Aggregation Of Bovine Platelets Research Articles

Inhibition of bovine platelets aggregation in response to Hyalomma anatolicum salivary gland proteins/peptides.

Aim:Ticks are obligate ectoparasites that have an impact on wide range of vertebrates and also act as a potential vector for the transmission of tropical theileriosis, babesiosis, etc., causing significant loss to livestock production worldwide. While feeding, they introduce their saliva containing different bioactive molecules into the host. These molecules have the capability to counteract the host hemostatic mechanism to suck host blood successfully. Therefore, the study was aimed to isolate anti-platelet aggregating peptides from salivary gland extract (SGE) of Hyalomma anatolicum ticks, a commonly available tick in India.Materials and Methods:Female H. anatolicum salivary glands were dissected out and SGE was prepared by homogenizing it in a suitable buffer under ice. Extract so obtained was fractionated by gel filtration chromatography using Sephacryl S-200 column. Total protein concentration in fractions was estimated and bovine platelets were isolated, stimulated with thrombin (positive control), treated with Gly-Pro-Arg-Pro amide (negative control) and with salivary gland fractions for identification of proteins/peptides having anti-platelet aggregating activities.Results:Proteins/peptides present in various salivary gland fractions inhibited the bovine platelet aggregation and the percent inhibition ranged between 33% and 35.8%.Conclusion:The results suggests that the fractions of H. anatolicum salivary glands possess thrombin-induced anti-platelet aggregating activity and which could be further exploited for raising anti-tick vaccine and also for therapeutic purpose.

Effects of imidazoline and nonimidazoline alpha-adrenergic agents, including xylazine, medetomidine, yohimbine, tolazoline, and atipamezole, on aggregation of bovine and equine platelets

To investigate effects of various imidazoline and nonimidazoline α-adrenergic agents on aggregation and antiaggregation of bovine and equine platelets. Blood samples obtained from 8 healthy adult cattle and 16 healthy adult Thoroughbreds. Aggregation and antiaggregation effects of various imidazoline and nonimidazoline α-adrenergic agents on bovine and equine platelets were determined via a turbidimetric method. Collagen and ADP were used to initiate aggregation. Adrenaline, noradrenaline, or α-adrenoceptor agents alone did not induce changes in aggregation of bovine or equine platelets or potentiate ADP- or collagen-induced platelet aggregation. Adrenaline and the α(2)-adrenoceptor agonist clonidine had an inhibitory effect on ADP- and collagen-induced aggregation of bovine platelets. The α(2)-adrenoceptor antagonists phentolamine and yohimbine also inhibited collagen-induced aggregation of bovine platelets. Noradrenaline, other α-adrenoceptor agonists (xylazine, oxymetazoline, and medetomidine), and α-adrenoceptor antagonists (atipamezole, idazoxan, tolazoline, and prazosin) were less effective or completely ineffective in inhibiting ADP- and collagen-induced aggregation of bovine platelets. The imidazoline α(2)-adrenoceptor agonist oxymetazoline submaximally inhibited collagen-induced aggregation of equine platelets, and the α(2)-adrenoceptor antagonist idazoxan, along with phentolamine and yohimbine, also inhibited collagen-induced aggregation of equine platelets. The imidazoline compound antazoline inhibited both ADP- and collagen-induced aggregation of equine platelets. Several drugs had effects on aggregation of platelets of cattle and horses, and effective doses of ADP and collagen also differed between species. The α(2)-adrenoceptor agonists (xylazine and medetomidine) and antagonists (tolazoline and atipamezole) may be used by bovine and equine practitioners without concern for adverse effects on platelet function and hemostasis.

