Study on antithrombogenicity of poly[beta-(acetylsalicylyloxy)ethyl methacrylate] relative to poly(hydroxyethyl methacrylate)

The antithrombogenicity of a polymer made of aspirin bound to hydroxyethyl methacrylate (HEMA), abbreviated as ASA-polymer, was compared with that of poly(hydroxyethyl methacrylate) (PHEMA). Platelet from platelet rich plasma (PRP) incubated with ASA-polymer surface exhibited noticeable decreases in adhesion and aggregation as compared to platelets incubated with PHEMA. Low molecular weight components other than aspirin, which may be released from ASA-polymer during the incubation with PRP, or contact with ASA-polymer causing denaturation of platelets without morphological changes could be responsible for the decrease of adhesion and aggregation. Both PRP and PPP exposed to ASA-polymer-coated surfaces exhibited a much smaller partial thromboplastin time (PTT) than if exposed to PHEMA-coated surfaces; the PTT of ASA-polymer was similar to that of glass exposed plasma. With respect to the in vivo antithrombogenicity, the ASA-polymer surface led to thrombus formation. This may be due to the partial hydrolysis of the acetyl groups resulting in the formation of a negatively charged surface which in turn accelerates the coagulation cascade despite its inhibitory effects on platelet adhesion and aggregation. On the other hand, neointima formed around a thrombus layer on PHEMA-coated sutures after 14 days.     

Critical evaluation of platelet aggregation in whole human blood

Platelet aggregation studies generally are performed in platelet-rich plasma (PRP) by the turbidometric method. The authors compared this technic with the recently introduced impedance aggregometry in PRP and whole blood (WB). In healthy controls there was a good correlation between the two technics when aggregation was induced by ADP or collagen. As compared with PRP, platelets in WB were more sensitive to the aggregating effect of thrombin, ristocetin, and arachidonic acid. Platelet sensitivity to prostacyclin was increased in WB. The anti-platelet effect of a single oral dose of aspirin could be detected for a longer period in WB than in PRP. Platelet aggregation tests in WB from patients with platelet dysfunctions showed the same response pattern to different aggregating agents as in PRP. In contrast to turbidometry, the impedance method in PRP and WB enabled registration of platelet aggregation in a dose-dependent fashion in a sample from a patient with severe hyperlipoproteinemia. It is concluded that platelet aggregation can be studied conveniently with the impedance method in the more physiologic medium of WB. Providing the same information as the well-established turbidometry, the time-sparing impedance method needs less citrated blood. Moreover, our results show an increased sensitivity of the WB system to some aggregating and anti-platelet agents.     

Platelet adhesion and adhesive proteins--how to measure the platelet adhesiveness]

We examined, platelet adhesive function in vivo by comparing the platelet retention rate (PRR) with other laboratory tests. In 71 patients, PRR did not correlate with bleeding time (BT), ristocetin-induced platelet agglutination (RIPA), vWF:AG, vWF:RCo, FVIII:C, fibrinogen, platelets counts or platelet aggregation tests. Among them, the coincidence rate of PRR with BT seems relatively better than that of other tests. In patients with Bernard-Soulier syndrome (BSS), PRR, RIPA and high shear-induced platelet aggregation (SIPA), measured using a cone-plate type instrument, were decreased, while platelet adhesion in a static system and SIPA at a low shear rate were normal. In patients with Glanzmann's thrombasthenia (TA), PRR and SIPA were decreased at both high and low shear rates, while RIPA and platelet adhesion were normal in a static system. In patients with deficient glycoprotein Ia/IIa complex which is one of the putative receptors for collagen, platelet adhesion in a static system is markedly decreased. In patients with von Willebrand disease, the pattern of platelet adhesion tests was similar to that of BSS patients, while in patients with afibrinogenemia, it was similar to that of TA patients except for SIPA at a high shear rate. The various experiments using monoclonal antibodies support these abnormalities. These findings, suggest that PRR is a suitable screening test, but further rapid and easy tests will be necessary to diagnose the defects of platelet adhesion at various shear rates.     

