Differences in the expression of cell cycle genes in osteoblasts and endothelial cells cultured on the surfaces of Ti6Al4V and Ti6Al7Nb alloys.

Three medically used alloys (Ti6Al4V, Ti6Al7Nb, and AISI 316 L) are compared due to proliferative potential and metabolic response of human cells (osteoblasts line Saos-2 and endothelial cells line EA.hy-926) seeded on the surfaces of these alloys. Although no statistically significant difference in the proliferative potential of the cells cultured on the surfaces of examined biomaterials was observed, it does not exclude relevant differences in metabolic response of these cells assessed as changes in genes' expression. As a result of our studies it was demonstrated that the changes in the expression of examined genes were very common. Our observation suggests the presence of the process of selective recognition of the contacted biomaterials by the cells seeded on their surfaces. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 1607-1617, 2017.

Adhesion, activation, and aggregation of blood platelets and biofilm formation on the surfaces of titanium alloys Ti6Al4V and Ti6Al7Nb.

Titanium alloys are still on the top list of fundamental materials intended for dental, orthopedics, neurological, and cardiovascular implantations. Recently, a special attention has been paid to vanadium-free titanium alloy, Ti6Al7Nb, that seems to represent higher biocompatibility than traditional Ti6Al4V alloy. Surprisingly, these data are not thoroughly elaborated in the literature; particularly there is a lack of comparative experiments conducted simultaneously and at the same conditions. Our study fills these shortcomings in the field of blood contact and microbiological colonization. To observe platelets adhesion and biofilm formation on the surfaces of compared titanium alloys, fluorescence microscope Olympus GX71 and scanning electron microscope HITACHI S-3000N were used. Additionally, flow cytometry analysis of platelets aggregation and activation in the whole blood after contact with sample surface, as an essential tool for biomaterial thrombocompatibility assessment, was proposed. As a result of our study it was demonstrated that polished surfaces of Ti6Al7Nb and Ti6Al4V alloys after contact with whole citrated blood and E. coli bacterial cells exhibit a considerable difference. Overall, it was established that Ti6Al4V has distinct tendency to higher thrombogenicity, more excessive bacterial biofilm formation and notable cytotoxic properties in comparison to Ti6Al7Nb. However, we suggest these studies should be extended for other types of cells and biological objects.

A new recombinant thrombolytic and antithrombotic agent with higher fibrin affinity--a staphylokinase variant. I. In vitro study.

We attempted to construct a new recombinant protein characterized by fibrin-specific properties of plasminogen activation combined with antithrombin and antiplatelet activities. To the C-terminal part of recombinant staphylokinase (r-SAK), which is a promising profibrinolytic agent, we assembled: (i) the Kringle 2 domain (K2) of tissue-type plasminogen activator (t-PA), containing a fibrin-specific binding site, (ii) the RGD sequence (Arg-Gly-Asp) for the prevention of platelet aggregation and (iii) the antithrombotic agent - hirudin. The cDNA for hybrid protein SAK-RGD-K2-Hir was cloned into pESP-3 yeast protein expression vector. The introduction of K2 t-PA, RGD sequence and hirudin into r-SAK molecule did not alter the SAK activity. The plasminogen activation rate (determined by K(M) and K(cat)) of SAK-RGD-K2-Hir was not significantly different from that of r-SAK. Affinity and binding strength of the recombinant protein to fibrin immobilized on the biosensor were higher than to r-SAK. We observed a higher clot lysis potency of SAK-RGD-K2-Hir as evidenced by a faster and more profound lysis of 125I-labeled human fibrin clots. The potency of thrombin inhibition by the hirudin part of the recombinant fusion protein SAK-RGD-K2-Hir was the same as that of r-Hir alone. In conclusion, the results of the in vitro study suggest that the SAK-RGD-K2-Hir construct can be a more potent and faster-acting thrombolytic agent with antithrombin and antiplatelet properties compared with standard r-SAK.

Calmodulin effect on purified rat cortical plasma membrane Ca(2+)-ATPase in different phosphorylation states.