Alteration of release and role of adenosine diphosphate and thromboxane A2 during collagen-induced aggregation of platelets from cattle with Chediak-Higashi syndrome

To compare the interaction of endogenous ADP with collagen and thromboxane A(2) (TXA(2)) during collagen-induced platelet aggregation between platelets from healthy cattle and those with Chediak-Higashi syndrome (CHS). POPULATION SAMPLE: Platelets harvested from blood samples from healthy Japanese Black cattle and those with CHS. Aggregation of gel-filtered platelets; release of ATP-ADP; and generation of thromboxane B(2) (TXB(2)), a metabolite of TXA(2), were measured. The potency of collagen to induce aggregation in platelets of cattle with CHS (ie, CHS platelets) was less than a tenth of that in platelets of healthy cattle (ie, control platelets). Platelet aggregation induced by collagen at an intermediate concentration depended on the coexistence of ADP and TXA(2), suggesting that released ADP cannot cause platelet aggregation by itself. Collagen-induced ADP release was markedly decreased, whereas TXB(2) production was slightly low in CHS platelets, compared with that in control platelets. A combination of subthreshold amounts of ADP and 9,11-dideoxy-9alpha, 11alpha-methano-epoxy-prostaglandin F(2) (U46619), a TXA(2) analogue, caused platelet aggregation. Similarly, a combination of subthreshold amounts of collagen and ADP caused platelet aggregation, whereas collagen and U46619 were not synergistic. Deficient ADP release ensuing from the delta-storage pool deficiency in platelets from cattle with CHS resulted in reduction of collagen-induced platelet aggregation, through attenuation of synergism between TXA(2) and ADP and between ADP and collagen. Furthermore, results of the study reported here indicated that TXA(2) was important for aggregation of bovine platelets.

Platelet dysfunction in Chediak-Higashi syndrome-affected cattle.

A serious symptom of cattle affected with Chediak-Higashi syndrome (CHS) is a bleeding tendency. This diathesis is characterized by insufficient platelet aggregation as a result of depressed response to collagen. One possible cause for the depression is a decrease in contribution of endogenous agonists such as ADP or thromboxane A(2), which are released following collagen stimulation. However, these endogenous agonists play only a minor role in collagen-induced aggregation of bovine platelets. More importantly, activation of phospholipase C as a result of a direct action of collagen is depressed, leading to a depression of Ca(2+) mobilization, in platelets from CHS-affected cattle. Several types of collagen receptor are proposed to work in concert to induce aggregation. Among them, glycoprotein VI (GPVI) and GPIa/IIa (integrin alpha2 beta1) have been supposed to play dominant roles in collagen-induced aggregation. However, there are arguments about the role of each receptor, especially the role of GPIa/IIa, and the crosstalk between receptors. Recently, we reported that the Ca(2+) signaling produced by rhodocytin, which had been first reported to be an agonist for the collagen receptor GPIa/IIa, produced much less Ca(2+) signaling in CHS platelets than in normal ones, whereas that produced by GPVI activators was normal. These suggest that GPIa/IIa or the rhodocytin-associated pathway is impaired in CHS platelets. CHS platelets are valuable to reassess the mechanism of collagen-dependent signal transduction system and to delineate the inter-relationship among collagen receptors.

In vitro inhibitory effect of SR 27417, a potent platelet-activating factor (PAF) receptor antagonist, on the PAF-induced bovine platelet aggregation

The in vitro inhibitory effect of SR 27417, an antagonist of the platelet-activating factor (PAF) receptor, on PAF-induced platelet aggregation was studied in blood collected from seven healthy Friesien calves. Inhibitory effects of SR 27417 were determined at thirteen different concentrations (0.1-400 nM) by using the dose-response curves of PAF on calf platelet aggregation. In the presence of SR 27417, the maximal slopes of aggregation (%/min) induced by low and high concentrations of PAF were significantly different from the control values obtained without an antagonist at p < or = 0.05 and p < or = 0.01 respectively. In vitro PAF-induced calf platelet aggregation was dose-dependently inhibited by SR 27417. The drug inhibited PAF-induced platelet aggregation in a competitive reversible manner (pA2 = 10.46 +/- 2.36 mol x L(-1)). In conclusion, the results of our study showed that addition of SR 27417 to bovine platelet in vitro inhibits PAF-induced platelet aggregation.

6,9-Dihydroxy-3(15)-caryophyllen-4,8-dione--a new antibiotic from a Marasmius species.

6,9-Dihydroxy-3(15)-caryophyllen-4,8-dione (1), a new cytotoxic caryophyllane sesquiterpene, was isolated from fermentations of a tropical Marasmius species. The structure was established by spectroscopic methods. 1 exhibits strong cytotoxic effects on different cell lines, but only weak antimicrobial activities. 1 weakly inhibits the incorporation of leucine and thymidine into proteins and DNA of mammalian cells and interferes with the aggregation of human and bovine platelets.