Platelet fibrinogen

Platelet fibrinogen has been studied in normal, thrombasthenic, and hypofibrinogenaemic subjects. It has been differentiated into adsorbed (plasma) and extractable (intraplatelet) fractions. Isotopic studies suggest that exchange does not occur between intraplatelet and plasma fibrinogen and it appears possible that the intra-platelet fraction may be derived from the megakaryocyte. Six of nine thrombasthenic patients were found to have a severe deficiency of both adsorbed and extractable fibrinogen. Since the remaining three had near-normal platelet fibrinogen and all nine failed to aggregate it is improbable that the failure to adsorb fibrinogen is responsible for the defect in aggregation. Magnesium partially corrects adhesion to fibrin and clot retraction by these platelets, but has not been found to influence their fibrinogen adsorption. It is considered that the basic platelet surface defect, of varying severity, is responsible for the abnormalities of adsorption, aggregation, and adhesion in thrombasthenia. In the case of congenital hypofibrinogenaemia, fibrinogen transfusion corrects the long bleeding time, platelet-adsorbed fibrinogen, and the ability of platelets to spread on glass. It is possible that fibrinogen influences the surface properties of human platelets, although the final mechanism is not determined.     

Abnormalities in platelet function and morphology in a case of thrombocythemia

A patient with "life-long" bleeding problems and thrombocythemia had normal results on routine coagulation and fibrinolysis tests, but platelet function tests, including bleeding time, platelet adhesiveness, aggregation, and uptake and release of labeled serotonin were markedly altered. In addition, several types of ultrastructural abnormalities not reported previously were observed in some of the patient's platelets. The relationship between thrombocythemia, platelet dysfunction, and abnormal morphology is discussed.     

Platelet aggregation in chronic idiopathic thrombocytopenic purpura.

A method for washing platelets by albumin density gradient separation has been modified to prepare platelet rich plasma of thrombocytopenic patients for platelet aggregation studies. The concentration procedure, consisting of centrifuging platelets into a specific gravity gradient between plasma and 40-45% aqueous solution of bovine albumin, does not affect platelet aggregation adversely. Platelet aggregation in eight patients with chronic idiopathic thrombocytopenic purpura was determined by this method. On the basis of the results the patients could clearly be divided into two groups: four patients with normal aggregation and four with a qualitative platelet defect. In contrast to the other patients, the group with an in vitro platelet functional defect all had more prolonged bleeding times and the presence of a serum antiplatelet antibody.     

The role of plasma proteins and stress in the assessment of hemocompatibility.

The physiological and psychological conditions of subjects supplying blood for hemocompatibility tests significantly affect the behavior of platelets in terms of both adhesion and activation. The responses of platelets to a standard biomaterial, polyethylene (PE), were examined with blood collected from male rabbits both in basal conditions and after stress. Different media were utilized. First, platelet-rich plasma (PRP) was used to obtain a PE response to contact with platelets. Then platelets drawn from PRP were isolated and washed with Krebs-Ringer solution. One aliquot was suspended in serum (Pw-S) where fibrinogen was absent, another aliquot in Krebs-Ringer solution (Pw-KR) (in order to avoid the influence of the plasma proteins on platelets), and a third aliquot in the original plasma from which the platelets were drawn (Pw-PPP) (in order to restore the initial condition of the plasma but with washed platelets). The analysis of platelet adhesion and morphology was performed by Scanning Electron Microscopy (SEM). Differences in platelet adhesion and morphology were observed with four different media in nonstressed animals, with Pw-PPP showing a higher number and Pw-S and PW-KR lower numbers. Platelet morphology indicated low levels of activation. The platelets drawn from stressed subjects could not be counted in either PRP or PPP medium because they were fully aggregated and adhered; in contrast, in Pw-KR and Pw-S, no significant differences were found with respect to nonstressed conditions, and there was little difference in platelet morphology. All of these factors underline the role of plasma proteins, in particular fibrinogen, in the stress-induced activation of platelets.     

