological conditions such as ischemic heart disease may induce changes in Ca(2+) homeostasis in cardiomyocytes and may induce contractile dysfunction. Furthermore, the sarcoplasmic reticulum involving a protein kinase C-mediated mechanism appears to be the main site of action of H(2)O(2) in cardiomyocytes.
J Cardiovasc Pharmacol Ther 1999 Jan
PMID:Mechanisms of Hydrogen Peroxide-Induced Increase in Intracellular Calcium in Cardiomyocytes. 1068 23

In this study, we examined the possibility that infarct-size limitation by repetitive preconditioning (PC) is achieved by activation of both protein kinase C (PKC) and tyrosine kinase. In addition, we assessed whether such kinase activation is triggered by angiotensin II type 1 (AT1) and alpha1-adrenergic receptors and whether sarcolemmal and mitochondrial adenosine triphosphate (ATP)-sensitive potassium (K(ATP)) channels play roles as effectors of cardioprotection in the rat. Under pentobarbital anesthesia, myocardial infarction was induced by 20-min coronary occlusion and 3-h reperfusion in the rat. Infarct size was determined by tetrazolium and expressed as a percentage of area at risk (%IS/AR). PC with one cycle of 5-min ischemia/5-min reperfusion before 20-min ischemia significantly reduced %IS/AR from the control value of 49.4 +/- 2.0 to 35.4 +/- 2.8, and repetitive PC with two cycles of 5-min ischemia/5-min reperfusion further limited %IS/AR to 3.2 +/-0.9. Infarct-size limitation by single-cycle PC was completely abolished by a PKC inhibitor, staurosporine (100 microg/kg; %IS/ AR, 45.7 +/- 5.0). In contrast, the cardioprotection by repetitive PC was only partially blocked by staurosporine (%IS/AR, 19.8 +/- 2.4), another PKC inhibitor, polymyxin B (5 mg/kg; %IS/AR, 16.2 +/- 3.1), or a tyrosine kinase inhibitor, genistein (5 mg/kg; %IS/AR, 21.8 +/- 1.4). However, a combined injection of genistein and staurosporine additively inhibited protection of repetitive PC (%IS/AR, 36.4 +/- 1.7). Staurosporine, polymyxin B, or genistein alone did not modify %IS/AR in nonpreconditioned rat hearts. Infarct-size limitation by repetitive PC was not attenuated by pretreatment with a selective AT1-receptor blocker (CV11974, 10 mg/kg), prazosin (0.6 mg/kg; %IS/AR, 6.4 +/- 3.2 and 1.6 +/- 0.5, respectively). A selective blocker of mitochondrial K(ATP) channels, 5-hydroxydecanoate (3 mg/kg), completely abolished the cardioprotective effect (%IS/AR, 50.8 +/-3.5), but HMR1883 (3 mg/kg), a selective blocker of sarcolemmal K(ATP) channels, failed to inhibit the preconditioning effect (%IS/AR, 4.4 +/- 0.7). These findings suggest that repetition of PC provokes activation of both PKC and tyrosine kinase, leading to enhanced antiinfarct tolerance by opening of mitochondrial but not sarcolemmal K(ATP) channels. It is unlikely that activation of either AT1 or alpha1-adrenergic receptor alone is crucial to trigger preconditioning. Key Words: Tyrosine kinase-Genistein-Angiotensin II-alpha1-Adrenergic receptor-Sarcolemmal K(ATP) channel-Mitochondrial K(ATP) channel.
J Cardiovasc Pharmacol 2000 Mar
PMID:Roles of tyrosine kinase and protein kinase C in infarct size limitation by repetitive ischemic preconditioning in the rat. 1071 Jan 17

Phosphoinositide turnover and protein kinase C (PKC) mediate the signaling of angiotensin II, which plays a pivotal role in ventricular remodeling after myocardial infarction (MI). To determine whether PKC is activated after MI, rat hearts after MI were subjected to in vitro quantitative autoradiography with [3H]phorbol 12,13-dibutyrate (PDBu), which is highly selective for PKC. [3H]PDBu binding in the infarcted area increased significantly compared with the non-infarcted region 7 and 21 days after MI, but not 1 and 3 days and 10 months after MI. [3H]PDBu binding in the noninfarcted area was similar to that in the sham-operated rats. Immunohistochemical analysis revealed that abundant macrophages (7 days after MI), fibroblasts, and myofibroblasts (7 and 21 days after MI) occupied the infarcted region. To investigate whether myocardial [3H]PDBu binding is affected by captopril, hearts were subjected to in vitro autoradiography with [3H]PDBu after 1- or 3-week captopril treatment or no treatment. Captopril treatment significantly suppressed [3H]PDBu binding in the infarcted area 3 weeks after MI, but not 1 week after MI nor in the noninfarcted areas. These results suggest that PKC is upregulated during the healing and fibrogenic process after MI and that captopril treatment suppresses the upregulation in the infarcted area.
J Cardiovasc Pharmacol 2000 Mar
PMID:Regional and temporal profiles of phorbol 12,13-dibutyrate binding after myocardial infarction in rats: effects of captopril treatment. 1071 Jan 18