Five new bioactive sesquiterpenes from the fungus Radulomyces confluens (Fr.) Christ.

The three protoilludanes radulone A (1), radulone B (2) and radudiol (3), the illudalane radulactone (4) and the illudane radulol (5) were isolated from the extracts of the culture fluids of the basidiomycete Radulomyces confluens. The structures of the five new compounds were determined by spectroscopic techniques. Radulone A (1) is a potent inhibitor of human and bovine platelet aggregation stimulated by different agonists, inhibiting preferentially the aggregation of human platelets induced by ADP with an IC50 value of 2 microM. In addition 1 exhibits cytotoxic and antimicrobial activities. The other four compounds exhibited weak antimicrobial and cytotoxic activity.

Effect of ketoprofen on PAF-induced bovine plateletaggregation

Effect of ketoprofen on PAF-induced bovine plateletaggregation

Combined effect of Web 2086 (Paf antagonist) and ketoprofen (Nsaid) on Paf-induced ex vivo platelet aggregation in bovine.

The effect of the specific PAF-antagonist WEB 2086, a thieno-triazolo-diazepine, and ketoprofen, a NSAID, was investigated on PAF-induced bovine platelet aggregation measured ex vivo in platelet-rich plasma (PRP). WEB 2086 was infused intravenously over 1 min followed immediately by ketoprofen administration over 1 s (both drugs = 3 mg/kg), in a group of six healthy male Friesian calves. Depending on the PAF concentration, a reversible (10(-8)-10(-9) mol/l) and irreversible (10(-5)-10(-7) mol/l) platelet aggregation was observed. The reversible aggregation was completely blocked by pretreatment of the animal with WEB 2086 and ketoprofen. The inhibitory effects observed during the irreversible aggregation were 47.22%, 54.00% and 88.00% at 10(-5), 10(-6) and 10(-7) mol/l PAF, respectively. Moreover, the aggregation obtained in these condition became reversible. Maximal inhibitory effect of WEB 2086 and ketoprofen on PAF-induced platelet aggregation in calves was observed within 30 min after administration of both drugs. This inhibition persisted even after 24 h and was significantly different from control with P < 0.05. The combined effect of both drugs exceeded the sum of the individual effects (synergism). It was concluded that WEB 2086 and ketoprofen very effectively blocked PAF-induced bovine platelet aggregation in platelet-rich plasma. The study also suggested a synergism between both substances.

Inhibition of PAF-induced platelet aggregation by WEB 2086 'in-vitro', an antagonist to the receptor for platelet-activating factor, in bovine.

The sensitivity of bovine platelet aggregation in response to PAF stimulation and the ability of WEB 2086 (a thieno-triazolodiazepine) to inhibit response to PAF-induced platelet aggregation were investigated in the blood from five healthy male Belgian Blue calves. The recorded response to PAF showed a plateau which was dependent on the PAF concentration. Platelet aggregation induced by PAF consists of two mechanisms: reversible and irreversible aggregations which are accompanied by the release of platelet granule contents. Reversible aggregation occurred above (2 . 10(-9) mol/l) PAF, and irreversible aggregation occurred above (2 . 10(-7) mol/l) PAF. Addition of WEB 2086 to bovine platelets in vitro induced a rightward shift in the dose-response curve to PAF. WEB 2086 inhibited PAF-induced aggregation in a competitive reversible manner (pA2 = 7.61). The results of our study show that PAF induces platelet aggregation in platelet-rich plasma (PRP) and that addition of WEB 2086 to bovine platelets in vitro inhibits PAF-induced Platelet Aggregation.

5-Hydroxy-3-vinyl-2(5H)-furanone -a New Inhibitor of Human Synovial Phospholipase A2 and Platelet Aggregation from Fermentations of a Calyptella Species (Basidiomycetes)

5-Hydroxy-3-vinyl-2(5H)-furanone, a potent and selective inhibitor of human synovial phospholipase A2 was isolated from fermentations of a Calyptella species. Its structure as identified by spectroscopic methods is identical to PA 147, an antibiotic previously isolated from a streptomycete. 5-hydroxy-3-vinyl-2(5H)-furanone inhibits the aggregation of human and bovine platelets stimulated by different inducers and exhibits weak antimicrobial activities.

