Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.3.1.21 (CPT)
 4,580 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Extracellular ATP suppressed the growth of HL-60 leukemia cells and induced their differentiation as revealed by N-formyl-methionyl-leucyl-phenylalanine-induced beta-glucuronidase release. ATP degraded to ADP, AMP, and adenosine, and the effect of ATP on cell growth was mimicked by these metabolites added to the cultures. The stable analog alpha,beta-methylene ATP, however, had only a weak inhibitory effect on cell growth. Adenine nucleotide-induced growth suppression was reversed by uridine, suggesting the involvement of intracellular pyrimidine starvation secondary to adenosine accumulation. Consistent with this, ATP induced intracellular starvation of pyrimidine nucleotides, and this effect was also prevented by pretreatment of cells with uridine. The order of effectiveness of ATP-induced differentiation of HL-60 cells, unlike that for growth suppression, was ATP > ADP > AMP, and adenosine had no effect. Furthermore, uridine had no effect and the stable analog, alpha,beta-methylene ATP also induced HL-60 cell differentiation, suggesting that differentiation was due to ATP per se. We tested the hypothesis that ATP-induced differentiation arises from activation of adenylyl cyclase by the novel P2Y(11) receptor using the cell-permeable inhibitor of protein kinase A, Rp-CPT-cAMPS (8-(4-chlorophenylthio)adenosine-3',5'-cyclic monophosphorothioate, Rp isomer). Rp-CPT-cAMPS (1-100 microM) prevented ATP-induced differentiation of HL-60 cells as assessed by fMLP-induced beta-glucuronidase release. However, Rp-CPT-cAMPS did not prevent ATP-induced growth suppression. Taken together, the data indicate that extracellular ATP suppresses HL-60 growth and induces their differentiation by distinct mechanisms. Growth suppression arises from adenosine generation and consequent pyrimidine starvation. Differentiation arises, at least in part, from a distinct mechanism involving the activation of cell surface P2 receptors coupled to cAMP generation and activation of protein kinase A.
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PMID:Extracellular ATP-dependent suppression of proliferation and induction of differentiation of human HL-60 leukemia cells by distinct mechanisms. 1107 40

Pigmented (PE) and nonpigmented (NPE) ciliary epithelial cells comprise the ciliary epithelium, the site of aqueous humor formation in the eye. In man, catecholamines increase the rate of aqueous humor formation, but the mechanism underlying these effects is not understood. Recent evidence suggests that Na-K-Cl cotransport plays a central role in blood-to-aqueous chloride transport across ciliary epithelium in cow and rabbit. We therefore investigated whether catecholamines stimulate Na-K-Cl cotransport in human PE cells. Na-K-Cl cotransporter protein was detected as a 170 kDa protein band on immunoblots. Immunofluorescence microscopy detected cotransporter on the basolateral membranes of the PE layer of ciliary epithelium from a human donor. Cotransporter immunofluorescence was also detected in cultured PE cells. Na-K-Cl cotransport activity measured as ouabain-insensitive bumetanide-sensitive(86)Rb uptake was stimulated by isoproterenol 1.6-fold, with an EC(50) = 28 n M and maximal stimulation at 1 microM. Other transport mechanisms involved in(86)Rb uptake were not affected. Stimulation by 1 microM isoproterenol was blocked by 10 n M ICI 118,551, a beta(2)-specific receptor antagonist, whereas the receptor subtype-specific antagonists yohimbine (alpha(2)), prazosin (alpha(1)) and atenolol (beta(1)) were ineffective. Norepinephrine stimulation (EC(50) = 280 n M) was also blocked by ICI 118,551. Dopamine stimulated Na-K-Cl cotransport 1.6-fold with an EC(50) = 14 microM. The dopamine effect could not be blocked by 10 microM SCH 23390, a D1-antagonist, but was abolished by ICI 118,551. Forskolin and CPT-cAMP stimulated Na-K-Cl cotransport 1.79- and 1.71-fold, respectively, whereas the inactive forskolin analogue 1,9-dideoxyforskolin had no effect. However, high concentrations of the PKA inhibitors PKI amide 14-22 and KT 5720 were needed to inhibit both PKA activity in cell lysates and isoproterenol stimulation of cotransport. This finding may indicate the presence of a novel PKA isoform in PE cells. Inhibitors of other protein kinases, including myosin light chain kinase, protein kinase G, calmodulin-dependent kinase and tyrosine kinase, were without effect on stimulated Na-K-Cl cotransport. When EC(50)s for catecholaminergic stimulations of Na-K-Cl cotransport in PE were compared to those in NPE, values within five-fold of one another were seen for isoproterenol and norepinephrine. In contrast, dopamine was 28-fold more potent in NPE than in PE. The data suggest that both PE and NPE possess beta(2)adrenergic receptors, but only NPE cells possess dopamine D1 receptors linked to Na-K-Cl cotransport.
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PMID:Catecholaminergic regulation of Na-K-Cl cotransport in pigmented ciliary epithelium: differences between PE and NPE. 1113 77

