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Biomedical subjects

A Ordinas

Publications and source records attributed to A Ordinas.

At least 91 records · Page 5Linked to original sources
PubMed1996

Analysis of platelet adhesion to a collagen-coated surface under flow conditions: the involvement of glycoprotein VI in the platelet adhesion.

Platelet adhesion to the exposed surface of the extracellular matrix in flowing blood is the first and critical reaction for in vivo thrombus formation. However, the mechanism of this in vivo platelet adhesion has yet to be studied extensively. One of the reasons for this is the lack of a practical assay method for assessing platelet adhesion under flow conditions. We have devised an assay method (the fluorescent adhesion assay) that is based on the technique originally reported by Hubbell and McIntire (Biomaterials 7:354, 1986) with some modifications to make it more amenable for assaying small samples and have developed an analysis method to quantify the extent of platelet adhesion and aggregation from fluorescence images by using a computer-assisted image analysis system. In our assay, platelet adhesion, expressed as the percentage of the area covered by adhered platelets, was found to increase biphasically as a function of time. In the first phase, platelets interacted with the coated collagen, transiently stopping on the surface; we called this reaction the temporary arrest. In the second phase, platelets adhered much more rapidly and permanently on the surface, and this adhesion was dependent on the shear rate; platelets formed aggregates in this phase. We used our assay to analyze the effects of platelet aggregation inhibitors on platelet adhesion. All three examined inhibitors, EDTA (10 mmol/L), antiglycoprotein (GP) IIb/IIIa, and GRGDS peptide (1 mmol/L), inhibited the second phase adhesion in flowing blood. Furthermore, GPVI-deficient platelets also showed defective second-phase adhesion under the same conditions. These results suggested that GPIIb/IIIa activation and GPVI contribute to the reaction inducing the second phase. The second-phase adhesion has been extensively investigated, and the consensus is that this reaction is mainly attributable to the platelet-platelet interaction. In this report, we were able to detect an earlier reaction, the temporary arrest. This temporary arrest would reflect the fast and weak interaction between platelet GPIb/IX and collagen-von Willebrand factor complexes on the collagen-coated surface.

Blood Platelets↗
PubMed1996

Effects of G-CSF administration and peripheral blood progenitor cell collection in 20 healthy donors.

The effects of both daily G-CSF administration and subsequent peripheral blood progenitor cell collection (PBPCC) by apheresis on 20 healthy adult donors were studied. All received daily G-CSF (filgrastim) 10 micrograms/kg for 5-7 days by subcutaneous injection. G-CSF administration was well tolerated, except for moderate bone pain and headache. Peak values of CD34+ cells were observed on days 5 (n = 12) or 6 (n = 8). In all donors a significant increase in CD3+, CD4+, CD8+, CD19+, and NK cells was observed on day 5 in relation to the baseline values. CD4/CD8 lymphocyte ratio was unmodified by G-CSF. None of the donors required a central venous line for PBPCC. Immediately after PBPCC, a platelet count below 100 x 10(9)/1 was observed in nine of 18 cases, although in all donors platelet counts were over 100 x 10(9)/1 7 days later. A lymphocytopenia on day 7 following PBPCC was observed, although there was a tendency to achieve baseline values 30-90 days after the procedure. Mean numbers ( +/- SD) of collected cells x 10(6)/kg after a median of two (1-4) apheresis sessions and a median of 20 1 (10-40) processed were: CD34+ 5.5 ( +/- 2.3), CD3+ 326 ( +/- 105), CD4+ 207 ( +/- 64), CD8+ 164 ( +/- 60), CD19+ 88 ( +/- 32), and NK cells 32 ( +/- 14). We conclude that G-CSF administration to healthy donors is a well-tolerated procedure which is associated with (a) obtaining a high number of hematopoietic progenitor cells, and (b) a significant increase in T, B, and NK cells in donors' blood. In addition, PBPCC by apheresis results in a moderate, rapidly reversible, and clinically irrelevant thrombocytopenia and a moderate lymphocytopenia, which tends to resolve within 3 months following the procedure.

Adolescent↗
PubMed1996

Increased induced monocyte tissue factor expression by plasma from patients with severe ovarian hyperstimulation syndrome.

