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J Vane

Publications and source records attributed to J Vane.

At least 19 recordsLinked to original sources
PubMed1995

Trumping the ACE.

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Angiotensin-Converting Enzyme Inhibitors↗
PubMed1994

Nitroblue tetrazolium inhibits oxidation of glyceryl trinitrate to nitric oxide in bovine aortic smooth muscle cells.

The effects of nitroblue tetrazolium (NBT), a well-known scavenger of superoxide anions and an inhibitor of nicotinamide adenine dinucleotide (NADPH)-dependent oxidations, were assessed on the metabolism of glyceryl trinitrate (GTN) to nitric oxide (NO) by bovine aortic smooth muscle cells (SMC). The extent of this metabolism was determined by measuring NO formed, using the inhibition of thrombin-induced platelet aggregation and relaxation of rabbit aortic strips as bioassay systems. In addition, NO produced from GTN by SMC was measured as nitrite (NO2-), one of its breakdown products. The antiplatelet effect of GTN (44 microM) was potentiated by SMC (0.12-0.46 x 10(5) cells) treated with indomethacin (10 microM) and this was inhibited in a concentration-dependent manner when the cells were pretreated with NBT (100 microM). NBT (3-100 microM) also reduced the formation of NO2- from GTN (600 microM) by SMC (3 x 10(5) cells). Furthermore, relaxations of endothelium-denuded strips of the rabbit aorta by GTN (10(-9)-10(-6) M) were attenuated when the strips were pretreated with NBT (100 or 500 microM). The formation of NO from L-arginine (L-Arg) by SMC was not affected by NBT. The hypotensive responses to GTN (0.25-1 mg/kg, i.v.) in anaesthetized rats were inhibited by pretreatment with NBT (1.25 mg/kg, i.v.) but NBT did not alter the hypotensive responses induced by SIN-1. Thus, NBT inhibited the bioconversion of GTN to NO both in vitro and in vivo. NBT may be a useful pharmacological tool to investigate the enzymic pathway(s) involved in the conversion of GTN to NO by smooth muscle cells or other cells.

Animals↗
PubMed1993

Tetrahydrobiopterin synthesis is induced by LPS in vascular smooth muscle and is rate-limiting for nitric oxide production.

GTPCH1 mRNA and BH4 synthesis is increased by LPS in vascular smooth muscle. Our data suggest that induction of GTPCH1 and NOS represent two arms of a common pathway required for immunostimulant-evoked NO synthesis. This conclusion is consistent with the view that the major function of immunostimulant-evoked BH4 is to support NOS. Moreover, GTPCH1 and other enzymes of the de novo BH4 synthetic pathway may prove to be important targets for therapy of clinical conditions arising from NO overproduction. As we begin to reveal the molecular events governing the induction and expression of GTPCH1 and NOS, additional therapeutic approaches for treating NO overproduction are certain to be revealed.

Amino Acid Oxidoreductases↗
PubMed1993

Conversion of glyceryl trinitrate to nitric oxide in tolerant and non-tolerant smooth muscle and endothelial cells.

1. Exposure of smooth muscle cells (SMC) to glyceryl trinitrate (GTN, 75-600 microM) for 30 min led to a concentration-dependent increase in nitrite (NO2-), one of the breakdown products of nitric oxide (NO). This was not affected by 30 min pretreatment of the cells with 0.5 mM of sulphobromophthalein (SBP) an inhibitor of glutathione-S-transferase (GST), by metyrapone or SKF-525A inhibitors of cytochrome P450. These experiments were confirmed by organ bath studies using rabbit aortic strips denuded of endothelium and contracted with phenylephrine. Thus, a 30 min incubation of the strips with 0.5 mM SPB, metyrapone or SKF-525A did not affect the relaxations in response to GTN (10(-10)-10(-6) M). 2. Potentiation of the anti-platelet effect of GTN (44 microM) by endothelial cells (EC, 40 x 10(3) cells) was not affected by prior incubation of EC with SBP, metyrapone or SKF-525A (all at 0.5 mM). 3. Potentiation of the antiplatelet activity of GTN (11-352 microM) by small numbers of SMC (24 x 10(3) cells) or EC (40 x 10(3) cells) treated with indomethacin (10 microM) was attenuated when the SMC or EC were treated in culture with a high concentration of GTN (600 microM) for 18 h beforehand (referred to as 'tolerant' cells). In addition, tolerant SMC produced far less NO2- than non-tolerant SMC. 4. Exposure of non-tolerant SMC or EC (10(5) cells) to GTN (200 microM) for 3 min increased (3-4 fold) the levels of guanosine 3':5'-cyclic monophosphate (cyclic GMP). This increase was much less (< I fold) in the tolerant SMC or EC (105 cells). The basal levels of cyclic GMP were similar in normal or tolerant SMC or EC. Sodium nitroprusside (80 JAM) or atrial natriuretic factor (ANF, I0- M) increased the levels of cyclic GMP in normal or tolerant SMC or EC to the same extent.5 The anti-platelet effects of GTN (44 JM) were potentiated by the sulphydryl donor N-acetylcysteine(NAC, 0.5mM). Incubation of GTN (150-1200fJM) for 30min with NAC (0.1-1mM) led to aconcentration-dependent increase in N02- formation. The reduced ability of tolerant SMC or EC to potentiate the anti-platelet activity of GTN was restored by NAC (0.5 mM). These anti-aggregatory effects were abolished by concurrent co-incubation with oxyhaemoglobin (10 JM) indicating that they were due to NO release.6 Thus, in SMC or EC, metabolism of GTN to NO does not depend on glutathione-S-transferase or the cytochrome P450 system. Furthermore, when compared to normal cells, tolerant SMC or EC metabolize GTN to NO less effectively as assessed by the reduced capacity to potentiate the antiplatelet effects of GTN, to release NO2- and to increase the level of cyclic GMP. This decrease in NO formation shows that tolerance to GTN is mainly due to impaired biotransformation of GTN to NO. NAC, by directly forming NO from GTN, compensates for this failing mechanism.

