From <em>D</em>ecember <em>2</em>019, a novel coronavirus, SARS-CoV-<em>2</em>, caused an outbreak of pneumonia in Wuhan city and rapidly spread throughout China and globally. However, the clinical characteristics and co-infection with other respiratory pathogens of patients with COVI<em>D</em>-19 and the factors associated with severity of COVI<em>D</em>-19 are still limited. In this retrospective cohort study, we included 354 inpatients with COVI<em>D</em>-19 admitted to Renmin Hospital of Wuhan University from February 4, <em>2</em>0<em>2</em>0 to February <em>2</em>8, <em>2</em>0<em>2</em>0. We found levels of interleukin-6, interleukin-10, C-reactive protein, <em>D</em>-<em>dimer</em>, white blood cell count and neutrophil count were clearly elevated in males and critical cases compared with females and severe and mild cases, respectively. However, lymphopenia was more severe in males than females and levels of tumor necrosis factor alpha were reduced significantly in critical cases than severe and mild cases. <em>2</em>3.5% of severe cases and <em>2</em>4.4% of critical cases were co-infected with other respiratory pathogens. Additionally, stepwise multivariable regression analysis suggested that co-infection, lymphocyte count and levels of <em>D</em>-<em>dimer</em> were associated with severity of COVI<em>D</em>-19.These findings provide crucial clues for further identification of the mechanisms, characteristics and treatments of patients with COVI<em>D</em>-19.

Keywords: COVID-19; Co-infection; Gender; Laboratory factors.

An outbreak of pneumonia caused by a novel coronavirus (COVI<em>D</em>-19) began in Wuhan, China in <em>D</em>ecember <em>2</em>019 and quickly spread throughout the country and world. An efficient and convenient method based on clinical characteristics was needed to evaluate the potential deterioration in patients. We aimed to develop a simple and practical risk scoring system to predict the severity of COVI<em>D</em>-19 patients on admission. We retrospectively investigated the clinical information of confirmed COVI<em>D</em>-19 patients from 10 February <em>2</em>0<em>2</em>0 to <em>2</em>9 February <em>2</em>0<em>2</em>0 in Wuhan Union Hospital. Predictors of severity were identified by univariate and multivariate logistic regression analysis. A total of 147 patients with confirmed SARS-CoV-<em>2</em> infection were grouped into non-severe (94 patients) and severe (53 patients) groups. We found that an increased level of white blood cells (WBC), neutrophils, <em>D</em>-<em>dimer</em>, fibrinogen (FIB), IL-6, C-reactive protein (CRP), erythrocyte sedimentation rate (ESR), alanine aminotransferase (ALT), aspartate aminotransferase (AST), α-hydroxybutyrate dehydrogenase (HB<em>D</em>H), serum amyloid A (SAA) and a decreased level of lymphocytes were important risk factors associated with severity. Furthermore, three variables were used to formulate a clinical risk scoring system named COVI<em>D</em>-19 index = 3 × <em>D</em>-<em>dimer</em> (µg/L) + <em>2</em> × lgESR (mm/hr) - 4 × lymphocyte (×10⁹ /L) + 8. The area under the receiver operating characteristic (ROC) curve was 0.843 (95% CI, 0.771-0.914). We propose an effective scoring system to predict the severity of COVI<em>D</em>-19 patients. This simple prediction model may provide healthcare workers with a practical method and could positively impact decision-making with regard to deteriorating patients.

<strong class="sub-title">Keywords:</strong> COVI<em>D</em>-19; SARS-CoV-<em>2</em>; predict; risk factor; score model.

Using the detergents n-dodecyl beta-<em>D</em>-maltoside and heptyl thioglycopyranoside, a subcore complex of photosystem II (PSII) has been isolated that contains the chlorophyll-binding protein, CP47, and the reaction center components, <em>D</em>1, <em>D</em><em>2</em>, and cytochrome b559. We have found, by using sucrose density centrifugation, that the resulting preparation consisted of a mixture of <em>dimer</em>ic and monomeric forms of the CP47 reaction center (RC) complex, having molecular masses of 410 +/- 30 and <em>2</em>00 +/- <em>2</em>8 k<em>D</em>a, respectively, as estimated by size exclusion chromatography. The level of the <em>dimer</em> in the preparation is significantly higher than the monomeric form. Both the monomer and <em>dimer</em> contain the proteins CP47, <em>D</em>1, and <em>D</em><em>2</em> and the alpha- and beta-subunits of cytochrome b559. Analyses by mass spectrometry and N-terminal sequencing showed that both forms of the CP47-RC complex contain the products of the psbI, psbTc (chloroplast gene), and psbW with molecular masses of 4195.5, 3849.6, and 59<em>2</em>7.4 <em>D</em>a, respectively. In contrast to the monomeric form, the CP47-RC <em>dimer</em> contained two extra proteins with low molecular weights, identified as the products of the psbL and psbK genes having molecular masses of 4365.5 and 4<em>2</em>9<em>2</em>.1, respectively. It was also found that the <em>dimer</em> contained slightly more molecules of chlorophyll a (<em>2</em>1 +/- <em>2</em>.5) than the monomer (18 +/- 1.5), a characteristic also observed in the room temperature absorption spectrum by comparing the ratio of absorption at 416 and 435 nm. Of particular note was the finding that the <em>dimer</em>, but not the monomer, contained plastoquinone-9 (estimated to be 1.5 +/- 0.3 molecules per RC). The results indicate that the CP47-RC monomer is derived from the <em>dimer</em>ic form of the complex, and therefore the latter is likely to represent an in vivo conformation. The PsbTc as well as the PsbI and PsbW proteins are identified as being intimately associated with the <em>D</em>1 and <em>D</em><em>2</em> proteins, and in the case of the <em>dimer</em>, importance is placed on the PsbL and PsbK proteins in sustaining plastoquinone binding and maintenance of the <em>dimer</em>ic organization. Assuming only one copy of the alpha- and beta-subunits of cytochrome b559, the monomeric and <em>dimer</em>ic forms of the complex would be expected to contain <em>2</em>1 and <em>2</em>3 x <em>2</em> transmembrane helices, respectively.

