Platelet-poor plasma not suitable for clinical endotoxin testing, demonstrated in horses.
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Animals↗
Biomedical subjects
A Sturk
Publications and source records attributed to A Sturk.
A comparative analysis of D-dimer assays in patients with clinically suspected pulmonary embolism.
To avoid angiography in patients with clinically suspected pulmonary embolism and non-diagnostic lung scan results, the use of D-dimer has been advocated. We assessed plasma samples of 151 consecutive patients with clinically suspected pulmonary embolism. Lung scan results were: normal (43), high probability (48) and non-diagnostic (60; angiography performed in 43; 12 pulmonary emboli). Reproducibility, cut-off values, specificity, and percentage of patients in whom angiography could be avoided (with sensitivity 100%) were determined for two latex and four ELISA assays. The latex methods (cut-off 500 micrograms/l) agreed with corresponding ELISA tests in 83% (15% normal latex, abnormal ELISA) and 81% (7% normal latex, abnormal ELISA). ELISA methods showed considerable within- (2-17%) and between-assay variation (12-26%). Cut-off values were 25 micrograms/l (Behring), 50 micrograms/l (Agen), 300 micrograms/l (Stago) and 550 micrograms/l (Organon). Specificity was 14-38%; in 4-15% of patients angiography could be avoided. We conclude that latex D-dimer assays appear not useful, whereas ELISA methods may be of limited value in the exclusion of pulmonary embolism.
Angiography↗
The effect of formula versus breast feeding and exogenous vitamin K1 supplementation on circulating levels of vitamin K1 and vitamin K-dependent clotting factors in newborns.
The influence of breast or formula feeding together with that of a single supplementation of vitamin K1 at birth, on the vitamin K1 level and vitamin K-dependent clotting factors were studied in 65 breast and 15 formula fed infants. All breast fed newborns without supplementation (n = 25) had very low serum vitamin K1 at weeks 1 and 6. Oral vitamin K supplementation (n = 22) or i.m. (n = 18) at birth resulted in high serum levels at week 1, while at week 6 the effect had disappeared. Formula fed infants had vitamin K1 values within the normal adult range at all study points. The low serum levels of vitamin K1 were not associated with haemorrhagic disorders or coagulation abnormalities. The mean values of vitamin K1 in maternal sera at weeks 1 and 6 were 2.3 nmol/l and 1.8 nmol/l and in breast milk 2.7 nmol/l and 2.0 nmol/l respectively. No correlation existed between the values in breast milk and maternal serum. To maintain serum levels of vitamin K1 within the adult physiological range, repeated administration of low doses is needed in breast fed newborns beyond 1 week of age.
Breast Feeding↗
Preeclampsia is not associated with altered platelet vasopressin binding and cytosolic Ca++ concentration.
OBJECTIVES: Preeclampsia is an important cause of fetal and maternal morbidity and mortality. Recently it was described that platelet cytosolic Ca++ levels could be used to screen for preeclampsia. The current study investigated platelet arginine vasopressin receptor characteristics, platelet cytosolic Ca++ levels, plasma- and platelet-bound arginine vasopressin in white pregnant women. STUDY DESIGN: In a cross-sectional study nine third-trimester nulliparous pregnant women with gestational hypertension (seven with proteinuria, two with excessive weight gain without proteinuria) were compared with nine healthy nulliparous pregnant women matched for gestation length and age and 10 healthy age-matched nonpregnant women. Determined were (1) platelet arginine vasopressin receptor number and affinity, (2) platelet cytosolic Ca++ levels, both basal and on arginine vasopressin or thrombin stimulation, and (3) plasma- and platelet-bound arginine vasopressin levels. RESULTS: None of the measured parameters differed significantly among the three groups studied. Mean arginine vasopressin receptor number and affinity ranged from 108 to 143 receptors per platelet and 0.35 to 0.40 nmol/L, respectively. A single population of binding sites was found (Hill number 0.96). Basal Ca++ levels ranged from 113.4 to 133.3 nmol/L, on arginine vasopressin stimulation from 199 to 250 nmol/L. Median arginine vasopressin levels in platelet-poor plasma were between 1.2 and 2.4 pg/ml, with circulating platelets being estimated to possess two to five molecules of arginine vasopressin per platelet. A significant correlation was found between platelet cytosolic Ca++ levels before and after arginine vasopressin stimulation (r = 0.69, p < 0.001) and a weak correlation between platelet receptor density and arginine vasopressin-stimulated platelet cytosolic Ca++ levels (r = 0.38, p < 0.05). CONCLUSIONS: The studied parameters, platelet cytosolic Ca++ levels, whether basal or after stimulation with arginine vasopressin and vasopressin platelet receptor density and affinity, do not discriminate already hypertensive or preeclamptic white women from nondiseased subjects. A valuable test to screen for preeclampsia awaits further prospective studies.