The plasma membrane Ca(2+)-ATPase in neuronal tissue plays an important role in fine tuning of the intracellular Ca(2+) concentration. The enzyme exhibits a high degree of tissue specificity and is regulated by several mechanisms. Here we analysed the relationship between separate modes of Ca(2+)-ATPase regulation, i.e., reversible phosphorylation processes mediated by protein kinases A and C, protein phosphatases PP1 and PP2A, and stimulation by calmodulin. The activity of PKA- or PKC-phosphorylated Ca(2+)-ATPase was influenced by the further addition of calmodulin, and this effect was more pronounced for PKC-phosphorylated calcium pump. In both cases the fluorescence study revealed the increased calmodulin binding, and for PKA-mediated phosphorylation it was correlated with a higher affinity of calcium pump for calmodulin. The incubation of Ca(2+)-ATPase with CaM prior to protein kinases action revealed that CaM presence counteracts the stimulatory effect of PKA and PKC. Under the in vitro assay cortical Ca(2+)-ATPase was a substrate for PP1 and PP2A. Protein phosphatases decreased both the basal activity of Ca(2+)-ATPase and its affinity for calmodulin. Fluorescence analysis confirmed the lowered ability of dephosphorylated Ca(2+)-ATPase for calmodulin binding. These results may suggest that interaction of CaM with calcium pump and its stimulatory action could be a partly separate phenomenon that is dependent on the phosphorylation state of Ca(2+)-ATPase.

Acute phase protein alpha 1-acid glycoprotein interacts with plasminogen activator inhibitor type 1 and stabilizes its inhibitory activity.

alpha(1)-Acid glycoprotein, one of the major acute phase proteins, was found to interact with plasminogen activator inhibitor type 1 (PAI-1) and to stabilize its inhibitory activity toward plasminogen activators. This conclusion is based on the following observations: (a) alpha(1)-acid glycoprotein was identified to bind PAI-1 by a yeast two-hybrid system. Three of 10 positive clones identified by this method to interact with PAI-1 contained almost the entire sequence of alpha(1)-acid glycoprotein; (b) this protein formed complexes with PAI-1 that could be immunoprecipitated from both the incubation mixtures and blood plasma by specific antibodies to either PAI-1 or alpha(1)-acid glycoprotein. Such complexes could be also detected by a solid phase binding assay; and (c) the real-time bimolecular interactions monitored by surface plasmon resonance indicated that the complex of alpha(1)-acid glycoprotein with PAI-1 is less stable than that formed by vitronectin with PAI-1, but in both cases, the apparent K(D) values were in the range of strong interactions (4.51 + 1.33 and 0.58 + 0.07 nm, respectively). The on rate for binding of PAI-1 to alpha(1)-glycoprotein or vitronectin differed by 2-fold, indicating much faster complex formation by vitronectin than by alpha(1)-acid glycoprotein. On the other hand, dissociation of PAI-1 bound to vitronectin was much slower than that from the alpha(1)-acid glycoprotein, as indicated by 4-fold lower k(off) values. Furthermore, the PAI-1 activity toward urokinase-type plasminogen activator and tissue-type plasminogen activator was significantly prolonged in the presence of alpha(1)-acid glycoprotein. These observations suggest that the complex of PAI-1 with alpha(1)-acid glycoprotein can play a role as an alternative reservoir of the physiologically active form of the inhibitor, particularly during inflammation or other acute phase reactions.

Comparison of platelet aggregability and P-selectin surface expression on platelets isolated by different methods.

Three methods commonly used for isolation of blood platelets from plasma were compared. Platelets were isolated by: 1) a washing method; 2) a method of metrizamide-gradient centrifugation; 3) a modified method of gel-filtration. The last method employed BSA-Sepharose gel instead of routinely used Sepharose gel saturated with BSA. BSA-Sepharose gel was prepared by covalent binding of thermally deactivated BSA to CNBr-activated Sepharose 2B. In contrast to platelets isolated by the other methods, an aggregability of the gel-filtered platelets and control platelets in plasma, both activated with ADP, were comparable. When expression of P-selectin on the surface of freshly isolated platelets was examined, the gel-filtered platelets exhibited the same extent of fluorescence signal as platelets in the citrated blood, whereas platelets isolated by the other methods exhibited twice the extent of the signal. The methods involving the centrifugation process cause a low but a significant platelet activation.

Partial purification and identification of cardiodepressant factor from the posterior pituitary lobe in rats.