OBJECTIVE

To determine the effects of soy milk consumption compared with cow's milk on inflammation, coagulation, and oxidative stress among patients with diabetic nephropathy.

METHODS

This randomized, crossover clinical trial was conducted on <em>2</em>5 type <em>2</em> diabetic patients with nephropathy. This study had two trial phases, each for 4 weeks and one washout period for <em>2</em> weeks. Patients were randomly assigned to consume a diet containing soy milk or a diet containing cow's milk.

RESULTS

Soy milk consumption resulted in a significant reduction in d-dimer level (percent change: -3.77 vs. 16.13%; P < 0.05). This significant effect remained even after adjusting for confounding factor (carbohydrate intake). However, soy milk consumption had no significant effects on tumor necrosis factor-α, interleukin-6, high-sensitivity C-reactive protein (hs-CRP), and malondialdehyde levels. The result was near to significance regarding the effect of soy milk consumption on hs-CRP (percent change: -35.45 vs. 36.76%; P = 0.05). However, this effect was not significant after adjusting for the confounding variable (carbohydrate intake).

CONCLUSIONS

Soy milk consumption could decrease serum d-dimer level among type <em>2</em> diabetic patients with nephropathy. However, markers of inflammation and oxidative stress did not change following soy milk intake among these patients.

<strong class="sub-title"> Background: </strong> Hypercoagulability seems to contribute to SARS-CoV-<em>2</em> pneumonia pathogenesis. However, age and metabolic syndrome are potential cofounders when assessing the value of coagulation biomarkers prediction of COVID-19 outcomes. We assessed whether coagulation biomarkers, including FVIII and VWF levels, measured at time of admission were predictive of COVID-19 adverse outcomes irrespective of age and major comorbidities associated with metabolic syndrome.

<strong class="sub-title"> Methods: </strong> Blood was sampled at admission in <em>2</em>43 adult COVID-19 patients for analysis of coagulation biomarkers including FVIII and VWF on platelet-poor plasma. The association between baseline CRP, aPTT ratio, PT ratio, D-dimers, fibrinogen, FVIII, VWF:Ag and FVIII/VWF:Ag ratio levels and adverse outcomes (increased oxygen requirements, thrombosis and death at day-30) was assessed by regression analysis after adjustment on age, sex, body mass index, diabetes and hypertension.

<strong class="sub-title"> Results: </strong> In univariable regression analysis increased CRP (SHR, 1.68; 95%CI, 1.<em>2</em>6 to <em>2</em>.<em>2</em>3), increased fibrinogen (SHR, 1.3<em>2</em>; 95% CI, 1.04 to 1.68) and decreased FVIII/VWF:Ag ratio (SHR, 0.70 ; 95% CI, 0.5<em>2</em>-0.96) levels at admission were significantly associated with the risk of increased oxygen requirement during follow-up. Leucocytes (SHR, 1.36; 95%CI, 1.04 to 1.76), platelets (SHR,1.71; 95%CI, 1.11 to <em>2</em>.6<em>2</em>), D-dimers (SHR, <em>2</em>.48; 95%CI, 1.66 to 3.78), FVIII (SHR, 1.78; 95%CI, 1.17 to <em>2</em>.68) were associated with early onset of thrombosis after admission. After adjustment for age, sex, BMI, hypertension and diabetes, these associations were not modified.

Conclusion: Coagulation biomarkers are early and independent predictors of increased oxygen requirement in COVID-19 patients.

<strong class="sub-title"> Keywords: </strong> Body Mass Index; Factor VIII; Oxygen; SARS‐CoV<em>2</em>; von Willebrand Factor.

OBJECTIVE

To investigate the markers of endogenous coagulation/fibrinolysis and vascular endothelial cell function, and to assess the relationships between hemostatic parameters and diabetic vascular complications in type <em>2</em> diabetic patients.

METHODS

Coagulation and fibrinolysis parameters were measured in 9<em>2</em> type <em>2</em> diabetic patients (43 male, 49 female, mean age 50.1 +/- 13.4 years) with (n = 44) and without (n = 48) vascular diabetic complications, and in 40 nondiabetic healthy subjects (<em>2</em>0 male, <em>2</em>0 female, mean age 49.8 +/- 15.1 years). Common lipid parameters were also measured.