OBJECTIVE: To investigate whether monocyte expression of tissue factor is increased by plasma from patients with severe ovarian hyperstimulation syndrome (OHSS). DESIGN: Prospective longitudinal study. SETTING: Assisted Reproduction Unit of the Hospital Clínic i Provincial in Barcelona, a tertiary care setting. PATIENTS: Nine IVF patients with severe OHSS. INTERVENTIONS: Pretreatment with leuprolide acetate was followed by gonadotropin treatment for ovarian follicular stimulation. After administration of hCG, a standard IVF-ET procedure was performed. MAIN OUTCOME MEASURES: Measurement during the syndrome and 4 to 5 weeks after recovery of induced monocyte tissue factor expression. RESULTS: In each of the nine patients, plasma obtained during the syndrome induced a significantly higher proportion of monocytes expressing tissue factor and a significantly higher intensity of tissue factor expression on monocytes than plasma obtained after recovery and control plasma. CONCLUSIONS: Procoagulant activity of blood monocytes, which is mediated principally by tissue factor expression, is increased in patients with severe OHSS. This fact may be important in thrombotic events associated with the syndrome.

Adult↗
PubMed1996

Clinical and laboratory factors associated with platelet transfusion refractoriness: a case-control study.

In recent years clinical factors have largely surpassed alloimmunization as the predominant cause of platelet refractoriness. This makes it necessary to properly identify and weigh the non-immune factors that have a major impact of refractoriness. A case-control study is suitable for such an analysis, and to our knowledge has not previously been performed to assess this issue. Fifty-two refractory patients were compared with 52 control patients who were transfused at the same time. Only one transfusion event was analysed per patient. Clinical and laboratory data were recorded at the time of selected transfusion, and their association with refractoriness was investigated by the contingency table method and the Cox stepwise logistic regression. There were 16 (31%) patients with HLA antibodies in the index group and only one in the control group. The corrected count increment in the group of patients refractory due to HLA antibodies was significantly lower than that in non-alloimmunized refractory patients [median (range): 48.5 (-3560, 4614) and 4058 (-4417, 6886), respectively; U = 493, P < 0.0001]. In the multivariate analysis, factors associated with refractoriness were the presence of HLA antibodies (odds ratio (OR) 50.7; 95% CI 5.5-463); fever (odds ratio 7.2; 95% CI 2.5-21) and BMT because of chronic myeloid leukaemia (odds ratio 7.3; 95% CI 1.8-30). The latter two were the only factors that remained independently associated with refractoriness after excluding alloimmunized patients and their controls. We conclude that HLA antibodies are strongly associated with platelet transfusion refractoriness, but account for less than a third of these patients. Fever and BMT because of chronic myeloid leukaemia were the only non-immune factors independently associated with refractoriness.

Adolescent↗
PubMed1996

Activated protein C inhibits thrombus formation in a system with flowing blood.

We studied the effect of increasing concentrations of protein C (PC) and activated protein C (APC) on haemostasis in an in vitro thrombosis model. Blood from healthy donors was anticoagulated with citrate-phosphate-dextrose (final citrate concentration 19 mM) or a low molecular weight heparin (LMWH, 20 IU/ml). Enzymatically denuded rabbit aorta segments were exposed to flowing blood for 10 min in an annular perfusion chamber. PC and APC were added to the perfusate immediately prior to exposure. In citrated blood at a shear rate of 800/s, PC and APC induced a statistically significant decrease in platelet deposit at 16 micrograms/ml and 32 micrograms/ml. In perfusions performed with blood anticoagulated with LMWH, there was no effect on platelet deposition at 16 and 32 micrograms/ml either at shear rates of 300/s or 800/s. Addition of PC showed no effect on fibrin deposition at a shear rate of 300/s; in contrast, a nonstatistically significant 40% reduction was seen at a shear rate of 800/s, compared to controls. Addition of APC caused a 100% reduction in fibrin formation at 16 and 32 micrograms/ml at both shear rates studied. PC and APC inhibited platelet deposition on the exposed subendothelial surface, in a dose-dependent manner. Effects of PC and APC on platelet function might be mediated through inhibition of thrombin generation at the platelet microenvironment.

Animals↗
PubMed1996

Red blood cell aggregability is increased by aspirin and flow stress, whereas dipyridamole induces cell shape alterations: measurements by digital image analysis.

