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PubMed1991

Heterogeneity of endothelium-dependent and endothelium-independent responses to aggregating platelets in porcine pulmonary arteries.

Experiments were designed to determine the endothelium-dependent and endothelium-independent responses to aggregating platelets in porcine pulmonary arteries. Isolated rings with and without endothelium from large (5-7-mm-diameter) and small (2-3-mm-diameter) pulmonary arteries were suspended in modified Krebs-Ringer bicarbonate solution bubbled with 95% O2-5% CO2 in the presence of indomethacin. Aggregating platelets caused relaxations in rings with endothelium but contractions in rings without endothelium, both of which were significantly larger in small versus large pulmonary artery rings. Serotonin and ADP caused concentration-dependent endothelium-augmented relaxations that were unaffected by ketanserin. Methiothepin, but not apyrase, significantly decreased the platelet-induced endothelium-dependent relaxations; the residual relaxation was abolished when rings were incubated with methiothepin, apyrase, and theophylline but was unaffected if apyrase was absent, indicating that ADP is responsible for the residual relaxation caused by aggregating platelets. Quiescent rings, with and without endothelium, contracted in a dose-dependent manner to norepinephrine and histamine but not to serotonin or vasopressin. The contraction to aggregating platelets was blocked by methiothepin, pyrilamine, and diphenhydramine but was unaffected by phentolamine, ketanserin, or incubation of the platelets with dazoxiben. These data indicate that, in large and small porcine pulmonary arteries, serotonin and ADP are the major contributors to the endothelium-dependent relaxation caused by aggregating platelets, while histamine appears to be responsible for the contraction that platelets cause in rings without endothelium.

Adenosine Diphosphate↗
PubMed1996

Effects of the novel water-soluble calcium antagonist (+/-)-3-(4-allyl-1-piperazinyl)-2,2-dimethylpropyl methyl 1,4-dihydro-2,6-dimethyl-4-(3-nitrophenyl)-3,5-pyridinedicarboxylate dihydrochloride on the endothelium-independent and endothelium-dependent contraction in isolated canine cerebral arteries.

The inhibitory effects of NKY-722 (+/-)-3-(4-allyl-1-piperazinyl)-2,2-dimethylpropyl methyl 1,4-dihydro-2,6-dimethyl-4-(3-nitrophenyl)-3,5-pyridinedicarboxylate dihydrochloride, CAS 117241-46-0) on endothelium-independent and endothelium-dependent contractions were examined in comparison with nicardipine in isolated canine cerebral arteries. NKY-722 relaxed the cerebral arteries contracted endothelium-independently by KCl, prostaglandin F2 alpha U-46619 (a thromboxane A2 agonist) and endothelin-1 with IC50 = 2.5, 3.4, 2.8 and 3.6 x 10(-10) mol/l, respectively. On the basis of IC50 values, NKY-722 was about 2 times more effective than nicardipine. Pretreatment of NKY-722 and nicardipine inhibited the endothelium-independent contraction induced by KCl and serotonin to a similar degree. NKY-722 attenuated the endothelium-dependent contractions caused by acetylcholine (ACh), adenosine diphosphate and KO2 with IC50 = 1.7, 3.8 and 1.4 x 10(-9) mol/l, respectively. NKY-722 was about 2 times less effective than nicardipine. NKY-722 and nicardipine inhibited dose-dependently the endothelium-dependent contractions induced by hemolysate. NKY-722 was nearly equipotent to nicardipine on the phasic contraction and slightly more potent on the tonic contraction. The inhibitory effect of NKY-722 on the endothelium-independent contraction induced by KCl and the endothelium-dependent contraction induced by ACh were sustained for a longer time than that of nicardipine after repetitive wash-out. In conclusion, NKY-722 inhibited effectively the endothelium-independent and endothelium-dependent contractions in the cerebral arteries. The effect of NKY-722 was similar to, but longer-lasting than that of nicardipine.

Acetylcholine↗
PubMedDate not supplied

Twenty-five years since the discovery of endothelium-derived relaxing factor (EDRF): does a dysfunctional endothelium contribute to the development of type 2 diabetes?

