Relation of depression to various markers of coagulation and fibrinolysis in patients with and without coronary artery disease

BACKGROUND: It has been suggested that changes in blood coagulation and fibrinolysis might explain the observed association between depression and coronary artery disease (CAD). So far, only a few coagulation factors have been investigated in this regard, and the results were not consistent. DESIGN: The aim of our study was to analyse a broad range of coagulation and fibrinolytic factors, with emphasis on factors directly involved in clot formation and degradation or reflecting coagulation activation, in patients with CAD and controls without CAD, as assessed by coronary angiography, who also underwent a diagnostic procedure for depression. METHODS: We screened 306 patients with CAD and controls without CAD for depression using the Hospital Anxiety and Depression Scale and Allgemeine Depressions Skala-L questionnaires. In participants with positive screening result, diagnosis of major depression was confirmed or excluded by a structured interview. We analysed the following coagulation and fibrinolytic factors: fibrinogen, prothrombin fragment F1+2, factor XIII A-subunit, factor XIII B-subunit, tissue plasminogen activator, plasminogen activator inhibitor-1, thrombin-activable fibrinolysis inhibitor, and D-dimer. RESULTS: We did not observe significant associations between depression and CAD, nor between depression and cardiovascular risk factors. Coagulation and fibrinolytic factors showed no differences between patients with CAD and controls, but they were associated with several cardiovascular risk factors. Depression was not associated with coagulation and fibrinolytic factors. No associations were found either when both CAD and depression were taken into account. CONCLUSION: Our study gives no evidence that there is a significant relation among depression, CAD, and blood coagulation and fibrinolysis.

Self-reactivity in the dimeric intravenous immunoglobulin fraction

Therapeutic intravenous immunoglobulin (IVIg) preparations contain antibodies reflecting the cumulative antigen experience of the donor population. IVIg contains variable amounts of monomeric and dimeric IgG, but there is little information available on their comparative antibody specificities. We have isolated highly purified fractions of monomeric and dimeric IgG by size-exclusion chromatography. Following treatment of all fractions at pH4, analyses by immunodot and immunocytology on human cell lines showed a preferential recognition of autoantigens in the dimeric IgG fraction. Investigation of the HEp-2 cytoplasmic proteome by 2D-PAGE, Western blot, and subsequent identification of IVIg reactive spots by mass spectrometry (LC-MS/MS) showed that IVIg recognized only a restricted set of the total proteins. Similar experiments showed that more antigens were recognized by the dimeric IgG fraction, especially when the dissociated dimer fraction was used, as compared to its monomeric counterpart. These observations are consistent with idiotype-anti-idiotype masking of auto-specific Abs in the dimeric fraction of IVIg.

Stability of coagulation assays performed in plasma from citrated whole blood transported at ambient temperature

Many preanalytical variables affect the results of coagulation assays. A possible way to control some of them would be to accept blood specimens shipped in the original collection tube. The aim of our study was to investigate the stability of coagulation assays in citrated whole blood transported at ambient temperature for up to two days after specimen collection. Blood samples from 59 patients who attended our haematology outpatient ward for thrombophilia screening were transported at ambient temperature (outdoor during the day, indoor overnight) for following periods of time: <1 hour, 4-6, 8-12, 24-28 and 48-52 hours prior to centrifugation and plasma-freezing. The following coagulation tests were performed: PT, aPTT, fibrinogen, FII:C, FV:C, FVII:C, FVIII:C, FIX:C, FX:C, FXI:C, VWF:RCo, VWF:Ag, AT, PC activity, total and free PS antigen, modified APC-sensitivity-ratio, thrombin-antithrombin-complex and D-dimer. Clinically significant changes, defined as a percentage change of more than 10% from the initial value, were observed for FV:C, FVIII:C and total PS antigen starting at 24-28 hours, and for PT, aPTT and FVII:C at 48-52 hours. No statistically significant differences were seen for fibrinogen, antithrombin, or thrombin-antithrombin complexes (Friedman repeated measures analysis of variance). The present data suggest that the use of whole blood samples transported at ambient temperature may be an acceptable means of delivering specimens for coagulation analysis. With the exception of factor V and VIII coagulant activity, and total PS antigen all investigated parameters can be measured 24-28 hours after specimen collection without observing clinically relevant changes.

