Inhibitory Effect of Hypocrellin A on Protein Kinase C in Liver and Skeletal Muscle of Obese Zucker Rats

2003 ◽  
Vol 31 (06) ◽  
pp. 871-878 ◽  
Author(s):  
Xianqin Qu ◽  
Lei Dang ◽  
J. Paul Seale
Keyword(s):  
Skeletal Muscle ◽  
Protein Kinase C ◽  
Protein Kinase ◽  
Inhibitory Effect ◽  
Zucker Rats ◽  
Dependent Manner ◽  
Kinase C ◽  
Hypocrellin A ◽  
Pkc Isoforms ◽  
Obese Zucker Rats

In this ex vivo study, the inhibitory activity of hypocrellin A (HA), a perylene quinonoid pigment isolated from the Chinese medicinal fungus Hypocrella bambuase, on protein kinase C (PKC) enzyme activity in insulin target tissues of obese Zucker rats was assessed. Pre-incubation with HA for 30 minutes significantly inhibited the activity of partially purified PKC enzyme from liver and soleus skeletal muscle in a dose-dependent manner ( IC 50=0.07 and 0.26 μg/ml, respectively). HA produced a greater inhibitory effect in enzyme prepared from the liver than enzyme prepared from soleus muscle. Since total PKC activity in these two insulin target tissues is the net result of several different isoforms of PKC, and PKC-θ is a major isoform expressed in the soleus skeletal muscle, the present data suggest that the naturally occurring compound, HA, may selectively inhibit certain PKC isoforms other than PKC-θ. Further investigations are required to determine which PKC isoforms are most susceptible to HA and whether changes in PKC signaling during treatment with HA can reverse abnormalities of glucose and lipid metabolism in insulin resistant and diabetic states.

scholarly journals Mechanism of inhibitory action of TMB-8 [8-(NN-diethylamino)octyl-3,4,5-trimethoxybenzoate] on aldosterone secretion in adrenal glomerulosa cells

1985 ◽  
Vol 232 (1) ◽  
pp. 87-92 ◽  
Author(s):  
I Kojima ◽  
K Kojima ◽  
H Rasmussen
Keyword(s):  
Protein Kinase C ◽  
Angiotensin Ii ◽  
Protein Kinase ◽  
Calcium Release ◽  
Calcium Influx ◽  
Inhibitory Effect ◽  
Dependent Manner ◽  
Aldosterone Secretion ◽  
Kinase C ◽  
Glomerulosa Cells

The mechanism of 8-(NN-diethylamino)octyl-3,4,5-trimethoxybenzoate (TMB-8) action was evaluated in isolated adrenal glomerulosa cells. TMB-8 inhibits both angiotensin II- and K+-stimulated aldosterone secretion in a dose-dependent manner. The ID50 for angiotensin II- and K+-stimulated aldosterone secretion is 46 and 28 microM, respectively. In spite of the fact that 100 microM-TMB-8 inhibits angiotensin II-stimulated aldosterone secretion almost completely, TMB-8 (100 microM) does not inhibit angiotensin II-induced 45Ca2+ efflux from prelabelled cells nor does it affect inositol 1,4,5-trisphosphate-induced calcium release from non-mitochondrial pool(s) in saponin-permeabilized cells. TMB-8 has no inhibitory effect on A23187-induced aldosterone secretion, but 12-O-tetradecanoylphorbol 13-acetate-induced aldosterone secretion is completely abolished. TMB-8 effectively inhibits both angiotensin II- and K+-induced increases in calcium influx but has no effect on A23187-induced calcium influx. TMB-8 inhibits the activity of protein kinase C dose-dependently. These results indicate that TMB-8 inhibits aldosterone secretion without inhibiting mobilization of calcium from an intracellular pool. The inhibitory effect of TMB-8 is due largely to an inhibition of plasma membrane calcium influx, but this drug also inhibits the activity of protein kinase C directly.