Twenty-five years ago, the discovery of endothelium-derived relaxing factor opened a door that revealed a new and exciting role for the endothelium in the regulation of blood flow and led to the discovery that nitric oxide (NO) multi-tasked as a novel cell-signalling molecule. During the next 25 years, our understanding of both the importance of the endothelium as well as NO has greatly expanded. No longer simply a barrier between the blood and vascular smooth muscle, the endothelium is now recognized as a complex tissue with heterogeneous properties. The endothelium is the source of not only NO but also numerous vasoactive molecules and signalling pathways, some of which are still not fully characterized such as the putative endothelium-derived relaxing factor. Dysfunction of the endothelium is a key risk factor for the development of macro- and microvascular disease and, by coincidence, the discovery that NO was generated in the endothelium corresponds approximately in time with the increased incidence of type 2 diabetes. Primarily linked to dietary and lifestyle changes, we are now facing a global pandemic of type 2 diabetes. Characterized by insulin resistance and hyperglycaemia, type 2 diabetes is increasingly being diagnosed in adolescents as well as children. Is there a link between dietary-related hyperglycaemic insults to the endothelium, blood flow changes, and the development of insulin resistance? This review explores the evidence for and against this hypothesis.

Animals↗
PubMed1997

Effect of chronic treatment with 17beta-estradiol and progesterone on endothelium-dependent and endothelium-independent relaxation in isolated aortic rings from ovariectomized rats.

OBJECTIVE: Our purpose was to study the influence of chronic treatment with sex hormones on endothelium-dependent and endothelium-independent relaxation of rat aortic rings. STUDY DESIGN: Rings of aortas, with and without endothelium, from rats treated with sex hormones or vehicle for 10 days were mounted in organ baths for isometric tension recording. Indomethacin (10(-5) mol/L) and N(omega)-nitro-L-arginine methyl ester (10(-4) mol/L), alone or in combination, were used to block cyclooxygenase and nitric oxide synthase, respectively. Mean data of contraction induced by potassium chloride (60 mmol/L), the relaxation by acetylcholine (10(-6) mol/L) in potassium chloride-contracted rings, tension induced by phenylephrine, and the negative logarithm of the concentration of acetylcholine or 3-morpholinosydnonimine producing a 50% relaxation, and area under the curve were calculated. RESULTS: Treatment with 17beta-estradiol (10 microg/rat/day) decreased the tension induced by 60 mmol/L potassium chloride and increased the relaxation by acetylcholine in the rings with endothelium precontracted with potassium chloride. Contraction induced by potassium chloride and relaxation induced by acetylcholine were not influenced by the treatment with progesterone (2 mg/rat/day) or estrogen-progesterone combination. Treatment with estradiol, progesterone, or both hormones had no effect on tension developed in intact rings in response to phenylephrine and did not influence endothelium-dependent relaxation to acetylcholine or endothelium-independent relaxation to 3-morpholinosydnonimine in rings contracted with phenylephrine. The inhibition by N(omega)-nitro-L-arginine methyl ester of endothelium-dependent relaxation by acetylcholine was attenuated after the treatment with the sex hormones. CONCLUSIONS: Chronic treatment with sex hormones did not increase production or release of endothelium-derived relaxing factor and did not change the sensitivity of rat aortic smooth muscle to nitric oxide. The treatment slightly counteracted the inhibition of endothelium-dependent relaxation produced by nitric oxide synthase blocker.

Acetylcholine↗
PubMed1994

Overnight storage of the porcine isolated splenic artery enhances endothelium-dependent contractions to NG-nitro-L-arginine methyl ester without impairing endothelium-dependent dilator function.