We evaluated the influence of pretreatment with aspirin in vitro, alone or combined with dipyridamole, on red cell aggregability. Samples were tested before and after being exposed to well-defined flow conditions. An aggregation rate was estimated through digital analysis of light microscopy images. Red cells of untreated blood that had been exposed to flow showed a higher aggregation rate (38.54 +/- 1.63 vs. 27.52 +/- 1.36; P < 0.05). This effect was not observed in the absence of platelets. Treatment with aspirin induced a high aggregation rate, with or without exposure to flow conditions. Dipyridamole alone or combined with aspirin provoked echinocytosis, disturbing the rouleaux arrangement (rates ranging from 11.25 +/- 1.91 - 13.62 +/- 1.62). Washing red cells after treatment restored about 90% of echinocytes to their biconcave shape, but aggregation rate did not recover in parallel. These results highlight the influence of red cell-platelet interactions in the regulation of haemostasis and show how therapeutic agents can interfere with rheological phenomena.

Aspirin↗
PubMed1996

Influence of the red blood cell preparation method on the efficacy of a leukocyte reduction filter.

The performance of a leukocyte reduction bedside filter with different types of RBC concentrates was analyzed. Three types of RBCs were prepared: buffycoat-depleted RBCs suspended in saline-adenine-glucose-mannitol (SAGM)-additive solution (BC-RBCs; n = 20), RBCs suspended in SAGM-additive solution without buffy coat removal (SAGM-RBCs; n = 20), and RBCs drawn in CPDA-I conservative solution and processed for component preparation by the platelet-rich plasma method (CPDA-RBCs; n = 20). The units were filtered within 8 h of collection. One filter was used for every 2 units. High numbers of residual WBCs were found even in the units filtered first. Filtration of CPDA-RBCs resulted in a higher residual WBC content than SAGM-RBCs or BC-RBCs (p = 0.0032 and p = 0.0002, respectively). The filter performance strikingly decreased when the WBC load per filter exceeded 4 x 10(9) or the platelet load was less than 100 x 10(9). We conclude that filter performance varies with the WBC and platelet content of the RBC concentrates. Under the experimental conditions assayed in this study CPDA-RBCs are the least appropriate ones to be used for bedside leukocyte reduction.

Blood Transfusion↗
PubMed1996

Procoagulant effect of incompatible platelet transfusions in alloimmunized refractory patients.

The clinical effectiveness of platelet transfusion in refractory patients is still a subject of debate. We have evaluated the possible hemostatic effect of platelet transfusion in 16 alloimmunized thrombocytopenic patients whose platelet counts were less than 20,000/microliters. Platelet concentrates were always obtained by apheresis procedures from incompatible donors. The posttransfusion platelet recovery was greater than 15% only in 3 cases. In the first 6 patients, measurements of bleeding time performed immediately before transfusion were in all cases longer than 30 min and did not change significantly 10 and 60 min after platelet transfusions. In all patients, ex vivo perfusion experiments with Baumgartner's platelet adhesion model, using native nonanticoagulated blood, were performed immediately before and 10 and 60 min after transfusion. No difference in platelet deposition onto the subendothelial surface was observed after platelet transfusion. Unexpectedly, the deposition of fibrin on the subendothelial surface was statistically augmented in the posttransfusion studies. Quantification of thrombin-antithrombin complexes (TAT) in plasma showed statistically significant elevations (p < 0.01) in the posttransfusion samples (31.9 +/- 12.6 vs. baseline 5.8 +/- 1.7 ng/ml), not justified by TAT levels in the transfused material (2.3 +/- 0.17 ng/ml). Transfusion of incompatible platelets to refractory patients may activate coagulation mechanisms in the absence of an increase in peripheral platelet count.

Blood Group Incompatibility↗
PubMed1996

Existence of a platelet-adhesion defect in patients with cirrhosis independent of hematocrit: studies under flow conditions.