It has been demonstrated previously that the cardiodepressant activity is present in the bovine hypothalamic extract and in the medium incubating the rat's posterior pituitary lobe "in situ". In this study medium incubating the posterior pituitary lobe was fractionated by a low pressure gel filtration procedure and the cardiodepressant fractions were pooled and further purified by the HPLC technique on C8 and TSK 3000 SW columns. It was shown, on the basis of mass spectrometry, that cardiodepressant activity is associated with substance(s) with molecular mass of about 500 d. Application of this fraction into the fluid used for incubation of isolated right auricle of the right heart atrium of a two-day-old rat, strongly decreased the frequency of spontaneous discharge of the pacemaker tissue.

Release of calcium and P-selectin from intraplatelet granules is hampered by procaine.

The inhibition of platelets by some local anaesthetics has been related to the modulation of platelet membrane lipid fluidity, and one of these compounds, procaine, has been proven to be particularly effective inhibitor. In the present study, we examined the effect of procaine on the mobilization of intracellular granule contents in isolated washed platelets. We revealed that the presence of 10 mg/ml procaine significantly hampered platelet release reaction, as demonstrated by the significant reduction in the expression of platelet P-selectin (CD62) on one hand, and significantly enhanced expression of GPIb alpha (CD42b) antigen on the other, following either 1 hour incubation of washed platelets at room temperature (%CD62: 37.1+/-6.8% of control incubated without procaine, p<<0.0001; %CD42b: 116.2+/-6.3% of control, p<0.0001) or activation of whole blood platelets with ADP, TRAP, or thrombin. Procaine, which acted as a rigidizer, significantly decreased platelet membrane fluidity (ESR h(+1)/h0 ratio of 5-DOXYL-Ste reduced down to 93.1+/-3.7% of control, p<0.001). In washed Fura-2-loaded platelets procaine not only brought about the significantly reduced Ca2+ release from intraplatelet storage pools after platelet stimulation with 15 micromol/l ADP (25.3+/-12.5% of control, p<0.001), but also it significantly increased the reduction in Ca2+ concentration upon the addition of Ca2+ chelator, EDTAK2 (48.9+/-13.5% vs. 40.9+/-12.1% of initial [Ca2+]i concentration, p(1,alpha)<0.025). Overall, procaine considerably reduced calcium mobilization from intraplatelet storage pools and Ca2+ efflux across platelet membrane. Based on these data, we suggest that the preventive effects of procaine on platelet release reaction and calcium mobilization might relate to the changes in the organization of membrane components embedded into a lipid bilayer, which are crucial in triggering of platelet release reaction. Procaine-mediated dislocations of some membrane components and/or distortion of lipid-protein interactions could generate a steric hindrance, which might interfere with platelet signal transduction, thus leading to impaired mobilization of Ca2+ and other components from intraplatelet storage pools.

Platelet membrane lipid fluidity and intraplatelet calcium mobilization in type 2 diabetes mellitus.

The aim of the present study was to relate the impairments in calcium mobilization and/or release to the altered membrane dynamics in platelets from patients with type 2 diabetes mellitus. Higher expression of P-selectin (1.4-fold, NS) and the reduction in GPIb alpha expression (by 27.8+/-16.7%, p < 0.0002), as well as the increased fractions of platelet microparticles (p < 0.03), reflected more intensified platelet release reaction in diabetic platelets. Overall, diabetic platelets appeared more vulnerable to stimuli facilitating calcium mobilization (by 41%, p < 0.01) and less susceptible to preventive effects of the agents hampering calcium release from intraplatelet storage pools (by 38%, p < 0.01). Both the increased calcium mobilization from intraplatelet storage pools and higher levels of intracellular free calcium in the presence of procaine in diabetic platelets correlated with the reduced platelet membrane lipid fluidity (resp. pR < 0.03 and pR < 0.015). We conclude that the biophysical state of platelet membrane components in diabetes mellitus is the crucial determinant of platelet hyperfunction and probably contributes to the intensified calcium mobilization in diabetic platelets. The depressed preventive effects of procaine on platelet release reaction and calcium mobilization in diabetic platelets may result from the primary dislocations and/or distortions of membrane components caused by the diabetic state.

Microplate reader--a convenient tool in studies of blood coagulation.

Results of platelet aggregation measured with a dual channel aggregometer and with a microplate reader are compared. Platelets in plasma were activated by ADP and by collagen, and thrombin was used for the aggregation study of gel filtered platelets. The results obtained with both instruments were quantitatively and qualitatively similar. A microplate reader allowed a simultaneous measurement of a high number of samples with a high degree of reproducibility. The same instrument can be useful in other coagulology studies. Results of citrated plasma clotting by thrombin or by recalcination together with results of platelet counting, both obtained with a microplate reader, are presented in this report as well.