Acetylcysteine↗
PubMed1992

The metabolism of glyceryl trinitrate to nitric oxide in the macrophage cell line J774 and its induction by Escherichia coli lipopolysaccharide.

The metabolism of glyceryl trinitrate (GTN) to nitric oxide (NO) was studied in the mouse macrophage cell line J774 and in the human monocytic cell line U937 in the absence or presence of Escherichia coli lipopolysaccharide (LPS). Two bioassay systems were used: inhibition of platelet aggregation and measurement of cGMP after stimulation by NO of guanylate cyclase in J774 cells. In addition, NO produced from GTN by cells or by cellular fractions was measured as nitrite (NO2-) one of its breakdown products. J774 cells (1.25 x 10(5) cells) treated with indomethacin (10 microM) enhanced the platelet inhibitory activity of GTN (22-352 microM) but not that of sodium nitroprusside (4 microM). This effect was abrogated by co-incubation with oxyhaemoglobin (oxyHb, 10 microM) indicating release of NO from GTN. U937 cells (up to 60 x 10(5)) did not metabolize GTN to NO. LPS (0.5 micrograms/mL for 18 hr) enhanced at least 2-fold the capacity of J774 cells but not that of U937 cells to form NO from GTN and this enhancement was attenuated when cycloheximide (10 micrograms/mL) was incubated together with LPS. In the absence of LPS stimulation, cycloheximide had no effect. Furthermore, when incubated with GTN (200 microM), J774 cells treated with LPS released more NO from GTN as indicated by a 3-fold greater increase in their level of cGMP which was prevented by oxyHb (10 microM). Incubation of J774 cells with GTN (75-600 microM) for 30 min led to a concentration-dependent increase in NO2- which was substantially reduced when the cells were boiled. The microsomal fraction was more potent than the cytosol in producing NO2- from GTN (1.2-2.4 mM). Release of NO2- from GTN by J774 cells was not affected by treating the cells with the NO synthase inhibitor, NG-monomethyl-L-arginine (MeArg, 300 microM). In J774 cells made tolerant to GTN, potentiation of the anti-platelet effects of GTN (11-352 microM) and release of NO2- from GTN was reduced. Thus, J774 cells but not U937 cells convert GTN to NO. This enzymic pathway (present mainly in the microsomal fraction of the J774 cells) is induced by LPS and is not regulated by endogenous NO released from L-Arg by the enzyme NO synthase. Furthermore, when compared to normal cells, tolerant J774 cells metabolize GTN to NO less effectively as assessed by a reduced capacity to potentiate the anti-platelet effect of GTN and to release NO2-.(ABSTRACT TRUNCATED AT 400 WORDS)

Amino Acid Oxidoreductases↗
PubMed1992

Metabolism of glyceryl trinitrate to nitric oxide by endothelial cells and smooth muscle cells and its induction by Escherichia coli lipopolysaccharide.