A defect in hemostasis has been repeatedly reported in patients with cirrhosis. However, the nature of this defect has not been fully characterized. We explored adhesive and cohesive functions of platelets from cirrhotic patients at different stages of disease development. The response of platelets to standard activating agents was tested by aggregometric procedures. The interaction of platelets with subendothelial components was explored in a perfusion system in which blood was exposed (shear rate, 800/s; 10 minutes) to denuded segments of rabbit aorta. Platelet interactions in these perfusions were analyzed morphometrically. Results were always compared with those obtained in similar studies using blood obtained from healthy subjects. Aggregation studies showed abnormal responses for single or several agonists. Abnormalities in aggregation were more evident in patients with severe disease (Child-Pugh class C), although they occasionally were abnormal for single agonists (ADP or U46619) in patients with less severe disease (Child-Pugh classes A or B). All the patient classes showed impaired platelet-subendothelial interactions (P < .01 vs. healthy subjects) that were not justified by the relative thrombocytopenia present in the more severely affected patients. Experimental increases in hematocrit in patients at stages B and C did not improve platelet-subendothelial interactions. Platelets from cirrhotic patients interact defectively with subendothelial components under flow conditions. The adhesion defect is more evident and consistent than the aggregation defects and may already be present in patients with mild liver failure. This adhesion defect may contribute to the defective hemostasis observed in cirrhotic patients.

Adult↗
PubMed1996

Redistribution of membrane glycoproteins in platelets activated under flow conditions.

A reduction in the ability of GPIb to bind specific MoAbs or ligands (vWF) has been reported in platelets exposed to thrombin in suspension. We have analyzed modifications in the presence of glycoproteins (GPs) on platelets activated under flow conditions in a system which allows limited thrombin and fibrin generation. Normal blood anticoagulated with low molecular weight heparin (LMWH, Dalteparin 20 IU/ml) was recirculated for up to 10 min at 800 s-1 through annular chambers containing denuded arterial segments. Aliquots of blood were removed from the reservoir at 0, 1, 5 and 10 min and immediately mixed with paraformaldehyde. Membrane glycoproteins: GPIb (CD42b), GPIIb-IIIa (CD41a), GPIV (CD36); and activation dependent antigens: P-selectin (CD62P) and lysosomal glycoprortein (CD63), were detected in whole blood by dual color flow cytometry. Circulation of through the perfusion system resulted in platelet activated as demonstrated by the increased percentage of platelets positive for antigens CD62P and CD63. A gradual increase in the binding of MoAbs directed against GPIb, GPIIb-IIIa, and GPIV epitopes was noted during the entire perfusion period. Observed differences in mean fluorescence intensities at all the observation times were statistically significant (P < 0.001). Our results obtained on platelets in an experimental thrombosis system indicate that GPIb, GPIIb-IIIa and GPIV remain on the surface of activated platelets and actually increase their expression. Alterations detected at the level of GPIb in platelets activated by thrombin in suspension may not take place under in vivo situations.

Antibodies, Monoclonal↗
PubMed1996

Platelet sensitivity to prostaglandin E1 inhibition is reduced in pre-eclampsia but not in nonproteinuric gestational hypertension.

OBJECTIVE: Platelet aggregometry was used to discriminate platelet sensitivity to prostaglandin E1 (PGE1) inhibition, to evaluate whether platelet behaviour in pre-eclamptic women was different in this respect than from that in nonproteinuric hypertensive women. METHODS: The amount of PGE1 required to inhibit in vitro platelet aggregation induced by arachidonic acid was determined in samples from 60 women: 20 nonpregnant controls, 20 women with normal pregnancies, 10 women with gestational hypertension and 10 with pre-eclampsia. RESULTS: The response to arachidonic acid was similar among the four groups. Amounts of PGE1 necessary to inhibit platelet aggregation were significantly higher in normal pregnant women compared with nonpregnant controls (P<0.001). Platelets from pre-eclamptic women required significantly higher concentrations of PGE1 to inhibit aggregation than the other groups studied (P<0.001). However, there was no significant difference between normal and nonproteinuric hypertensive pregnant women. CONCLUSIONS: Our findings support the notion that increased platelet reactivity during late pregnancy is exacerbated in pre-eclamptic women but not in nonproteinuric hypertensive women. This is in agreement with the hypothesis that pre-eclampsia and gestational hypertension are different conditions. Prospective studies are required to confirm if this simple test may be useful in the early identification of pregnant women at risk for pre-eclampsia.

Adult↗
PubMed1996

Hypercoagulable state in patients with antiphospholipid syndrome is related to high induced tissue factor expression on monocytes and to low free protein s.