RESULTS

The plasma levels of fibrinogen, antithrombin III (AT III), plasminogen activator inhibitor-1 (PAI-1), von Willebrand factor (vWF) activity and prothrombin time were found to be significantly increased in the type <em>2</em> diabetic patients compared with the healthy subjects. Glycosylated hemoglobin lc was inversely correlated with plasma protein S and activated prothrombin time. Protein C and S activities were positively correlated with plasma vWF activity, and were negatively correlated with plasma t-PA levels. vWF activity was negatively correlated with plasma t-PA levels. AT III levels were positively correlated with plasma total cholesterol levels, plasma low density lipoprotein cholesterol levels, plasma triglycerides and D-dimer levels. Plasma PAI-1 levels and factor V activity in diabetic patients with microvascular complications were significantly higher than those of the diabetic patients without microvascular complications. The plasma PAI-1 and platelet count were increased in patients with diabetic retinopathy compared with the diabetic patients without retinopathy. Plasma PAI-1 levels and factor VII activity were significantly higher in the diabetic patients with nephropathy than in diabetic patients without nephropathy. Plasma concentrations of fibrinogen and PAI-1 were significantly higher in the diabetic patients with neuropathy than the diabetic patients without neuropathy.

CONCLUSIONS

The data demonstrated that patients with type <em>2</em> diabetes mellitus had a hypercoagulable state and hypofibrinolysis, thereby indicating that activation of coagulation with a reduced fibrinolytic activity may contribute to the increased risk of vascular disease in type <em>2</em> diabetic patients.

OBJECTIVE

To investigate the thrombotic tendency in patients with systemic lupus erythematosus (SLE) by evaluating congenital or acquired abnormalities associated with an increased risk of venous and/or arterial thrombosis.

METHODS

A total of 57 patients with SLE were included in the study. Twenty-one patients (37%) had a history of arterial and/or venous thrombosis and 36 patients (63%) did not have such a history. Sera from 50 healthy controls were examined. Protein C, protein S, antithrombin, D-dimer, fibrinogen, homocysteine, anticardiolipin antibodies (aCL), lupus anticoagulant (LAC), prothrombin G20210A, and methylenetetrahydrofolate reductase (MTHFR) C677T gene mutation were evaluated.

RESULTS

Protein C, antithrombin, fibrinogen, D-dimer, and homocysteine levels were significantly higher in patients with SLE than in controls. A prothrombin mutation was observed in 2 (4%) of 50 controls and in 6 (11%) of 57 patients. A significantly higher prevalence (P = 0.036) of MTHFR homozygous mutation was observed in patients with SLE (14 [25%] of 57) in comparison with controls (4 [8%] of 50). IgG-aCL and IgM-aCL levels were significantly higher in patients with SLE than in controls (P < 0.0001). The presence of medium-high >> or = 20 IgG phospholipid units/ml) IgG-aCL antibody titers was significantly higher (P = 0.005) in patients with thrombosis (11 [52%] of 21) than in patients without (5 [14%] of 36) thrombosis. LAC was present in 22 (38.5%) of 57 patients and in none of 50 controls.

CONCLUSIONS

In this study, we confirm the association between thrombosis and IgG-aCL at medium-high titers and suggest that the coexistence of other risk factors can affect the expression of thrombosis in patients with SLE.

Coagulation and fibrinolytic activities were studied in 18 subjects with Behçet's disease and compared with results from 14 matched control patients suffering from sero-negative arthritis. Significantly higher plasma concentrations (median and range) were found in Behçet's patients for the following variables: fibrinogen 3.7 (1.7-6.9) vs 3.0 (<em>2</em>.0-5.1) g/l, p less than 0.05; von Willebrand factor antigen, 115 (7<em>2</em>-344) vs 74 (60-119)%, p less than 0.00<em>2</em>; plasminogen activator activity (10(6)/ECLT<em>2</em>) <em>2</em>19 (94-3<em>2</em>9) vs 137 (78-197) units, p less than 0.00<em>2</em>; tissue plasminogen activator inhibitor (t-PA-I) activity, 9.1 (5.5-19.3) vs 5.1 (1.8-1<em>2</em>.0) IU/ml, p less than 0.00<em>2</em>; and PAI-1 antigen, 13.9 (4.5-<em>2</em>0.9) vs 6.4 (<em>2</em>.4-11.1) ng/ml, p less than 0.00<em>2</em>. Protein C antigen was significiantly lower: 97 (70-183) vs 1<em>2</em>6 (96-<em>2</em><em>2</em>0)%, p less than 0.0<em>2</em>. No differences were observed in antithrombin III activity or antigen, factor VIII coagulant activity, fibrinopeptides A and B beta 15-4<em>2</em>, plasminogen, alpha-<em>2</em>-antiplasmin, functional and immunological tissue-plasminogen activator, thrombin-antithrombin complexes and <em>D</em>-<em>dimer</em>. Levels of tissue plasminogen activator inhibitor (activity and antigen) correlated with disease activity while fibrinogen and von Willebrand factor concentrations did not. Seven of the 18 subjects with Behçet's disease had suffered thrombotic events but it was not possible to distinguish these from the 11 patients without thrombosis using the assays performed. The results suggest the abnormal fibrinolytic activity in Behçet's disease is due to increased inhibition of tissue plasminogen activator. No abnormality of coagulation or fibrinolytic activity specific to Behçet's disease was detected.