This study examined the influence of overnight storage on endothelium-independent contractions to 5-hydroxytryptamine (5-HT), endothelium-dependent contractions to NG-nitro-L-arginine methyl ester (L-NAME), and endothelium-dependent relaxations to substance P (SP) and L-arginine, using the porcine isolated splenic artery. In endothelium-intact (E+) segments from fresh porcine isolated splenic arteries or segments from the same vessels stored overnight at 4 degrees C, either in Krebs-Henseleit saline or in Krebs-Henseleit saline containing 1 mM L-arginine, 5-HT caused concentration-related contractions that were similar under all three conditions. Overnight storage enhanced contractions of the splenic artery to L-NAME, an effect not observed if the vessels were co-stored with 1 mM L-arginine. L-NAME failed to contract endothelium-denuded (E-) segments from fresh tissues or tissues stored overnight, indicating that its constrictor effects were endothelium-dependent. SP caused concentration-related, endothelium-dependent relaxations of the splenic artery that were inhibited by 100 microM L-NAME, indicating that the relaxations could be attributed to the stimulated release of NO from endothelial cells. Established contractions to 100 microM L-NAME in E+ segments from fresh tissues, or segments from the same tissues stored overnight at 4 degrees C, either in Krebs-Henseleit saline or in Krebs-Henseleit saline containing 1 mM L-arginine, were all reversed by 1 mM L-arginine to similar extents, indicating that overnight storage did not affect endothelium-dependent dilator responses to L-arginine.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1997

Both endothelium-dependent and endothelium-independent function is impaired in patients with angina pectoris and normal coronary angiograms.

BACKGROUND: The aim of this study was to investigate both endothelium-dependent and endothelium-independent vasodilatation in syndrome X patients. Recently selective impairment of endothelium-dependent function has been reported in a small number of syndrome X patients. However, other investigators have reported impaired endothelium-independent function. METHODS: We infused the endothelium-independent vasodilators papaverine and glyceryl trinitrate, and endothelium-dependent vasodilator acetylcholine in the left coronary artery of 35 patients with syndrome X and in 17 control subjects (atypical chest pain, negative exercise test, and normal coronary angiograms). Coronary blood flow was measured with an intracoronary Doppler catheter positioned in the proximal left anterior descending coronary artery, and the artery diameter was assessed using quantitative coronary angiography. RESULTS: The mean increase in coronary blood flow in response to a 12 mg dose of papaverine was significantly less in the syndrome X group (185 +/- 74% vs 411 +/- 59%, P < 0.001). The increase in coronary blood flow in response to acetylcholine, at doses of 1, 3, 10, and 30 micrograms.min-1, was also significantly lower in the syndrome X group (12 +/- 13 (P < 0.05), 41 +/- 33, 57 +/- 68, and 124 +/- 87% (P < 0.001)) as compared to the control group (76 +/- 49, 214 +/- 116, 355 +/- 115, and 361 +/- 74%). CONCLUSION: These findings demonstrate that both endothelium-dependent and endothelium-independent dilatation of the coronary microvasculature is impaired in syndrome X.

Acetylcholine↗
PubMed1994

Endothelium-derived nitric oxide attenuates neutrophil adhesion to endothelium under arterial flow conditions.

Nitric oxide (NO) synthesized from cultured endothelial cells inhibits platelet aggregation and adhesion to subendothelial extracellular matrix and may contribute to the thromboresistance of the endothelium. NO has also been shown to inhibit neutrophil aggregation and adherence to postcapillary venules. Whether NO derived from the intact endothelium of an arterial wall can influence platelet and neutrophil adhesion under whole-blood arterial flow conditions was evaluated in this study. Porcine aortic segments with intact endothelium were exposed to flowing porcine arterial blood for 5 minutes at a shear rate of 424 sec-1. Pretreatment of the endothelium with the physiological precursor of NO, L-arginine (2 mmol/L), reduced 111In-labeled neutrophil adhesion by 32% from 10.2 +/- 1.6 to 6.9 +/- 1.3 x 10(3)/cm2 (P < .05), relative to control. This effect was reversed by the inhibitor of NO synthesis, N omega-nitro-L-arginine methyl ester (L-NAME, 5 mmol/L) (8.2 +/- 3.0 versus 8.6 +/- 3.2 x 10(3)/cm2 for control; P = NS). Pretreatment of the endothelium with D-arginine (2 mmol/L) did not influence neutrophil adhesion (8.7 +/- 2.0 versus 8.6 +/- 2.0 x 10(3)/cm2 for control; P = NS). The intact endothelium, which is normally thromboresistant, shows a low basal level of 51Cr activity, corresponding to a platelet adhesion less than 0.5 x 10(6)/cm2, and this thromboresistance was not significantly influenced by L-arginine. These results indicate that NO derived from an intact arterial endothelium under whole-blood arterial flow conditions may be an important modulator of neutrophil interaction with the intact endothelium.

