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ABSTRACT: The adhesion of cells is normally studied in the absence of flow. In a classic setup, ELISA wells are coated with an adhesive
protein and subsequently incubated with a cell suspension. The gravity is responsible for the transport of the cells to the
adhesive surface. This setup is not appropriate for blood cells, such as platelets, because platelet adhesion occurs physiologically
under flow conditions. The shear stress induced by the flow causes qualitative and quantitative differences in adhesion (1). Blood platelets have a relatively low density and do not settle easily under gravity. Under flow conditions, convective
diffusion of platelets to the vessel wall is responsible for a strong increase of platelet number near the reactive surface,
resulting in a strongly enhanced platelet adhesion. A second important effect of flow is mediated by the shear stresses exerted
on the platelets. To overcome these stresses, a special protein, the von Willebrand factor, is necessary, which has a very
high affinity for the platelet membrane receptor glycoprotein Ib. In the absence of flow, the presence of von Willebrand factor
is not necessary for platelet adhesion, but with increasing shear rates, the presence of von Willebrand factor becomes essential.
Another disadvantage of studying platelet adhesion under static conditions is that owing to the presence of red blood cells,
these experiments must be performed with washed platelets, and the required centrifugation steps cause unwanted platelet activation.
03/2008: pages 159-170;
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ABSTRACT: Vessel wall injury induces the formation of a haemostatic plug. Restoration of vascular integrity should involve cessation of further platelet and fibrin deposition and subsequent removal of these thrombi by both the fibrinolytic system and proteases delivered by infiltrating inflammatory cells. We hypothesized that adhesion of platelets and inflammatory cells [polymorphonuclear leucocyte (PMN)] to fibrin is differently supported after exposure of fibrin during fibrinolysis. Fibrin surfaces were exposed to fibrinolytic agents, and platelet and PMN adhesion was studied under conditions of flow. Specific adhesion of platelets to preformed fibrin was reduced by fibrinolytic treatment of the fibrin. PMN adhesion to fibrin was only slightly affected even after 180 min exposure to plasmin. With fibrin still present after fibrinolytic treatment, the impaired platelet adhesion seems explained by loss of the primary platelet adhesion site gamma400-411 on fibrin. PMN binding to fibrin clearly depends on other sites that are less degraded by fibrinolysis. We have shown that PMN adhesion in flowing blood to lysed fibrin was still present, whereas platelet adhesion was impaired due to the loss of the primary platelet adhesion site gamma400-411. Based on our in-vitro perfusion model, we conclude that fibrinolysis specifically interferes with the thrombogenicity of fibrin in the haemostatic plug, whereas the inflammatory response is preserved. The latter may participate in the long-term removal and restructuration of the plug.
Blood Coagulation and Fibrinolysis 08/2006; 17(5):421-4. · 1.24 Impact Factor
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ABSTRACT: Acute thrombotic arterial occlusion is the leading cause of morbidity and mortality in the Western world. Von Willebrand factor is thought to be the only indispensable adhesive substrate to promote thrombus formation in high shear environments. We found that thrombospondin-1, a glycoprotein enriched in arteriosclerotic plaques, might function as an alternative substrate for thrombus formation. Platelets adhered to thrombospondin-1 in a shear dependent manner with an optimum shear as found in stenosed arteries. Adhesion is extremely firm, with no detachment of platelets up to a shear rate of 4000 s(-1). Experiments using platelets from a patient completely lacking von Willebrand factor showed that von Willebrand factor is not involved in platelet binding to thrombospondin-1. Platelet adhesion to thrombospondin-1 is not mediated via beta3-integrins or GPIa. CD36 partially mediates the adhesion of pre-activated platelets. We identified GPIb as high shear adhesion-receptor for thrombospondin-1. Soluble GPIb, as well as antibodies against the GPIb, blocked platelet adhesion almost completely. The new discovered thrombospondin-1-GPIb adhesion axis under arterial shear conditions might be important, not only during thrombus formation but also for pathological processes where other cells bind to the endothelium or subendothelium, including arteriosclerosis, inflammation and tumor metastasis, and a promising therapeutic target.
