Summary
Combined oral contraceptives (OC) are known to increase the risk of venous thromboembolism. The aim of this randomized, cycle-controlled, cross-over study in 28 healthy volunteers was to ...assess potential differences between the effects of an OC containing 150 µg levonorgestrel (as representative of the so-called second generation OC) and an OC containing 150 µg desogestrel (as representative of the third generation OC) in combination with 30 µg ethinylestradiol on several coagulation factors and markers of thrombin formation. All participants used each OC for two cycles, and were switched to the other OC after a washout period of two menstrual cycles. The plasma concentrations of factors II, VII, X, and fibrinogen significantly increased during use of both the levonorgestrel- and desogestrel-containing OC’s. The plasma concentrations of factor VIII increased, and of factor V decreased, changes which only reached statistical significance during the use of the desogestrel-containing OC. During exposure to the desogestrel-containing OC, as compared with the levonorgestrel-containing OC, both factor VII and factor II showed a greater increase (FVII: 32% and 12% respectively; p <0.0001; FII: 16% and 12% respectively; p = 0.048), whereas factor V showed a greater decrease (–11% and –3% respectively; p = 0.010). Only one of the markers for ongoing coagulation (prothrombin fragment 1+2) showed a significant increase during OC use, whereas concentrations of thrombin-antithrombin complexes and soluble fibrin remained unchanged. For these markers, there was no difference between the tested OC’s. We conclude that there are differences between the effects of levonorgestrel and desogestrel-containing OC’s on some coagulation factors. Whether these changes provide a biological explanation for the reported differences in venous thromboembolic risk is as yet unclear. The real challenge now becomes to define a pattern of changes in the various systems which, if affected simultaneously, may tip the hemostatic balance towards a prethrombotic state and may lead to overt clinical venous thromboembolism.
The effect of oral contraceptives (OC) on fibrinolytic parameters was investigated in a cycle-controlled cross-over study in which 28 non-OC using women were randomly prescribed either a ...representative of the so-called second (30 microg ethinylestradiol, 150 microg levonorgestrel) or third generation OC (30 microg ethinylestradiol, 150 microg desogestrel) and who switched OC after a two month wash out period. During the use of OC, the levels of tissue-type plasminogen activator (tPA) activity, plasminogen, plasmin-alpha2-antiplasmin complexes and D-dimer significantly increased (by 30 to 80%), while the levels of plasminogen activator inhibitor- (PAI-1) antigen, PAI-1 activity and tPA antigen significantly decreased (25 to 50%), suggesting an increase in endogenous fibrinolytic activity. These OC-induced changes were not different between the two contraceptive pills. TAFI (thrombin-activatable fibrinolysis inhibitor) levels increased on levonorgestrel, and even further increased on desogestrel. A clot lysis assay that probes both fibrinolytic activity and the efficacy of the coagulation system to generate thrombin necessary to down regulate fibrinolysis via TAFI showed no change of the clot lysis time during OC use. This finding suggests that the OC-induced increase in endogenous fibrinolytic activity is counteracted by an increased capacity of the coagulation system to down regulate fibrinolysis via TAFI. Indeed we observed that during OC use there was a significant increase of F1+2 generation during clot formation. When these assays were performed in the presence of an antibody against factor XI, we observed that the clot lysis time was significantly increased during OC use and that the increase in F1+2 generation during OC therapy was due to a factor XI-independent process, which was significantly higher on desogestrel than on levonorgestrel. These data indicate that the OC-induced inhibition of endogenous fibrinolysis takes place in a factor XI-independent way and is more pronounced on desogestrel than on levonorgestrel-containing OC.
BACKGROUNDLipopolysaccharide (LPS), the major glycolipid component of Gram-negative bacterial outer membranes, is a potent endotoxin responsible for many of the directly or indirectly induced ...symptoms of infection. Lipoproteins (in particular, high-density lipoproteins) sequester LPS, thereby acting as a humoral detoxification mechanism.