Animals↗
PubMed2004

The endothelium in health and disease: a discussion of the contribution of non-nitric oxide endothelium-derived vasoactive mediators to vascular homeostasis in normal vessels and in type II diabetes.

Endothelial dysfunction is considered as a major risk factor of cardiovascular complications of type I and types II diabetes. Impaired endothelium-dependent vasodilatation can be directly linked to a decreased synthesis of the endothelium-derived nitric oxide (NO) and/or an increase in the production of reactive oxygen species such as superoxide. Administration of tetrahydrobiopterin, an important co-factor for the enzyme nitric oxide synthase (NOS), has been demonstrated to enhance NO production in prehypertensive rats, restore endothelium-dependent vasodilatation in coronary arteries following reperfusion injury, aortae from streptozotocin-induced diabetic rats and in patients with hypercholesterolemia. Tetrahydrobiopterin supplementation has been shown to improve endothelium-dependent relaxation in normal individuals, patients with type II diabetes and in smokers. These findings from different animal models as well as in clinical trials lead to the hypothesis that tetrahydrobiopterin, or a precursor thereof, could be a new and an effective therapeutic approach for the improvement of endothelium function in pathophysiological conditions. In addition to NO, the endothelium also produces a variety of other vasoactive factors and a key question is: Is there also a link to changes in the synthesis/action of these other endothelium-derived factors to the cardiovascular complications associated with diabetes? Endothelium-derived hyperpolarizing factor, or EDHF, is thought to be an extremely important vasodilator substance notably in the resistance vasculature. Unfortunately, the nature and, indeed, the very existence of EDHF remains obscure. Potentially there are multiple EDHFs demonstrating vessel selectivity in their actions. However, until now, identity and properties of EDHF that determine the therapeutic potential of manipulating EDHF remains unknown. Here we briefly review the current status of EDHF and the link between EDHF and endothelial dysfunction associated with diabetes.

Animals↗
PubMed1991

Pharmacological evidence that captopril possesses an endothelium-mediated component of vasodilation: effect of sulfhydryl groups on endothelium-derived relaxing factor.

Captopril, an angiotensin-converting enzyme inhibitor, reportedly can scavenge superoxide anion (O2-), a property attributed to its sulfhydryl group. The present investigation, using rabbit aortic rings precontracted with either norepinephrine or clonidine, was designed to determine whether captopril possesses an endothelium-dependent component of vasodilation related to its ability to protect endothelium-derived relaxing factor (EDRF) from superoxide-mediated destruction. Also studied were enalaprilat, a nonsulfhydryl angiotensin-converting enzyme-inhibitor, superoxide dismutase, and the sulfhydryl compounds glutathione (GSH), N-2-mercaptopropionylglycine (MPG) and N-acetylcysteine (NAC). Captopril, but not enalaprilat, caused dose-dependent relaxations in preconstricted aortic rings containing an intact endothelium. Rings denuded of endothelium were unresponsive to any dose of captopril. Captopril's vasodilation was not related to prostaglandin influence but was associated with an increase in cyclic GMP. Superoxide dismutase, GSH, MPG and NAC also produced endothelium-dependent relaxations similar to captopril. It was also demonstrated that endothelium-dependent relaxations to acetylcholine were enhanced by captopril, GSH, MPG and NAC but not by enalaprilat. In another set of experiments, the ability of captopril to inhibit superoxide-mediated inactivation of EDRF was examined. Pyrogallol, a potent generator of O2-, and superoxide dismutase, a scavenger of O2-, were used as a basis for comparing a possible scavenging effect of captopril. In preconstricted rings, pyrogallol elicited endothelium-dependent contractions that were attenuated by both captopril and superoxide dismutase. Similar effects were found with GSH, MPG and NAC but not with enalaprilat. These results suggest that captopril's endothelium-dependent vasodilation is due to its sulfhydryl group and the ability of the latter to scavenge O2-, thereby protecting EDRF.