scholarly journals Protein kinase C isoforms ɛ, η, δ and ζ in murine adipocytes: expression, subcellular localization and tissue-specific regulation in insulin-resistant states

1996 ◽  
Vol 316 (3) ◽  
pp. 865-871 ◽  
Author(s):  
Ernst U. FREVERT ◽  
Barbara B. KAHN
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Subcellular Localization ◽  
Response To Treatment ◽  
Dependent Manner ◽  
Insulin Resistant ◽  
Kinase C ◽  
Specific Distribution ◽  
Pkc Isoforms ◽  
Brown Adipose

The Ca2+-insensitive protein kinase C (PKC) isoforms ε, η, δ and ζ are possible direct downstream targets of phosphatidylinositol 3-kinase (PI3-K), and might therefore be involved in insulin signalling. Although isoform-specific changes in PKC expression have been reported for skeletal muscle and liver in insulin-resistant states, little is known about these isoforms in adipocytes. Therefore we studied (1) expression and subcellular localization of these isoforms in murine adipocytes, (2) translocation of specific isoforms to membranes in response to treatment with insulin and phorbol 12-myristate 13-acetate (PMA) and (3) regulation of expression in insulin-resistant states. The PKC isoforms ε, η, δ and ζ are expressed in adipocytes. Immunoreactivity for all isoforms is higher in the membranes than in the cytosol, but subcellular fractionation by differential centrifugation shows an isoform-specific distribution within the membrane fractions. PMA treatment of adipocytes induces translocation of PKC-ε and -δ from the cytosol to the membrane fractions. Insulin treatment does not alter the subcellular distribution of any of the isoforms. 3T3-L1 adipocytes express PKC-ε and -ζ, and PKC-ε expression increases with differentiation from preadipocytes to adipocytes. PKC-ε expression decreases in an adipose-specific and age/obesity-dependent manner in two insulin-resistant models, the brown-adipose-tissue-deficient mouse and db/db mouse compared with control mice. We conclude that, although none of the isoforms investigated seems to be activated by insulin, the decrease in PKC-ε expression might contribute to metabolic alterations in adipocytes associated with insulin resistance and obesity.

scholarly journals Tissue and isoform-selective activation of protein kinase C in insulin-resistant obese Zucker rats - effects of feeding

1999 ◽  
Vol 162 (2) ◽  
pp. 207-214 ◽  
Author(s):  
X Qu ◽  
JP Seale ◽  
R Donnelly
Keyword(s):  
Insulin Resistance ◽  
Protein Kinase C ◽  
Protein Kinase ◽  
Membrane Fraction ◽  
Zucker Rats ◽  
Kinase C ◽  
Pkc Epsilon ◽  
Obese Rats ◽  
Obese Zucker Rats ◽  
Fasted Rats

The mechanisms of insulin resistance in the obese Zucker rat have not been clearly established but increased diacylglycerol-protein kinase C (DAG-PKC) signalling has been associated with decreased glucose utilisation in states of insulin resistance and non-insulin-dependent diabetes mellitus. The purpose of this study was to characterise tissue- and isoform-selective differences in DAG-PKC signalling in insulin-sensitive tissues from obese Zucker rats, and to assess the effects of feeding on DAG-PKC pathways. Groups of male obese (fa/fa, n=24) and lean (fa/-, n=24) Zucker rats were studied after baseline measurements of fasting serum glucose, triglycerides, insulin and oral glucose tolerance tests. Liver, epididymal fat and soleus muscle samples were obtained from fed and overnight-fasted rats for measurements of DAG, PKC activity and individual PKC isoforms in cytosol and membrane fractions. Obese rats were heavier (488+/-7 vs 315+/-9 g) with fasting hyperglycaemia (10.5+/-0.8 vs 7.7+/-0.1 mM) and hyperinsulinaemia (7167+/-363 vs 251+/-62 pM) relative to lean controls. In fasted rats, PKC activity in the membrane fraction of liver was significantly higher in the obese group (174+/-16 vs 108+/-12 pmol/min/mg protein, P<0.05) but there were no differences in muscle and fat. The fed state was associated with increased DAG levels and threefold higher PKC activity in muscle tissue of obese rats, and increased expression of the major muscle isoforms, PKC-theta and PKC-epsilon: e.g. PKC activity in the membrane fraction of muscle from obese animals was 283+/-42 (fed) vs 107+/-20 pmol/min/mg protein (fasting) compared with 197+/-27 (fed) and 154+/-21 pmol/min/mg protein (fasting) in lean rats. In conclusion, hepatic PKC activity is higher in obese rats under basal fasting conditions and feeding-induced activation of DAG-PKC signalling occurs selectively in muscle of obese (fa/fa) rats due to increased DAG-mediated activation and/or synthesis of PKC-theta and PKC-epsilon. These changes in PKC are likely to exacerbate the hyperglycaemia and hypertriglyceridaemia associated with obesity-induced diabetes.

