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S Heptinstall

Publications and source records attributed to S Heptinstall.

At least 19 recordsLinked to original sources
PubMed1996

Contact-induced modulation of neutrophil elastase secretion and phagocytic activity by platelets.

It has been reported that platelets stimulate generation of reactive oxygen species in neutrophils and monocytes by a mechanism that requires mutual cell-cell contact and the presence of P-selectin on the platelet surface. In the present study we investigated the effect of platelet-neutrophil contacts on neutrophil elastase secretion and phagocytic activity. Non-activated or thrombin-activated platelets were fixed with formaldehyde, washed and incubated with neutrophils in the absence or presence of various neutrophil agonists. Elastase secretion was determined by measuring the enzyme activity in cell-free supernatants using a chromogenic substrate. Platelet-neutrophil adhesion and ingestion of zymosan particles by neutrophils were quantitated by light microscopy. Platelets significantly reduced elastase secretion from neutrophils but had no effect on the elastase activity in the supernatant of neutrophil lysates. When neutrophils were stimulated with the ionophore A23187 or the chemotactic peptide FMLP, thrombin-activated platelets were more potent to inhibit elastase secretion when compared with non-activated platelets. Neutrophils that were not able to bind platelets to their surface had a significantly lower phagocytic activity when compared with neutrophil with adherent platelets or neutrophils that were incubated in the absence of platelets. The results indicate that platelet-neutrophil contacts may also lead to an inhibition of neutrophil functions and that such inhibition could be due to a transient contact rather than due to a firm platelet-neutrophil adhesion.

Blood Platelets↗
PubMed1996

Changes in the composition of the platelet cytoskeleton in response to ADP: effects of MK-852 and ARL 66096.

Platelet activation by adenosine diphosphate (ADP) results in an alteration in the composition of the cytoskeleton. Here we have determined the effects of MK-852 and ARL 66096 on the cytoskeletal changes that occur. MK-852 is a GPIIb/IIIa antagonist that inhibits aggregation by interfering with fibrinogen binding ARL 66096 is a P2T antagonist that selectively inhibits ADP-induced aggregation. Neither agent inhibits the shape change response. Experiments were performed in hirudinized platelet-rich plasma. Platelet activation led to a significant and sustained increase in the cytoskeletal content of actin binding protein (ABP), myosin, alpha-actinin, a 66K protein and actin, and a significant decrease in a 31K protein. In the presence of MK-852 there was no increase in ABP or the 66K protein and no decrease in the 31K protein. The increase in myosin and alpha-actinin became reversible but there was still incorporation of actin into the cytoskeleton. In the presence of ARL 66096 there was no increase in ABP or the 66K protein and no decrease in the 31K protein. ARL 66096 also prevented incorporation of alpha-actinin and actin. As with MK-852, myosin incorporation became reversible. The results suggest that (1) myosin is incorporated into the cytoskeleton transiently during shape change, (2) ADP interaction with the ADP aggregation receptor (but not that for shape change) is associated with alpha-actinin and actin incorporation into the cytoskeleton, and (3) further changes that occur are consequent to fibrinogen binding and platelet aggregation.

Actinin↗
PubMed1996

Studies on the effects of agonists and antagonists on platelet shape change and platelet aggregation in whole blood.

The shape change that occurs when platelets are stimulated with an agonist can be quantitated by monitoring changes in their forward-scatter/side-scatter profile using a flow cytometer. Here we have stimulated platelets in citrated whole blood with several agonists and determined the time-course and extent of the shape change that occurs. In some experiments parallel investigations of shape change and aggregation were performed. Aggregation was measured by monitoring the fall in number of single platelets using a Whole Blood Platelet Counter. Some agents (ADP, PAF, U46619 and 5HT) produced a strong and rapid change in platelet forward-scatter/side-scatter that was maximal within 10 s. Others (A23187 and collagen) produced a strong but slower response. Adrenaline produced only a weak response that was also slow to develop, and PMA did not produce any response. The concentrations of each of ADP, PAF, U46619 and 5HT needed to induce a shape change were lower than those required for aggregation. Selective PAF, TXA2 and 5HT antagonists (WEB 2086, sulotroban and MCI-9042) clearly inhibited both the shape change and the aggregation induced by the appropriate agonist; in each case the effect of the antagonist was to move the dose-response curve to the right. These results are consistent with the shape change and aggregation brought about by each of these agonists being mediated via a single receptor. In contrast, a selective P2T purinoceptor antagonist (ARL 66096) markedly inhibited the aggregation induced by ADP but was found to have little or no effect on shape change. This is consistent with these platelet responses to ADP being mediated by different receptors, with P2T receptors mediating only the aggregation response.

