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J L Gordon

Publications and source records attributed to J L Gordon.

At least 55 records · Page 3Linked to original sources
PubMed1983

Stimulation of prostaglandin production through purinoceptors on cultured porcine endothelial cells.

ATP (approx. 1-300 microM) induces dose-dependent prostacyclin secretion from perfused columns of microcarrier beads with cultured endothelial monolayers attached. The response is transient, shows little tachyphylaxis and can reach approx. 100 times control values. 2-Methylthio-ATP is more potent, ADP slightly less potent, AMP much less potent and adenosine is ineffective. These results are consistent with the presence of a purinoceptor on endothelium linked to the prostacyclin synthetic pathway.

Adenine Nucleotides↗
PubMed1983

Differential calcium dependence of contractile responses and 86Rb efflux from the rabbit aorta induced by vasoactive stimuli.

86Rb was used to monitor potassium movements in strips of rabbit aorta simultaneously with measurements of tension. Histamine, noradrenaline, the prostaglandin endoperoxide analogue U46619, angiotensin II, and 144 mM K+ each induced an increase in 86Rb efflux concomitantly with contraction. For the first four agonists there was a rank-order correlation between the contractile response and 86Rb efflux, but 144 mM K+ induced a massive increase in 86Rb efflux although it was the weakest contractile stimulus. Contraction and increase in 86Rb efflux-induced K+ were both reduced by verapamil, which blocks voltage-sensitive calcium channels, implying that both effects of K+ were mediated mainly by a depolarisation-induced influx of calcium. Noradrenaline increased both tension and 86Rb efflux through an action on alpha-adrenoceptors, but its effect on efflux, unlike its effect on tension, was apparently totally dependent on the presence of extracellular calcium. Experiments performed in the presence of lanthanum, which blocks calcium influx, showed that the intracellular store of calcium released by noradrenaline apparently played no role in inducing 86Rb efflux, although it could trigger contraction. Lanthanum also blocked contraction induced by K+ but had less effect on the increase in 86Rb efflux induced by K+. Thus, agonist-induced vascular contraction and 86Rb efflux can be dissociated, but under normal conditions all the contractile stimuli tested induced 86Rb efflux.

15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5↗
PubMed1983

Spontaneous and agonist-induced 86Rb efflux from rabbit aortic smooth muscle cells in culture: a comparison with fresh tissue.

Potassium efflux was measured in rabbit aortic strips and smooth muscle cells cultured from them by monitoring the release of isotope from preparations preloaded with 86Rb. The basal rate of 86Rb efflux from rabbit subcultured aortic smooth muscle cells was eightfold higher than from freshly isolated strips, but calculations of reuptake of isotope in the tissue indicated that the measured rate constant for efflux from aortic strips underestimated the true rate by about fourfold. The rate constant for efflux from freshly dispersed cells was less than half that of subcultured cells and remained unchanged for 5 days in culture. It then rose and by around day 10 had reached the value for subcultured cells. The increase in efflux coincided with the onset of cell division. The increased rate of efflux was accompanied by an increased rate of uptake so that the internal potassium content of the cells remained constant. Heparin decreased the efflux of 86Rb from subcultured cells to that of freshly isolated cells concomitant with a reduction in the rate of proliferation. The onset of cell division and increased basal efflux of potassium was associated with a loss of responsiveness to noradrenaline and histamine as assessed by monitoring 86Rb efflux, although depolarising solutions of potassium chloride were still able to elicit a response. Responsiveness to noradrenaline and histamine could be restored by the addition of heparin. These results suggest that the lack of responsiveness of subcultured cells is not due to irreversible dedifferentiation but to a reversible loss in proliferating cells of receptors for vasoactive agents or of a coupling mechanism between receptor occupation and ion gating.

Animals↗
PubMed1983

Regulation of prostaglandin production and ectoenzyme activities in cultured aortic endothelial cells.