The cellular mechanisms of coronary vasospasm are unclear, and a role for protein kinase C (PKC) activation by the endogenous vasoconstrictors endothelin-1 (ET-1) and prostaglandin F2alpha (PGF2alpha) has been suggested. In this study, we developed a phorbol ester-induced PKC downregulation protocol to investigate the relation between the amount and activity of specific PKC isoforms in coronary arterial smooth muscle and coronary vasoconstriction by ET-1 and PGF2alpha. Isometric tension was measured in deendothelialized porcine coronary artery strips, [Ca2+]i was monitored in single coronary smooth muscle cells loaded with fura-2, and the whole tissue, cytosolic, and particulate fractions were examined for PKC activity and reactivity with isoform-specific anti-PKC antibodies using Western blot analysis. In Ca(2+)-free (2 mM EGTA) Krebs solution, ET-1 (10(-7) M), PGF2alpha (10(-5) M) and PKC activator phorbol 12,13-dibutyrate (PDBu) (10(-6) M) caused significant contractions that were completely inhibited by the PKC inhibitors staurosporine and calphostin C, no significant change in [Ca2+]i, and significant activation and translocation of the Ca(2+)-independent epsilon-PKC but not the Ca(2+)-dependent alpha-PKC. In Ca(2+)-free Krebs, a single application of PDBu produced maximal contraction and PKC activity after 30 min, which declined to basal levels in 3 h and remained steady for 24 h, but did not prevent subsequent increases in contraction and PKC activity with a new addition of PDBu and did not significantly decrease the amount of alpha- or epsilon-PKC. Repeated (five to eight) applications of PDBu in Ca(2+)-free Krebs at 3-h intervals completely inhibited subsequent increases in contraction and PKC activity to PDBu, ET-1, or PGF2alpha, and significantly decreased the amount of epsilon-PKC but not that of alpha-PKC. These results provide evidence that a Ca(2+)-independent coronary vasoconstriction induced by ET-1 and PGF2alpha is associated with activation of the epsilon-PKC isoform. The results suggest that, in coronary artery smooth muscle, downregulation of PKC is isoform specific and is more dependent on the frequency rather than the duration of PKC activation. The results also suggest that repeated downregulation of epsilon-PKC might play a role in preconditioning of the coronary artery against vasoconstriction by ET-1 and PGF2alpha.
J Cardiovasc Pharmacol 2000 Mar
PMID:Preconditioning of coronary artery against vasoconstriction by endothelin-1 and prostaglandin F2alpha during repeated downregulation of epsilon-protein kinase C. 1071 Jan 37

This study investigated the vasorelaxant action of the sesquiterpene polygodial, isolated from the bark of Drymis winteri, on rat portal vein in vitro, contracted by various agonists. Polygodial (21-342 microM) preincubated 20 min before, produced graded antagonism of the contractile responses caused by bradykinin, endothelin-1, noradrenaline, the stable analogue of thromboxane A2 U46619, substance P, neurokinin B, and senktide (an NK3-selective agonist). Polygodial, at the same concentration, also produced graded inhibition of the contractile response induced by potassium chloride and by phorbol ester. At the median inhibitory concentration (IC50) level, polygodial was approximately 114- to 177-fold more active in inhibiting mediated contractions to senktide and phorbol ester. When assessed in the tonic contraction induced by endothelin-1 (0.5 nM) or by phorbol (3 microM), polygodial (0.1-100 microM) produced concentration-dependent relaxation, with maximal inhibition (E(max)) of 62 +/- 2% and 100%, respectively. Finally, polygodial (0.1-100 microM) inhibited the rhythmic spontaneous contractions of the rat portal vein (E(max) of 75 +/- 2%). Taken together, these results suggest that the vasorelaxant actions caused by polygodial in rat portal vein are, at least in part, associated with inhibition of calcium influx through voltage-sensitive channels and interaction with protein kinase C-dependent mechanisms. In addition, these data confirm and extend our previous suggestion that polygodial preferentially antagonizes tachykinin-mediated contraction, especially the NK3-mediated responses.
J Cardiovasc Pharmacol 2000 Apr
PMID:Action of polygodial on agonist-induced contractions of the rat portal vein in vitro. 1077