15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5↗
PubMed1996

ADP-induced P-selectin expression on platelets as a predictor of successful thrombolysis.

Unlike coronary thrombolysis, the role of platelet activity in the outcome of local thrombolytic therapy for peripheral ischaemia is not well understood. In the present study ten patients undergoing local pulse spray thrombolysis (PST) with recombinant tissue-type plasminogen activator (rt-PA), six patients undergoing conventional infusion thrombolysis (CT) with rt-PA and another six patients undergoing arteriography with iopamidol were studied. Venous blood samples obtained before and after the procedure were analysed using a flow cytometric technique for detection of platelet activation after labelling platelets with VH10, a monoclonal antibody against P-selectin. In the present study no significant differences were observed in P-selectin expression before and after any of the procedures, except that P-selectin expression following ADP stimulation was reduced in patients who had received conventional thrombolysis. Unexpectedly, we observed relatively greater P-selectin expression, particularly after ADP stimulation, both before and following thrombolysis in ten patients in whom thrombolysis was successful compared with six patients in whom thrombolysis was unsuccessful. ADP-induced P-selectin expression on platelets may therefore be a useful predictor of outcome of peripheral intra-arterial thrombolysis.

Adenosine Diphosphate↗
PubMed1996

Platelet reactivity in vitro in relation to thromboxane in healthy pregnancy.

There is substantial evidence of increased platelet reactivity in vivo and in vitro during pregnancy. Previous in vitro studies suggest that platelets from pregnant women show increased sensitivity to agonists, the response to which has a thromboxane dependent component. The aim of this study was to determine whether this is due to increased activity of the thromboxane biosynthetic pathway or to increased platelet sensitivity to the effects of thromboxane. During pregnancy, platelets were more sensitive to the pro-aggregatory effects in vitro of the thromboxane mimetic U46619, in whole blood and in platelet rich plasma, compared to those from non-pregnant controls. The difference in extent of U46619-induced platelet aggregation between groups was abolished in the presence of a high concentration of the specific thromboxane antagonist ICI 192605, but not by prior incubation of blood with aspirin. Platelets from pregnant women were significantly less sensitive to inhibition of arachidonic acid induced activation by the thromboxane synthetase inhibitor dazmegrel, but there was no change in platelet cyclic AMP accumulation under these conditions. Arachidonic acid induced platelet thromboxane B2 production was similar in pregnant and non-pregnant subjects. In conclusion, platelets are more sensitive to the activating effects of thromboxane during pregnancy, but there is no change in the intrinsic reactivity of the thromboxane biosynthetic pathway.

15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5↗
PubMed1995

Longitudinal studies of platelet cyclic AMP during healthy pregnancy and pregnancies at risk of pre-eclampsia.

1. Platelet behaviour in vitro in relation to cyclic AMP was studied longitudinally during pregnancy and in the same women when they were not pregnant. Subjects comprised a group of healthy primigravidae and a group of women deemed at risk of pre-eclampsia, on the basis of a previous history of the condition. 2. There was a progressive decline during pregnancy in sensitivity of platelets to inhibition of the arachidonic acid-induced release reaction by agents which act via cyclic AMP. This effect was maximum at 36 weeks' gestation. 3. Basal platelet cyclic AMP levels, and those in the presence of a phosphodiesterase inhibitor, did not change throughout the period of the study. 4. By contrast, platelet cyclic AMP accumulation in response to a variety of adenylate cyclase stimulators was reduced from early pregnancy, throughout the gestational period, compared with post-natally. This effect was noted when platelets were incubated with prostaglandins acting via different surface receptors or with forskolin and was most marked on co-incubation with a phosphodiesterase inhibitor. 5. Compared with healthy women, platelets from women with a previous history of pre-eclampsia tended to accumulate less cyclic AMP in response to adenylate cyclase stimulators. This was the case both during pregnancy and post-natally. Further investigation of adenylate cyclase activity in platelets in relation to pre-eclampsia is required.