Acetylcholine↗
PubMed1992

Effects of hydrodynamics and leukocyte-endothelium specificity on leukocyte-endothelium interactions.

In vivo microscopy was used to assess the relative contribution of hydrodynamic forces (network topography and shear rate) and the specificity for leukocytes to interact with venular endothelium as determinants of leukocyte-endothelium interactions. To ascertain this, microvascular networks in the rat and rabbit mesentery were examined under normograde and mechanically induced retrograde flows to determine the effect of reversed flow on leukocyte-endothelium interactions in arterioles and venules. The data indicate that retrograde perfusion under hemodynamic (red blood cell velocity and shear rate) states equivalent to normograde flow significantly increased leukocyte marginating flux in arterioles (from 0 to 0.5 cells/5 sec) and decreased flux significantly in venules (from 1.0 to 0.2 cells/5 sec). The increased flux in arterioles under retrograde conditions, however, was significantly lower than the flux in venules under normograde conditions and the decreased flux in venules during retrograde flow was significantly greater than the flux in arterioles during normograde flow. This apparent discrepancy appears to be the result of a heterogeneous distribution of adhesive receptors on vascular endothelium. Furthermore, marginating leukocytes in arterioles made only brief contact with the endothelium before being swept away while marginating leukocytes in venules during normal and retrograde perfusion rolled along the vascular wall, with similar velocities in both directions. In conclusion, although hydrodynamic forces are important in facilitating leukocyte margination through mechanisms of radial migration, it is leukocyte-endothelium specificity in venules that ultimately determines leukocyte-endothelium interactions.

Animals↗
PubMed2000

SERCA pump isoform expression in endothelium of veins and arteries: every endothelium is not the same.

Endothelium from rat aorta expresses sarco/endoplasmic reticulum Ca2+(SERCA) pump gene SERCA3 where as the smooth muscle expresses SERCA2. This has led to the postulate that vascular endothelium expresses SERCA3. To test this postulate, we examined the SERCA2 and SERCA3 mRNA expression in endothelium and smooth muscle dissected from coronary artery, coronary vein, aorta and vena cava of pig. Smooth muscle from all arteries and veins expressed only the SERCA2 mRNA. Endothelium from coronary artery, coronary vein and aorta expressed both SERCA2 and SERCA3 mRNA but the endothelium from vena cava did not express SERCA3 mRNA although it expressed SERCA2. These observations support the postulate that vascular endothelium expresses SERCA3 but the affirmation is equivocal because vena cava endothelium does not express SERCA3.

Animals↗
PubMed1993

Receptor mechanism of thrombin-induced endothelium-dependent and endothelium-independent coronary vascular effects in dogs.