Here, we demonstrate that the metabolism of glyceryl trinitrate (GTN) to nitric oxide (NO) occurs not only in bovine aortic smooth muscle cells (SMCs) but also in endothelial cells (ECs) and that this biotransformation is enhanced by pretreatment with Escherichia coli lipopolysaccharide (LPS). Two bioassay systems were used: inhibition of platelet aggregation and measurement of cGMP after stimulation by NO of guanylate cyclase in SMCs or ECs. In addition, NO produced from GTN by cells was measured as nitrite (NO2-), one of its breakdown products. Indomethacin (10 microM)-treated SMCs or ECs enhanced the platelet inhibitory activity of GTN. This effect was abrogated by coincubation with oxyhemoglobin (oxyHb; 10 microM), indicating release of NO from GTN. LPS (0.5 microgram/ml; 18 h) enhanced at least 2- to 3-fold the capacity of SMCs or ECs to form NO from GTN, and this enhancement was attenuated when cycloheximide (10 micrograms/ml) was incubated together with LPS. Furthermore, when incubated with GTN (200 microM) SMCs or ECs treated with LPS (0.5 microgram/ml; 18 h) released more NO from GTN than nontreated cells as indicated by a much higher (8- to 9-fold) increase in the levels of cGMP. Exposure of SMCs to GTN (600 microM) for 30 min led to an increase in the levels of NO2- dependent on cell numbers, which was enhanced when SMCs were treated with LPS. Incubation of nontreated or LPS-treated cells with NG-monomethyl-L-arginine (300 microM; 60 min) did not influence the metabolism of GTN to NO. SMCs failed to enhance the antiplatelet activity of sodium nitroprusside. Anesthetized rats treated with an intraperitoneal injection of LPS (20 mg/kg) 18 h beforehand showed enhanced hypotensive responses to GTN (0.25-1 mg/kg). These effects were blocked by methylene blue (10 mg/kg) but not by indomethacin (3 mg/kg). LPS did not alter the hypotensive responses induced by phentolamine, verapamil, or SIN-1. Thus, both in vitro and in vivo, LPS induces the enzyme(s) metabolizing GTN to NO.

Animals↗
PubMed1992

Cultured astrocytoma cells generate a nitric oxide-like factor from endogenous L-arginine and glyceryl trinitrate: effect of E. coli lipopolysaccharide.

1. The inhibitory activity of astrocytoma cells (0.25-3 x 10(5)) treated with indomethacin (10 microM) on platelet aggregation was enhanced by incubating the cells with E. coli lipopolysaccharide (LPS, 0.5 micrograms ml-1) for 18 h. This effect was attenuated when cycloheximide (10 micrograms ml-1) was incubated together with LPS. The inhibition of platelet aggregation by cells treated with LPS was potentiated by superoxide dismutase (60 u ml-1) and ablated by oxyhaemoglobin (oxyHb, 10 microM) or NG-monomethyl-L-arginine (L-NMMA, 30-300 microM). The effects of L-NMMA were reversed by co-incubation with L-arginine (L-Arg, 100 microM) but not D-arginine (D-Arg, 100 microM). LPS also increased the levels of nitrite in the culture media and this increase was ablated by co-incubation with L-NMMA (300 microM) or cycloheximide (10 micrograms ml-1). 2. Astrocytoma cells (0.5 x 10(5)) treated with indomethacin (10 microM) enhanced the platelet inhibitory activity of glyceryl trinitrate (GTN, 11-352 microM) but not that of sodium nitroprusside (4 microM). Furthermore, when incubated with GTN (200 microM) a 4 fold increase in the levels of guanosine 3':5'-cyclic monophosphate (cyclic GMP) was observed. These effects were abrogated by co-incubation with oxyHb (10 microM) but not with L-NMMA (300 microM). Treatment of the cells with LPS (0.5 micrograms ml-1) for 18 h did not enhance their capacity to form NO from GTN. 3. Thus, in cultured astrocytoma cells, LPS enhances the formation of nitric oxide from endogenous L-arginine.In addition, these cells can metabolize GTN to nitric oxide but this process is not enhanced by LPS stimulation.

Arginine↗
PubMed1991

Studies on the importance of the proposed release of nitric oxide from platelets.

The importance of the presence of the L-arginine (L-Arg)-nitric oxide (NO) pathway in platelet function was investigated. This was achieved by studying the interactions between anti-platelet agents such as glyceryl trinitrate (GTN) or iloprost (Ilo) and the NO precursor L-Arg. Thrombin or collagen-induced aggregation of human washed platelets was inhibited in a concentration-dependent fashion by a 3 min incubation with GTN (40-200 microM). This effect was reversed by oxyhaemoglobin (oxyHb, 10 microM). GTN at the lowest concentration tested (40 microM) potentiated the anti-aggregatory activity of subthreshold concentrations of Ilo (0.2 nM). GTN did not potentiate the action of L-arginine (100 microM) and L-Arg by itself failed to influence thrombin or collagen-induced platelet aggregation. The platelet responses to thrombin or collagen were not affected by a 3 min or 1 h treatment with the NO synthesis inhibitor NG-monomethyl-L-arginine (MeArg, 300 microM). This treatment did not alter the platelet inhibitory effects of GTN and in the presence of MeArg, exogenously applied L-Arg did not potentiate the anti-platelet activity of GTN. These results indicate that under our experimental conditions human washed platelets do not generate sufficient amounts of NO to modify (1) platelet aggregation or (2) the anti-platelet activity of GTN or iloprost.