Dosing lepirudin in patients with heparin-induced thrombocytopenia and normal or impaired renal function: a single-center experience with 68 patients

The recommended dose (bolus 0.4 mg/kg followed by 0.15 mg/kg per hour) of lepirudin, a direct thrombin inhibitor licensed for treatment of heparin-induced thrombocytopenia (HIT), is too high. Starting in 2001, we omitted the bolus and reduced maintenance dose by at least one-third. Analyzing 53 HIT patients treated between January 2001 and February 2007, we observed that therapeutic anticoagulation intensity already 4 hours after lepirudin start had been reached with the following initial lepirudin doses (median): 0.078 mg/kg per hour [creatinine clearance (CrCl) more than 60 mL/min], 0.040 mg/kg per hour (CrCl 30-60 mL/min), and 0.013 mg/kg per hour (CrCl < 30 mL/min). The efficacy of this treatment was documented by increasing platelets and decreasing D-dimers. Based on this experience, we derived a lepirudin dosing regimen, which was prospectively evaluated treating 15 HIT patients between March 2007 and February 2008. We show that omitting the initial lepirudin bolus and administering 0.08 mg/kg per hour in patients with CrCl more than 60 mL/min, 0.04 mg/kg per hour in patients with CrCl 30-60 mL/min, and 0.01 to 0.02 mg/kg per hour in those with CrCl less than 30 mL/min is efficacious and safe, as documented by increasing platelet counts, decreasing D-dimer levels, and rare thrombotic (1 of 46) and major bleeding (4 of 46) complications.

Surfactant proteins SP-B and SP-C and their precursors in bronchoalveolar lavages from children with acute and chronic inflammatory airway disease

BACKGROUND: The surfactant proteins B (SP-B) and C (SP-C) are important for the stability and function of the alveolar surfactant film. Their involvement and down-regulation in inflammatory processes has recently been proposed, but their level during neutrophilic human airway diseases are not yet known. METHODS: We used 1D-electrophoresis and Western blotting to determine the concentrations and molecular forms of SP-B and SP-C in bronchoalveolar lavage (BAL) fluid of children with different inflammatory airway diseases. 21 children with cystic fibrosis, 15 with chronic bronchitis and 14 with pneumonia were included and compared to 14 healthy control children. RESULTS: SP-B was detected in BAL of all 64 patients, whereas SP-C was found in BAL of all but 3 children; those three BAL fluids had more than 80% neutrophils, and in two patients, who were re-lavaged later, SP-C was then present and the neutrophil count was lower. SP-B was mainly present as a dimer, SP-C as a monomer. For both qualitative and quantitative measures of SP-C and SP-B, no significant differences were observed between the four evaluated patient groups. CONCLUSION: Concentration or molecular form of SP-B and SP-C is not altered in BAL of children with different acute and chronic inflammatory lung diseases. We conclude that there is no down-regulation of SP-B and SP-C at the protein level in inflammatory processes of neutrophilic airway disease.

Thrombophilic risk factors in patients with cranial and spinal dural arteriovenous fistulae