Platelet membrane fluidity and intraplatelet Ca2+ mobilization are affected in uraemia.

In present investigations, platelet membrane fluidity and intraplatelet Ca2+ mobilization were analysed in uraemic platelets by fluorescence techniques. Thirteen non-dialyzed uraemic patients and 16 control subjects were examined. Anisotropy of DPH-probe, measured at 37 degrees C, was significantly higher in control (0.2236 +/- 0.0050) than in uraemic platelets (0.1969 +/- 0.0082; p < 0.01). There was no difference between control (109.8 +/- 6.0 nM) and uraemic platelets (100.0 +/- 7.3 nM) when the basal [Ca2+]i in resting platelets was determined. Activation of platelets by ADP (12.5 microM) or by thrombin (0.1 U/ml) resulted in an increase in [Ca2+]i. It was significantly higher (p* < 0.003 for ADP and p* < 0.009 for thrombin, respectively) in control platelets (383.6 +/- 56.3 nM and 2031.0 +/- 298.8 nM, respectively) than in uraemic ones (191.0 +/- 21.3 nM and 838.7 +/- 144.1 nM, respectively). The amount of released Ca2+ was higher in control platelets activated by both ADP and thrombin (157.6 +/- 21.4 nM and 409.3 +/- 71.0 nM, respectively) than in uraemic platelets (76.7 +/- 15.7 nM and 203.0 +/- 29.3 nM, respectively) and the differences were significant (p < 0.01 and p* < 0.01, respectively). These results indicate an abnormal intracellular Ca2+ mobilization in uraemic platelets. Both increased membrane fluidity and decreased Ca2+ mobilization should be considered as a possible reason of reduced fibrinogen receptor exposure on uraemic platelets.

A new variant of the viral haemorrhagic disease of rabbits virus.

A new variant of viral haemorrhagic disease of rabbits (VHD) virus, recently detected in Poland and called Blaszki (BLA), gives positive results in enzyme-linked immunosorbent assay (ELISA) and exhibits viral protein of 60 kilodaltons (VP 60), as detected by Western blot analysis. This BLA variant of VHD virus has caused high morbidity and mortality in rabbits, as have other reported variants with similar clinical signs and pathological lesions, but-in contrast to other variants-the BLA variant gave negative results in the haemagglutination test. This development indicates the limitations of haemagglutination testing in the diagnosis of VHD.

Platelet membrane fluidity and intraplatelet Ca2+ homeostasis are affected in uremia.

Recently we have described a dependence of platelet disability in thrombosis upon the progression in renal failure and an elevated level of RGDS-containing degradation products in uremic plasma, which is also correlated with progression in renal failure. Based on fluorescence techniques, our present investigations concerned possible changes in platelet membrane fluidity and intraplatelet calcium homeostasis in uremic platelets. Washed platelets loaded with DPH or with Fura-2 were examined with LS-50 luminescence spectrometer. Light anisotropy of DPH measured at 37 degrees C was significantly higher in control platelet membranes than in uremic ones. It can be considered as more fluidic membranes of uremic platelets. No difference between the basal intraplatelet calcium level was found for uremic and control platelets, but in the presence of 5 mM EGTA, the basal level was reduced significantly deeper in uremic platelets. Activation of platelets by both ADP (12.5 microM) and thrombin (0.1 U/ml) resulted in rapid increase in the intraplatelet calcium level in the examined platelets, but this increase was significantly higher for control platelets. The results indicate an abnormal intracellular calcium homeostasis in uremic platelets, which is associated with an increased fluidity of platelet membranes in uremia.

Changes in platelet response to adenosine 5'-diphosphate caused by verapamil.