Coronavirus disease <em>2</em>019 (COVI<em>D</em>-19) is a new human infectious disease. The etiology for this outbreak is a novel coronavirus named severe acute respiratory syndrome coronavirus <em>2</em> (SARS-CoV-<em>2</em>). Thus far, related research on COVI<em>D</em>-19 is still in preliminary stage. This paper summarized the latest outcomes of corresponding study from Chinese centers and clarified the hematopoietic abnormality caused by SARS-CoV-<em>2</em> and potential mechanism. Lymphopenia was common in the early stage after the onset of COVI<em>D</em>-19. A significant decrease was observed in peripheral C<em>D</em>4+ and C<em>D</em>8+ T lymphocytes. As the illness progressed, neutrophilia emerged in several cases, and patients with severe critical pulmonary conditions showed higher neutrophils than common type. Thrombocytopenia was resulting from the consumption and/or the reduced production of platelets in damaged lungs. Anemia was not observed notably, but the decrease in hemoglobin was frequent. The activation of monocyte-macrophage system aggravates the immune damage of lung and other tissues, which leads to the increase of <em>D</em>-<em>dimer</em>, prothrombin time, and platelet consumption.

<strong class="sub-title">Keywords:</strong> COVI<em>D</em>-19; Hemoglobin; Lymphopenia; Neutrophilia; SARS-CoV-<em>2</em>; Thrombocytopenia.

<strong class="sub-title"> Introduction: </strong> Emerging data have described poor clinical outcomes from infection with the novel severe acute respiratory syndrome coronavirus <em>2</em> (SARS-CoV <em>2</em>) among African American patients and those from underserved socioeconomic groups. We sought to describe the clinical characteristics and outcomes of acute kidney injury (AKI) in this special population.

<strong class="sub-title"> Methods: </strong> This is a retrospective study conducted in an underserved area with a predominance of African American patients with coronavirus disease <em>2</em>019 (COVID-19). Descriptive statistics were used to characterize the sample population. The onset of AKI and relation to clinical outcomes were determined. Multivariate logistic regression was used to determine factors associated with AKI.

<strong class="sub-title"> Results: </strong> Nearly half (49.3%) of the patients with COVID-19 had AKI. Patients with AKI had a significantly lower baseline estimated glomerular filtration rate (eGFR) and higher FiO<em>2</em> requirement and D-dimer levels on admission. More subnephrotic proteinuria and microhematuria was seen in these patients, and the majority had a pre-renal urine electrolyte profile. Patients with hospital-acquired AKI (HA-AKI) as opposed to those with community-acquired AKI (CA-AKI) had higher rates of in-hospital death (5<em>2</em> vs. <em>2</em>3%, p = 0.005), need for vasopressors (4<em>2</em> vs. <em>2</em>5%, p = 0.0<em>2</em>4), and need for intubation (55 vs. <em>2</em>5%, p = 0.006). A history of heart failure was significantly associated with AKI after adjusting for baseline eGFR (OR 3.38<em>2</em>, 95% CI 1.1<em>2</em>1-13.<em>2</em>31, p = 0.03<em>2</em>).

Conclusion: We report a high burden of AKI among underserved COVID-19 patients with multiple comorbidities. Those who had HA-AKI had worse clinical outcomes compared to those who with CA-AKI. A history of heart failure is an independent predictor of AKI in patients with COVID-19.

Keywords: Acute kidney injury; COVID-19; Heart failure; Novel coronavirus.

OBJECTIVE

We investigated the plasma levels of molecular markers for platelet activity and the thrombotic and fibrinolytic status in patients with hypertrophic cardiomyopathy and dilated cardiomyopathy to determine the activating site of coagulation in these disorders.

BACKGROUND

A thromboembolic event is a serious complication in patients with idiopathic cardiomyopathy. However, the activating site of the coagulation system in idiopathic cardiomyopathy has not been fully investigated.

METHODS

We determined the plasma levels of molecular markers for platelet activity (platelet factor 4 and beta-thromboglobulin), thrombotic status (fibrinopeptide A and thrombin-antithrombin III complex) and fibrinolytic status (<em>D</em>-<em>dimer</em> and plasmin-alpha <em>2</em>-plasmin inhibitor complex) in 13 patients with hypertrophic cardiomyopathy, 17 patients with dilated cardiomyopathy and <em>2</em>0 normal subjects.

RESULTS

Plasma levels of platelet factor 4, beta-thromboglobulin and plasmin-alpha <em>2</em>-plasmin inhibitor complex did not differ significantly among the three groups, whereas plasma levels of fibrinopeptide A and thrombin-antithrombin III complex in both patient groups were significantly higher than those in normal subjects. Plasma levels of <em>D</em>-<em>dimer</em> in patients with dilated cardiomyopathy were significantly higher than those in patients with hypertrophic cardiomyopathy and normal groups. In patients with hypertrophic cardiomyopathy, both fibrinopeptide A and thrombin-antithrombin III complex levels were significantly correlated with left atrial diameter. In patients with dilated cardiomyopathy, fibrinopeptide A and thrombin-antithrombin III complex levels showed a positive correlation with left ventricular end-diastolic volume and a negative correlation with fractional shortening of the left ventricle.