Adenylyl Cyclases↗
PubMed1995

ADP-induced platelet aggregation and actin polymerization. Involvement of GpIIb/IIIa and the effect of Mg2+.

We have investigated the effects of Mg2+ (added to platelet rich plasma [PRP] as 10mM MgCl2 or MgSO4) on the platelet aggregation and actin polymerization that occurs in response to adenosine diphosphate (ADP). The PRP was prepared from blood containing hirudin as anticoagulant. Mg2+ added before 1 microM ADP completely inhibited aggregation and markedly inhibited actin polymerization. Mg2+ (10mM) added before 10 microM ADP converted irreversible aggregation into a reversible response; similarly, apparently irreversible actin polymerization was converted to a reversible response in which polymerization was followed by some actin depolymerization. Mg2+ added after inducing platelet aggregation with 10 microM ADP produced parallel disaggregation of platelets and actin depolymerization. Actin polymerization occurs immediately on adding ADP to PRP (in association with shape change) and further polymerization occurs in association with platelet aggregation. When aggregation (and the actin polymerization associated with this) was prevented by adding M148, a monoclonal antibody directed at the GpIIb/IIIa complex, or simply by avoiding stirring the sample, Mg2+ had no effect on actin polymerization/depolymerization. Thus Mg2+ only affected the changes in actin that were associated with the aggregation response. This was in contrast to iloprost (which acts at the PGI2 receptor to stimulate adenylate cyclase) which induced rapid actin depolymerization when added after ADP stimulation of platelets under circumstances where platelet aggregation was avoided. These results show that Mg2+ affects actin polymerization as well as platelet aggregation, and that it affects the actin polymerization associated with aggregation but not that associated with shape change (in contrast to iloprost which inhibits both).(ABSTRACT TRUNCATED AT 250 WORDS)

Actins↗
PubMed1995

Factors that contribute to spontaneous platelet aggregation and streptokinase-induced aggregation in whole blood.

When whole blood is stirred there is a "spontaneous" platelet aggregation (SPA) which is presumed to be caused by proaggregatory factors released from platelets and other blood cells. Adding streptokinase (SK) to stirred whole blood frequently increases the rate and extent of the platelet aggregation that occurs; this is likely to be via immune complex formation between SK and natural anti-SK antibodies leading to increased release of pro-aggregatory factors. In this investigation we have examined the effects of several inhibitors and antagonists in an attempt to identify the proaggregatory factors that contribute to both SPA and SK-induced aggregation (SKA) and to evaluate different means of inhibiting both processes. The effects of the inhibitors/antagonists were determined in vitro after adding them to citrated whole blood obtained from healthy volunteers. Platelet aggregation was measured using a platelet counting technique. Inhibition of both SPA and SKA by apyrase and by FPL 66096 (a P2T receptor antagonist) demonstrated the involvement of ADP in both processes. Inhibition by chlorpromazine indicated that the most likely source of the ADP is red cells. The effects of sulotroban (a TXA2 antagonist) indicated involvement of TXA2 in SKA but not in SPA. The lack of effect of specific antagonists at S2, alpha 2 and PAF receptors suggested lack of involvement of serotonin, catecholamines and platelet-activating factor in either SPA or SKA. Both SPA and SKA were potently inhibited by low concentrations of iloprost (a PGI2 analogue), but a high concentration of SIN-1 (a NO donor) was much less effective.(ABSTRACT TRUNCATED AT 250 WORDS)

Adenosine Diphosphate↗
PubMed1995

Magnesium modifies the responses of platelets to inhibitory agents which act via cAMP.