OBJECTIVE: Numerous studies have reported the technical aspects and results of surgical and/or endovascular treatment of cranial dural arteriovenous fistulae (cDAVF) and spinal dural arteriovenous fistulae (sDAVF). Only a few of them have addressed the question of thrombophilic conditions, which may be relevant as pathogenetic factors or can increase the risk for venous thromboembolic events. Therefore, the objective of this study is to compare thrombophilic risk factors in patients with cDAVF and sDAVF with no history of trauma. METHODS: A total of 43 patients (25 with cDAVF and 18 with sDAVF) were included in this study. Blood samples were analyzed for G20210A mutation of the prothrombin gene and factor V Leiden mutation. In all patients, prothrombin time, international normalized ratio, fibrinogen, antithrombin, protein C and S activity, von Willebrand factor antigen, ristocetin cofactor activity, D-dimer, coagulation factor VIII activity, and tissue factor pathway inhibitor were determined. Screening was performed for the occurrence of lupus antiphospholipid and cardiolipin antibodies. RESULTS: The prevalence of G20210A mutation of the prothrombin gene was significantly higher in patients with cDAVF (n = 6) compared with patients with sDAVF (n = 0; P < 0.05, Fisher's exact test). A factor V Leiden mutation was found in 3 patients with sDAVF and in 1 patient with cDAVF (P = 0.29, Fisher's exact test). No significant difference was found for other parameters, except for fibrinogen, but decreased protein C activity was more frequent in patients with cDAVF compared with patients with sDAVF (4 versus 1). Decreased protein S activity was encountered in 3 patients (2 with sDAVF and 1 with cDAVF). Cardiolipin antibodies were found in 2 patients with cDAVF but in none with sDAVF, whereas only 1 patient with sDAVF had lupus antiphospholipid antibodies. CONCLUSION: In both groups of patients with dural arteriovenous fistulae, genetic thrombophilic abnormalities occurred in a higher percentage than in the general population. The differences of the genetic abnormalities may be involved in different pathophysiological mechanism(s) in the development of these distinct neurovascular entities.

Monomerization of dimeric IgG of intravenous immunoglobulin (IVIg) increases the antibody reactivity against intracellular antigens

Intravenous immunoglobulin (IVIg) preparations are derived from pooled plasma from up to 60,000 healthy human donors and reflect the immunologic experience of the donor population. IVIg contains monomeric and dimeric IgG populations which are in a dynamic equilibrium depending on concentration, pH, temperature, donor pool size, time and stabilizers added in order to keep the portion of dimeric IgG below a certain level. In the present study, monomeric and dimeric fractions were isolated by size exclusion chromatography. The dimeric fractions, however, showed a dynamic instability and tended to dissociate. Both dimeric and monomeric IgG fractions were acid treated (pH 4) in order to dissociate the dimeric IgG. Western-blot analysis identified a sub-population of SDS resistant IgG dimers. Furthermore, the reactivities of the fractions were tested against a panel of self- and exo-antigens. There was a marked increase in activity of the dimeric compared to the monomeric IgG fraction against various intracellular self-antigens. Our data indicates that the increased reactivities of pH 4-treated fractions can mainly be attributed to dimer dissociation, as pH 4-treated monomers do not show significantly increased activities against a range of antigens.

The crystal structure of the platelet activator aggretin reveals a novel (alphabeta)2 dimeric structure

Aggretin is a C-type lectin purified from Calloselasma rhodostoma snake venom. It is a potent activator of platelets, resulting in a collagen-like response by binding and clustering platelet receptor CLEC-2. We present here the crystal structure of aggretin at 1.7 A which reveals a unique tetrameric quaternary structure. The two alphabeta heterodimers are arranged through 2-fold rotational symmetry, resulting in an antiparallel side-by-side arrangement. Aggretin thus presents two ligand binding sites on one surface and can therefore cluster ligands in a manner reminiscent of convulxin and flavocetin. To examine the molecular basis of the interaction with CLEC-2, we used a molecular modeling approach of docking the aggretin alphabeta structure with the CLEC-2 N-terminal domain (CLEC-2N). This model positions the CLEC-2N structure face down in the "saddle"-shaped binding site which lies between the aggretin alpha and beta lectin-like domains. A 2-fold rotation of this complex to generate the aggretin tetramer reveals dimer contacts for CLEC-2N which bring the N- and C-termini into the proximity of each other, and a series of contacts involving two interlocking beta-strands close to the N-terminus are described. A comparison with homologous lectin-like domains from the immunoreceptor family reveals a similar but not identical dimerization mode, suggesting this structure may represent the clustered form of CLEC-2 capable of signaling across the platelet membrane.