CONCLUSIONS

The activated coagulation system in patients with hypertrophic and dilated cardiomyopathy may be triggered by left atrial dilation in hypertrophic cardiomyopathy and left ventricular enlargement and dysfunction in dilated cardiomyopathy.

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).

CONCLUSIONS

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.

OBJECTIVE

We aimed to characterize the pharmacokinetics and pharmacodynamics of drotrecogin alfa (activated) (recombinant human activated protein C) in patients with severe sepsis.

METHODS

Patients (N = 1690) in a randomized, double-blind, placebo-controlled phase 3 trial received a 96-hour infusion of placebo (n = 840) or drotrecogin alfa (activated) (n = 850), 24 microg x kg(-1) x h(-1). Plasma samples from 680 patients were collected for pharmacokinetic assessment. Pharmacodynamic effects on activated partial thromboplastin time, D-dimer, protein C, and interleukin 6 were analyzed by drotrecogin alfa (activated) steady-state plasma concentration (C(ss)) quartile.

RESULTS

Transient endogenous activated protein C concentrations above 10 ng/mL were observed in 11 placebo-treated patients (3.3%). In drotrecogin alfa (activated)-treated patients, the median C(ss) was 44.9 ng/mL and the median plasma clearance (CL(p)) was 40.1 L/h. C(ss) was reached within 2 hours after the infusion was started. Plasma concentrations were below the assay quantitation limit of 10 ng/mL within 2 hours after the infusion was stopped in 92% of patients. CL(p) increased with increasing body weight, so infusion rates should be based on predose body weight. Mean CL(p) associated with age, sex, or baseline hepatic or renal function differed by less than 30% from the mean CL(p) in all patients and resided within the interquartile range of CL(p) in all patients. Dose adjustment is not required on the basis of these factors alone or in combination. No correlation was detected between C(ss) quartile and bleeding risk or the magnitudes of effect on biomarkers of coagulopathy (D-dimers and protein C) and inflammation (interleukin 6).

CONCLUSIONS

Plasma concentrations of drotrecogin alfa (activated) attain steady state rapidly after the infusion is started and decline rapidly after the infusion is stopped. The infusion rate should be based on predose body weight and not on any other demographic or baseline clinical covariate.

BACKGROUND

It may be safe to omit additional diagnostic testing in selected patients with suspected pulmonary embolism (PE) who have a negative D-dimer test, but this approach has never been evaluated in a randomized, controlled trial.

OBJECTIVE

To determine if additional diagnostic testing can be safely withheld in patients with suspected PE who have negative erythrocyte agglutination D-dimer test results.

METHODS

Randomized comparisons in 2 subgroups of a prospective multicenter study.

METHODS

7 university hospitals.

METHODS

1126 outpatients or inpatients with suspected PE; of these, 456 patients with negative erythrocyte agglutination D-dimer test results were randomly assigned to the intervention groups. Patients were classified into 2 clinical probability groups: those with a low clinical probability of PE (low-probability group) and those with a moderate or high clinical probability of PE, a nondiagnostic ventilation-perfusion lung scan, and no evidence of proximal deep venous thrombosis on bilateral ultrasonography (moderate- or high-probability group).

METHODS

The experimental intervention for both probability groups was no further diagnostic testing for PE. The control intervention for the low-probability group was a ventilation-perfusion lung scan followed by ultrasonography of the proximal deep veins of the legs on the same day. If the lung scan was nondiagnostic, ultrasonography of the legs was repeated 7 and 14 days later. The control intervention for the moderate- or high-probability group was ultrasonography of the proximal deep veins of the legs after 7 and 14 days. In the control and experimental groups, anticoagulation was withheld or withdrawn if PE was not diagnosed.

METHODS

Symptomatic venous thromboembolism (VTE) during 6 months of follow-up.

RESULTS

Prevalence of VTE was 15.2% in the 1126 enrolled patients. In the low-probability group, VTE occurred during follow-up in 0 of 182 patients who had no additional diagnostic testing and in 1 of 185 patients who had additional testing (difference, -0.5 percentage point [95% CI, -3.0 to 1.6 percentage points]). In the moderate- or high-probability group, VTE occurred during follow-up in 1 of 41 patients who had no additional diagnostic testing and in 0 of 41 patients who had additional testing (difference, 2.4 percentage points [CI, -6.4 to 12.6 percentage points]).

CONCLUSIONS

The authors could not enroll 2000 patients as originally planned; 3 randomly assigned patients did not receive the allocated intervention, and 7 received inadequate follow-up. Personnel who performed follow-up evaluations were not blinded to the results of diagnostic testing at enrollment or to allocation group assignments.

CONCLUSIONS

In patients with a low probability of PE who have negative D-dimer results, additional diagnostic testing can be withheld without increasing the frequency of VTE during follow-up. Low clinical probability and negative D-dimer results occur in 50% of outpatients and in 20% of inpatients with suspected PE.

OBJECTIVE

The aim of this study was to investigate whether risk factors for embolism would promote thrombus formation in patients with nonvalvular atrial fibrillation (NVAF).

METHODS

Hemostatic markers for platelet activity (ie, platelet factor-4 an<em>d</em> beta-thromboglobulin [TG]), thrombotic status (ie, prothombin fragments 1 an<em>d</em> <em>2</em>), an<em>d</em> fibrinolytic status (ie, <em>d</em>-<em>dimer</em>) were <em>d</em>etermine<em>d</em> in <em>2</em>46 patients with NVAF (mean age, 66.1 years) an<em>d</em> 111 control subjects without NVAF (68.3 years).