The FASEB Journal 09/2003; 17(11):1490-2. · 5.71 Impact Factor
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ABSTRACT: Thrombopoietin (TPO) is known to sensitize platelets to other agonists at 20 ng/ml, and above 100 ng/ml it is an independent activator of aggregation and secretion. In studies with a perfusion chamber, TPO, between 0.01 ng/ml and 1 ng/ml, increased platelet adhesion to surface-coated fibrinogen, fibronectin and von Willebrand Factor (VWF) but not to a collagen-coated surface. Increased adhesion was observed at shear rates of 300/s and 800/s in perfusions with whole blood as well as in suspensions of platelets and red blood cells reconstituted in plasma. The by the cyclooxygenase inhibitor, indomethacin, and the thromboxane A2-receptor blocker, SQ30741, abolished the stimulation by TPO. The effect of TPO was mimicked by a very low concentration (10 nmol/l) of the thromboxane TxA2 analogue, U46619. Real-time studies of platelet adhesion to a VWF-coated surface at a shear of 1000/s showed that about 20% of the platelets were in a rolling phase before they became firmly attached. TPO (1 ng/ml) pretreatment reduced this number to < 5%, an effect again abolished by indomethacin. Thus, TPO potentiates the direct and firm attachment of platelets to surface-coated ligands for alphaIIbbeta3, possibly by increasing the ligand affinity of the integrin.
British Journal of Haematology 05/2003; 121(3):482-90. · 4.94 Impact Factor
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ABSTRACT: The multimeric glycoprotein von Willebrand factor (VWF) mediates platelet adhesion to collagen at sites of vascular damage. The binding site for collagen types I and III is located in the VWF-A3 domain. Recently, we showed that His(1023), located near the edge between the "front" and "bottom" faces of A3, is critical for collagen binding (Romijn, R. A., Bouma, B., Wuyster, W., Gros, P., Kroon, J., Sixma, J. J., and Huizinga, E. G. (2001) J. Biol. Chem. 276, 9985-9991). To map the binding site in detail, we introduced 22 point mutations in the front and bottom faces of A3. The mutants were expressed as multimeric VWF, and binding to collagen type III was evaluated in a solid-state binding assay and by surface plasmon resonance. Mutation of residues Asp(979), Ser(1020), and His(1023) nearly abolished collagen binding, whereas mutation of residues Ile(975), Thr(977), Val(997), and Glu(1001) reduced binding affinity about 10-fold. Together, these residues define a flat and rather hydrophobic collagen-binding site located at the front face of the A3 domain. The collagen-binding site of VWF-A3 is distinctly different from that of the homologous integrin alpha(2) I domain, which has a hydrophilic binding site located at the top face of the domain. Based on the surface characteristics of the collagen-binding site of A3, we propose that it interacts with collagen sequences containing positively charged and hydrophobic residues. Docking of a collagen triple helix on the binding site suggests a range of possible engagements and predicts that at most eight consecutive residues in a collagen triple helix interact with A3.
Journal of Biological Chemistry 05/2003; 278(17):15035-9. · 4.77 Impact Factor
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ABSTRACT: Transient interactions of platelet-receptor glycoprotein Ibalpha (GpIbalpha) and the plasma protein von Willebrand factor (VWF) reduce platelet velocity at sites of vascular damage and play a role in haemostasis and thrombosis. Here we present structures of the GpIbalpha amino-terminal domain and its complex with the VWF domain A1. In the complex, GpIbalpha wraps around one side of A1, providing two contact areas bridged by an area of solvated charge interaction. The structures explain the effects of gain-of-function mutations related to bleeding disorders and provide a model for shear-induced activation. These detailed insights into the initial interactions in platelet adhesion are relevant to the development of antithrombotic drugs.
Science 09/2002; 297(5584):1176-9. · 31.20 Impact Factor
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ABSTRACT: Transient interactions of platelet-receptor glycoprotein Ibα (GpIbα) and the plasma protein von Willebrand factor (VWF) reduce
platelet velocity at sites of vascular damage and play a role in haemostasis and thrombosis. Here we present structures of
the GpIbα amino-terminal domain and its complex with the VWF domain A1. In the complex, GpIbα wraps around one side of A1,
providing two contact areas bridged by an area of solvated charge interaction. The structures explain the effects of gain-of-function
mutations related to bleeding disorders and provide a model for shear-induced activation. These detailed insights into the
initial interactions in platelet adhesion are relevant to the development of antithrombotic drugs.
Science 08/2002; 297(5584):1176-1179. · 31.20 Impact Factor
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ABSTRACT: Evidence from animal experiments suggests that endothelial cell activation plays a pathogenetic role in the development of cerebral ischemia after subarachnoid hemorrhage (SAH). We measured plasma concentrations of two markers of endothelial cell activation, i.e., ED1-fibronectin (ED1-fn) and von Willebrand factor (vWf), among patients with aneurysmal SAH. We analyzed the relationships of concentrations to initial clinical conditions, treatment modalities, and the occurrence of delayed cerebral ischemia.