Metabolic predictors of inflammation, adhesion, and coagulability in healthy younger-aged adults

Elevated levels of inflammatory biomarkers are associated with the pathophysiology of cardiovascular diseases and are predictors of cardiovascular events. The objective of this study was to determine the unique contributions of metabolic factors as predictors of inflammation (C-reactive protein (CRP) and interleukin-6 (IL-6)), adhesion (soluble intercellular adhesion molecule-1 (sICAM-1)), and coagulation (D-dimer) in healthy younger-aged adults. Participants were 83 women and 92 men (mean age 30.04 years, s.d. +/- 4.8, range 22-39) of normal weight to moderate obese weight (mean BMI 24.4 kg/m(2), s.d. +/- 3.35, range 17-32). The primary data analytical approaches included Pearson correlation and multiple linear regression. Circulating levels of CRP, IL-6, sICAM-1, and D-dimer were determined in plasma. Higher levels of CRP were independently associated with higher BMI, a greater waist-to-hip ratio, female gender, and higher triglycerides (P < 0.001). Higher IL-6 levels were independently associated with a greater waist-to-hip ratio (P < 0.01). Higher levels of sICAM-1 were independently associated with higher BMI, higher triglycerides, and lower insulin resistance (P < 0.001). Higher D-dimer levels were independently associated with higher BMI and being female (P < 0.001). Having a higher BMI was most consistently associated with elevated biomarkers of inflammation, adhesion, and coagulation in this sample of healthy younger-aged adults, although female gender, insulin resistance, and lipid levels were also related to the biomarkers. The findings provide insight into the adverse cardiovascular risk associated with elevated body weight in younger adults.

The effects of aspirin and nonselective beta blockade on the acute prothrombotic response to psychosocial stress in apparently healthy subjects

We hypothesized that the 2 cardiovascular drugs aspirin and propranolol attenuate the prothrombotic response to acute psychosocial stress relative to placebo medication. We randomized 56 healthy subjects, double-blind, to 5-day treatment with an oral dose of either 100 mg of aspirin plus 80 mg of propranolol combined, single aspirin, single propranolol, or placebo medication. Thereafter, subjects underwent a 13-minute psychosocial stressor. Plasma levels of von Willebrand factor antigen (VWF:Ag), fibrinogen, coagulation factor VII (FVII:C) and XII (FXII:C) activity, and D-dimer were determined in blood samples collected immediately pre- and post-stress and 45 minutes post-stress. The stress-induced changes in prothrombotic measures were adjusted for gender, age, body mass index, mean arterial blood pressure, smoking status, and sleep quality. There was an increase in VWF:Ag levels from immediately pre-stress to 45 minutes post-stress in the placebo group relative to the 3 subject groups with verum medication (P's dimer were similar in all 4 medication groups. The combination of aspirin with propranolol, single aspirin, and single propranolol all attenuated the acute response in plasma VWF:Ag levels to psychosocial stress. This suggests that these cardiovascular drugs might exert limited protection from the development of stress-triggered coronary thrombosis.

Effects of gender and dementia severity on Alzheimer's disease caregivers' sleep and biomarkers of coagulation and inflammation