Verapamil is widely used in the treatment of patients with coronary artery disease. The effect of verapamil on vascular smooth muscle cells is well documented. This effect is mediated by the inhibition of calcium fluxes across plasma membranes. Some data suggest that verapamil may affect platelet functions in thrombosis, but those observations were made for much higher verapamil concentration than could be achieved in vivo. Our current investigations are focused on an influence of low doses of verapamil (0.1-1.0 microM) on platelet response to ADP. We have found that verapamil at concentration of 0.1 microM can inhibit platelet aggregation (by 10%) evoked in PRP by 1.0-1.5 microM ADP. Moreover, the inhibitory effect is potentiated by prolonged time of platelet preincubation with verapamil. On the other hand, we have found a significant reduction in the number of fibrinogen receptors exposed on the platelet surface of patients (n = 21) treated with therapeutic doses (240 mg/day) of verapamil during two weeks of drug administration. The mean number of exposed receptors was reduced from 75,000 to 40,000 per platelet, with significance p < 0.0001. In vitro platelet preincubation with verapamil, even in much higher concentrations, did not affect fibrinogen binding to ADP activated platelets. It suggests, that in vitro exposure of platelets to verapamil for a short time has no effect on the expression of fibrinogen receptors on platelets, but prolonged in vivo interaction of this drug with platelets results in reduction of the fibrinogen receptor exposition. Thus, observed inhibition of platelet aggregation does not relay on a simple reduction of the number of exposed receptors, but intraplatelet signalling has to be affected. In fact, we have observed, in platelets pretreated with low doses of verapamil, significantly reduced release of calcium ions upon activation by ADP, whereas the calcium influx under such conditions does not seem to be affected.

Diabetes mellitus alters the effect of peptide and protein ligands on membrane fluidity of blood platelets.

The increased nonenzymatic glycosylation of platelet membrane proteins has been suggested to underlie platelet hypersensitivity in diabetes and the relationship of this to the reduced membrane lipid fluidity has been reported. As the modulation in membrane fluidity may determine the degree of accessibility of membrane receptors, the consequent alterations in membrane lipid-protein interactions in diabetes mellitus may also underlie the differentiated effects of various thrombotic and fibrinolytic agents on platelet membrane lipid bilayer. In the present study we employed electron paramagnetic resonance and fluorescence spectroscopy to explore the ligand-induced platelet membrane fluidity changes in diabetic state, i.e. under conditions when the membrane architecture is considerably altered. The yield of the excimer formation of pyrenemaleimide (PM), which depends directly upon the collisional rate and distances between molecules, was elevated in diabetic platelet membranes, thus pointing to the occurrence of some constraints in the structure/conformation of platelet membrane proteins in diabetes mellitus. Such an immobilization of PM was accompanied by the significant elevation in membrane protein glycation in diabetic platelets. The effects of various interacting ligands on platelet membrane fluidity were significantly lower in diabetic platelets, and the differences were much more distinct at the lower depths of a lipid bilayer. Nevertheless, the alterations in membrane lipid fluidity observed upon the interaction of a given ligand occurred with an approximately equal frequency in control and diabetic platelets. Moreover, the probability that these alterations were less profound in diabetic platelets was the same for all types of ligands studied. In diabetic patients the interaction of RGDS and tissue-type plasminogen activator with platelet membranes resulted in much smaller reductions of the h+/h0 parameters in 5-DOXYL-Ste acid-labelled platelets, thus indicating a lesser rigidization of membrane lipid bilayer in diabetes. Likewise, the fluidizing effect of both fibrinogen itself and fibrinogen-derived peptides containing gamma-chain carboxy-terminal sequence H-12-V was less pronounced in diabetic platelet membranes.

Does reduced membrane lipid fluidity underlie the altered thrombin-induced expression of integrin α(IIb)ß(3) and PADGEM-140 in membranes of platelets from diabetic juveniles?

In diabetic patients, where the membrane lipid microviscosity of blood platelets is altered, the availability of platelet membrane receptors may change concomitantly. Platelet hypersensitivity in diabetic subjects was previously hypothesized to result from the nonenzymatic glycosylation-induced loss in platelet membrane fluidity. In our present study juvenile type 1 diabetic subjects were compared with their relevant controls with respect to thrombin-stimulated platelet activation in relation to glycation-induced impairments of platelet membrane dynamics. Our results indicate that: (a) the mean steady-state fluorescence polarization (p) of both 1,6-diphenyl-1,3,5-hexatriene (DPH) and 1-anilino-8-naphthalenesulphonate (ANS) in membranes from diabetic subjects were significantly greater than for control subjects, thus indicating reduced membrane lipid fluidity in diabetic platelets in various membrane regions; (b) the significantly higher [(3)H]NaBH(4) reduction, indicating the increased attachment of glucose to protein amino groups, was attributed to the proteins extracted from diabetic platelet membranes; (c) CD62-positive resting platelets were not significantly more abundant in diabetic patients; (d) basically, unaltered amounts of PADGEM-140 membrane antigen (CD62) copies were detected in resting diabetic platelets; (e) significantly higher numbers of membrane glycoprotein ß(3) were found in diabetic platelets; (f) thrombin-induced elevations in the expression of CD61 (ß(3)) and CD62 (PADGEM-140) occurred to much higher extent in platelets of diabetic patients, thus pointing to more profound activation of diabetic platelets by thrombin; (g) the total amounts of platelet membrane glycoprotein ß(3) was significantly reduced in platelet lysates from diabetic subjects. We conclude that glycation-induced rigidization of platelet membranes might hypersensitize diabetic platelets to aggregating agents by rendering platelet membrane receptors more exposed to the external environment. Thus, thrombin may bind more efficiently to the exposed glycoprotein receptors (due to glycation) in diabetic platelets. Such excessive exposure and displacements toward the external environment might favour the accelerated shedding of some membrane proteins in diabetic platelets. We further suggest that their subsequent replacements would render platelet intrinsic storage pools exhausted and thus, might explain the diminished total amount of ß(3) found in platelets of diabetic patients.