Antiphospholipid antibodies (aPLs) are associated with thrombosis, but the mechanisms of this thrombotic tendency are unknown. We studied 56 patients 612 with systemic lupus erythematosus [SLE] and aPLs and previous thrombosis, 12 with SLE and aPLs but no thrombosis, 15 with SLE without aPLs or thrombosis, 11 with primary antiphospholipid syndrome with thrombosis, and 6 asymptomatic subjects with aPLs) to investigate the ability of aPLs to induce tissue factor (TF) expression on human normal monocytes. A double direct immunofluorescence technique (anti-CD14 and anti-TF) was used, and procoagulant activity in viable and disrupted cells was measured after plasma incubation for 6 hours at 37 degrees C with normal mononuclear cells. Hemostasis regulatory proteins, prothrombin fragment 1 + 2, and thrombin-antithrombin III complex levels were determined. Increased TF expression and procoagulant activity were observed using plasma samples from SLE patients with aPLs and thrombosis (P < .01) and from primary antiphospholipid syndrome patients (P < .01) but not from patients with SLE and aPLs but no thrombosis, patients with SLE without aPLs, or asymptomatic patients with aPLs. Purified aPL immunoglobulins from one primary antiphospholipid syndrome and two SLE patients added to normal plasma showed a significant increase in both TF expression and procoagulant activity (P < .05) compared with purified aPL from two SLE patients without thrombosis. The addition of nonspecific IgG from three SLE patients without aPLs and from three control subjects did not increase TF expression. Low free protein S was seen in eight patients. Increased TF expression and low free protein S correlated with thrombosis (P < .01) and with higher prothrombin fragment 1 + 2 and thrombin-antithrombin III values (P < .01). These observations may contribute to a further understanding of the thrombotic risk in aPL patients.

Antiphospholipid Syndrome↗
PubMed1996

Antibodies to CD36 (GPIV) inhibit platelet adhesion to subendothelial surfaces under flow conditions.

The membrane glycoprotein CD36 (glycoprotein [GP] IV) has previously been shown to accelerate the initial interaction of platelets with purified type I collagen in both static and flow systems. In the present study, the role of CD36 on platelet interaction with physiologically relevant collagenous surfaces was addressed. Using arterial subendothelium (SE) and endothelial cell extracellular matrix (ECM), studies were performed under flow conditions with annular and parallel-plate perfusion chambers, respectively, at a shear rate of 800 s-1 for 2, 5, and 10 minutes. Perfusates consisted of citrated normal blood samples incubated with Fab fragments of a monospecific polyclonal anti-CD36 antibody or with each of three new anti-CD36 monoclonal antibodies (MoAbs) that inhibit platelet adhesion to purified type I collagen in a static system (131.4, 131.5, and 131.7). Perfusions over SE were also carried out using citrated blood samples from a Naka-negative donor, whose platelets lack CD36. Morphometric evaluation of the perfused samples showed that polyclonal anti-CD36 Fab and the three monoclonal anti-CD36 antibodies inhibited platelet adhesion to the two substrates by 40% after 2 minutes of perfusion and by 30% after 5 minutes (P < .005 on SE and P < .01 on ECM), but at 10 minutes, significant inhibition was seen only on SE with polyclonal anti-CD36 Fab. Similar inhibitions were seen with Naka-negative platelets on SE. These studies demonstrate that CD36 plays a role in the early stages of platelet adhesion to physiologically relevant subendothelial surfaces.

Antibodies, Monoclonal↗
PubMed1996

[Erythrocytes modulate the anti-aggregation effect of aspirin and dipyridamole. Study under static conditions and in a flow system].