We collected 123 blood samples from 27 patients with aneurysmal SAH. Aneurysms were treated surgically in 19 cases, were treated endovascularly in 7 cases, and remained untreated in 1 case. Twelve patients developed symptomatic delayed cerebral ischemia.
Initial concentrations of ED1-fn (4.3 +/- 3.7 microg/ml) and vWf (17.8 +/- 8.2 microg/ml) were higher than the reference values (ED1-fn, 1.7 +/- 0.9 microg/ml, P < 0.001; vWf, 11.5 +/- 5.2 microg/ml, P = 0.003). Concentrations were higher among patients in poor clinical condition at admission, compared with patients in good clinical condition (mean difference, ED1-fn, 5.7 microg/ml, P = 0.04; vWf, 10.4 microg/ml, P = 0.02). Levels of both markers increased significantly after surgery (mean increase, ED1-fn, 7.5 microg/ml, P = 0.01; vWf, 13.2 microg/ml, P = 0.05) and after ischemic episodes (mean increase, ED1-fn, 8.3 microg/ml, P = 0.02; vWf, 5.0 microg/ml, P = 0.04).
Plasma concentrations of markers of endothelial cell activation were increased early after SAH and were significantly associated with the clinical condition at admission. We also observed a significant increase in concentrations after surgery and after ischemic episodes. Whether endothelial cell activation is a causal or indirectly related factor in the pathogenesis of delayed cerebral ischemia after SAH is still uncertain.
Neurosurgery 06/2002; 50(6):1223-9; discussion 1229-30. · 2.79 Impact Factor
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ABSTRACT: The interaction of platelets with fibrinogen is a key event in the maintenance of a haemostatic response. It has been shown that the 12-carboxy-terminal residues of the gamma-chain of fibrinogen mediate platelet adhesion to immobilized fibrinogen. These studies, however, did not exclude the possibility that other domains of fibrinogen are involved in interactions with platelets. To obtain more insight into the involvement of other domains of fibrinogen in platelet adhesion, we studied platelet adhesion in flowing blood to patient dysfibrinogen Vlissingen/Frankfurt IV (V/FIV), to several variant recombinant fibrinogens with abnormalities in the gamma-chain segments gamma318-320 and gamma408-411. Perfusion studies at physiological shear rates showed that platelet adhesion was absent to gammaDelta408-411, slightly reduced to the heterozygous patient dysfibrinogen V/FIV and strongly reduced to the homozygous recombinant fibrinogens: gammaDelta319-320, gamma318Asp-->Ala and gamma320Asp-->Ala. Furthermore, antibodies raised against the sequences gamma308-322 and gamma316-333 inhibited platelet adhesion under shear conditions. These experiments indicated that the overlapping segment gamma316-322 contains amino acids that could be involved in platelet adhesion to immobilized fibrinogen under flow conditions. In soluble fibrinogen, this sequence is buried inside the fibrinogen molecule and becomes exposed after polymerization. In addition, we have shown that this fibrin-specific sequence also becomes exposed when fibrinogen is immobilized on a surface.
British Journal of Haematology 06/2002; 117(3):650-7. · 4.94 Impact Factor
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ABSTRACT: The interaction of platelets with fibrinogen is a key event in the maintenance of a haemostatic response. It has been shown that the 12-carboxy-terminal residues of the γ-chain of fibrinogen mediate platelet adhesion to immobilized fibrinogen. These studies, however, did not exclude the possibility that other domains of fibrinogen are involved in interactions with platelets. To obtain more insight into the involvement of other domains of fibrinogen in platelet adhesion, we studied platelet adhesion in flowing blood to patient dysfibrinogen Vlissingen/Frankfurt IV (V/FIV), to several variant recombinant fibrinogens with abnormalities in the γ-chain segments γ318–320 and γ408–411. Perfusion studies at physiological shear rates showed that platelet adhesion was absent to γΔ408-411, slightly reduced to the heterozygous patient dysfibrinogen V/FIV and strongly reduced to the homozygous recombinant fibrinogens: γΔ319-320, γ318Asp→Ala and γ320Asp→Ala. Furthermore, antibodies raised against the sequences γ308–322 and γ316–333 inhibited platelet adhesion under shear conditions. These experiments indicated that the overlapping segment γ316–322 contains amino acids that could be involved in platelet adhesion to immobilized fibrinogen under flow conditions. In soluble fibrinogen, this sequence is buried inside the fibrinogen molecule and becomes exposed after polymerization. In addition, we have shown that this fibrin-specific sequence also becomes exposed when fibrinogen is immobilized on a surface.