This study has investigated the interaction of raised extracellular magnesium and agents which act via cAMP with respect to inhibition of platelet aggregation and effects on platelet cAMP accumulation. Iloprost (3 ng/ml) and PGD2 (0.2 microgram/ml) each caused time dependent increases in platelet cAMP which were significantly greater in the presence of 3 mM added MgSO4 (p < 0.01). Addition of ADP (5 microM) resulted in a fall in cAMP which remained higher in the presence of MgSO4 (p < 0.01). Forskolin (5 micrograms/ml) and DN9693 (100 microM) also caused increments in platelet cAMP but these responses were not influenced by added MgSO4. Addition of ADP resulted in a further increase in cAMP which was augmented by MgSO4 (p < 0.03). This increase was abolished by adenosine deaminase (1.2 U/ml). These experiments show that MgSO4 can modify the cAMP responses produced by iloprost and PGD2 and by forskolin and DN9693 when ADP is present. It appears that as well as inhibiting, ADP can also stimulate cAMP production under certain experimental conditions. This appears to be due to breakdown of ADP to adenosine.

Blood Platelets↗
PubMed1995

Effects of ticlopidine administered to healthy volunteers on platelet function in whole blood.

Ticlopidine is thought to be a selective inhibitor of ADP-induced platelet function. Here we have investigated the effects of ticlopidine on platelet function in whole blood induced by ADP and by other platelet agonists. Whole blood was used because it was considered that ADP derived from red cells might act synergistically with other platelet agonists to enhance platelet responses, and that ticlopidine might interfere with this process. Measurements were performed using blood from 16 healthy volunteers before ticlopidine administration, after taking ticlopidine 250 mg daily for 10 days, after taking ticlopidine 250 mg twice daily for a further 10 days, and after 14 days off treatment. Ticlopidine proved to be a very effective inhibitor of the platelet aggregation induced by ADP; it was most effective in enhancing the reversibility of the aggregation response. The drug modestly but significantly reduced streptokinase, adrenaline, collagen, sodium arachidonate, PAF and U46619 - induced platelet aggregation. The drug significantly reduced the extent of the release reaction (14C-5HT release) induced by ADP, streptokinase, PAF, ristocetin and sodium arachidonate, and also reduced the extent of the synergistic 14C-5HT release induced by combinations of ADP and PAF, ADP and adrenaline and PAF and adrenaline. The various inhibitory effects of ticlopidine were evident after treatment with 250 mg daily but were more pronounced after 250 mg twice daily. All values had returned to normal after 14 days off treatment. Ticlopidine had no effect on serum thromboxane B2 production nor on several parameters of coagulation and fibrinolysis. We conclude that ticlopidine is an effective inhibitor of ADP-induced platelet aggregation and also the platelet aggregation and 14C-5HT release induced in whole blood by a number of platelet agonists and combinations of agonists. These latter effects are probably mainly via a selective effect on ADP. The inhibitory effects of the drug are dose-related.

15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5↗
PubMed1994

Neonatal platelet reactivity and serum thromboxane B2 production in whole blood: the effect of maternal low dose aspirin.

OBJECTIVES: Concern has been expressed about possible neonatal side effects after the use of maternal anti-platelet agents in pregnancy, particularly low dose aspirin treatment. We have studied neonatal platelet behaviour using whole blood techniques, and assessed the neonatal effect of the maternal ingestion of 60 mg aspirin daily. DESIGN: Cross sectional and randomised, double-blind, placebo-controlled. SETTING: University hospital. SUBJECTS: 1. Eight normal women, studied before conception, and their infants. 2. Twenty-four infants whose mothers had been randomised to receive either 60 mg aspirin daily, or placebo, in double-blind fashion. METHODS: The Clay Adams Ultra Flo 100 whole blood single platelet counter was employed to measure platelet aggregation in response to various agonists. The platelet release reaction was also measured in whole blood, and serum thromboxane B2 (TxB2) production was measured by radio-immunoassay. Umbilical cord blood samples were obtained at delivery. RESULTS: 1. Neonatal platelet aggregation induced by adrenaline, ADP and platelet activating factor was reduced in comparison with their mothers (P < 0.01), whereas the neonatal platelet release reaction was reduced when stimulated by collagen and U46619 (a thromboxane mimetic) (P < 0.01). Serum TxB2 production was similar in mothers and babies. 2. Neonatal platelet aggregation, release reaction and serum TxB2 production were not significantly reduced in infants exposed to maternal aspirin in comparison with those neonates exposed to maternal placebo. This is in contrast to the effect on maternal platelets. CONCLUSIONS: Although only a small number of patients were studied, we interpret this as a relative sparing of neonatal platelet reactivity due to the presystemic action of low dose aspirin.

