Aggregation Of Gel-filtered Platelets Research Articles

Activation of human platelets by lysophosphatidic acid.

The biochemical mechanisms of platelet activation by lysophosphatidic acid were investigated. Lysophosphatidic acid interacts with a membrane receptor coupled to the inhibitory GTP-binding protein Gi and produces a rapid decrease of the intracellular concentration of cAMP. Aggregation of gel-filtered platelets by lysophosphatidic acid requires the presence of extracellular CaCl2, as this phospholipid does not induce secretion of platelet dense granules. Platelet activation by lysophosphatidic acid in the absence of extracellular CaCl2 does not involve phospholipase C activation, as evaluated by measuring mobilization of Ca2+ from internal stores and pleckstrin phosphorylation, but causes the rapid tyrosine phosphorylation of several intracellular proteins. Our results indicate that activation of intracellular tyrosine kinases is not secondary to Ca2+ mobilization and protein kinase C activation in lysophosphatidic acid-stimulated platelets.

Effect of albumin on the inhibition of platelet aggregation by beta- lactam antibiotics

Platelet aggregation and bleeding time abnormalities are reported in patients receiving beta-lactam antibiotics (beta LAs), although clinical bleeding most frequently occurs in chronically ill, malnourished patients. Although most beta LAs bind to serum albumin, the relative influence of bound versus unbound beta LAs on platelet function is unknown. We examined the effect of beta LAs on the aggregation of gel-filtered platelets from normal subjects and on platelet-rich plasma (PRP) from hypoalbuminemic patients. Therapeutic concentrations of five beta LAs were added to normal platelets at different albumin concentrations (1.5 to 4.5 g/dL). Inhibition of aggregation by the beta LAs was inversely proportional to the albumin concentration, and most antibiotic-treated samples showed more than 50% inhibition at albumin levels below 2.0 g/dL. When PRPs from hypoalbuminemic patients were incubated with cephalothin, aggregation was completely inhibited, in contrast to samples from patients with normal albumin levels, and this decreased platelet aggregation was partially restored (25% to 75%) by increasing the albumin concentration above 4.0 g/dL. Specific binding of [35S]-benzylpenicillin to normal platelets decreased proportionately as the albumin concentration increased in the range of 1.0 to 5.0 g/dL. The inhibitory effects of beta LAs on platelets in vitro appear to be influenced by albumin concentration. Plasma albumin concentration may influence bleeding in patients receiving beta LAs.

Effect of Albumin on the Inhibition of Platelet Aggregation by β-Lactam Antibiotics

Platelet aggregation and bleeding time abnormalities are reported in patients receiving beta-lactam antibiotics (beta LAs), although clinical bleeding most frequently occurs in chronically ill, malnourished patients. Although most beta LAs bind to serum albumin, the relative influence of bound versus unbound beta LAs on platelet function is unknown. We examined the effect of beta LAs on the aggregation of gel-filtered platelets from normal subjects and on platelet-rich plasma (PRP) from hypoalbuminemic patients. Therapeutic concentrations of five beta LAs were added to normal platelets at different albumin concentrations (1.5 to 4.5 g/dL). Inhibition of aggregation by the beta LAs was inversely proportional to the albumin concentration, and most antibiotic-treated samples showed more than 50% inhibition at albumin levels below 2.0 g/dL. When PRPs from hypoalbuminemic patients were incubated with cephalothin, aggregation was completely inhibited, in contrast to samples from patients with normal albumin levels, and this decreased platelet aggregation was partially restored (25% to 75%) by increasing the albumin concentration above 4.0 g/dL. Specific binding of [35S]-benzylpenicillin to normal platelets decreased proportionately as the albumin concentration increased in the range of 1.0 to 5.0 g/dL. The inhibitory effects of beta LAs on platelets in vitro appear to be influenced by albumin concentration. Plasma albumin concentration may influence bleeding in patients receiving beta LAs.