BACKGROUND: Being a caregiver for a spouse with Alzheimer's disease is associated with increased risk for cardiovascular illness, particularly for males. This study examined the effects of caregiver gender and severity of the spouse's dementia on sleep, coagulation, and inflammation in the caregiver. METHODS: Eighty-one male and female spousal caregivers and 41 non-caregivers participated (mean age of all participants 70.2 years). Full-night polysomnography (PSG) was recorded in each participants home. Severity of the Alzheimer's disease patient's dementia was determined by the Clinical Dementia Rating (CDR) scale. The Role Overload scale was completed as an assessment of caregiving stress. Blood was drawn to assess circulating levels of D-dimer and Interleukin-6 (IL-6). RESULTS: Male caregivers who were caring for a spouse with moderate to severe dementia spent significantly more time awake after sleep onset than female caregivers caring for spouses with moderate to severe dementia (p=.011), who spent a similar amount of time awake after sleep onset to caregivers of low dementia spouses and to non-caregivers. Similarly, male caregivers caring for spouses with worse dementia had significantly higher circulating levels of D-dimer (p=.034) than females caring for spouses with worse dementia. In multiple regression analysis (adjusted R(2)=.270, p<.001), elevated D-dimer levels were predicted by a combination of the CDR rating of the patient (p=.047) as well as greater time awake after sleep onset (p=.046). DISCUSSION: The findings suggest that males caring for spouses with more severe dementia experience more disturbed sleep and have greater coagulation, the latter being associated with the disturbed sleep. These findings may provide insight into why male caregivers of spouses with Alzheimer's disease are at increased risk for illness, particularly cardiovascular disease.

Coagulation activity before and after acute psychosocial stress increases with age

OBJECTIVE: To assess whether stress further increases hypercoagulation in older individuals. We investigated whether acute stress-induced changes in coagulation parameters differ with age. It is known that hypercoagulation occurs in response to acute stress and that a shift in hemostasis toward a hypercoagulability state occurs with age. However, it is not yet known whether acute stress further increases hypercoagulation in older individuals, and thus may increase their risk for cardiovascular disease (CVD). METHODS: A total of 63 medication-free nonsmoking men, aged between 20 and 65 years (mean +/- standard error of the mean = 36.7 +/- 1.7 years), underwent an acute standardized psychosocial stress task combining public speaking and mental arithmetic in front of an audience. We measured plasma clotting factor VII activity (FVII:C), fibrinogen, and D-dimer at rest, immediately, and 20 minutes after stress. RESULTS: Increased age predicted greater increases in fibrinogen (beta = 0.26, p = 0.041; DeltaR(2) = 0.05), FVII:C (beta = 0.40, p = .006; DeltaR(2) = 0.11), and D-dimer (beta = 0.51, p < .001; DeltaR(2) = 0.18) from rest to 20 minutes after stress independent of body mass index and mean arterial blood pressure. General linear models revealed significant effects of age and stress on fibrinogen, FVII:C, and D-dimer (main effects: p < .04), and greater D-dimer stress reactivity with older age (interaction age-by-stress: F(1.5/90.4) = 4.36, p = .024; f = 0.33). CONCLUSIONS: Our results suggest that acute stress might increase vulnerability in the elderly for hypercoagulability and subsequent hemostasis-associated diseases like CVD.

Effect of oral melatonin on the procoagulant response to acute psychosocial stress in healthy men: a randomized placebo-controlled study

Acute mental stress is a potent trigger of acute coronary syndromes. Catecholamine-induced hypercoagulability with acute stress contributes to thrombus growth after coronary plaque rupture. Melatonin may diminish catecholamine activity. We hypothesized that melatonin mitigates the acute procoagulant stress response and that this effect is accompanied by a decrease in the stress-induced catecholamine surge. Forty-five healthy young men received a single oral dose of either 3 mg melatonin (n = 24) or placebo medication (n = 21). One hour thereafter, they underwent a standardized short-term psychosocial stressor. Plasma levels of clotting factor VII activity (FVII:C), FVIII:C, fibrinogen, D-dimer, and catecholamines were measured at rest, immediately after stress, and 20 min and 60 min post-stress. The integrated change in D-dimer levels from rest to 60 min post-stress differed between medication groups controlling for demographic and metabolic factors (P = 0.047, eta(p)(2) = 0.195). Compared with the melatonin group, the placebo group showed a greater increase in absolute D-dimer levels from rest to immediately post-stress (P = 0.13; eta(p)(2) = 0.060) and significant recovery of D-dimer levels from immediately post-stress to 60 min thereafter (P = 0.007; eta(p)(2) = 0.174). Stress-induced changes in FVII:C, FVIII:C, fibrinogen, and catecholamines did not significantly differ between groups. Oral melatonin attenuated the stress-induced elevation in the sensitive coagulation activation marker D-dimer without affecting catecholamine activity. The finding provides preliminary support for a protective effect of melatonin in reducing the atherothrombotic risk with acute mental stress.