We previously reported that thrombin produces endothelium-dependent relaxation and endothelium-independent constrictions in canine coronary arteries. To determine whether these opposing vascular effects of thrombin are mediated by the same receptor mechanism, but at different cell types, we investigated the effects of thrombin receptor agonist peptide (TRAP) on isolated canine coronary arteries with and without intact endothelium. In coronary arteries with intact endothelium, addition of 0.01-3.0 microM TRAP, a 14-amino acid residue peptide (SFLLRNPNDKYEPF) homologous to the newly exposed N-terminus after cleavage of the cloned human thrombin receptor, produced rapid, dose-dependent relaxation (Emax = -89.6 +/- 2.3%, n = 26). Threshold concentration was 0.03 microM, and IC50 value was 0.3 microM. Mechanical disruption of the endothelium completely abolished the TRAP-induced relaxation; instead a dose-dependent contraction was observed. Expressed as a percentage of the maximum 70 mM KCl-induced contraction, the maximum contraction observed with 3 microM TRAP was 62.0 +/- 4.1% (n = 32). Pretreatment of endothelium-intact coronary arteries with either 3 microM hemoglobin or 0.25 mM NG-monomethyl-L-arginine (L-NMMA), specific inhibitors of endothelium-derived relaxing factor or nitric oxide (EDRF/NO), also inhibited the relaxation and unmasked the constrictor effect. The pharmacokinetic characteristics of the opposing coronary vascular effects of TRAP are similar to those observed with thrombin, but specific thrombin inhibitors, such as hirudin and D-phenylalanyl-prolyl-L-arginine chloromethyl ketone (PPACK), which inhibit both thrombin relaxant and constrictor effects, had no effect on TRAP-induced responses.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Chloromethyl Ketones↗
PubMed1992

Endothelium-dependent and endothelium-independent effects of adenosine diphosphate in renovascular hypertension.

Norepinephrine-induced responses in isolated perfused mesenteric vascular bed from normotensive and renovascular hypertensive rats were examined in the presence of adenosine diphosphate (ADP, 2 x 10(-6) M). Responses to norepinephrine were significantly greater in vessels from hypertensive rats. Norepinephrine-induced contractions increased after the removal of endothelium. N omega-Nitro-L-arginine (L-NOARG), a potent inhibitor of nitric oxide formation, similarly increased contractions. The greatest responses were obtained, however, after treatment of the vascular segments with methylene blue. The presence of ADP caused significant endothelium-dependent decreases in contractions. Although decreases caused by ADP in vessels with endothelium after treatment with L-NOARG were not statistically significant, a tendency to decreased responses seems to suggest that L-NOARG diminishes but does not completely prevent the effect of ADP in mesenteric vessels. Methylene blue partially reduced the endothelium-dependent ADP-induced relaxant effects in sham-operated nephrectomized rats. A tendency to increased contractions to norepinephrine was observed in the presence of ADP after removal of endothelium. Thus, in the mesenteric resistance arteries of the rat under stimulation by ADP, it appears that nitric oxide released from L-arginine and the activity of soluble guanylate cyclase account only in part for the endothelium-dependent decreased responses to norepinephrine. When nitric oxide formation or soluble guanylate cyclase activity are depressed simultaneously with endothelium damage, ADP released from platelets or red blood cells may be an important factor that acts synergically with vasoconstrictor stimuli.(ABSTRACT TRUNCATED AT 250 WORDS)

Adenosine Diphosphate↗
PubMed1986

Effect of hypercholesterolemia on vascular reactivity in the rabbit. I. Endothelium-dependent and endothelium-independent contractions and relaxations in isolated arteries of control and hypercholesterolemic rabbits.

We studied the effects of hypercholesterolemia on vascular responsiveness in different arteries isolated from rabbits: control groups of rabbits and groups receiving the atherogenic diet consisted of eight animals each. In the arteries, 16 weeks of cholesterol-rich (0.3%) diet evoked intimal lesions which were more pronounced than those noted after 8 weeks of hypercholesterolemia; the aortic arch was affected significantly more by the lesions than the abdominal aorta and the pulmonary artery. Segments of the arteries were mounted in organ chambers for isometric tension recording or for measurement of the endothelium-derived relaxant factor. Contractions caused by acetylcholine and prostaglandin F2 alpha were not altered by the hypercholesterolemia; those evoked by serotonin were moderately augmented only in the aortic arch of hypercholesterolemic rabbits. As the degree of intimal lesion formation increased, the contractions to norepinephrine and clonidine were progressively inhibited. The endothelium-independent relaxations to nitroglycerin were inhibited in only the most severely affected arteries; the endothelium-dependent relaxations to acetylcholine and adenosine triphosphate were progressively inhibited as the degree of fatty streak formation augmented. Thus, in the aortic arch, the relaxations to 3 X 10(-6) M acetylcholine, expressed as percent of the initial contraction, decreased from 86.7 +/- 3.3% in control tissues to 16.3 +/- 8.6% in the 16-week hypercholesterolemic vessels; in the abdominal aortas these relaxations averaged 93.5 +/- 2.2% in control vessels and 72.0 +/- 6.9% in the hypercholesterolemic tissues. The acetylcholine-induced release of endothelium-derived relaxant factor from the abdominal aorta was not significantly affected by the hypercholesterolemia. We conclude from these studies that in arteries obtained from hypercholesterolemic rabbits: the contractions caused by serotonergic mechanisms tend to be augmented, while those to alpha-adrenergic activation are decreased, the endothelium-independent relaxations are modified only in the more severely affected arteries, and the endothelium-dependent relaxations are progressively inhibited as the degree of fatty streak formation augments, probably because a step subsequent to the release of endothelium-derived relaxant factor is altered.