Adult↗
PubMed1994

Inhibition of platelet aggregation by transdermal glyceryl trinitrate.

OBJECTIVE: To determine the optimum conditions for the demonstration of an antiplatelet effect of nitric oxide and to use these conditions to elucidate the effects of a transdermal glyceryl trinitrate patch on platelet aggregation in normal volunteers. METHODS: An open prospective crossover study. The effects of nitric oxide on platelet aggregation in whole blood and platelet rich plasma as stimulated by adenosine diphosphate and U46619 was assessed in the presence and absence of iloprost and MB22948. Optimum conditions for the demonstration of an antiplatelet effect of nitric oxide were then applied to whole blood from normal volunteers in the presence and absence of a transdermal glyceryl trinitrate patch. SETTING: University hospital. SUBJECTS: Eight normal volunteers. MAIN OUTCOME MEASURES: Platelet aggregation in the presence and absence of transdermal glyceryl trinitrate. RESULTS: The optimum conditions for the demonstration of an antiplatelet effect of nitric oxide in whole blood were collecting blood into a tube containing MB22948 and citrate and stimulating platelet aggregation with adenosine diphosphate in the presence or absence of iloprost. Using this method a significant effect of transdermal glyceryl trinitrate on platelet aggregation was shown (P < 0.03) in the presence and absence of iloprost. CONCLUSIONS: These results are consistent with an inhibitory effect on platelet aggregation of nitric oxide liberated by transdermal glyceryl trinitrate. Optimum test conditions are needed to show this effect. The clinical significance of the antiplatelet effect of transdermal glyceryl trinitrate is unknown.

Adenosine Diphosphate↗
PubMed1993

A study of the use of the thromboxane A2 antagonist, sulotroban, in combination with streptokinase for local thrombolysis in patients with recent peripheral arterial occlusions: clinical effects, platelet function and fibrinolytic parameters.

In peripheral thrombolysis adjuvant anti-platelet therapy may help to lyse otherwise resistant thrombus, thereby increasing the number of patients successfully treated and reducing the "time to lysis". If continued after lysis it may help to prevent early rethrombosis. In this pilot study 21 patients undergoing peripheral thrombolysis with streptokinase were randomised to receive the thromboxane receptor antagonist sulotroban or placebo. The dose of sulotroban given was 2 mg/min (four patients), 4 mg/min (five patients) or 8 mg/min (four patients), eight patients received placebo. The clinical and laboratory effects of the treatment were monitored. Thrombolysis was achieved more quickly in patients receiving sulotroban, however, there was no difference between groups in the number of patients in whom recanalisation was achieved (six of eight receiving placebo and eight of 13 receiving sulotroban) or in the number of cases of early rethrombosis. During lysis there was an increase in plasma beta-thromboglobulin with similar levels being found in patients receiving sulotroban and streptokinase and those receiving streptokinase alone. No other major changes in platelet function during lysis were seen in patients receiving streptokinase alone. Sulotroban significantly reduced platelet aggregation and 14C-5HT release in response to several platelet agonists. With the thromboxane mimetic U46619 the degree of inhibition of aggregation and 14C-5HT release depended on the dose of sulotroban used. High levels of inhibition were associated with an excess of haemorrhagic complications especially in combination with a low plasma fibrinogen level. We conclude that the use of low dose sulotroban in combination with streptokinase merits further study and may hve a role in accelerating lysis.(ABSTRACT TRUNCATED AT 250 WORDS)

Aged↗
PubMed1992

Changes in G-actin after platelet activation in platelet rich plasma.

We have used the DNase I inhibition assay to study changes in G-actin after platelet activation in platelet-rich plasma (PRP) induced by ADP. Because of problems associated with depolymerization of F-actin after lysis of ADP-activated platelets in the presence of plasma, G-actin was measured using a lysis buffer that contained formaldehyde to prevent any depolymerization of F-actin. Different patterns of response were seen depending on the concentration of ADP used, and these were modified by avoiding aggregation by either not stirring the sample or by adding EDTA. The results show rapid conversion of G-actin to F-actin in association with shape change, and there is a further decrease in G-actin associated with irreversible platelet aggregation. Thus evidence is presented that actin polymerization occurs in two phases after ADP stimulation.

Actins↗
PubMed1992

A bioassay for inhibition of serotonin release from bovine platelets.