Oral melatonin reduces blood coagulation activity: a placebo-controlled study in healthy young men

Melatonin has previously been suggested to affect hemostatic function but studies on the issue are scant. We hypothesized that, in humans, oral administration of melatonin is associated with decreased plasma levels of procoagulant hemostatic measures compared with placebo medication and that plasma melatonin concentration shows an inverse association with procoagulant measures. Forty-six healthy men (mean age 25 +/- 4 yr) were randomized, single-blinded, to either 3 mg of oral melatonin (n = 25) or placebo medication (n = 21). One hour thereafter, levels of melatonin, fibrinogen, and D-dimer as well as activities of coagulation factor VII (FVII:C) and VIII (FVIII:C) were measured in plasma. Multivariate analysis of covariance and regression analysis controlled for age, body mass index, mean arterial blood pressure, heart rate, and norepinephrine plasma level. Subjects on melatonin had significantly lower mean levels of FVIII:C (81%, 95% CI 71-92 versus 103%, 95% CI 90-119; P = 0.018) and of fibrinogen (1.92 g/L, 95% CI 1.76-2.08 versus 2.26 g/L, 95% CI 2.09-2.43; P = 0.007) than those on placebo explaining 14 and 17% of the respective variance. In all subjects, increased plasma melatonin concentration independently predicted lower levels of FVIII:C (P = 0.037) and fibrinogen (P = 0.022) explaining 9 and 11% of the respective variance. Melatonin medication and plasma concentration were not significantly associated with FVII:C and D-dimer levels. A single dose of oral melatonin was associated with lower plasma levels of procoagulant factors 60 min later. There might be a dose-response relationship between the plasma concentration of melatonin and coagulation activity.

Rifampicin-activated human pregnane X receptor and CYP3A4 induction enhance acetaminophen-induced toxicity

Acetaminophen (APAP) is safe at therapeutic levels but causes hepatotoxicity via N-acetyl-p-benzoquinone imine-induced oxidative stress upon overdose. To determine the effect of human (h) pregnane X receptor (PXR) activation and CYP3A4 induction on APAP-induced hepatotoxicity, mice humanized for PXR and CYP3A4 (TgCYP3A4/hPXR) were treated with APAP and rifampicin. Human PXR activation and CYP3A4 induction enhanced APAP-induced hepatotoxicity as revealed by hepatic alanine aminotransferase (ALT) and aspartate aminotransferase (AST) activities elevated in serum, and hepatic necrosis after coadministration of rifampicin and APAP, compared with APAP administration alone. In contrast, hPXR mice, wild-type mice, and Pxr-null mice exhibited significantly lower ALT/AST levels compared with TgCYP3A4/hPXR mice after APAP administration. Toxicity was coincident with depletion of hepatic glutathione and increased production of hydrogen peroxide, suggesting increased oxidative stress upon hPXR activation. Moreover, mRNA analysis demonstrated that CYP3A4 and other PXR target genes were significantly induced by rifampicin treatment. Urinary metabolomic analysis indicated that cysteine-APAP and its metabolite S-(5-acetylamino-2-hydroxyphenyl)mercaptopyruvic acid were the major contributors to the toxic phenotype. Quantification of plasma APAP metabolites indicated that the APAP dimer formed coincident with increased oxidative stress. In addition, serum metabolomics revealed reduction of lysophosphatidylcholine in the APAP-treated groups. These findings demonstrated that human PXR is involved in regulation of APAP-induced toxicity through CYP3A4-mediated hepatic metabolism of APAP in the presence of PXR ligands.