Acetylcholine↗
PubMed1990

Comparative behavior of thrombin and an inactive derivative, FPR-thrombin, toward the rabbit vascular endothelium. Heparin liberates FPR-thrombin from the endothelium in vivo.

Thrombin rapidly binds to and saturates rabbit aorta endothelium in vitro, a process that depends on pericellular glycosaminoglycans and that is inhibited by heparin. To characterize the initial adsorption of thrombin to the endothelium in vivo, an enzymatically inactive derivative, FPR-thrombin (i.e., thrombin inactivated by D-Phe-Pro-Arg-chloromethyl ketone), was prepared. The binding characteristics of thrombin and FPR-thrombin to heparin-Sepharose and to the endothelial surface of rabbit aorta segments in vitro were compared. From these experiments, we concluded that FPR-thrombin mimicked, qualitatively, the binding of thrombin to the endothelium. When injected intravenously, 125I-FPR-thrombin was removed rapidly from the rabbit circulation (T1/2, approximately 1.4 minutes) and simultaneously was adsorbed by the vascular endothelium, particularly in the lung. By injecting heparin (1,000 units/kg i.v.) before 125I-FPR-thrombin, adsorption by the aorta endothelium at 30 minutes after injection was reduced by 90%, and T1/2 was increased to approximately 3.4 minutes. Heparin, administered at various times after 125I-FPR-thrombin, liberated a significant proportion of 125I-FPR-thrombin from the endothelial surface into the plasma compartment as shown by a pronounced "spike" on the plasma curve, a concomitant loss of radioactivity from the lung and from the aorta endothelium, and analysis of the radioactive components of plasma taken before and after heparin injection. Thus, FPR-thrombin was cleared rapidly from the circulation, and endothelium-bound FPR-thrombin was released into the circulation by heparin.

Amino Acid Chloromethyl Ketones↗
PubMed1991

Endothelium-independent and endothelium-dependent contractions mediated by P2X- and P2Y-purinoceptors in canine basilar arteries.

Both alpha, beta-methylene ATP and beta, gamma-methylene ATP (P2X selective agonists) were shown to induce transient contraction in intact and endothelium-removed preparations of canine basilar arteries. 2-Methylthio ATP (a P2Y selective agonist) caused transient contraction of intact arteries and this response was nearly abolished by removal of the endothelium. In the presence of alpha, beta-methylene ATP (10(-6) M), the endothelium-independent contractions induced by alpha, beta-methylene ATP itself (10(-6) M) and by beta, gamma-methylene ATP (10(-5) M) were both abolished. The endothelium-dependent contraction induced by 2-methylthio ATP (10(-7) M) was not attenuated by alpha, beta-methylene ATP. The contraction induced by 2-methylthio ATP (10(-7) M) was attenuated markedly by reactive blue 2 (a P2Y antagonist) (3 x 10(-6) M), aspirin (5 x 10(-5) M), OKY-046 (thromboxane A2 synthetase inhibitor) (10(-5) M) and ONO-3708 (thromboxane A2 antagonist) (10(-8) M). However, these agents did not affect the endothelium-independent contraction induced by alpha, beta-methylene ATP (10(-6) M). Neither TMK-777 (a 5-lipoxygenase inhibitor) (10(-7) M) nor superoxide dismutase (100 U/ml) plus catalase (1,000 U/ml) affected either contraction. The present experiments demonstrate that P2X-purinoceptors mediate endothelium-dependent contraction in the canine basilar artery, and that the endothelium-derived contracting factor in this system is probably thromboxane A2.