A bioassay was developed to study agents capable of inhibiting the release of serotonin from bovine blood platelets. It is a simple, inexpensive, and reproducible high-throughput bioassay suitable for quality control of feverfew, Tanacetum parthenium, a crude drug with proven migraine prophylactic activity that is being considered for governmental registration and regulation. The bioassay, which requires no experimental animals or human subjects, was used to assess the in vitro activity of T. parthenium samples grown from seed obtained from 10 different regions of Europe. The activity was found to vary significantly within and between samples, with no geographical correlation. Serotonin release inhibition was shown to be significantly correlated with the content of the germacranolide sesquiterpene lactone, parthenolide, although other sesquiterpene lactones from this plant and other members of the Asteraceae were also shown to be active. The activities of six other species of Tanacetum, as well as of Artemisia absinthium (wormwood) and Zingiber officinale (ginger), and two commercial drugs for migraine prophylaxis, verapamil hydrochloride and propranolol hydrochloride, were also assessed. The relevance of the bovine platelet serotonin release inhibition bioassay to antimigraine research is discussed.

Animals↗
PubMed1992

A randomized placebo-controlled study of the effect of low dose aspirin on platelet reactivity and serum thromboxane B2 production in non-pregnant women, in normal pregnancy, and in gestational hypertension.

OBJECTIVE: To investigate the effect of 60 mg aspirin daily on platelet reactivity and prostaglandin production in various groups of patients. Similar regimens, which are thought to act through inhibition of platelet thromboxane production, are currently undergoing clinical assessment for the prevention of pre-eclampsia and intrauterine growth retardation. DESIGN: A prospective randomized placebo controlled study. SETTING: University Hospital, Nottingham. SUBJECTS: 12 non-pregnant female volunteers, 18 normal primigravidae before 16 weeks gestation and 16 pregnant women admitted with gestational hypertension (GH) at a mean gestation of 38 weeks. INTERVENTIONS: In the non-pregnant women blood samples were taken before and after a 10-day course of 60 mg aspirin daily. The primigravidae had blood samples taken at 16 weeks and then they were randomized to receive either 60 mg aspirin daily or a matched placebo. Further blood samples were obtained at 28, 32 and 36 weeks. MAIN OUTCOME MEASURES: Changes in platelet reactivity and release reaction, and serum thromboxane production, were estimated in whole blood. RESULTS: 60 mg aspirin daily significantly inhibited cyclo-oxygenase dependent platelet aggregation, release reaction and serum thromboxane production in non-pregnant and pregnant women, and in women with GH (P less than 0.01). When adrenaline was used as the aggregating agent, the cyclo-oxygenase pathway was recruited in the increased reactivity seen in the third trimester of normal pregnancy, and was sensitive to inhibition by low dose aspirin. CONCLUSION: Low dose aspirin would appear to be an appropriate agent for the inhibition of platelet reactivity associated with hypertensive pregnancy.

Adult↗
PubMed1992

Parthenolide content and bioactivity of feverfew (Tanacetum parthenium (L.) Schultz-Bip.). Estimation of commercial and authenticated feverfew products.

Three physicochemical methods (HPLC, NMR spectroscopy, and HPLC of a derivative) have been used to measure parthenolide in authenticated Tanacetum parthenium (feverfew) and in several commercial purported feverfew products. A bioassay based on inhibition of the secretory activity of blood platelets by extracts of feverfew in comparison with parthenolide was also used. Similar results were obtained for all three physicochemical assays and also for the bioassay. Thus different methodologies yield consistent values for parthenolide content of feverfew preparations. Parthenolide appears to be mainly responsible for the antisecretory effects of extracts of feverfew. Authenticated Tanacetum parthenium grown in the UK contained a high level of parthenolide in leaves, flowering tops and seeds but a low level in stalks and roots. The level of parthenolide in powdered leaf material fell during storage. The purported feverfew products varied widely in their parthenolide content and in some products parthenolide was not detected. Possible reasons for the variation in parthenolide content are discussed. Since therapeutic efficacy has only been demonstrated for preparations of feverfew that contain parthenolide, it is suggested that manufacturers of feverfew products should use measurements of parthenolide as a means of standardization and quality control.

Capsules↗