RESULTS

The beta-TG level was higher in NVAF patients than in control subjects. D-<em>dimer</em> levels were higher in NVAF patients having risk factors (mean [+/- SE] <em>d</em>-<em>dimer</em> level, 158.6 +/- 9.<em>2</em> ng/mL) than in those without risk factors (mean <em>d</em>-<em>dimer</em> level, 9<em>2</em>.1 +/- 6.7 ng/mL; p < 0.01) an<em>d</em> in control subjects (mean <em>d</em>-<em>dimer</em> level: control subjects with risk factors, 79.1 +/- 10.3 ng/mL; control subjects without risk factors, 31.0 +/- 7.4 ng/mL; p < 0.01). NVAF (o<em>d</em><em>d</em>s ratio [OR], 3.94; 95% confi<em>d</em>ence interval [CI], 1.87 to 8.30; p = 0.0003) an<em>d</em> age of>>/= 75 years (OR, 5.68; 95% CI, <em>2</em>.87 to 11.<em>2</em>3; p < 0.0001) emerge<em>d</em> as pre<em>d</em>ictors of elevate<em>d</em> levels of <em>d</em>-<em>dimer</em>, an<em>d</em> only NVAF (OR, 10.30; 95% CI, 5.67 to 18.7<em>2</em>; p < 0.0001) emerge<em>d</em> as a pre<em>d</em>ictor of elevate<em>d</em> levels of beta-TG.

CONCLUSIONS

NVAF patients whose conditions were complicated with risk factors for embolism could be in the prothrombotic state. Advanced age is a strong predictor of the prothrombotic state in NVAF patients.

Normal pregnancy is associated with alterations of the hemostatic system towards a hypercoagulable state and an increased risk of venous thromboembolism. The risk of venous thrombosis is higher in pregnant women with factor V Leiden (FVL) than in those with wildtype factor V. Routine laboratory assays are not useful to detect hypercoagulable conditions. A prospective and systematic evaluation of hemostatic system activation in women with and without FVL during an uncomplicated pregnancy employing more sensitive markers of hypercoagulability, such as prothrombin fragment 1+<em>2</em> (F1+<em>2</em>), thrombin-antithrombin complex (TAT), <em>D</em>-<em>Dimer</em>, or the endogenous thrombin potential (ETP), an indicator of the plasma's potential to generate thrombin, has not been performed. We prospectively followed 113 pregnant women with (n = 11) and without (n = 10<em>2</em>) FVL and measured F1+<em>2</em>. TAT, <em>D</em>-<em>Dimer</em> and the ETP at the 1<em>2</em>th, <em>2</em><em>2</em>nd and 34th gestational week as well as 3 months after delivery (baseline) in each subject. None of the women developed clinical signs of venous thromboembolism during pregnancy or postpartum. Pregnant women with and without FVL exhibited substantial activation of the coagulation and fibrinolytic system as indicated by a gradual increase of F1+<em>2</em>, TAT and <em>D</em>-<em>Dimer</em> throughout uncomplicated pregnancy up to levels similar to those found in acute thromboembolic events (p < 0.0001 by analysis of variance for each parameters). Levels of F1+<em>2</em> and TAT were comparable between women with and without FVL, but levels of <em>D</em>-<em>Dimer</em> were significantly higher in women with FVL than in those without the mutation (p = 0.0005). The ETP remained unchanged in both women with and without FVL at all timepoints. Our data demonstrate a substantial coagulation and fibrinolytic system activation in healthy women with and without FVL during uncomplicated pregnancy. An elevated F1+<em>2</em>, TAT or <em>D</em>-<em>Dimer</em> level during pregnancy is not necessarily indicative for an acute thromboembolic event. The normal ETP in both women with and without FVL suggests that the capacity of the plasma to generate thrombin after in vitro activation of the clotting system is not affected by pregnancy. Higher levels of <em>D</em>-<em>Dimer</em> in women with FVL than in women with wildtype factor V at baseline as well as during pregnancy indicate increased fibrinolytic system activation in carriers of the mutation.

The <em>delta</em> subunit of Escherichia coli ATP synthase has been expressed and purified, both as the intact polypeptide and as <em>delta</em>', a proteolytic fragment composed of residues 1-134. The solution structure of <em>delta</em>' as a five-helix bundle has been previously reported (Wilkens, S., <em>D</em>unn, S. <em>D</em>., Chandler, J., <em>D</em>ahlquist, F. W., and Capaldi, R. A. (1997) Nat. Struct. Biol. 4, 198-<em>2</em>01). The <em>delta</em> subunit, in conjunction with <em>delta</em>-depleted F1-ATPase, was fully capable of reconstituting energy-dependent fluorescence quenching in membrane vesicles that had been depleted of F1. A complex of <em>delta</em> with the cytoplasmic domain of the b subunit of F0 was demonstrated and characterized by analytical ultracentrifugation using bST34-156, a form of the b domain lacking aromatic residues. Molecular weight determination by sedimentation equilibrium supported a b<em>2</em><em>delta</em> subunit stoichiometry. The sedimentation coefficient of the complex, <em>2</em>.1 S, indicated a frictional ratio of approximately <em>2</em>, suggesting that <em>delta</em> and the b <em>dimer</em> are arranged in an end-to-end rather than side-by-side manner. These results indicate the feasibility of the b<em>2</em><em>delta</em> complex reaching from the membrane to the membrane-distal portion of the F1 sector, as required if it is to serve as a second stalk.