Prostaglandin production, angiotensin-converting enzyme, and 5'-nucleotidase were measured in porcine aortic endothelial cells in situ (with a multi-well template on an opened aorta), in primary culture and in subcultures. Changes during culture were monitored and the effects of culture conditions were investigated by growing cells on a biological matrix or on plastic, by adding different sera to the growth medium, and by harvesting cells enzymically or mechanically. Prostacyclin production by endothelium in primary culture is highest immediately after cell isolation and subsequently declines; this pattern is repeated each time the cells are subcultured. The level at which production stabilises is approximately 200 pg X 10(6) cells-1 X h-1. Detaching cells by physical means stimulates production much more than enzymic dispersion; the type of serum or the presence of a biological matrix does not alter prostaglandin production. The relative amount of prostaglandin E produced increases with time, from approximately 20% of the prostacyclin production shortly after isolation to greater than 100% in subcultured cells. None of the culture conditions that we tested altered this trend. Angiotensin-converting enzyme activity decreases during primary culture, but activity can be sustained by including homologous serum (from whole blood or from platelet-free plasma) in the culture medium. The method of harvesting cells, or the presence of a matrix, did not affect enzyme activity. 5'-Nucleotidase also declines during culture, with a progressive decrease in both Km and Vmax from template to primary culture to subcultures. None of the variations in culture conditions prevented this change. Ecto-adenosine-deaminase activity, not detectable in cultured cells, can be measured in the template. Part of this activity was released by the vascular wall and could be due to plasma diffusing from the interstitial space.

5'-Nucleotidase↗
PubMed1983

Effects of arachidonic acid and its cyclo-oxygenase and lipoxygenase products on lymphatic vessel contractility in vitro.

Lymphatic vessels exhibit rhythmical contractility in vivo and in vitro and this activity appears to regulate lymph flow. A technique for measuring the circular muscle contractions of isolated bovine mesenteric lymphatic vessel segments has been devised and utilized to study the pharmacological properties of these vessels. Non-contracting lymphatic vessels can be induced to contract rhythmically with a variety of mediators, the most potent being a stable PGH2 analogue (compound U46619), and the leukotrienes B4, C4 and D4 (threshold concentrations in the nanomolar range). Prostaglandin F2 alpha, noradrenaline, serotonin and histamine also elicited rhythmical activity but much higher concentrations were required. PGE2 and PGE1 were potent inhibitors of spontaneous contractions or those induced with U46619. In keeping with the diverse pharmacological effects of the metabolites of arachidonic acid, the addition of arachidonate to an isolated lymphatic vessel generated both stimulatory and inhibitory activities. It is concluded that arachidonic acid products (produced in the lymphatic vessel or entering the vessel in lymph draining the tissues) regulate lymph flow through their effects on lymphatic smooth muscle.

Alprostadil↗
PubMed1983

Endothelium-dependent relaxation of the pig aorta: relationship to stimulation of 86Rb efflux from isolated endothelial cells.

Bradykinin, adenosine triphosphate (ATP) and acetylcholine each relaxed histamine-contracted strips of pig aorta in a dose-dependent manner. These relaxations were abolished when the endothelium was removed. Relaxation induced by ATP was mimicked by adenosine diphosphate (ADP) but adenosine monophosphate (AMP) and adenosine were about 120 times less potent. Relaxation induced by acetylcholine was antagonized by atropine in a competitive manner, and carbachol induced the same degree of relaxation as acetylcholine, but was about 10 times less potent. The calcium ionophore, A23187, also induced a dose-dependent relaxation of pig aortic strips provided the endothelium was present, suggesting that a rise in the level of ionized calcium within the endothelial cells is one means by which vascular smooth muscle relaxation can be triggered. Bradykinin, ATP, ADP, AMP, adenosine and A23187 each induced a dose-dependent increase in 86Rb efflux from preloaded pig aortic endothelial cells. The dose-response curves for stimulation of 86Rb efflux and for endothelium-dependent relaxation were similar for each individual compound. ADP was equipotent with ATP, but AMP and adenosine were about 120 times less potent. Neither acetylcholine nor carbachol, in concentrations that induce endothelium-dependent relaxation, had any effect on 86Rb efflux from isolated aortic endothelial cells. Lanthanum, which blocks calcium influx, abolished the increases in 86Rb efflux induced by bradykinin and ATP, and the calcium ionophore A23187 was the most effective stimulant of 86Rb efflux, suggesting that the potassium transport induced by these agents is calcium-activated. It is concluded that endothelial responses to bradykinin and ATP can be assessed by monitoring 86Rb efflux, which probably reflects a calcium-activated efflux of potassium associated with the endothelium-dependent vascular relaxation induced by these agents. This pathway is apparently not involved in endothelial responses to acetylcholine.