British Journal of Haematology 05/2002; 117(3):650 - 657. · 4.94 Impact Factor
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ABSTRACT: ADP plays a central role in regulating platelet function. It induces platelet aggregation via the activation of 2 major ADP receptors, P2Y(1) and P2Y(12). We have investigated the role of P2Y(12) in platelet adhesion and thrombus formation under physiological flow by using blood from a patient with a defect in the gene encoding P2Y(12). Anticoagulated blood from the patient and from healthy volunteers was perfused over collagen-coated coverslips. The patient's thrombi were smaller and consisted of spread platelets overlying platelets that were not spread, whereas control thrombi were large and densely packed. Identical platelet surface coverage, aggregate size, and morphology were found when a P2Y(12) antagonist, N(6)-(2-methylthioethyl)-2-(3,3,3-trifluoropropylthio)-beta,gamma-dichloromethylene ATP (also known as AR-C69931 MX), was added to control blood. The addition of a P2Y(1) antagonist (adenosine-3',5'-diphospate) to control blood resulted in small, but normally structured, thrombi. Thus, the ADP-P2Y(12) interaction is essential for normal thrombus buildup on collagen. The patient's blood also showed reduced platelet adhesion on fibrinogen, which was not due to changes in morphology. Comparable results were found by using control blood with AR-C69931 MX and also with adenosine-3',5'-diphospate. This suggested that P2Y(12) and P2Y(1) were both involved in platelet adhesion on immobilized fibrinogen, thereby revealing it as ADP dependent. This was confirmed by complete inhibition on the addition of creatine phosphate/creatine phosphokinase.
Arteriosclerosis Thrombosis and Vascular Biology 05/2002; 22(4):686-91. · 6.37 Impact Factor
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ABSTRACT: In order to study the functional importance of the collagen, heparin and glycoprotein-Ib-binding domain, we deleted the A1 domain of von Willebrand factor (vWF), corresponding to residues 478–716, by oligonucleotide-directed mutagenesis. The resulting ΔA1-vWF cDNA was expressed in COS-1 monkey kidney cells and compared to wild-type vWF. The higher-molecular-mass multimers were decreased in ΔA1 recombinant von Willebrand factor (ΔA1-rvWF) compared to plasma vWF and rvWF. The reactivity of ΔA1-rvWF and rvWF with monoclonal antibodies directed against the collagen-binding domain (residues 969–992), the vessel-wall-binding domain, and the binding site for glycoprotein IIb-III a on platelets was identical. The interaction with vWF of the monoclonal antibody directed against the glycoprotein Ib binding domain was abolished for ΔA1-rvWF, and similar to plasma vWF for rvWF. The binding of factor VIII to ΔA1-rvWF and rvWF was similar. ΔA1-rvWF and rvWF bound similarly to collagen, but the binding of ΔA1-rvWF to heparin and to platelets in the presence of ristocetin were abolished. These data indicate that the heparin-binding site in the A1 domain is essential. There is no second binding domain for glycoprotein Ib outside the A1 domain. The collagen-binding domain in the A1 domain is either not active or its action can be compensated by the second collagen-binding domain.
European Journal of Biochemistry. 02/1991; 196(2):369 - 375.