Effects of magnesium sulphate on leptin-dependent platelet aggregation: an ex vivo study.

Magnesium sulphate has well known antiplatelet properties. Its effect on leptin-dependent platelet aggregation has not been studied previously. Thus, we performed this ex vivo study to investigate whether magnesium sulphate is able to inhibit leptin-dependent aggregation of human platelets. We obtained platelet rich plasma (PRP) from venous blood samples of 16 healthy male volunteers, and we measured ADP-induced platelet aggregation in the presence of leptin alone (5-500 ng/mL) or leptin and magnesium sulphate (0.25-8 mM). Platelet pre-incubation with leptin led to a significant and dose-dependent increase in ADP-induced platelet aggregation. Magnesium sulphate was able to inhibit the pro-aggregating effect of leptin in a dose-dependent manner. The inhibitory effect was apparent at 1 mM of magnesium sulphate concentration (% maximal aggregation=38.1 +/- 12.2) and reached its maximum at 8 mM (% maximal aggregation=20.0 +/- 7.8). Our results demonstrate that leptin-dependent platelet aggregation is inhibited by magnesium sulphate in a dose-dependent manner. It seems conceivable that the blocking of hydrolysis of phosphoinositide and of intracellular calcium mobilization by magnesium sulphate may be involved in these findings.     

The role of Fbg in platelet adhesion to polymeric materials in conditions of psychological stress.

The effect of psychological stress on platelet adhesion to five polymeric materials (polyurethane, polyurethane filled with BaSO4, polyethyleneterephthalate, silicone and low-density polyethylene) was studied. The platelets were obtained from non-stressed and stressed rabbits as platelet-rich plasma (PRP) and, once washed (Pw), were suspended in different media, i.e. in platelet poor plasma (Pw-PPP), in serum (Pw-S) and in Krebs-Ringer solution (Pw-KR). Scanning electron microscopy of platelet adhesion and morphology revealed differences in the platelet activating power of the various materials. The washing procedure and resuspension in PPP generally resulted in an increased number of adherent platelets, compared with the number of platelets adherent to the same material in PRP. However, platelets washed and suspended in Pw-KR or Pw-S showed the same shape distribution as in PRP. When platelets from stressed rabbits were used, there was very strong aggregation and activation of the platelets in both PRP and Pw-PPP, independent of the chemical nature and surface structure of the material. In contrast, in Pw-KR and Pw-S (in which Fbg is absent) a general picture of single, not very modified platelets was observed. Their number and shapes changed according to the nature of the different materials. On the whole, the present results confirm our original hypothesis of a key role of the psychological condition of the blood donor and strongly indicate Fbg as the determinant factor in the pattern of platelet adhesion.     

Calcium-diacylglycerol guanine nucleotide exchange factor I gene mutations associated with loss of function in canine platelets

Calcium-Diacylglycerol Guanine Nucleotide Exchange Factor I (CalDAG-GEFI) has been implicated in platelet aggregation signaling in CalDAG-GEFI knockouts. Functional mutations were identified in the gene encoding for CalDAG-GEFI in 3 dog breeds. Affected dogs experienced epistaxis, gingival bleeding, and petechiation. Platelet number, von Willebrand factor, clot retraction, and coagulation screening assays were normal, whereas bleeding time tests were prolonged. Platelet aggregation and release responses to all agonists, except thrombin, were markedly impaired. Platelet membranes had normal concentrations of integrin alphaIIb-beta3; however, ADP-induced fibrinogen binding by activated platelets was markedly impaired. Forskolin-stimulated platelets exhibited a marked increase in intraplatelet cAMP associated with impaired phosphodiesterase (PDE) activity, whereas levels of extractable phosphoinositides were 1.5-fold to 2-fold higher in thrombin-stimulated affected platelets. DNA analysis of the CalDAG-GEFI gene in affected dogs documented the existence of 3 distinct mutations within portions of the CalDAG-GEFI gene encoding for structurally conserved regions within the catalytic domain of the protein. The mutations are predicted to result in either lack of synthesis, enhanced degradation, or marked impairment of protein function. The dysfunctional profile of canine platelets observed in mutant dogs putatively links CalDAG-GEFI and its target Rap1 or other Ras family member, for the first time, to a role in pathways that regulate cAMP PDE activity and thrombin-stimulated phosphoinositide anchoring or metabolism. The finding of distinct functional mutations in 3 dog breeds suggests that mutations in the CalDAG-GEFI gene may be implicated in similar defects in human patients with congenital platelet disorders having primary secretion defects of unknown etiology.     