PURPOSE: The modulating role played by red cells on platelet function is well known, yet its mechanism of action in the presence of antiaggregating agents is not clearly defined. We tried to assess the influence of aspirin (ASA) or dipyridamole (DIP) treated red cells on platelet aggregation in plasma, with the samples in static conditions and under flow in a perfusion system. MATERIAL AND METHODS: Platelet-rich plasma (PRP) was adjusted to 4 x 10(9)/L. Red cells were washed with a 0.9% sodium chloride and 5mM glucose solution, then resuspended in PRP until a 40% haematocrit was attained. The drugs were added separately or together and the mixture was incubated for 10 minutes at 37 degrees C. Three series of tests were carried out: a) with reconstituted blood; b) with red cells alone; c) with the red cells washed in a 10-fold volume so as to eliminate all the drugs. The samples were divided in two parts: one was let to rest and the other was circulated through a perfusion system with a peristaltic pump. Simultaneous tests with untreated red cells and PRP were run as control in each session. A multichannel aggregameter was used to assess platelet aggregation according to Born's turbidimetric method. Variance analysis (ANOVA) or Student's t test were used for the statistical evaluation. RESULTS: Platelets attained from control samples under flow conditions showed partial inhibition of the maximal aggregation (58.02 +/- 9.92%). The inhibitory power of ASA was higher than that of DIP in all cases. Aggregation was totally inhibited by ASA from the beginning in treated whole blood, but the inhibitory effect appeared later when working with red cells alone, suggesting that red cells must retain part of the drug, which cannot exert its effect until it has been released after several minutes. The inhibition of platelet aggregation when working with washed treated red cells was more effective under flow conditions, chiefly with combined treatment with ASA+DIP. CONCLUSION: These studies stress the importance of flow conditions in the red cells-platelets interaction and in the regulation of the therapeutic agents used to modify platelet function.

Adenosine Diphosphate↗
PubMed1996

Management of plasmapheresis in Spain.

In Spain, there is no specific regulation for plasmapheresis. At the present time, there are two decrees issued by the central government that regulate blood donation and blood banks operation. The seventeen autonomous regions have independent powers including health care. Regulation at this level is possible, but not currently in place. In 1993, 1,266,704 units of whole blood were collected in Spain. This whole blood donation was enough to cover the need for red blood cells, but not to cover the needs of plasma derivatives. It has been calculated that in order to fulfil the requirements of factor and albumin about 400,0001 of plasma are needed. In 1994, there were over 10 centres in Spain where there were ongoing plasmapheresis programmes. Other centres are preparing plasmapheresis programmes scheduled to start in 1995. Spain needs an extra amount of plasma in between 250,000 and 300,0001 to cover the needs of plasma derivatives. It is concluded that it is probably best to meet plasma derivatives requirements with plasma obtained from a plasmapheresis programme. This approach has mainly two advantages: -the plasma generated is of the best quality available in order to produce coagulation factor concentrates; -it does not produce a "by-product" that in principle is not needed and would be unethical to throw away. We believe that to achieve self-sufficiency in plasma donation throughout a plasmapheresis programme requires flexibility in the current law and a substantial investment in promotion programmes.

Humans↗
PubMed1996

[Identification of a variant D(IVa) with anti-D in serum].

The first D(IVa) variant described so far in Spain is presented. This variant was identified, by means of a commercial monoclonal antibodies panel, in a RhD positive patient with anti-D in her serum. Differential diagnosis is discussed and the clinical relevance of identifying these variants is commented.

Antibodies, Monoclonal↗
PubMed1995

[Hemorrhagic incidents during prolonged ambulatory treatment with acenocoumarol].

BACKGROUND: To analyze hemorrhagic complications in a series of outpatients treated with acenocoumarol in an anticoagulant specialized unit by a prospective observational clinical study. PATIENTS AND METHODS: 1,200 outpatients (682 women/518 men, mean age 54.6 +/- 15.8 yrs.) treated with acenocoumarol for at least 6 weeks, with a total follow-up of 2,795 patients-yr. Prevalence and incidence of bleeding was analyzed. The episodes that were potentially life-threatening, or forced to blood transfusion or hospital patient admittance were considered as major bleedings, and the remainder episodes were minor. RESULTS: There were 379 minor bleedings in 258 patients (incidence 13.56/100 patients-yr.), 45 major bleedings (1.61/100 patients-yr.) and 2 lethal bleedings (0.07/100 patients-yr.). Minor bleedings correlated with more advanced age (57.3 +/- 11.8 vs. 53.9 +/- 16.7 yrs., p = 0.002), with the first two months in treatment in the 511 patients who start the treatment during the study (31.09 vs. 13.04/100 patients-yr., p < 0.001), and with a worse achievement of the desired anticoagulation (72.4% vs. 81.6%; p = 0.002). Major bleeding was associated with local causes in 48.9%, and with an excessive anticoagulation in 35.6%. CONCLUSIONS: Bleeding is relatively frequent during acenocoumarol therapy, mainly in patients with worse control, but only in a few of these episodes is severe, and is usually associated with local lesions.

Acenocoumarol↗