Failing cardiac hypertrophy is associated with an inadequate sarcoplasmic reticulum (SR) function. The hypothesis was examined that pressure overloaded hearts fail to increase SR Ca(2+) uptake rate proportionally to the hypertrophy and that carnitine palmitoyltransferase-1 inhibition by etomoxir ((+/-)-ethyl 2[6(4-chlorophenoxy)hexyl] oxirane-2-carboxylate) can counteract this process. Severe left ventricular pressure overload was induced in rats by constricting the ascending aorta for 8, 10, 14 and 28 weeks leading to cardiac hypertrophy (+62 - +103% of sham-operated rats) and pulmonary congestion. Homogenate oxalate-facilitated SR Ca(2+) uptake rate g wet wt(-1) was reduced (P<0.05) by 29.9+/-1.8% irrespective of phospholamban phosphorylation (in the presence of catalytic subunit of protein kinase A) and inhibition of SR Ca(2+) release channel by ruthenium red. SERCA2 protein level was reduced (P<0.05) by 30.4+/-0.8%. SR Ca(2+) uptake rate was inversely correlated (P<0.05) with left ventricular weight but was not affected by the occurrence of pulmonary congestion. Because SR Ca(2+) uptake rate of whole ventricles was not reduced, a hypertrophy proportional dilution of SR Ca(2+) uptake has to be inferred which precedes pulmonary congestion. Treatment with etomoxir (15 mg kg body wt(-1) day(-1) for 10 weeks) did not affect left ventricular weight but decreased (P:<0.05) the right ventricular hypertrophy related to pulmonary congestion. In parallel, SR Ca(2+) uptake rate of left ventricle and myosin isozyme V(1) were increased (P<0.05). Etomoxir represents a candidate approach for prevention of heart failure by inducing a hypertrophy proportional increase in SR Ca(2+) uptake rate.
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PMID:Sarcoplasmic reticulum function and carnitine palmitoyltransferase-1 inhibition during progression of heart failure. 1113 55

Effects of cGMP on voltage-gated currents in the somatic membrane of isolated newt olfactory receptor cells were investigated using the whole-cell mode of the patch-clamp technique. Under voltage clamp, membrane depolarization generated time- and voltage-dependent current responses, a transient inward current and a sustained outward current. When cGMP or a membrane permeant analog of cGMP, 8-p-chlorophenylthio-cGMP (CPT-cGMP), was applied to the recorded cell, the amplitude of the transient inward current increased markedly, but that of the sustained outward current did not change significantly. When each current was isolated by pharmacological agents, 0.1 mM CPT-cGMP increased the peak amplitude of a Na(+) current (I(Na)) by approximately 40%, a T-type Ca(2+) current (I(Ca,T)) by approximately 40%, and an L-type Ca(2+)current (I(Ca,L)) by approximately 10%; however it did not change significantly the amplitude of a delayed rectifier K(+) current (I(K)). A selective cGMP-dependent protein kinase inhibitor, KT5823, blocked the enhancement by cGMP of I(Na) and I(Ca,T), suggesting that cGMP increases these currents via cGMP-dependent phosphorylation. Under current-clamp conditions, application of CPT-cGMP lowered the current threshold of action potentials induced by current injection, and increased the maximum spike frequency in response to strong stimuli. We suggest that cGMP may lower the threshold in olfactory perception by decreasing the current threshold to generate spikes, and also prevent the saturation of odor signals by increasing the maximum spike frequency.
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PMID:Modulation by cGMP of the voltage-gated currents in newt olfactory receptor cells. 1124 73