Adult↗
Sequence characterization of a sheep cDNA for antithrombin III.
We determined the cDNA sequence of the mRNA for antithrombin III (AT III) from sheep liver. It encodes a protein of 465 amino acids, including a signal peptide of 32 amino acids. The amino acid sequence of the mature protein shows a sequence identity of 89.1%, 95.6% and 85.0% to the human, bovine and rabbit equivalents, respectively. Cysteine residues involved in disulfide bonds as well as potential glycosylation sites are conserved between the four species. In contrast, the amino acid sequence of the signal peptide shows a smaller identity, i.e., 68.7% and 56.3% compared to the human and rabbit preprotein, respectively.
Amino Acid Sequence↗
Acquired protein S deficiency might be associated with a prethrombotic state during estrogen treatment for tall stature.
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Adolescent↗
A missense mutation Pro157 Arg in lipoprotein lipase (LPLNijmegen) resulting in loss of catalytic activity.
Here we report on the molecular defect that leads to a deficiency of lipoprotein lipase (LPL) activity in a proband of Dutch descent. Southern-blot analysis of the LPL gene from the patient did not reveal any major DNA rearrangements. Sequencing of polymerase-chain-reaction-amplified DNA revealed that the proband is a homozygote for G725C, resulting in a substitution of Pro157 for Arg. This substitution alters a restriction site for PvuII, which allowed rapid identification of the mutant allele in family members. Site-directed mutagenesis and transient expression of the mutant LPL in COS cells produced an enzymatically inactive protein, establishing the functional significance of this mutation. This naturally occurring mutation which alters the Pro157 adjacent to Asp156 of the proposed catalytic triad, indicates that this region of the protein is indeed crucial for LPL catalytic activity.
Adult↗
Interleukin-6 release and the acute-phase reaction in patients with acute myocardial infarction: a pilot study.
We investigated the potential role of interleukin-6 as a mediator of the acute-phase reaction (APR) in patients with acute myocardial infarction. Of the six patients studied, five demonstrated increased plasma interleukin-6 levels. Interleukin-6 levels began to increase at 14 hours (mean; range = 8 to 20 hours) after the initial complaints and reached maximal levels of 28 to 250 U/mL (normal values less than 10 U/mL) after 36 hours (mean; range = 24 to 52 hours). No correlation was seen between the size of the interfaction as indicated by creatine kinase MB assays and the extent of the interleukin-6 increases (r = 0.44; p = 0.38). As an indicator of the APR, plasma C-reactive protein (CRP) levels were measured. CRP levels began to increase after 16 hours (mean; range = 8 to 24 hours) and reached maximum levels of 56 to 322 mg/L (normal values less than 3 mg/L) after 65 hours (mean; range = 48 to 92 hours). The increase of the interleukin-6 level preceded the increase of the CRP level in three patients and was simultaneous in two patients. Maximal interleukin-6 levels correlated significantly with maximal CRP levels (r = 0.96; p = 0.002). Thus these findings indicate that interleukin-6 is an important endogenous mediator for the APR in patients with acute myocardial infarction.
Acute Disease↗
Variable effects of radiological contrast media on thrombus growth in a rabbit jugular vein thrombosis model.
We studied the effect of an ionic high osmolar contrast medium (Ioxitalamate), an ionic low osmolar contrast medium (Ioxaglate) and various nonionic low osmolar contrast media (Iopamidol, Iopromide and Iohexol) on thrombus growth in a rabbit jugular vein thrombosis model. Thrombus growth was determined by the accretion of 125I-labeled fibrinogen onto autologous preformed thrombi in rabbit jugular veins at various time-intervals from 15 min up to 10 h after infusion of the study solution. The ionic low osmolar contrast medium markedly inhibited thrombus growth whereas all nonionic low osmolar contrast media promoted thrombus growth. The ionic high osmolar, contrast medium inhibited thrombus growth, but less than the ionic low osmolar contrast medium. Within the group of nonionic contrast media, the Iopamidol associated promotion of thrombus growth was significantly higher than the Iopromide or Iohexol associated effects. The simultaneous administration of the apparently most potent thrombus growth promoting contrast medium (i.e. Iopamidol) and heparin resulted in complete abolishment of the increase in thrombus growth. These results support the claims of prothrombotic properties of nonionic as compared to ionic contrast media and could explain the clinically encountered thromboembolic complications after the use of nonionic low osmolar contrast media.
Animals↗
Subcellular distribution and phosphorylation of vinculin isoforms in human blood platelets.