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ABSTRACT: Endothelial cells synthesize and store von Willebrand factor. We have studied the storage and secretion of von Willebrand factor in cultured human umbilical vein endothelial cells. In particular, we were interested in the nature of the storage compartment and the effects of perturbation on the storage and secretion processes. The storage compartment for von Willebrand factor was isolated from homogenates of endothelial cells. By an immunostaining technique the isolated vesicles stained for von Willebrand factor. The staining pattern was similar to that of Weibel-Palade bodies in intact endothelial cells. We concluded that the storage compartment containing von Willebrand factor is identical to the Weibel-Palade body. The von Willebrand factor of the isolated storage vesicles is predominantly constructed of polypeptide chains with a Mr of 220 kD. On the other hand, von Willebrand factor continuously secreted by endothelial cells is constructed of both a 220 kD and a larger precursor (apparent Mr of 275 kD) subunit. The storage vesicles contain von Willebrand factor that supports ristocetin-induced platelet aggregation. Thus, endothelial cells store fully processed, biologically active von Willeband factor within Weibel-Palade bodies. Short-term ( < 1 h) treatment of endothelial cells with the perturbing phorbol ester 4β-phorbol-12-myristate-13-acetate (PMA) results in release of cellular stored von Willebrand factor. 24–48 h after exposure to PMA the endothelial cell distribution of von Willeband factor is changed distinctly. While the contents of the von Willebrand factor storage sites in the cells are gradually restored within 48 h, enhanced amounts of von WiUebrand factor are secreted into the medium. The number as well as the size of von Willebrand factor storage granules in the endothelial cells increase after exposure to phorbol ester, as determined by immunofiuorescence microscopy. Phorbol ester treated cells release stored von Willebrand factor 48 h after they have been stimulated. PMA decreases the von Willebrand factor contents of the extracellular matrix; the deposition of von Willebrand factor in the subendothelium is blocked by PMA, whereas the degradation of matrix von Willebrand factor is not affected. Thus, perturbation of endothelial cells changes the cellular distribution of von Willebrand factor.
Pathophysiology of Haemostasis and Thrombosis 08/1970; 18(4-6):246-261. · 2.23 Impact Factor
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ABSTRACT: 1.1.|Human platelet 6-phosphofructokinase (ATP:d-fructose 6-phosphate 1-phosphotransferase, EC 2.7.1.11) was partially purified. The final preparation had a specific activity of 7.1 μmoles Fru-1,6-P2 formed per min per mg protein at 25 °C.2.2.|SO42− activated the enzyme activity especially at low Fru-6-P concentrations. This effect is due to diminished cooperativity with respect to Fru-6-P.3.3.|SO42− diminished the allosteric inhibition by ATP. Other nucleotide phosphates caused no inhibition when tested in the presence of 3 mM SO42−. In the absence of SO42− CTP also inhibited enzyme activity.
Biochimica et Biophysica Acta (BBA) - Enzymology.
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ABSTRACT: Administration of Ticlopidine to human volunteers resulted in a prolonged bleeding time and decreased or absent aggregation of platelets with collagen and epinephrine. Adenosine diphosphate (ADP) induced platelet aggregation was initiated by a normal shape change, but the rate of the first wave of aggregation had decreased. The second wave of aggregation was absent. ADP-binding to platelets of volunteers, consisted of two classes of binding sites:one with high affinity and one with low affinity, giving a curvilinear and a linear part in a concentration dependency curve. After Ticlopidine, the low affinity part of the curve had disappeared. Evidence will be presented that this is a specific Ticlopidine effect.
Thrombosis Research.
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ABSTRACT: 1.1. Human platelets were isolated by gel filtration on Sepharose 2B. The suspension was concentrated in a dialysis bag place in polyethyleneglycol 20 000.2.2. In these cells levels of glycolytic intermediates were measured. Caculations of mass action ratiosn for various glycolytic reactions indicated that the reaction catalyzed by 6-phosphofructokinase was far displace from equilirium while those catalyzed by phosphoglucose isomerase, aldolase and triosephosphate isomerase approached equilibrium. This observation holds for platelets isolated in Ca2+ buffer as well as for cells isolated in Ca2+-free tyrode. The platelets in the former buffer contained five times less Fru-1,6-P2 and 2–3 times less dihydroxyacetone-P than cells in the later.3.3. Platelets in Ca2+ buffer accumulated Fru-1,6-P2 and dihydroxyacetone-P upon addition of collagen. Accumulation was maximal after about 10 min and decreased to control values again after about 30 min. Lactate accumulated faster than in paralle controls, reflecting an increased glycolytic flux during platelet-collagen interaction. In contrast platelets in Ca2+-free tyrode showed no accumulation of Fru-1,6-P2 and dihydroxyacetone-P after collagen addition. In several suspensions these levels were even lower than in parallel controls. Differences were most pronounced after 10–20 min, but diminished thereafter. Glycolytic flux was not increased in these platelets and in several suspensions lactate production was even lower than in controls.4.4. It is concluded that the variations in Fru-1,6-P2 concentration reflect the activity of phosphofructokinase. Acute changes in Fru-1,6-P2 levels parallel alterations in glycolytic flux, but flux itself is not related to the absolute Fru-1,6-P2 concentration in the cell. Whether phosphofructokinase is activated or inhibited depends on the suspending medium which effects platelet metabolism before and/or during the interaction with collagen.