Adenosine Triphosphate↗
PubMed1990

Fluoride produces endothelium-dependent relaxation and endothelium-independent contraction in coronary artery.

NaF produced endothelium-dependent relaxation and endothelium-independent contraction in porcine, bovine, canine and human coronary artery rings precontracted with either KCl or prostaglandin F2 alpha. For practical reasons the porcine coronary artery was selected to investigate the mechanisms responsible for these responses. Methylene blue, indomethacin, N-ethylmaleimide, pertussis toxin and cholera toxin all significantly attenuated the endothelium-dependent relaxation caused by fluoride. Pretreatment with deferoxamine had no effect on relaxation and superoxide dismutase/catalase potentiated the relaxation produced by fluoride. Fluoride also contracted vessels with or without the endothelium to equal tension levels and had no apparent relaxing effect on basal tone. The contraction produced by fluoride was significantly attenuated by pertussis toxin and cholera toxin; however, none of the other agents examined significantly altered contraction. Bradykinin also caused endothelium-dependent relaxation and this response was significantly attenuated by methylene blue but not indomethacin. Therefore, fluoride appears to relax the arteries by releasing an endothelium-derived relaxing factor similar to that released by bradykinin (methylene blue sensitive) and one or more prostanoid type endothelium-derived relaxing factor(s) (indomethacin sensitive). Furthermore, fluoride relaxation and contraction may be guanine nucleotide-binding regulatory protein-mediated based on sensitivity to the guanine nucleotide-binding regulatory protein modulators.

Animals↗
PubMed1988

Pharmacological evidence that endothelium-derived relaxing factor is nitric oxide: use of pyrogallol and superoxide dismutase to study endothelium-dependent and nitric oxide-elicited vascular smooth muscle relaxation.

The principal objective of this study was to elucidate the influence of superoxide anion on both endothelium-dependent arterial relaxation elicited by acetylcholine and endothelium-independent arterial relaxation produced by nitric oxide (NO). Pyrogallol was used to generate superoxide in the oxygenated bathing medium, and superoxide dismutase was used to scavenge superoxide. Pyrogallol caused endothelium-dependent contractions of bovine intrapulmonary arterial and venous smooth muscle after precontraction of muscle by phenylephrine. Acetylcholine- and NO-elicited arterial relaxations were promptly converted to marked contractions upon addition of pyrogallol. Moreover, pyrogallol markedly inhibited the development of arterial relaxant responses to acetylcholine and NO. However, isoproterenol- and glyceryl trinitrate-elicited arterial relaxations were unaffected by pyrogallol. Both pyrogallol and oxyhemoglobin enhanced arterial contractile responsiveness to phenylephrine in an endothelium-dependent manner, whereas indomethacin was without effect. Similarly, both pyrogallol and oxyhemoglobin inhibited acetylcholine- and NO-elicited arterial cyclic GMP accumulation, whereas indomethacin was without effect. Uncontracted arterial rings maintained under tension showed endothelium-dependent contraction and decreased cyclic GMP levels in response to oxyhemoglobin but not pyrogallol. Superoxide dismutase enhanced arterial relaxation and cyclic GMP accumulation in response to both acetylcholine and NO. Using a bioassay superfusion cascade system in which intact perfused artery was the source of endothelium-derived relaxing factor (EDRF) and three endothelium-denuded arterial strips mounted in series served as the detector of EDRF, superfusion of strips with pyrogallol blocked relaxation caused by perfusion of artery with acetylcholine. Superoxide dismutase enhance the relaxations produced by arterial perfusion with acetylcholine and prevented the effects of pyrogallol.

Animals↗