The three-<em>d</em>imensional structure of the catalytically efficient beta-xylosi<em>d</em>ase from Selenomonas ruminantium in complex with competitive inhibitor 1,3-bis[tris(hy<em>d</em>roxymethyl)methylamino]propane (BTP) was <em>d</em>etermine<em>d</em> by using X-ray crystallography (1.3A resolution). Most H bon<em>d</em>s between inhibitor an<em>d</em> protein occur within subsite -1, inclu<em>d</em>ing one between the carboxyl group of E186 an<em>d</em> an N group of BTP. The other N of BTP occupies subsite +1 near K99. E186 (pK(a) 7.<em>2</em>) serves as catalytic aci<em>d</em>. The pH (6-10) profile for 1/K(i)((BTP)) is bell-shape<em>d</em> with pK(a)'s 6.8 an<em>d</em> 7.8 on the aci<em>d</em>ic limb assigne<em>d</em> to E186 an<em>d</em> inhibitor groups an<em>d</em> 9.9 on the basic limb assigne<em>d</em> to inhibitor. Mutation K99A eliminates pK(a) 7.8, strongly suggesting that the BTP monocation bin<em>d</em>s to the <em>d</em>ianionic enzyme D14(-)E186(-). A se<em>d</em>imentation equilibrium experiment estimates a K(<em>d</em>) ([<em>dimer</em>](<em>2</em>)/[tetramer]) of 7 x 10(-9)M. Similar k(cat) an<em>d</em> k(cat)/K(m) values were <em>d</em>etermine<em>d</em> when the tetramer/<em>dimer</em> ratio changes from 0.00<em>2</em>8 to <em>2</em>6 suggesting that <em>dimers</em> an<em>d</em> tetramers are equally active forms.

Patients with essential thrombocythemia (ET) have an increased frequency of thrombosis, but the relationship of both thrombosis and JAK<em>2</em> V617F allele burden with platelet turnover, acquired activated protein C resistance (aAPCR), and levels of coagulation factors and soluble markers of platelet, and endothelial activation is not well known. In 53 ET patients (<em>2</em>6 with a history of thrombosis), reticulated platelets (RP) percentage, aAPCR, platelet tissue factor (TF) expression, and plasma levels of TF, coagulation factors, soluble P-selectin (sP-selectin), soluble C<em>D</em>40 ligand (sC<em>D</em>40L), von Willebrand factor antigen (VWF:Ag), soluble thrombomodulin (sTM), <em>D</em>-<em>dimer</em> and prothrombin fragment 1 + <em>2</em> were compared with those in matched healthy individuals and correlated with thrombosis occurrence and JAK<em>2</em> mutational load. ET patients with thrombosis had significantly higher values for RP percentage, aAPCR, and levels of factors V and VIII, VWF:Ag, sP-selectin, and sC<em>D</em>40L than patients without thrombosis and controls. At multivariate study, RP percentage, factor V levels, and aAPCR were independently associated with an increased risk of thrombosis. Patients with JAK<em>2</em> mutation had significantly lower levels of free protein S (PS) and higher levels of TF, sP-selectin, sC<em>D</em>40L, VWF:Ag, and sTM than those with wild-type allele. A mutant allele dosage effect >>or= 1<em>2</em>%) was observed for TF, sP-selectin, sC<em>D</em>40L, VWF:Ag, and PS levels. These results support a role for platelet turnover, factor V, and aAPCR in the thrombosis of ET as well as the association between JAK<em>2</em> V617F allele burden and either decreased free PS or increased TF and soluble markers of platelet and endothelial activation.

Lipid-lowering by postmenopausal hormone therapy (HRT) explains only partly the assumed coronary risk reduction associated with therapy. To explore other possible mechanisms, we studied associations of HRT use with inflammation and hemostasis risk markers in women>>/=65 years of age. Subjects were selected from 3393 participants in the fourth year examination of the Cardiovascular Health Study, an observational study of vascular disease risk factors. After excluding women with vascular disease, we compared levels of inflammation and hemostasis variables in the <em>2</em>30 women using unopposed estrogen and 60 using estrogen/progestin, with those of 196 nonusers selected as controls. Compared with nonusers, unopposed estrogen use was associated with 59% higher mean C-reactive protein (P<0.001), but with modestly lower levels of other inflammation indicators, fibrinogen, and alpha-1 acid glycoprotein (P<0.001). Factor VIIc was 16% higher among estrogen users (P<0.001), but this was not associated with higher thrombin production (prothrombin fragment 1-<em>2</em>), or increased fibrin breakdown (<em>D</em>-<em>dimer</em>). Concentration of plasminogen activator inhibitor-1 was 50% lower in both using groups (P<0.001) compared with nonusers, and this was associated with higher plasmin-antiplasmin complex: 8% higher in estrogen and 18% higher in estrogen/progestin users (P<0. 05). Relationships between the markers and hormone use were less pronounced in estrogen/progestin users, with no association for C-reactive protein except in women in upper <em>2</em> tertiles of body mass index (P for interaction, 0.0<em>2</em>). The direction and strength of the associations of HRT use with inflammation markers differed depending on the protein, so it is not clear whether HRT confers coronary risk reduction through an inflammation-sensitive mechanism. Associations with hemostasis markers indicated no association with evidence of procoagulation and a possible association with increased fibrinolytic activity.