Biochimica et Biophysica Acta (BBA) - General Subjects.
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ABSTRACT: In the reaction catalyzed by human platelet phosphofructokinase, Mg2+ is required for optimal activity. Maximal MG2− activation was obtained at [Mg2+] = [MgITP2−] or higher. At high MgATP2− concentrations there is an increase in the allosteric inhibition by ATP4−.
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ABSTRACT: Human platelet 6-phosphofructokinase (EC 2.7.1.11) shows cooperativity towards Fru-6-P and is allosterically inhibited by high Mg . ATP2− concentrations. No relation could be demonstrated between the cooperativity towards Fru-6-P and the inhibition by Mg . ATP2−. Increasing the concentrations of Mg . ATP2− only raised the apparent Km values for Fru-6-P, but did not change the Hill constants. a possible formation of a Mg . ATP2− : enzyme . Fru-6-P complex during catalysis was investigated. Our calculations suggest that such a ternary complex is indeed formed during the reaction.
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ABSTRACT: Study objective – Balloon angioplasty produces mechanical vessel wall injury that leads to substantial blood platelet deposition at the angioplasty site. The aim of the study was to determine whether thermal angioplasty might, by contrast, reduce platelet adhesion by denaturation of subendothelial adhesive proteins. Design – Native and cultured human subendothelium was briefly heated to ≥ 100°C by laser irradiation or to 50-100°C by immersion in preheated phosphate buffered saline (PBS). Subsequently, the subendothelium was exposed for 5 min to flowing human blood at a shear rate of 1300 s−1. Blood platelet adhesion to the subendothelium was determined quantitatively. Experimental material — Human umbilical arteries were used and the subendothelial matrix derived from cultured umbilical vein endothelial cells. Measurements and results – After heating arterial subendothelium by laser irradiation to ≥100°C, zero platelet adhesion was found v 36(SD 2)% adhesion to the non-heated surface (p<0.00l). After laser heating of the subendothelial matrix to ≥100°C, platelet adhesion was absent in 10/10 experiments (p < 0.01). After heating the matrix to 100°C by immersion in PBS, platelet adhesion was reduced to 5(5)% v 31(7)% at 37°C (p<0.00l). Conclusions – These in vitro results, if extrapolated to catheter interventions, suggest that thermal injury to the vessel wall by laser angioplasty or other thermal angioplasty methods may provide a basic and clinically relevant advantage over mechanical angioplasty modalities, because of a potentially reduced risk of complications related to platelet adhesion.
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ABSTRACT: 1.1. The relation between energy production and energy consumption was studied in resting human platelets. Platelets were gel filtered in a medium with 1 mM CN− and without glucose and then incubated at 37°C for 60 min. Platelet glycogen was converted into lactate at a rate of 0.79 ± 0.23 (mean ± S.D., n = 6) μmol glycosyl residues/min per 1011 cells. This conversion was linear with time for 40 min and then ceased abruptly. During a subsequent 20 min of incubation, glycogen and lactate levels remained constant, indicating a halt in energy production; the cells remained intact during this period.2.2. During the first 40 min, 14C-labeled ATP and adenylate energy charge also decreased linearly with time. Thereafter, constant levels were maintained for another 20 min.3.3. Addition of glucose stimulated lactate production which originated primarily from endogenous glycogen at 0–50 μM glucose but which at higher glucose concentrations, originated predominantly from glucose.4.4. At lactate production rates of 3 μmol/min per 1011 cells, a stable ATPm concentration and adenylate energy charge were maintained. At a lower flux, both parameters decreased and a linear relationship was observed between the decrease in ATPm level and adenylate energy charge versus lactate formation. When the lactate production was 1 μmol lactate formed/min per 1011 cells, the adenylate energy charge fell 0.008 unit/min and ATPm level fell 1.1% of total radioactivity/min.5.5. From these data, the balance between production and consumption of ATPm was calculated. A decrease in production was almost completely matched by a decrease in consumption. A slight discrepancy remained which increased as the rate of ATPm formation fell. When ATPm formation was 2 μmol/min per 1011 cells consumption exceeded production by 15 nmol ATPm/ min per 1011 cells. The slight excess of ATPm consumption over production at suboptimal rates of ATPm production provides a tool for feedback control of ATPm-consuming processes on glycolysis and glycogenolysis.
Biochimica et Biophysica Acta (BBA) - General Subjects.