Suppression of platelet activity on microdomain surfaces of 2-hydroxyethyl methacrylate-polyether block copolymers

Block copolymers constructed from chains of poly(2-hydroxyethyl methacrylate) (PHEMA) and either poly-ethyleneoxide (PEO) or poly-propyleneoxide (PPO) were synthesized. These block copolymers exhibited microdomain structure. Platelet adhesion on their surfaces was investigated by a column elution method to examine the effect of microdomain structure. The number of platelets adhered from whole blood was smaller for the block copolymer systems than for the homopolymers. Minimum points of platelet adhesion appeared at approximately 0.38 mol fraction of HEMA in the HEMA-PO system. Both block copolymer surfaces showed microdomains of alternate lamellar structure. Furthermore, the percent of platelets released from the column after incubation was investigated using PRP. In the case of homopolymers, released platelet percentages decreased with an increase of incubation time. Released platelet percentages from the block copolymers, however, were nearly constant with changing incubation time. These results show that HEMA-EO and HEMA-PO block copolymers had the ability to suppress both reversible and irreversible adhesion of platelets to their respective microdomain surfaces.     

Platelet and growth factor concentrations in activated platelet-rich plasma: a comparison of seven commercial separation systems

Platelet-rich plasma (PRP) is blood plasma that has been enriched with platelets. It holds promise for clinical use in areas such as wound healing and regenerative medicine, including bone regeneration. This study characterized the composition of PRP produced by seven commercially available separation systems (JP200, GLO PRP, Magellan Autologous Platelet Separator System, KYOCERA Medical PRP Kit, SELPHYL, MyCells, and Dr. Shin’s System THROMBO KIT) to evaluate the platelet, white blood cell, red blood cell, and growth factor concentrations, as well as platelet-derived growth factor-AB (PDGF-AB), transforming growth factor beta-1 (TGF-β1), and vascular endothelial growth factor (VEGF) concentrations. PRP prepared using the Magellan Autologous Platelet Separator System and the KYOCERA Medical PRP Kit contained the highest platelet concentrations. The mean PDGF-AB concentration of activated PRP was the highest from JP200, followed by the KYOCERA Medical PRP Kit, Magellan Autologous Platelet Separator System, MyCells, and GLO PRP. TGF-β1 and VEGF concentrations varied greatly among individual samples, and there was almost no significant difference among the different systems, unlike for PDGF. The SELPHYL system produced PRP with low concentrations of both platelets and growth factors. Commercial PRP separation systems vary widely, and familiarity with their individual advantages is important to extend their clinical application to a wide variety of conditions.     

Selective serotonin reuptake inhibitors: measurement of effect on platelet function

Selective serotonin reuptake inhibitors (SSRIs) reduce platelet serotonin and are associated with increased gastrointestinal bleeding, an effect that is enhanced when taken with NSAIDs or aspirin. The best method to evaluate hemorrhagic events in patients taking SSRIs has not been determined. Platelet aggregation, which is not widely available, shows SSRI inhibition of platelet function; we tested whether a platelet function analyzer could detect SSRI inhibition of platelet function. Two groups of outpatients with mood disorders were recruited; each patient was taking a stable dose of either an SSRI or bupropion for at least 6 weeks. They were tested using the platelet function analyzer-100 (PFA-100; Dade International Inc, Miami, Fla) concomitantly with platelet aggregation. Fifty-eight patients were analyzed. We detected significant differences between the groups using aggregation methods with arachidonic acid (aggregation, P = 0.00001; release, P = 0.009) and collagen (aggregation, P = 0.016; release, P = 0.006). The PFA-100 did not detect differences between the groups or results outside the reference range. The PFA-100 does not detect the inhibitory effects of SSRIs on platelet function, but it can be used to direct evaluation of bleeding in a patient taking an SSRI. Abnormal PFA-100 results suggest additional evaluation for von Willebrand disease, other platelet inhibitory medications, or underlying intrinsic platelet dysfunction.     