14-epidihydrocochlioquinone B and 14-epicochlioquinone B, antibiotics from fermentations of the ascomycete Neobulgaria pura: structure elucidation and effects on platelet aggregation.

14-Epicochlioquinone B (1) and 14-epidihydrocochlioquinone B (2) were isolated from submerged cultures of Neobulgaria pura (Pers. ex Fr.) Petrak. 14-Epicochlioquinone B is a potent inhibitor of human and bovine platelet aggregation stimulated by different inducers. 14-Epidihydrocochlioquinone B does not inhibit the aggregation of platelets. In addition, both 1 and 2 exhibited cytotoxic and antimicrobial activities.

Two inhibitors of platelet aggregation from a Panus species (Basidiomycetes).

Two inhibitors of platelet aggregation were isolated from fermentations of Panus sp. 9096. One inhibitor proved to be identical to naematolon (2), an antibiotic previously isolated by S. Backens et al. from several Hypholoma species. The other metabolite, panudial (1), is a new nordrimane (cis-annelation of the bicyclus) lacking the carbon atom in position 10 of the drimane skeleton. Panudial is a potent inhibitor of bovine and human platelet aggregation stimulated by different inducers.

Aggregation of bovine platelets by Fusobacterium necrophorum

Washed cell suspensions of biovar A strains of Fusobacterium necrophorum aggregated cattle platelets, but similar suspensions of biovar B strains did not. Platelets were also aggregated by heat-treated bacterial cells or the lipopolysaccharide of biovar A. No platelet aggregation occurred in the presence of the cell-free culture supernatant of biovar A and of all samples prepared from biovar B. Scanning electron microscopy revealed that aggregated platelets were not damaged. Platelet aggregation was inhibited by EDTA, aspirin and quinacrine, and lag time was retarded by these inhibitors, indicating the reaction was a Ca 2+-dependent, cyclo-oxygenase sensitive event. Platelet aggregation may be a virulence marker, probably mediated by the lipopolysaccharide of F. necrophorum biovar A strains.

Influence of Aspirin and Carbacyclin on Bovine Platelet Function

Cows, calves, and sheep are the animals of choice for in vivo studies on total artificial hearts. In this study, the response of human and bovine platelets to agonists and antagonists was followed. Epinephrine and arachidonate failed to cause aggregation of bovine platelets. Exposure of bovine platelets to prostaglandin E1 (PGE1) resulted in inhibition of response to the action of adenosine diphosphate (ADP). Although epinephrine restored the response of PGE1 treated human platelets, it failed to restore the sensitivity of bovine platelets to the action of ADP. Exposure of bovine platelets to aspirin (100.0 mumol/L) or administration to calves intravenously (10.0 mg/kg) inhibited platelet cyclo-oxygenases. Infusion of carbacyclin (U55185) inhibited the ex vivo platelet response to the action of ADP. Results of this study demonstrate that the response of bovine platelets to agonists such as epinephrine, arachidonate, and the endoperoxide mimetic, U46619, is severely compromised. The authors' observations in this study, as well as earlier findings demonstrating the inability of bovine platelets to fully spread on a surface, suggests that the cow may not be an appropriate "model" for evaluating thrombogenicity caused by artificial organs and implants.

Effects of apocynin, a drug isolated from the roots of Picrorhiza kurroa, on arachidonic acid metabolism

Apocynin is a constituent of root extracts of the medicinal herb Picrorhiza kurroa and has been shown to possess anti-inflammatory properties. We investigated the effects of apocynin on the production of arachidonic acid-derived inflammatory mediators by guinea pig pulmonary macrophages. Apocynin concentration-dependently inhibited the formation of thromboxane A 2, whereas the release of prostaglandins E 2 and F 2α was stimulated. Apocynin potently inhibited arachidonic acid-induced aggregation of bovine platelets, possibly through inhibition of thromboxane formation. The present results suggest that apocynin might, beside its therapeutic effects in inflammatory conditions when given in a root extract of P. kurroa, also be a valuable tool in the development of new anti-inflammatory or anti-thrombic drugs.