Acetylcholine↗
PubMed1983

Stimulation of endothelial prostacyclin production plays no role in endothelium-dependent relaxation of the pig aorta.

Stimulation of prostacyclin production by pig aortic endothelial cells adhering to microcarrier beads superfused in columns, or 3H release from cells prelabelled with [3H]-arachidonate, was studied in response to a range of agents that induce endothelium-dependent vascular relaxation. Bradykinin, adenosine triphosphate (ATP) and ionophore A23187 each stimulated release of prostacyclin from unlabelled cells and of 3H from prelabelled cells but acetylcholine did not. Bradykinin induced a parallel, dose-dependent increase in 3H release and 86Rb efflux, measured simultaneously from columns of aortic endothelial cells preloaded with 86Rb and [3H]-arachidonate. The rank-order of effectiveness at inducing both 3H and 86Rb release, measured simultaneously from columns of aortic endothelial cells prelabelled with 86Rb and [3H]-arachidonate and challenged with maximal doses of each agonist, was: A23187 greater than bradykinin greater than ATP. The similarity between agonist-induced 3H release (from cells prelabelled with [3H]-arachidonate) and 86Rb efflux indicates that a common mechanism may be responsible, and the effectiveness of ionophore A23187 suggests that a rise in the intracellular level of calcium may be involved. The lack of effect of acetylcholine on release of prostacyclin from unlabelled cells or of 3H from cells prelabelled with [3H]-arachidonate provides further evidence that acetylcholine acts on endothelial cells by a mechanism that does not involve calcium mobilisation. Although bradykinin, ATP and ionophore A23187 each induced release of prostacyclin from aortic endothelial cells, prostacyclin did not relax the pig aorta. Furthermore, endothelium-dependent relaxation was unaffected by pretreating aortic strips with aspirin. It therefore appears that neither prostacyclin nor any other cyclo-oxygenase product mediates endothelium-dependent relaxation of the pig aorta.

Acetylcholine↗
PubMed1982

Effects of isolation and culture on prostaglandin synthesis by porcine aortic endothelial and smooth muscle cells.

Freshly isolated neonatal porcine aortic tissue (smooth muscle with or without endothelium present) produced approximately 30 ng/mg wet tissue of 6-oxo-prostaglandin F1 alpha (the stable hydrolysis product from prostacyclin) and approximately 15 ng/mg of prostaglandin E2, as measured by radioimmunoassay after 24 h incubation in culture medium. Primary cultures of porcine endothelial and smooth muscle cells (isolated by enzymic digestion of aortic tissue) exhibited the same pattern of prostaglandin production, but absolute values were greater than for fresh tissue, particularly in the case of endothelium. Subcultures of endothelium produced smaller amounts of prostaglandins, although the pattern remained similar. In contrast, subcultures of smooth muscle cells produced a greater total amount of prostaglandins than did primary cultures, and the main product was prostaglandin E2. Experiments with [14C] prostaglandin H2 or [14C]arachidonic acid confirmed that aortic tissue, cultured endothelium, and primary cultures or aortic smooth muscle cells synthesized prostacyclin, and demonstrated that subcultured smooth muscle cells enzymically isomerised prostaglandin H2 to prostaglandin E2. Kinetic studies showed that prostaglandin production by cultured vascular cells was transiently increased by subculture or changing the growth medium, and that production per cell declined with increasing cell density. The change in pattern of prostaglandin production during culture was shown to be due to a rapid decline in the rate of prostacyclin production (which apparently began immediately after tissue isolation), together with a more gradual rise in prostaglandin E2 production. These results indicate that the amounts and ratios of prostaglandins produced by vascular endothelial and smooth muscle cells are greatly affected by the conditions used to isolate and culture the cells; vascular cells in vivo may similarly alter their pattern of prostaglandin production in response to local changes in their environment.

Animals↗
PubMed1980

Metabolism of adenine nucleotides by ectoenzymes of vascular endothelial and smooth-muscle cells in culture.