Leptin, a circulating hormone secreted mainly from adipose tissues, is involved in the control of body weight. The plasma concentrations are correlated with body mass index, and are reported to be high in patients with insulin resistance, which is one of the major risk factors for cardiovascular disease. However, the direct effect of leptin on vascular wall cells is not fully understood. In this study, we investigated the effects of leptin on reactive oxygen species (ROS) generation and expression of monocyte chemoattractant protein-1 (MCP-1) in bovine aortic endothelial cells (BAEC). We found that leptin increases ROS generation in BAEC in a dose-dependent manner and that its effects are additive with those of glucose. Rotenone, thenoyltrifluoroacetone (TTFA), carbonyl cyanide m-chlorophenylhydrazone (CCCP), Mn(III)tetrakis (4-benzoic acid) porphyrin (MnTBAP), uncoupling protein-1 (UCP1) HVJ-liposomes, or manganese superoxide dismutase (MnSOD) HVJ-liposomes completely prevented the effect of leptin, suggesting that ROS arise from mitochondrial electron transport. Leptin increased fatty acid oxidation by stimulating the activity of carnitine palmitoyltransferase-1 (CPT-1) and inhibiting that of acetyl-CoA carboxylase (ACC), pace-setting enzymes for fatty acid oxidation and synthesis, respectively. Leptin-induced ROS generation, CPT-1 activation, ACC inhibition, and MCP-1 overproduction were found to be completely prevented by either genistein, a tyrosine kinase inhibitor, H-89, a protein kinase A (PKA) inhibitor, or tetradecylglycidate, a CPT-1 inhibitor. Leptin activated PKA, and the effects of leptin were inhibited by the cAMP antagonist Rp-cAMPS. These results suggest that leptin induces ROS generation by increasing fatty acid oxidation via PKA activation, which may play an important role in the progression of atherosclerosis in insulin-resistant obese diabetic patients.
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PMID:Leptin induces mitochondrial superoxide production and monocyte chemoattractant protein-1 expression in aortic endothelial cells by increasing fatty acid oxidation via protein kinase A. 1134 29

We examined whether mitogen-activated protein (MAP) kinase was activated by stimulation of the cAMP pathway and whether MAP kinase activation was involved in synthesis of PRL and GH in GH(3) cells. Treatment of the cells with a cAMP analog, 8-(4-chlorophenylthio)cAMP (CPT-cAMP), activated MAP kinase and increased PRL at both the protein and messenger RNA levels. The protein and messenger RNA of GH were decreased by the treatment. We constructed the luciferase reporter genes after the promoters of PRL and GH and found the activation of both promoters by the CPT-cAMP treatment. We confirmed that overexpression of the catalytic subunit of cAMP-dependent protein kinase had essentially the same effects on MAP kinase activation and synthesis of PRL and GH as the CPT-cAMP treatment. Furthermore, treatment of the cells with pituitary adenylate cyclase-activating polypeptide 27 activated MAP kinase. The activation of PRL promoter by CPT-cAMP and pituitary adenylate cyclase-activating polypeptide 27 was abolished by pretreatment with PD098059 and H89. Although the increase in PRL and GH secretion by CPT-cAMP was inhibited by H89, PD098059 had no effect on secretion. These results suggest that cAMP-induced MAP kinase activation is essential for PRL gene expression, but not for secretion of PRL and GH.
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PMID:Involvement of mitogen-activated protein kinase in cyclic adenosine 3',5'-monophosphate-induced hormone gene expression in rat pituitary GH(3) cells. 1141