This study was undertaken to explore possible signal-transduction mechanisms involved in the Ca2+-sensitizing effects of carbachol and endothelin-1 (ET-1) by using beta-escin-skinned smooth muscle of porcine coronary artery. Pretreatment with C3 exoenzyme of Clostridium botulinum, which selectively inactivates rho p21 by adenosine diphosphate (ADP) ribosylation, resulted in a significant inhibition of ET-1-induced Ca2+ sensitization, but had no effect on carbachol-induced Ca2+ sensitization. Whereas the protein kinase C (PKC) inhibitors calphostin C and staurosporine did not affect the Ca2+-sensitizing effect of carbachol, the tyrosine kinase inhibitors genistein and tyrphostin 25 greatly but incompletely suppressed it. In contrast, the Ca2+-sensitizing effect of ET-1 was significantly inhibited by either calphostin C or genistein. Although the inhibitory effect of calphostin C on ET-1-induced Ca2+ sensitization was less than that of genistein, the effects of calphostin C and genistein were additive. The genistein-sensitive component of ET-1-induced Ca2+ sensitization appeared to include the C3-sensitive one. However, a substantial enhancement by ET-1 of the Ca2+-induced contraction was observed even in the presence of the two inhibitors. In beta-escin-skinned smooth muscle of rabbit mesenteric artery, ET-1-induced Ca2+ sensitization was marginally affected by C3 pretreatment, calphostin C, and genistein. We conclude that, although PKC activation and rho p21 protein-dependent and -independent tyrosine phosphorylation each plays an important role in an increase in myofilament Ca2+ sensitivity, the contributions of these signaling pathways to Ca2+ sensitization are different depending on receptor agonists and tissues used. Furthermore, these data suggest the existence of an as yet undefined signal-transduction mechanism involved in Ca2+ sensitization caused by receptor agonists.
J Cardiovasc Pharmacol 2000 May
PMID:Agonist-dependent difference in the mechanisms involved in Ca2+ sensitization of smooth muscle of porcine coronary artery. 1081 86

In this study we report about the modulation of connexin45 (Cx45) gap junction channel properties by phosphorylation of the connexin molecules through different protein kinases. Phosphorylation of Cx45 was studied in HeLa cells transfected with mouse Cx45 (mCx45). Using Western blotting (WB) and immunocytochemistry, these cells were found exclusively positive for Cx45 and the protein was separated as a doublet of bands with a calculated mass of 46 and 48 kD. After dephosphorylation using calf intestine phosphatase (CIP), the 48 kD band disappeared almost completely leaving a single band at 46 kD. This effect can be prevented by including phosphatase inhibitors during CIP treatment. These results indicate that the 48 kD signal represents a phosphorylated form of Cx45. To investigate the effects of (de)phosphorylation of Cx45 on the conductive properties of gap junction channels built of this connexin, cell pairs were subjected to dual voltage clamp experiments and coupling was determined before and after addition of PMA, 4alpha-PDD, cAMP, cGMP, and pervanadate to the superfusate. 100 nM of the PKC activating phorbol ester PMA increased normalized junctional conductance by 50.9+/-28%. 100 nM of the inactive phorbol ester 4alpha-PDD had no significant effect. Activation of PKA with 1 mM 8-Br-cAMP decreased coupling by 20.9+/-5.7% while 1 mM 8-Br-cGMP (PKG-activation) was ineffective. 100 microM pervanadate, a tyrosine phosphatase inhibitor, reduced coupling by 43.7+/-11.1%. Single channel measurements, under identical phosphorylating conditions, were not significantly different from each other and all frequency histograms exhibited two conductance peaks at approximately 20 and 40 pS. WB analysis revealed, as compared to control conditions, a relative increase of the 48 kD signal upon stimulation with pervanadate (142+/-42%) and 8-Br-cAMP (50+/-23%) whereas neither stimulation with PMA nor 8-Br-cGMP had a significant effect. These experiments show that electrical intercellular conductance via Cx45 gap junction channels is differentially regulated by phosphorylation. However, regulation does not act by changing single channel conductance, but most likely by modulation of the open probability of Cx45 gap junction channels.
Cardiovasc Res 2000 Jun
PMID:Electrical conductance of mouse connexin45 gap junction channels is modulated by phosphorylation. 1091 60