PATIENTSDifferences in the lipoprotein composition in human plasma and lymph of a control patient group (n = 5) without systemic inflammatory response syndrome (non-SIRS/MOF) and patients with SIRS and multiple organ failure (MOF, n = 9) were studied. The LPS binding capacity of the lipoproteins in SIRS/MOF and non-SIRS/MOF patients was investigated by rechallenge of the plasma and lymph with fluorescently labeled LPS ex vivo. The lipoprotein composition was analyzed using immunochemical techniques and high-performance gel permeation chromatography.
RESULTSIn the non-SIRS/MOF patient group, plasma and lymph levels of apolipoprotein A-I (600 and 450 mg/L, respectively), apolipoprotein B (440 and 280 mg/L, respectively), total cholesterol (2.88 and 1.05 mM, respectively), and total triglycerides (0.67 and 0.97 mM, respectively) were observed. In the SIRS/MOF group, a decrease of apolipoprotein A-I (−55% in plasma and lymph), a decrease of apolipoprotein B (−43% in plasma and −38% in lymph), and a decrease of total cholesterol levels (−54% in plasma and −37% in lymph) were demonstrated. However, the triglyceride levels in the SIRS/MOF group showed a 30% increase in plasma and a 47% decrease in lymph compared with the non-SIRS/MOF patients. In SIRS/MOF patients, a 2.8-fold increase in plasma and a 1.8-fold increase in lymph of the LPS low-density lipoprotein/high-density lipoprotein ratio was observed, indicating that the relative LPS binding capacity of the lipoproteins in the SIRS/MOF patient group showed a trend to be shifted mainly toward low-density lipoproteins. Furthermore, in plasma and lymph of four SIRS/MOF patients, a novel cholesterol-containing high-density lipoprotein–like particle was found that barely had LPS binding capacity (<5%).
CONCLUSIONSIn the SIRS/MOF patients, the changes in lipoprotein composition in lymph are a reflection of those in plasma, except for the triglyceride levels. In comparison with the non-SIRS/MOF patients, the SIRS/MOF patients show a shifted LPS binding capacity of high-density lipoproteins toward low-density lipoproteins in plasma and in lymph. Moreover, in plasma and lymph, novel cholesterol-containing particles, resembling high-density lipoprotein, were identified in the SIRS/MOF patient group.
Lipopolysaccharide (LPS), the major outer membrane component of gram-negative bacteria, is a potent endotoxin that triggers cytokine-mediated systemic inflammatory responses in the host. Plasma ...lipoproteins are capable of LPS sequestration, thereby attenuating the host response to infection, but ensuing dyslipidemia severely compromises this host defense mechanism. We have recently reported that Escherichia coli J5 and Re595 LPS chemotypes that contain relatively short O-antigen polysaccharide side chains are efficiently redistributed from high-density lipoproteins (HDL) to other lipoprotein subclasses in normal human whole blood (ex vivo). In this study, we examined the role of the acute-phase proteins LPS-binding protein (LBP) and phospholipid transfer protein (PLTP) in this process. By the use of isolated HDL containing fluorescent J5 LPS, the redistribution of endotoxin among the major lipoprotein subclasses in a model system was determined by gel permeation chromatography. The kinetics of LPS and lipid particle interactions were determined by using Biacore analysis. LBP and PLTP were found to transfer LPS from HDL predominantly to low-density lipoproteins (LDL), in a time- and dose-dependent manner, to induce remodeling of HDL into two subpopulations as a consequence of the LPS transfer and to enhance the steady-state association of LDL with HDL in a dose-dependent fashion. The presence of LPS on HDL further enhanced LBP-dependent interactions of LDL with HDL and increased the stability of the HDL-LDL complexes. We postulate that HDL remodeling induced by LBP- and PLTP-mediated LPS transfer may contribute to the plasma lipoprotein dyslipidemia characteristic of the acute-phase response to infection.