1. The spasmolytic and anti-spasmogenic activity of beta-adrenoceptor agonists on airways smooth muscle is thought to involve activation of the cyclic AMP/cyclic AMP-dependent protein kinase (PKA) cascade. Here we have tested the hypothesis that PKA mediates the anti-spasmogenic activity of isoprenaline and other cyclic AMP-elevating agents in guinea-pig isolated trachea by utilizing a number of cell permeant cyclic AMP analogues that act as competitive 'antagonists' of PKA. 2. Anion-exchange chromatography of guinea-pig tracheae resolved two peaks of PKA activity that corresponded to the type I ( approximately 5%) and type II ( approximately 93%) isoenzymes. 3. Pre-treatment of tracheae with zardaverine (30 microM), vasoactive intestinal peptide (VIP) (1 microM) and the non-selective activator of PKA, Sp-8-CPT-cAMPS (10 microM), produced a non-parallel rightwards shift in the concentration-response curves that described acetylcholine (ACh)-induced tension generation. The type II-selective PKA inhibitor, Rp-8-CPT-cAMPS (300 microM), abolished this effect. 4. Pre-treatment of tracheae with Sp-8-Br-PET-cGMPS (30 microM) produced a non-parallel rightwards shift of the concentration-response curves that described ACh-induced tension generation. The selective cyclic GMP-dependent protein kinase (PKG) inhibitor, Rp-8-pCPT-cGMPS (300 microM), abolished this effect. 5. Pre-treatment of tracheae with isoprenaline (1 microM) produced a 10 fold shift to the right of the ACh concentration-response curve by a mechanism that was unaffected by Rp-8-Br-cAMPS (300 microM, selective inhibitor of type I PKA), Rp-8-CPT-cAMPS (300 microM) and Rp-8-pCPT-cGMPS (300 microM). 6. We conclude that the anti-spasmogenic activity of Sp-8-CPT-cAMPS, zardaverine and VIP in guinea-pig trachea is attributable to activation of the cyclic AMP/PKA cascade whereas isoprenaline suppresses ACh-induced contractions by a mechanism(s) that is independent of PKA and PKG.
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PMID:Evidence that the anti-spasmogenic effect of the beta-adrenoceptor agonist, isoprenaline, on guinea-pig trachealis is not mediated by cyclic AMP-dependent protein kinase. 1149 3

1. Urocortin is an endogenous vasodilator although the mechanism of vasorelaxation is not completely understood. The hypothesis that an alteration of smooth muscle calcium concentration is involved was tested using isometric tension recording and calcium fluorimetry. The relationship between contraction and intracellular calcium was also estimated. 2. Urocortin produced a concentration dependent relaxation (pD(2) 8.59+/-0.06, n=6) of vessels pre-contracted with a physiological salt solution containing 42 mM KCl (42 mM K-PSS). 3. Removal of the endothelium did not alter the effect of urocortin, pD(2) was 8.49+/-0.11, n=5. 4. Corticotropin-releasing factor relaxed 42 mM K-PSS pre-contracted vessels with less potency compared to urocortin (pD(2) 6.99+/-0.28, n=5). 5. Urocortin at 100 nM relaxed vessels pre-contracted with 42 mM K-PSS by 59.6+/-4.6% (n=8) and vessels pre-contracted with 500 nM noradrenaline by 25.2+/-6.8% (n=6). Both effects were not accompanied by a change in the intracellular calcium concentration. 6. Urocortin at 100 nM produced a significant rightward shift of 0.33+/-0.07 units of normalized intracellular calcium (n=5) of the relationship between tension and intracellular calcium. 7. The urocortin-induced relaxation was considerably reduced in the presence of 0.3 mM Rp-8-CPT-cAMPS, a cyclic AMP-dependent protein kinase (PKA) inhibitor. 8. The PKA-activator Sp-5,6-DCl-cBIMPS relaxed 42 mM K-PSS pre-contracted vessels (pD(2) 4.98+/-0.07, n=6). Sp-5,6-DCl-cBIMPS at 0.1 mM relaxed vessels by 85.3+/-2.5% (n=5), but did not change the intracellular calcium concentration. 9. In conclusion, the data show that urocortin is a potent, endothelium-independent dilator of rat tail arteries and suggest that this effect is mediated by PKA causing a reduction of the sensitivity of the contractile apparatus for calcium.
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PMID:Urocortin relaxes rat tail arteries by a PKA-mediated reduction of the sensitivity of the contractile apparatus for calcium. 1172 64