1. Pig aortic endothelial and smooth-muscle cells in culture rapidly catabolize exogenous ATP, ADP or AMP. 2. In both cell types catabolism is due to Mg2+-stimulated ectoenzymes. 3. Inhibition and substrate-specificity studies suggest that both cell types possess three distinct ectonucleotidases, namely nucleoside triphosphatase (EC 3.6.1.15), nucleoside diphosphatase (EC 3.6.1.6) and 5'-nucleotidase (EC 3.1.3.5), as well as nucleoside diphosphate kinase (EC 2.7.4.6). 4. These ectonucleotidase systems could be of importance in the regulation of neurotransmission, blood platelet function and vasodilation.

Acid Anhydride Hydrolases↗
PubMed1980

Effects of vasoactive and inflammatory agents on cyclic AMP levels in W138 fibroblasts, endothelial and vascular smooth muscle cells in culture.

The vasoactive amines noradrenaline, histamine and 5-hydroxytryptamine and the beta-adrenergic agonist, isoproterenol, had no effect on porcine aortic endothelial or smooth muscle cell cyclic AMP, but these amines increased the level of cyclic AMP in W138 fibroblasts. PGE1, PGE2 and PGI2 were also without effect on endothelial or smooth muscle cells, while increasing WI38 cell cyclic AMP. PGF2 alpha and 6-oxo-PGF1 alpha had no effect on all three types of cell. Inhibition of endogenous prostaglandin production by the cells, using aspirin, did not affect the basal levels of cyclic AMP or the subsequent responses to added prostaglandins in all three cell types.

Animals↗
PubMed1980

Regulation of macrophage lysosomal enzyme secretion: role of arachidonate metabolites, divalent cations and cyclic AMP.

We have investigated the role in macrophage lysosomal enzyme release of arachidonate metabolites, extracellular divalent cations and cyclic AMP (cAMP) which modulate secretion in other cell types. Lysosomal enzyme secretion induced by zymosan was accompanied by release of malondialdehyde (MDA), which is derived from arachidonic acid via prostaglandin synthase. Blockade of MDA formation, by aspirin or indomethacin, was associated with only a small inhibitory effect on lysosomal enzyme release by zymosan: arachidonate metabolites thus play only a minor role in mediating macrophage lysosomal enzyme release. Zymosan-induced secretion of lysosomal enzymes from macrophages did not require extracellular magnesium or calcium although release was enhanced by magnesium and inhibited by calcium. These effects may be related to an influence of the ions on phagocytosis. Elevation of intracellular divalent cation concentrations, by ionophore A23187, induced release of lysosomal enzymes but this was a result of cell lysis. Adenylate cyclase stimulants and dibutyryl cAMP produced slight inhibition of zymosan-induced lysosomal enzyme release. Aminophylline and papaverine caused more marked inhibition but their effects may be due to actions independent of phosphodiesterase inhibition. Our data indicate that arachidonate metabolites and cAMP do not play a major role in regulating zymosan-induced enzyme release from macrophages. Extracellular calcium and magnesium may modulate secretion but the role of intracellular divalent cations remains to be established. We conclude that macrophage lysosomal enzyme secretion is controlled by regulatory mechanisms different from those which control similar degranulation processes in other cell types.

Animals↗
PubMed1979

Vascular endothelial and smooth muscle cells in culture selectively release adenine nucleotides.

Endothelial cells in culture can modulate platelet aggregation and vascular tone, in part by producing prostacyclin (PGI2), a powerful vasodilator and inhibitor of platelet aggregation, but also by their ecto-ADPase activity, which initiates the conversion of pro-aggregating ADP to adenosine, a potent vasodilator and platelet inhibitor. We have now demonstrated that cultured aortic endothelial cells exposed to trypsin, thrombin or other stimuli can liberate a high proportion of their adenine nucleotides without substantial loss of lactate dehydrogenase. ADP rapidly accumulates extracellularly, reaching biologically active concentrations before there is further breakdown to adenosine. Whether this selective release of nucleotides is a response to damage, or whether it represents a specific secretory mechanism remains to be resolved. Cultured aortic smooth muscle cells can secrete adenine nucleotides in a similar manner, but extracellular conversion to adenosine occurs much faster.

Adenine Nucleotides↗