12 [formula omitted]-Hydroxyeicosatetraenoic acid plays a central role in the regulation of platelet activation

When platelets are activated by the recognition of exposed collagen fibers, they start synthesizing two major arachidonic acid metabolites, i.e. thromboxane A 2 and 12 S -hydroxyeicosatetraenoic acid (12-HETE) via cyclooxygenase and 12-lipoxygenase pathways, respectively. Although the physiological role of the former is well established, that of the latter has not been fully elucidated. Recently, we have revealed that 12-HETE interferes with collagen-induced platelet aggregation [ Sekiya, F. et al. (1990) Biochim.Biophys.Acta 1044, 165–168]. In the present paper, we show that this substance enhances thrombin-induced aggregation of bovine platelets, in sharp contrast with the case of collagen. Additionally, 12-HETE is able to prevent the prostaglandin E 1-induced elevation of platelet cAMP level and counteracts its inhibitory effect on platelet aggregations. With these observations, we propose a novel self-regulatory mechanism of platelets where 12-HETE plays a key role; it switches sensitivity of platelets from the primary agonist (collagen) to the secondary one (thrombin), and cancels the inhibitory effect of cAMP elevators.

Inhibitory mechanism of cis-polyunsaturated fatty acids on platelet aggregation: the relation with their effects on Ca 2+ mobilization, cyclic AMP levels and membrane fluidity

The in vitro inhibitory effects of cis-polyunsaturated fatty acids, linolenic (18:2 Δ 9,12), α-linoleic (18:3 Δ 9,12,15) and eicosatrienoic (20:3 Δ 11,14,17) acid, on bovine platelet aggregation and their inhibitory mechanism were investigated. These fatty acids inhibited platelet aggregation induced by ADP and thrombin to similar extent. Fluorescence analyses with fura-2-loaded platelets showed that, in the concentration ranges that inhibited aggregation, they also inhibited agonist-induced increase in cytoplasmic Ca 2+. According to radioimmunoassay study, addition of these fatty acids increased cyclic AMP contents in the presence of theophylline corresponded with their inhibitory effects on aggregation. These fatty acids induced a 1.6–1.8-fold increase over basal concentration of cyclic AMP in the concentration ranges that fully inhibited aggregation. On the other hand, saturated fatty acid, stearic acid, affected neither aggregation nor cyclic AMP levels. As reported previously ((1985) Biochim. Biophys. Acta 818, 391), these unsaturated fatty acids induced increase in membrane fluidity in the same concentration range. These results suggest that inhibition of platelet aggregation by cis-polyunsaturated fatty acids is due to the increase in cyclic AMP levels. This increase seems to be due to stimulation of adenylate cyclase which is mediated by membrane perturbation.

Cooperativity between platelet-activating factor and collagen in aggregation of bovine platelets III

In cooperative aggregation of bovine platelets induced by simultaneous addition of PAF and collagen at subthreshold concentrations the following observations were made. (i) Formation of phosphatidic acid and arachidonic acid metabolites, which characterize PAF and collagen alone-induced aggregation, respectively, was observed very obviously. (ii) A thromboxane antagonist did not completely block the cooperative aggregation. The extent of residual aggregation activity was dependent on concentration of collagen used in the simultaneous administration with PAF. These results suggest that both signal transduction pathways activated by PAF and collagen alone at high concentrations are attained by simultaneous addition of both agonists at subthreshold concentrations through unknown mechanisms.

Cooperativity between platelet-activating factor and collagen in aggregation of bovine platelets, II

When subthreshold amounts of platelet-activating factor (PAF) and collagen were added simultaneously, strong aggregation of platelets was induced. However, each agonist alone at these concentrations could not induce aggregation at all (S. Kojima et al , (1987) Biochem. Biophys. Res. Commun. 145, 915–920). This cooperativity was observed not only in aggregation but also in changes of intracellular Ca 2+ concentration, 47 kDa protein phosphorylation, and formation of thromboxane. These findings suggest that various steps of signal transduction pathways are enhanced during their cooperativity.