Use of native or platelet count adjusted platelet rich plasma for platelet aggregation measurements

BACKGROUND: It is still not clear whether native or platelet count adjusted platelet rich plasma (PRP) should be used for platelet aggregation measurements.AIM: To evaluate the necessity of using adjusted PRP in platelet function testing. METHODS: Platelet aggregation with native PRP and adjusted PRP (platelet count: 250/nl, obtained by diluting native PRP with platelet poor plasma) was performed on the Behring Coagulation Timer (BCT(R)) using ADP, collagen, and arachidonic acid as agonists. Healthy subjects, patients on antiplatelet treatment, and patients with thrombocytosis (platelet counts in PRP > 1250/nl) were investigated. RESULTS: No significant differences in the maximum aggregation response were seen when using either native or adjusted PRP from healthy subjects and patients on antiplatelet treatment. Nevertheless, some patients taking aspirin or clopidogrel showed reduced inhibition of ADP and arachidonic acid induced aggregation in adjusted PRP but not in native PRP. The maximum velocity of healthy subjects and patients on antiplatelet treatment varied significantly as a result of the degree of dilution of the adjusted PRP. Surprisingly, the BCT provided good results when measuring platelet aggregation of native PRP from patients with thrombocytosis, whereas commonly used aggregometers could not analyse platelet aggregation of native PRP in these patients. CONCLUSION: The time consuming process of PRP adjustment may not be necessary for platelet aggregation measurements. Moreover, using adjusted PRP for monitoring aspirin or clopidogrel treatment may falsify results. Therefore, it may be better to use native PRP for platelet aggregation measurements, even in patients with thrombocytosis.     

Aggregation of human platelets and adhesion of Streptococcus sanguis.

Platelet vegetations or thrombi are common findings in subacute bacterial endocarditis. We investigated the hypothesis that human platelets selectively bind or adhere strains of Streptococcus sanguis and Streptococcus mutans and aggregate, as a result, into an in vitro thrombus. Earlier ultrastructural studies suggested that aggregation of platelets over time by Staphylococcus aureus was preceded in order by adhesion and platelet activation. We uncoupled the adhesion step from activation and aggregation in our studies by incubating streptococci with platelet ghosts in a simple, quantitative assay. Adhesion was shown to be mediated by protease-sensitive components on the streptococci and platelet ghosts rather than cell surface carbohydrates or dextrans, plasma components, or divalent cations. The same streptococci were also studied by standard aggregometry techniques. Platelet-rich plasma was activated and aggregated by certain isolates of S. sanguis. Platelet ghosts bound the same strains selectively under Ca2+- and plasma-depleted conditions. Fresh platelets could activate after washing, but Ca2+ had to be restored. Aggregation required fresh platelets in Ca2+-restored plasma and was inducible by washed streptococcal cell walls. These reactions in the binding and aggregometry assays were confirmed by transmission electron microscopy. Surface microfibrils on intact S. sanguis were identified. These appendages appeared to bind S. sanguis to platelets. The selectivity of adhesion of the various S. sanguis strains to platelet ghosts or Ca2+- and plasma-depleted fresh washed platelets was similar for all donors. Thus, the platelet binding site was expressed widely in the population and was unlikely to be an artifact of membrane aging or preparation. Since selective adhesion of S. sanguis to platelets was apparently required for aggregation, it is suggested that functionally defined receptors for ligands on certain strains of S. sanguis may be present on human platelets. Some differences in the selectivity and rate of the aggregation response were noted among platelet donors, although the meaning of the variability requires further study. Nonetheless, these interactions may contribute to platelet accretion in the initiation and development of vegetative lesions in the subacute bacterial endocarditis.     