Functional Involvement of Thrombospondin in Platelet Aggregation Induced by Low Versus High Concentrations of Thrombin

Thrombospondin (TSP) is a major platelet secretory glycoprotein. Earlier studies of various investigators demonstrated that TSP is the endogenous platelet lectin and is responsible for the hemagglutinating activity expressed on formaldehyde-fixed thrombin-treated platelets. The direct effect of highly purified TSP on thrombin-induced platelet aggregation was studied. It was observed that aggregation of gel-filtered platelets induced by low concentrations of thrombin (less than or equal to 0.05 U/ml) was progressively inhibited by increasing concentrations of exogenous TSP (greater than or equal to 60 micrograms/ml). However, inhibition of platelet aggregation by TSP was not observed when higher than 0.1 U/ml thrombin was used to activate platelets. To exclude the possibility that TSP inhibits platelet aggregation by affecting thrombin activation of platelets, three different approaches were utilized. First, by using a chromogenic substrate assay it was shown that TSP does not inhibit the proteolytic activity of thrombin. Second, thromboxane B2 synthesis by thrombin-stimulated platelets was not affected by exogenous TSP. Finally, electron microscopy of thrombin-induced platelet aggregates showed that platelets were activated by thrombin regardless of the presence or absence of exogenous TSP. The results indicate that high concentrations of exogenous TSP (greater than or equal to 60 micrograms/ml) directly interfere with interplatelet recognition among thrombin-activated platelets. This inhibitory effect of TSP can be neutralized by anti-TSP Fab. In addition, anti-TSP Fab directly inhibits platelet aggregation induced by a low (0.02 U/ml) but not by a high (0.1 U/ml) concentration of thrombin.(ABSTRACT TRUNCATED AT 250 WORDS)

Fibrinogen fragment D is a recognition site for release-related platelet aggregation

The present study probes for the segment of the fibrinogen molecule which interacts with the platelet surface upon induction of release-related aggregation. The capability of platelets affixed with fibrinogen fragment D or fibrinogen fragment E to enhance aggregation of gel-filtered platelets (GFP) was compared with that of platelets affixed with fibrinogen. It is shown that release-related aggregation induced by either A23187, 10 μM ADP or thrombin is enhanced by the addition of fixed platelets bearing covalently bound fragment D by as much as 60–70% of the augmentation obtained by platelets bearing fibrinogen. On the other hand, fixed platelets bearing covalently-bound fragment E have no effect on the aggregation of GFP. It is concluded that fragment D bears the site for the interaction of fibrinogen with activated platelets, apparently, with surface-bound thrombospondin.

Influence of β2-Glycoprotein-l Upon the Content of cAMP and cGMP in Human Blood Platelets

The influence of beta 2-glycoprotein-I on the cyclic nucleotide content in human gel filtered platelets (GFP) was studied: beta 2-G-I up to a concentration of 5 mg/ml does not alter the basal cAMP level in resting GFP and also does not influence the action of PGE1 which strongly inhibits platelet aggregation by elevation of cAMP. Collagen induced aggregation of GFP leads to an increase of the platelet cGMP content. Preincubation of GFP with 0.2 mg/ml of beta 2-G-I enhances significantly the cGMP increase by collagen. In a similar manner beta 2-G-I also amplifies the increase in cGMP concentration induced by ADP. In this case, a concomitant alteration of the aggregation pattern is observed. Thrombin produces a much higher cGMP elevation in GFP than ADP or collagen. The admixture of 0.2 mg/ml of beta 2-G-I is not able to enhance further the thrombin effect on cGMP. The results are discussed in view of the possible significance of these observations for platelet aggregation in vivo.

Aggregation of Gel-Filtered Guinea-Pig Platelets by Lipoproteins

Very low density lipoproteins (VLDL) and low density lipoproteins (LDL) were isolated from serum of hypercholesterolemic guinea-pigs, and the effect of these lipoproteins on guinea-pig platelets was studied. VLDL (greater than 100 microgram/ml) and LDL (greater than 400 microgram/ml) were found to cause aggregation of gel-filtered platelets (GFP), although the extent of GFP aggregation by LDL was smaller than that by VLDL. In platelet-rich plasma, however, lipoproteins could not induce platelet aggregation. VLDL and LDL even at the low concentrations at which lipoproteins alone could not induce aggregation potentiated ADP-induced aggregation of GFP. VLDL-induced aggregation of GFP was inhibited by apyrase (0.2--1.0 mg/ml) in a concentration-related manner. Prostaglandin E1, dipyridamole, potassium cyanide and ethylenediaminetetraacetic acid inhibited VLDL- and ADP-induced aggregation of GFP in the almost same degree. Inhibitions of VLDL-induced GFP aggregation by acetylsalicylic acid and albumin were slightly stronger than that of ADP-induced aggregation. These findings suggest that lipoproteins modulate platelets so that endogenous ADP can be released from platelets.