Arginine↗
PubMed1991

Impairment of the L-arginine-nitric oxide pathway in mast cells from spontaneously hypertensive rats.

Serosal mast cells (MC) from 6 month old spontaneously hypertensive rats (SHR) were compared to MC from 6 month old Wistar Kyoto rats (WKYR) for their ability to release nitric oxide (NO). The relationship between histamine release and NO-like activity from these cells was also investigated. MC from SHR released less NO-like factor than MC from WKYR as assessed by the use of platelet aggregation and soluble guanylate cyclase activation as bioassays for NO. Sodium nitroprusside elevated the concentrations of cGMP to a similar extent in MC from SHR or WKYR. No changes in the levels of cAMP were observed. The release of histamine from MC induced by compound 48/80 or the calcium ionophore A23187 was greater in MC from SHR than in MC from WKYR. Thus, MC from SHR show a decreased production of NO-like activity which is reflected by a decreased ability to inhibit platelet aggregation. The decreased production of cGMP in the MC leads to an increased stimulated release of histamine.

Animals↗
PubMed1991

Diphenylene iodonium, an inhibitor of free radical formation, inhibits platelet aggregation.

Diphenylene iodonium is an inhibitor of the enzyme NADPH-oxidase and prevents the generation of oxygen-derived free radicals in neutrophils (Cross and Jones, 1986). Here we show that diphenylene iodonium (0.25-2 microM) inhibited, according to the dose, thrombin-induced platelet-aggregation in human washed platelets and ADP-induced platelet aggregation in platelet-rich plasma. At the concentrations which inhibited platelet aggregation diphenylene iodonium did not alter platelet concentrations of cAMP or cGMP but enhanced the anti-platelet activity of iloprost, sodium nitroprusside or cultured endothelial cells. These findings highlight the importance of free radicals as platelet pro-aggregatory agents.

Adenosine Diphosphate↗
PubMed1991

Nitric oxide from vascular smooth muscle cells: regulation of platelet reactivity and smooth muscle cell guanylate cyclase.

1. Incubation of smooth muscle cells (SMC) from bovine aorta for 3 min with human washed platelets treated with indomethacin (10 microM) promoted a cell number-related inhibition of platelet aggregation induced by thrombin (40 mu ml-1). This inhibition was not attributable to products of the cyclo-oxygenase pathway for the SMC were also treated with indomethacin (10 microM). 2. The inhibitory activity of the SMC on platelet aggregation was enhanced by incubating the SMC with E. coli lipopolysaccharide (LPS, 0.5 micrograms ml-1) for a period of 9 to 24 h. This effect was attenuated when cycloheximide (10 micrograms ml-1) was incubated together with LPS. Cycloheximide did not prevent the inhibitory activity of the non-treated cells. 3. The inhibition of platelet aggregation obtained with non-treated or LPS-treated SMC was potentiated by superoxide dismutase (SOD, 60 u ml-1) and ablated by oxyhaemoglobin (OxyHb, 10 microM). Preincubation of the SMC with NG-monomethyl-L-arginine (L-NMMA, 30-300 microM) for 60 min prevented their antiaggregatory activity. This effect was reversed by concurrent incubation with L-arginine (L-Arg, 100 microM) but not with D-arginine (D-Arg, 100 microM). 4. Exposure of the non-treated SMC (5 x 10(5) cells) to stirring (1000 r.p.m., 37 degrees C) for 10 min led to a significant increase in their levels of guanosine 3':5'-cyclic monophosphate (cyclic GMP) but not adenosine 3':5'-cyclic monophosphate (cyclic AMP). L-NMMA (300 microM) attenuated the increase in cyclic GMP induced by stirring but did not affect the basal levels of cyclic GMP in the cells.5. These findings support the idea that non-treated or LPS-treated cultured SMC can produce an NO-like factor. Production by the latter requires protein synthesis as evidenced by blockade with cycloheximide. This NO-like factor may play a role in the auto-regulation of smooth muscle cell reactivity through a cyclic GMP-dependent mechanism.

1-Methyl-3-isobutylxanthine↗