Platelet adhesion onto protein-coated and uncoated polyetherurethaneurea having tertiary amino groups in the substituents and its derivatives

Interactions of platelet with novel polyetherurethaneurea and its heparinized derivative were investigated. Platelet adhesion onto the material and release of serotonin or adenosine phosphate from platelet-rich plasma (PRP) were suppressed by an introduction of amino groups to polyetherurethaneurea, by quaternization of the polymer, and further by heparinization of the polymer. When the material was precoated with one of major plasma proteins and the protein-coated materials were taken to contact with washed platelet suspension (WP), the dependence of platelet adhesion and activation on the properties of polymers was different from that observed for PRP interaction. Platelet adhesion and activation were promoted according to the nature of coating proteins in the order albumin less than gamma-globulin less than fibrinogen and with increasing degree of denaturation of coating proteins. When the polymer materials were coated with proteins by immersing in aqueous solution containing two kinds of plasma proteins, adhesion behaviors of platelet were similar to those observed for PRP-uncoated material interaction. These experimental facts indicate that the selectivity of platelet for protein-coated material cannot be assessed by the interaction of WP with materials coated with a single kind of protein. It was concluded that material surface to which albumin is selectively adsorbed without denaturation does not stimulate adhering platelets for release reactions.     

Platelet components today

Platelet concentrates (PC) continues to be the only resource available to treat patients with qualitative and/or quantitative platelet defects. Currently we have three methods to prepare PC: platelet rich plasma (PRP), buffy coat (BC), and apheresis. In recent years, the characteristics of the products have improved significantly: greater yield of better platelets with less RBC and leucocyte content. At the present, BC and apheresis derived PC offer some advantages in comparison to PRP, as it can been prepared in additive solutions, culture them using sensitive methods or apply pathogen reduction technologies.     

Optimized preparation method of platelet-concentrated plasma and noncoagulating platelet-derived factor concentrates: maximization of platelet concentration and removal of fibrinogen

Platelet-rich plasma (PRP) has been clinically used as an easily prepared growth factor cocktail that can promote wound healing, angiogenesis, and tissue remodeling. However, the therapeutic effects of PRP are still controversial, due partly to the lack of optimized and standardized preparation protocols. We used whole blood (WB) samples to optimize the preparation protocols for PRP, white blood cell-containing (W-PRP), platelet-concentrated plasma (PCP), and noncoagulating platelet-derived factor concentrate (PFC). PRP and W-PRP were most efficiently collected by 10 min centrifugation in a 15-mL conical tube at 230-270 g and 70 g, respectively. To prepare PCP, platelets were precipitated by centrifugation of PRP at >2300 g, 90% of supernatant plasma was removed, and the platelets were resuspended. For preparation of noncoagulating PFC, the supernatant was replaced with one-tenth volume of saline, followed by platelet activation with thrombin. Platelet (before activation) and platelet-derived growth factor (PDGF)-BB (after activation) concentrations in PCP were approximately 20 times greater than those in WB, whereas PFC contained a 20-times greater concentration of platelets before platelet activation and a 50-times greater concentration of PDGF-BB without formation of a fibrin gel after platelet activation than WB. Surprisingly, total PDGF-BB content in the PFC was twice that of activated WB, which suggested that a substantial portion of the PDGF-BB became trapped in the fibrin glue, and replacement of plasma with saline is crucial for maximization of platelet-derived factors. As an anticoagulant, ethylene di-amine tetra-acetic acid disodium inhibited platelet aggregation more efficiently than acid citrate dextrose solution, resulting in higher nonaggregated platelet yield and final PDGF-BB content. These results increase our understanding of how to optimize and standardize preparation of platelet-derived factors at maximum concentrations.     