In this study we investigated human blood platelet vinculin microheterogeneity, the subcellular localization and phosphorylation of the different isoforms before and after platelet stimulation. At least 5 vinculin isoforms could be detected, as well as meta-vinculin. These isoforms did not demonstrate a specific subcellular localization, i.e. their relative content was similar in cytoskeleton, membrane skeleton and cytosol. Upon platelet stimulation with thrombin a small increase in alpha-vinculin was noted in all platelet subfractions. The cytoskeleton of non-stimulated platelets contained a minor quantity of vinculin. Upon thrombin stimulation of the platelets the cytoskeletal vinculin content increased significantly; previously we already reported a maximal 10% incorporation of the total platelet vinculin content into the cytoskeleton upon stimulation. A phosphorylation of a minor vinculin-isoform, i.e. at the alpha'/alpha location was mainly detected in the cytoskeleton. This phosphorylation was observable in the non-stimulated platelet cytoskeletal vinculin. These findings argue against a regulatory role for vinculin phosphorylation in the uptake of the main isoforms of this protein in the platelet cytoskeleton upon thrombin stimulation. The function of the phosphorylated cytoskeletal vinculin remains to be established.
Blood Platelets↗
Studies on the selectivity of enzymes involved in platelet-activating factor formation in stimulated cells.
The present studies were undertaken to obtain further insight into the selectivities of the enzymes, i.e., phospholipase A2 and acetyltransferase, involved in platelet-activating factor (PAF) production upon stimulation of human polymorphonuclear leukocytes (PMN) and platelets. After appropriate stimulation of the cells in the presence of [3H]acetate the total PAF and analogs, i.e., 1-alkyl-2-acetyl-, 1-alkenyl-2-acetyl-, and 1-acyl-2-acetyl-glycero-3- phosphocholine were isolated by high performance liquid chromatography. The isolated mixture was subjected to treatment with phospholipase A1 to differentiate acetate incorporation into 1-ether linked and 1-ester linked species. The ratio of acetate incorporation into 1-ether linked vs 1-ester linked PAF analogs amounted to 13.8 +/- 1.0 and 1.3 +/- 0.1 for PMN and platelets, respectively. When compared to the ratio of 1-ether linked and 1-ester linked species in the diradylglycerophosphocholine precursors in each cell type, i.e., 1.13 for PMN and 0.22 for platelets, these data suggested a pronounced selectivity for the phospholipase A2 and/or acetyltransferase in the process of PAF production. When the experiments were repeated with cells that had been pretreated with phenylmethanesulfonylfluoride (PMSF) to block the acetylhydrolase, the most dramatic effects were observed on acetate incorporation into 1-acyl-2-acetyl-glycero-3-phosphocholine, which increased much more than that into 1-alk(en)yl-2-acetyl-glycero-3-phosphocholine. Under these conditions, the ratio of acetate incorporation into 1-ether linked vs 1-ester linked PAF analogs became 1.4 +/- 0.2 and 0.17 +/- 0.02 for PMN and platelets, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)
1-Alkyl-2-acetylglycerophosphocholine Esterase↗
Comparison of the early dynamics of systemic prostacyclin release after administration of tumor necrosis factor and endotoxin to healthy humans.
Excessive production of prostaglandins may be of importance for the development of organ damage in generalized infection. To investigate the role of tumor necrosis factor (TNF) in systemic prostacyclin release in gram-negative septicemia, the plasma concentrations of its stable metabolite 6-keto-prostaglandin F1 alpha (6-keto-PGF1 alpha) were sequentially measured after intravenous bolus injections of recombinant human TNF (50 micrograms/m2; n = 6) and Escherichia coli endotoxin (2 ng/kg; n = 3) in healthy men. TNF induced a rapid increase in plasma 6-keto-PGF1 alpha from 0.11 +/- 0.01 to 0.44 +/- 0.15 ng/ml after 30 min (P less than .001). Endotoxin also elicited a rise in plasma 6-keto-PGF1 alpha, but peak values were reached only after 90 min (from 0.07 +/- 0.01 to 0.19 +/- 0.04 ng/ml; P less than .002). These results indicate that TNF may serve as an intermediate factor in systemic elaboration of prostacyclin in endotoxemia and gram-negative septicemia.
6-Ketoprostaglandin F1 alpha↗
Multicentre comparison of the Novo Biolabs immunometric CK-MB assay with four established methods.
The analytical performance of a new immunometric CK-MB assay from Novo Biolabs was evaluated in three hospitals. Comparisons were made with another immunometric assay, an immune inhibition/precipitation technique, an immune inhibition method, and a column separation method. The Novo assay exhibited good precision (CV 3-7%) at all concentration levels tested (6-150 micrograms/L) and was found to be linear over the range 0-200 micrograms/L. Comparison of the Novo kit with the other methods using patients' sera showed acceptable correlation between the methods over a wide concentration range (r = 0.96-0.99). We conclude that the assay is a useful adjunct to the specific measurement of CK-MB, particularly in routine laboratories with a high throughput of such determinations.