The UT-A1 urea transporter plays an important role in maintaining the hyperosmolar milieu of the inner medulla. Vasopressin increases urea permeability in rat terminal inner medullary collecting ducts (IMCDs) within 5-10 min. To elucidate the mechanism, IMCD suspensions were radiolabeled with [(32)P]orthophosphate. UT-A1 was immunoprecipitated and analyzed by autoradiogram and Western blot. Both the 97- and 117-kDa UT-A1 proteins were phosphorylated. Vasopressin treatment increased the phosphorylation of both UT-A1 proteins at 2 min, which peaked at 5-10 min and remained elevated for up to 30 min. There was a discernable increase in UT-A1 phosphorylation with 10 pM and a 50% increase with 10-100 nM vasopressin. 1-Desamino-8-D-arginine vasopressin (dDAVP) or 8-(4-chlorophenylthio)-cAMP (CPT-cAMP) also increased UT-A1 phosphorylation. The vasopressin-stimulated increase in UT-A1 phosphorylation was blocked by H-89 or a specific peptide inhibitor of protein kinase A. Phosphatase inhibitors (okadaic acid, calyculin) increased UT-A1 phosphorylation. We conclude that vasopressin increases UT-A1 phosphorylation via protein kinase A within 2-5 min in rat IMCDs. This suggests that phosphorylation of UT-A1 may be the mechanism by which vasopressin rapidly increases urea permeability in vivo.
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PMID:Vasopressin rapidly increases phosphorylation of UT-A1 urea transporter in rat IMCDs through PKA. 1173 16

Acute promyelocytic leukemia (APL) is characterized by the specific chromosome translocation t(15;17) with promyelocytic leukemia-retinoic acid receptor-alpha (PML-RARA) fusion gene and the ability to undergo terminal differentiation as an effect of all-trans retinoic acid (ATRA). Recently, arsenic trioxide (As(2)O(3)) has been identified as an alternative therapy in patients with both ATRA-sensitive and ATRA-resistant APL. At the cellular level, As(2)O(3) triggers apoptosis and a partial differentiation of APL cells in a dose-dependent manner; both effects are observed in vivo among patients with APL and APL animal models. To further explore the mechanism of As(2)O(3)-induced differentiation, the combined effects of arsenic and a number of other differentiation inducers on APL cell lines (NB4 and NB4-R1) and some fresh APL cells were examined. The data show that a strong synergy exists between a low concentration of As(2)O(3) (0.25 microM) and the cyclic adenosine monophosphate (cAMP) analogue, 8-CPT-cAMP, in fully inducing differentiation of NB4, NB4-R1, and fresh APL cells. Furthermore, cAMP facilitated the degradation of As(2)O(3)-mediated fusion protein PML-RARalpha, a process considered to play a key role in overcoming the differentiation arrest of APL cells. On the other hand, cAMP could significantly inhibit cell growth by modulating several major players in G(1)/S transition regulation. Interestingly, H89, an antagonist of protein kinase A, could block the differentiation-inducing effect of As(2)O(3) potentiated by cAMP. These results thus support the existence of a novel signaling cross-talk for APL maturation, which may deepen understanding of As(2)O(3)-induced differentiation in vivo, and thus furnish insights for new therapeutic strategies.
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PMID:Synergic effects of arsenic trioxide and cAMP during acute promyelocytic leukemia cell maturation subtends a novel signaling cross-talk. 1180 7


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