Summary
The effect of oral contraceptives (OC) on fibrinolytic parameters was investigated in a cycle-controlled cross-over study in which 28 non-OC using women were randomly prescribed either a ...representative of the so-called second (30 µg ethinylestradiol, 150 µg levonorgestrel) or third generation OC (30 µg ethinylestradiol, 150 µg desogestrel) and who switched OC after a two month wash out period. During the use of OC, the levels of tissue-type plasminogen activator (tPA) activity, plasminogen, plasmin-α
2
-antiplasmin complexes and D-dimer significantly increased (by 30 to 80%), while the levels of plasminogen activator inhibitor-1 (PAI-1) antigen, PAI-1 activity and tPA antigen significantly decreased (25 to 50%), suggesting an increase in endogenous fibrinolytic activity. These OC-induced changes were not different between the two contraceptive pills. TAFI (thrombin-activatable fibrinolysis inhibitor) levels increased on levonorgestrel, and even further increased on desogestrel. A clot lysis assay that probes both fibrinolytic activity and the efficacy of the coagulation system to generate thrombin necessary to down regulate fibrinolysis via TAFI showed no change of the clot lysis time during OC use. This finding suggests that the OC-induced increase in endogenous fibrinolytic activity is counteracted by an increased capacity of the coagulation system to down regulate fibrinolysis via TAFI. Indeed we observed that during OC use there was a significant increase of F1+2 generation during clot formation. When these assays were performed in the presence of an antibody against factor XI, we observed that the clot lysis time was significantly increased during OC use and that the increase in F1+2 generation during OC therapy was due to a factor XI-independent process, which was significantly higher on desogestrel than on levonorgestrel. These data indicate that the OC-induced inhibition of endogenous fibrinolysis takes place in a factor XI-independent way and is more pronounced on desogestrel than on levonorgestrel-containing OC.
Summary
High-density lipoproteins (HDL) can bind and neutralize lipopoly- saccharides (LPS) in vitro and in vivo. HDL can also affect fibrinolytic activity and can directly influence platelet ...function by reducing platelet aggregation. In this study, the effects of reconstituted HDL (rHDL) on LPS-induced coagulation, fibrinolysis and platelet activation in humans were investigated. In a double-blind, randomized, placebo-controlled, cross-over study, eight healthy male volunteers were injected with LPS (4 ng/kg) on two occasions, once in conjunction with rHDL (40 mg/kg, given as a 4 h infusion starting 3.5 h prior to LPS injection), and once in conjunction with placebo. rHDL significantly reduced LPS-induced activation of coagulation (plasma levels of prothrombin fragment F1+2) and fibrinolysis (plasma levels of tissue type plasminogen activator antigen, t-PA). No effect was observed on LPS-induced inhibition of the fibrinolytic pathway (PAI-1) or on the transient thrombocytopenia elicited by LPS. Furthermore, rHDL treatment significantly enhanced the inhibition of collagen-stimulated inhibition of platelet aggregation during endotoxemia, but had no such effect on arachido- nate-stimulated platelet aggregation. rHDL treatment per se also reduced collagen-induced platelet aggregation. These results indicate that rHDL modifies the procoagulant state associated with endotoxemia.
A combination of thin-layer chromatography (TLC) and in situ spectrofluorimetry for the determination of free bile acids and bile acids conjugated with glycine or taurine is described. This method ...makes it possible to determine bile acids concentrations as low as 0.15-0.25 nmol (0.05-0.1 microgram) in a simple and reproducible way. Moreover, information can be obtained about conjugation patterns and relative concentrations of mono-, di-, and trihydroxy bile acids as well as about the presence of abnormal bile acids. After TLC the bile acids are made visible in uv light by dipping the layer in sulfuric acid in diethyl ether and warming it under well-described conditions. The fluorescence of the bile acids on the thin layer can be measured and makes it possible to quantitate them. The method presented here is applicable to bile acid-containing extracts from serum, bile, and feces, and the results are in good agreement with those obtained by enzymatic and gas-liquid chromatographic techniques.
A simplified extraction procedure for bile acids from wet faeces, using methanol/hydrochloric acid is described. Extracts were analyzed by gas-liquid chromatography, thin-layer chromatography and an ...enzymatic assay, with 3 alpha-hydroxysteroid dehydrogenase. Recoveries of some stable bile acids, added to faeces, were studied; extraction efficiency was also investigated with a procedure using radioactive labelled bile acids given orally to patients. Resin treatment of faecal extracts, because of the sometimes hard colour of the extracts, resulted in a slightly lower recovery as determined by the enzymatic method. Recoveries were higher, using the proposed extraction procedure, than those obtained with extracts prepared by the standard procedure of Grundy et al 6.
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