scholarly journals Different roles of protein kinase C α and δ isoforms in the regulation of neutral sphingomyelinase activity in HL-60 cells

1999 ◽  
Vol 344 (3) ◽  
pp. 921-928 ◽  
Author(s):  
Dora VIŠNJIĆ ◽  
Drago BATINIŠ ◽  
Hrvoje BANFIŠ
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Inhibitory Effect ◽  
Interferon Γ ◽  
Neutral Sphingomyelinase ◽  
Kinase C ◽  
Pkc Isoforms ◽  
Ifn Γ ◽  
The Difference ◽  
Specific Regulation

The signalling mechanisms responsible for the hydrolysis of sphingomyelin mediated by 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] and interferon γ (IFN-γ) in HL-60 cells were investigated. IFN-γ was found to increase selectively the activity of cytosolic, Mg2+-independent, neutral sphingomyelinase. The treatment of HL-60 cells with the combination of 1,25(OH)2D3 and IFN-γ had an additive effect on sphingomyelin hydrolysis, ceramide release and the activity of cytosolic, Mg2+-independent, neutral sphingomyelinase. The pretreatment of HL-60 cells with staurosporine, chelerythrine chloride and bisindolylmaleimide abolished the activity of sphingomyelinase in response to 1,25(OH)2D3 and IFN-γ. Calphostin C, which acts on the regulatory site of protein kinase C (PKC), and Gö 6976, a selective inhibitor of Ca2+-dependent PKC isoforms, inhibited the effect of 1,25(OH)2D3 but had no effect on the IFN-γ-mediated increase in activity of sphingomyelinase. Isoform-specific antibodies were used to deplete different PKC isoforms from cytosol before the treatment of the cytosolic fraction with 1,25(OH)2D3, arachidonic acid (AA) and PMA. The depletion of PKC isoforms β1, β2, ϵ, η, μ, ζ and λ had no effect on the activation of sphingomyelinase induced by 1,25(OH)2D3 or by AA. The depletion of PKC α from the cytosol completely abolished the effect of 1,25(OH)2D3 on sphingomyelinase activity but had no effect on the AA-induced activity of sphingomyelinase. PMA had no effect on the activity of sphingomyelinase in either untreated or α-depleted cytosol but significantly increased the activity of sphingomyelinase when added to cytosol depleted of PKC ∆. Moreover, PMA inhibited the effect of 1,25(OH)2D3 on sphingomyelinase activation but the inhibitory effect was abolished by prior depletion of PKC ∆ from the cytosol. These studies demonstrate that 1,25(OH)2D3-induced activation of sphingomyelinase is mediated by PKC α. Furthermore, PKC ∆ had an inhibitory effect on sphingomyelinase, suggesting that the difference between the 1,25(OH)2D3- and PMA-mediated effects on sphingomyelin turnover depends on the specific regulation of the PKC α and PKC ∆ isoforms.