Creatine Kinase↗
Comparison of the early dynamics of coagulation activation after injection of endotoxin and tumor necrosis factor in healthy humans.
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Adult↗
Experimental endotoxemia in humans: analysis of cytokine release and coagulation, fibrinolytic, and complement pathways.
Endotoxemia was evoked by bolus injection of Escherichia coli endotoxin (2 ng/kg body weight) in six healthy subjects to investigate the early kinetics of cytokine release in relation to the development of clinical and hematologic abnormalities frequently seen in gram-negative septicemia. The plasma concentration of tumor necrosis factor (TNF) increased markedly after 30 to 45 minutes, and reached a maximal level after 60 to 90 minutes. In each volunteer, the initial increase of plasma interleukin 6 (IL-6) concentrations occurred 15 minutes after the initial TNF increase, and maximal IL-6 concentrations were reached at 120 to 150 minutes. A transient increase in body temperature and pulse rate occurred simultaneously with the initial TNF and IL-6 increases, whereas a significant decrease in blood pressure occurred after 120 minutes. These changes were proportional to the changes in TNF and IL-6 concentrations. Coagulation activation, as assessed by a rise of prothrombin fragments and thrombin-antithrombin III complexes, was noted after 120 minutes, in the absence of activation of the contact system. A two- to sixfold increase in the concentrations of tissue plasminogen activator (t-PA) and von Willebrand factor antigen indicated endothelial cell activation. This increase started at 120 and 90 minutes, respectively. The release of t-PA coincided with activation of the fibrinolytic pathway, as measured by plasmin-alpha 2-antiplasmin complexes. The fibrinolytic activity of t-PA was subsequently offset by release of plasminogen activator inhibitor, observed 150 minutes after the endotoxin injection, and reaching a peak at 240 minutes. No complement activation was detected. These results show that in humans endotoxin induces an early, rapidly counteracted fibrinolytic response, and a more long-lasting activation of thrombin by a mechanism other than contact system activation. In addition, our data suggest that endotoxin-induced leukopenia and endothelial cell activation are mediated by TNF.
Adult↗
Deficiencies of coagulation-inhibiting and fibrinolytic proteins in outpatients with deep-vein thrombosis.
BACKGROUND: Isolated deficiencies of antithrombin III, protein C, protein S, and plasminogen have been implicated as a cause of deep-vein thrombosis. It is assumed that patients with recurrent, familial, or juvenile thrombosis are very likely to have such a deficiency. METHODS: We studied the prevalence of isolated deficiencies of these proteins in 277 consecutive outpatients with venographically proved acute deep-vein thrombosis, as compared with 138 age-matched and sex-matched controls without deep-vein thrombosis, and calculated the positive predictive value of a history of recurrent, familial, or juvenile venous thromboembolism for the presence of such a deficiency. RESULTS: The overall prevalence of deficiencies of any of these proteins in the patients with venous thrombosis was 8.3 percent (23 of 277 patients) (95 percent confidence interval, 5.4 to 12.4), as compared with 2.2 percent in the controls (3 of 138 subjects) (95 percent confidence interval, 0.5 to 6.1; P less than 0.05 for the comparison between groups). The positive predictive values for the presence of an isolated protein deficiency in patients with recurrent, familial, or juvenile deep-vein thrombosis, defined as the proportion of patients with the clinical finding who had a deficiency of one or more of the proteins, were 9, 16, and 12 percent, respectively. CONCLUSIONS: The cause of acute venous thrombosis in most outpatients (91.7 percent) cannot be explained by abnormalities of coagulation-inhibiting and fibrinolytic proteins. The information obtained from the medical history concerning recurrent or familial venous thrombosis or the onset of the disease at a young age is not useful for the identification of patients with protein deficiencies.
Acute Disease↗
Vinculin is a permanent component of the membrane skeleton and is incorporated into the (re)organising cytoskeleton upon platelet activation.
Vinculin, a 130-kDa protein discovered in chicken gizzard smooth-muscle cells and subsequently also described in platelets, is believed to be involved in membrane-cytoskeleton interactions. In this study we investigated vinculin distribution in human blood platelets. Two skeletal fractions and a remaining cytosolic fraction were prepared with a recently described Triton X-100 lysis buffer causing minimal post-lysis breakdown by proteolysis. The presence of vinculin was demonstrated in the membrane skeleton and cytosol of resting and thrombin-activated human platelets. Upon thrombin stimulation vinculin also appeared in the cytoskeleton. this cytoskeletal incorporation was completed during the early stages of platelet aggregation and secretion, when the uptake of myosin, actin-binding protein and talin was still not maximal. We conclude therefore, that vinculin may play an important role in the structural (re)organisation of the human platelet cytoskeleton upon platelet activation.
Blood Platelets↗
Endotoxin-induced cytokines in human septicemia.
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Adult↗