Platelet volume parameters as a diagnostic tool: The influence of anticoagulation and storage conditions on platelet impedance volume

Summary Rapid progress has been made in the design of aperture impedance cell counters, and parameters such as mean platelet volume and platelet distribution width have become routinely available to most physicians. Platelet volume is influenced by both platelet production in the bone marrow and platelet activation or sequestration in the circulation. In thrombocytopenic patients, it is often possible to differentiate between megakaryocytic and amegakaryocytic disease states on the basis of platelet volume analysis. In patients with thrombocytosis, a myeloproliferative disorder may be suspected if the platelet distribution width is high. However, the conditions of sample preparation and storage still give rise to considerable inaccuracy in the determination of platelet volume parameters. In this study, platelet impedance volume was strongly influenced by anticoagulation, storage time, and incubation temperature. Changes in platelet volume were more pronounced in whole blood than in platelet rich plasma. However, mainly large platelets were lost during the preparation of platelet rich plasma. Collecting blood directly into a mixture of citrate and low dose glutaraldehyde stabilized platelet volume for up to 2 h after venipuncture at room temperature. This method reduces platelet volume changes in vitro and is in this respect superior to the usual EDTA blood count or the use of platelet-inhibitory agents.Abbreviations CO₂ Carbon dioxide - EDTA Ethylene diamine tetraacetic acid - GTA Glutaraldehyde - LDH Lactate dehydrogenase - MPV Mean platelet volume - PRP Platelet rich plasma - WB Whole blood     

Spontaneous platelet aggregation in a hereditary giant platelet syndrome (MPS)

The characteristics of spontaneous platelet aggregation (SPA) in a hereditary giant platelet syndrome (Montreal platelet syndrome, MPS) are examined. SPA was quantitated by microscopy from the decrease in single platelets in platelet-rich plasma (PRP). In contrast to normal donors, a significant proportion (20-50%) of platelets in MPS whole blood and PRP occurred in microaggregates typically containing 2-6 disk-shaped platelets. Stirring MPS-PRP at 1000 rpm for 10 minutes further increased the fraction of platelets in aggregates by 10-170%, the percentage increase not being correlated to the donor's platelet count (5000-220,000 microliters-1). Normal platelets resuspended in MPS platelet-poor plasma (PPP) did not undergo SPA, whereas MPS platelets resuspended in normal PPP or Ca2+-free, fibrinogen-free Tyrode's continued to show SPA. The increase in SPA could be inhibited by 10 microM prostaglandin (PG) E1, 150 mM ASA or glutaraldehyde or formaldehyde fixation; however, it was not inhibited by 10 nM PGI2 and was only partially inhibited by 1 microM 2-chloroadenosine and 1-10 units/ml apyrase. SPA in Acid-citrate-dextrose-PRP was much less than in PRP; however, SPA reoccurred on returning the platelets to platelet-free plasma or Tyrode's. Platelet aggregation (PA) could be increased over that due to SPA alone by the addition of adenosine diphosphate, adrenaline, collagen, ionophore A-23187, arachidonic acid and ristocetin, with results suggesting that the response to these agents is normal. The ristocetin-induced increase in PA was completely blocked by an IgG specific for Bernard-Soulier syndrome. In contrast, MPS platelets had a reduced sensitivity to thrombin, which appeared to be more pronounced at low platelet counts. There was no correlation between the thrombin insensitivity and the extent of SPA. Total adenosine triphosphate (ATP) and thrombin-induced release of ATP and platelet factor 4 appeared normal for MPS platelets. The ultrastructural features of MPS platelets were within normal limits except for an increased frequency of giant granules. SPA was observed for 5/5 MPS donors, but only one of three MPS donors' platelets evaluated for glycoprotein I and sialic acid content showed any measurable reduction as compared with normal controls. The above observations point to the existence of an as yet undetermined anomaly of MPS plasma membrane related to a fibrinogen and Ca2+ independent form of platelet aggregation.