Stimulation of erythrocyte membrane Mg(2+)-ATPase activity by glutathione S-conjugates.

Pig erythrocyte membrane Mg2+-ATPase activity was stimulated by various glutathione S-conjugates. For alkyl S-conjugates, the Km for the stimulation was lower, the more hydrophobic was the conjugate. 2,4-Dinitrophenyl-S-(N-acetyl)cysteine also stimulated the Mg2+-ATPase activity, suggesting a low specificity of the ¿glutathione S-conjugate pump¿. The Km values for the stimulation by 2,4-dinitrophenyl conjugates were lower than predictable on the basis of hydrophobicity which indicates a high affinity of the transporter for these conjugates.

Concentration of RGDS-containing degradation products in uremic plasma is correlated with progression in renal failure.

A concentration of protein degradation products containing the RGDS sequence, which could contribute to a lower reactivity of uremic platelets, has been estimated in both uremic (n = 16) and control (n = 7) plasmas. Degradation products and other small molecules were separated from plasma by filtration through AMICON YM-10 filter. RGDS antigen was determined in filtered material using the radioimmunoassay method based on monospecific anti-RGDS rabbit polyclonal antibodies. The concentration of RGDS-containing degradation products in uremic plasma ranged from 0.8 to 353 nM with mean value 58.6 +/- 24.9 nM and was higher than in control (0.7 to 5.9 nM, mean value 2.1 +/- 0.9 nM). Moreover, the level of RGDS-antigen positively correlated with plasma creatinine concentration (R = 0.87, p < 0.001). The filtered material showed an inhibitory effect on fibrinogen binding to control platelets in respect to RGDS-antigen concentration. We conclude that the elevated concentration of RGDS-containing degradation products in uremic plasma is partially responsible for bleeding tendency in renal failure.

Expression of fibrinogen receptors on platelets of uremic patients is correlated with the content of GPIIb and plasma level of creatinine.

Platelets of uremic patients, activated with ADP, exposed less fibrinogen receptors than control platelets, i.e. 24612 +/- 5541 and 33400 +/- 4302 receptors per platelet, respectively. However, this difference was not statistically significant. When compared with the total number of GPIIb/IIIa complexes, quantified from platelet glycoprotein IIb (GPIIb) contents, active receptors on the platelet surface represented 13.6% and 35.1% of total pool of fibrinogen receptors in uremic and control platelets, respectively. The number of exposed fibrinogen receptors was positively correlated with the amount of GPIIb copies in both uremic and normal platelets. In uremic platelets, both the number of exposed receptors and the number of GPIIb copies were correlated with the plasma creatinine concentration suggesting, that binding of fibrinogen to uremic platelets depends upon the degree of renal failure. Uremic platelets contain similar amounts of fibrinogen as control ones i.e. 13.2 +/- 2.3 micrograms and 17.6 +/- 2.2 micrograms per 1 x 10(8) platelets, respectively. Whereas for beta-thromboglobulin (beta-TG) there was a significant difference of 392 +/- 102 ng and 803 +/- 202 ng per 1 x 10(8) platelets, respectively. Reduced beta-TG content in uremic platelets suggests limited platelet activation in vivo. These results support the concept that uremic platelets have impaired functions and indicate that there is a relationship between the progression in renal failure and disability of platelets in thrombosis.