scholarly journals Purification of catalytic domain of rat spleen p72syk kinase and its phosphorylation and activation by protein kinase C

1998 ◽  
Vol 331 (2) ◽  
pp. 649-657 ◽  
Author(s):  
Peter BOROWSKI ◽  
Max HEILAND ◽  
Lutz KORNETZKY ◽  
Sven MEDEM ◽  
Rainer LAUFS
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Tyrosine Kinase ◽  
Kinase Activity ◽  
Catalytic Domain ◽  
Assay Method ◽  
Dependent Manner ◽  
Tyrosine Kinase Activity ◽  
Kinase C ◽  
Pkc Isoforms

The catalytic domain of p72syk kinase (CDp72syk) was purified from a 30000 g particulate fraction of rat spleen. The purification procedure employed sequential chromatography on columns of DEAE-Sephacel and Superdex-200, and elution from HA-Ultrogel by chloride. The analysis of the final CDp72syk preparation by SDS/PAGE revealed a major silver-stained 40 kDa protein. The kinase was identified by covalent modification of its ATP-binding site with [14C]5´-fluorosulphonylbenzoyladenosine and by immunoblotting with a polyclonal antibody against the ‘linker ’ region of p72syk. By using poly(Glu4, Tyr1) as a substrate, the specific activity of the enzyme was determined as 18.5 nmol Pi/min per mg. Casein, histones H1 and H2B and myelin basic protein were efficiently phosphorylated by CDp72syk. The kinase exhibited a limited ability to phosphorylate random polymers containing tyrosine residues. CDp72syk autophosphorylation activity was associated with an activation of the kinase towards exogenous substrates. The extent of activation was dependent on the substrates added. CDp72syk was phosphorylated by protein kinase C (PKC) on serine and threonine residues. With a newly developed assay method, we demonstrated that the PKC-mediated phosphorylation had a strong activating effect on the tyrosine kinase activity of CDp72syk. Studies extended to conventional PKC isoforms revealed an isoform-dependent manner (α > βI = βII > γ) of CDp72syk phosphorylation. The different phosphorylation efficiencies of the PKC isoforms closely correlated with the ability to enhance the tyrosine kinase activity.

Dopamine-induced translocation of protein kinase C isoforms visualized in renal epithelial cells

2000 ◽  
Vol 279 (6) ◽  
pp. C1812-C1818 ◽  
Author(s):  
Susana Nowicki ◽  
Maria Sol Kruse ◽  
Hjalmar Brismar ◽  
Anita Aperia
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Epithelial Cells ◽  
Subcellular Fractionation ◽  
Confocal Imaging ◽  
Receptor Subtype ◽  
Dependent Manner ◽  
Renal Epithelial Cells ◽  
Kinase C ◽  
Pkc Isoforms

Short-term regulation of sodium metabolism is dependent on the modulation of the activity of sodium transporters by first and second messengers. In understanding diseases associated with sodium retention, it is necessary to identify the coupling between these messengers. We have examined whether dopamine, an important first messenger in tubular cells, activates and translocates various protein kinase C (PKC) isoforms. We used a proximal tubular-like cell line, LLCPK-1 cells, in which dopamine was found to inhibit Na+-K+-ATPase in a PKC-dependent manner. Translocation of PKC isoforms was studied with both subcellular fractionation and confocal microscopy. Both techniques revealed a dopamine-induced translocation from cytosol to plasma membrane of PKC-α and -ε, but not of PKC-δ, -γ, and -ζ. The process of subcellular fractionation resulted in partial translocation of PKC-ε. This artifact was eliminated in confocal studies. Confocal imaging permitted detection of translocation within 20 s. Translocation was abolished by a phospholipase C inhibitor and by an antagonist against the dopamine 1 subtype (D1) but not the 2 subtype of receptor (D2). In conclusion, this study visualizes in renal epithelial cells a very rapid activation of the PKC-α and -ε isoforms by the D1 receptor subtype.

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