The purpose of this study was to test whether extracellular Na+ differentially regulates agonist-induced contraction in vascular smooth muscle. Exposure of rat aorta to 20 nM extracellular Na+ by substitution of 123 mM Na+ with N-methyl-D-glucamine or choline, inhibited norepinephrine-induced contraction to a greater magnitude than contraction to prostaglandin F2alpha. In the absence of extracellular Ca2+ and in 20 mM Na+ solution containing 123 mM N-methyl-D-glucamine, the norepinephrine and prostaglandin F2alpha contraction remained unaltered. In contrast, in the absence of extracellular Ca2+ and in 20 mM Na+ solution containing 123 mM choline, the norepinephrine and prostaglandin F2alpha contraction were decreased and increased, respectively. Contraction to the phorbol ester, phorbol dibutyrate, was inhibited in 20 mM extracellular Na+ solution containing N-methyl-D-glucamine. Removal of extracellular Ca2+ inhibited the phorbol dibutyrate contraction, and 20 mM extracellular Na+ solution containing N-methyl-D-glucamine did not inhibit the phorbol dibutyrate contraction elicited in the absence of extracellular Ca2+. Complete replacement of extracellular Na+ with choline, and concomitant treatment with nifedipine to reduce the elevated basal tone after Na+ replacement, also resulted in greater inhibition of norepinephrine- as compared with prostaglandin F2alpha-induced contraction. Ethylisopropylamiloride, a Na+/H+ exchange inhibitor, did not alter norepinephrine contraction, as determined in the presence of nifedipine to reduce the elevated basal tone due to ethylisopropylamiloride. Acidification, which may result from decreased Na+/H+ exchange, inhibited the prostaglandin F2alpha-induced contraction to a greater magnitude than contraction to norepinephrine. These results demonstrate that extracellular Na+ selectively regulates agonist-induced contraction. The study further suggests that the selectivity may be related to an extracellular Na+-dependent process that is activated by protein kinase C, such as Na+/Ca2+ exchange, and is unrelated to the release of intracellular Ca2+ and Na+/H+ exchange.
J Cardiovasc Pharmacol 2000 Sep
PMID:Differential regulation of norepinephrine- and prostaglandin F2alpha-induced contraction by extracellular Na+ in rat aorta. 1097 84

Apolipoprotein (apo) A-I generates high-density lipoprotein (HDL) by removing cellular cholesterol and phospholipid on the interaction with cells as a main source of plasma HDL. The reaction is induced by dibutylyl cyclic (dbc) adenosine monophosphate (AMP) in RAW 264, mouse macrophage cell line cells, and we investigated its pharmacologic modulation using this cell model. Release of cellular cholesterol and choline phospholipid by apoA-I was increased 9.9 and 4.2 times, respectively, by pretreatment of the cells with 300 microM dbcAMP for 24 h. Calmodulin inhibitors, W7 (N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide) and W5 (N(6-aminohexyl)-1-naphthalenesulfonamide), increased the apoA-I-mediated lipid release by 3 times from the dbcAMP-treated cells. The optimal drug concentrations (80 and 160 microM for W7 and W5, respectively) were not parallel with those reported for in vitro calmodulin inhibition (IC50, 28 and 240 microM for W7 and W5, respectively, toward phosphodiesterase activity), and in fact 40 microM W7 showed much stronger intracellular calmodulin inhibition than did 300 microM W5 using S7AAS2, a fluorescent peptide probe. Other calmodulin inhibitors such as amitriptyline, chlorpromazine, and trifluoperazine showed no effect on the apoA-I-mediated cholesterol release. In contrast to these results, neither dbcAMP nor W7 influenced the diffusion-mediated nonspecific cholesterol efflux to lipid microemulsion. We concluded that W7 and W5 increased the interaction of apoA-I with RAW 264 cells to generate more HDL. The effect did not seem directly correlated to their cal modulin inhibition or modulation of cAMP and protein kinase C.
J Cardiovasc Pharmacol 2000 Nov
PMID:Enhancement of the cAMP-induced apolipoprotein-mediated cellular lipid release by calmodulin inhibitors W7 and W5 from RAW 264 mouse macrophage cell line cells. 1106 21


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