The activation of the clotting cascade leading to deep venous thrombosis begins during total hip arthroplasty, but few studies have assessed changes in coagulation during surgery. A better understanding of thrombogenesis during total hip arthroplasty may provide a more rational basis for treatment. In 3 separate studies, the following observations were made. Circulating indices of thrombosis and fibrinolysis: prothrombin F1.<em>2</em>, thrombin-antithrombin complexes, fibrinopeptide A, and <em>D</em>-<em>dimer</em>, did not increase during osteotomy of the neck of the femur or during insertion of the acetabular component, but rose significantly during insertion of the femoral component. Thrombin-antithrombin complexes, fibrinopeptide A, and <em>D</em>-<em>dimer</em> were higher after insertion of a cemented component than insertion of a noncemented femoral component. A significant decline in central venous oxygen tension was observed after relocation of the hip joint and after insertions of cemented and noncemented femoral components, providing evidence of femoral venous occlusion during insertion of the femoral component. In patients receiving a cemented femoral component, mean pulmonary artery pressure increased after relocation of the hip joint, indicating intraoperative pulmonary embolism. No changes in mean pulmonary artery pressure were noted with noncemented total hip arthroplasty. Administration of 1000 units of unfractionated heparin before insertion of a cemented femoral component blunted the rise of fibrinopeptide A. The results of these studies suggest that (1) the greatest risk of activation of the clotting cascade during total hip arthroplasty occurs during insertion of the femoral component; (<em>2</em>) femoral venous occlusion and use of cemented components are factors in thrombogenesis during total hip arthroplasty; and (3) measures to prevent deep venous thrombosis during total hip arthroplasty (such as intraoperative anticoagulation) should begin during surgery rather than during the postoperative period and be applied during insertion of the femoral component.

We examined the properties of membrane-spanning channels formed by gramicidin analogues that differ from [Val1]gramicidin A by having a single residue deletion or insertion at the formyl-NH terminus and of hybrid channels formed between such 14-, 15-, and 16-residue analogues. The channels' backbone structure, and helix sense, are not affected by the sequence modifications, because hybrid channels were observed for all combinations tested, and there was no excess energetic cost associated with hybrid channel formation. When hybrid channels form between analogues of different length the hybrid channel stability depends on the nature of the sequence dissimilarity. If two analogues differ by one residue (<em>delta</em> n = 1), the hybrid channels are destabilized by approximately 10 kJ/mol, because there is a defect (a "gap" in the peptide backbone) at the join between the two beta 6.3-helical monomers such that the <em>dimer</em> is stabilized by only five intermolecular C = O ... H-N hydrogen bonds rather than the usual six. This defect also alters the hybrid channels' permeability characteristics: the single-channel conductances are decreased, as if there were an additional barrier to ion movement through the channel. If the formyl-NH-terminal residue is Gly (and <em>delta</em> n = 1), the hybrid channels show multi-state behavior with voltage-dependent transitions between two conductance levels. If two analogues differ by two residues (<em>delta</em> n = <em>2</em>), the hybrid channels are stabilized by 3 kJ/mol, indicating that structural continuity at the join between the monomers has been restored, as have the hybrid channels' permeability characteristics. The increased hybrid channel stability (when <em>delta</em> n = <em>2</em>) may arise from altered membrane-channel interactions.

OBJECTIVE

We sought to determine:1) whether normal <em>D</em>-<em>dimer</em> enzyme-linked immunosorbent assay (ELISA) assays predicted the absence of pulmonary embolism (PE) in the high-volume emergency department (E<em>D</em>) of the Brigham and Women's Hospital, and <em>2</em>) whether E<em>D</em> physicians accepted normal <em>D</em>-<em>dimer</em> levels as confirmation of no PE without further diagnostic testing such as lung scanning, chest computed tomography (CT) scanning, or pulmonary angiography.

BACKGROUND

Although the plasma D-dimer ELISA is a sensitive screening test for excluding acute PE, this laboratory marker has not been widely integrated into clinical algorithms such as creatine kinase-MB fraction or troponin testing for acute myocardial infarction.

METHODS

We mandated that E<em>D</em> physicians order <em>D</em>-<em>dimer</em> ELISA tests on all patients suspected of acute PE. We reviewed the clinical record of each E<em>D</em> patient initially evaluated for suspected PE during the year <em>2</em>000. We determined whether additional imaging tests for PE were obtained and whether the final diagnosis was PE.

RESULTS

Of 1,106 <em>D</em>-<em>dimer</em> assays, 559 were elevated and 547 were normal. Only <em>2</em> of 547 had PE despite a normal <em>D</em>-<em>dimer</em>. The sensitivity of the <em>D</em>-<em>dimer</em> ELISA for acute PE was 96.4% (95% confidence interval [CI]: 87.5% to 99.6%), and the negative predictive value was 99.6% (95% CI: 98.7% to >99.9%). Nevertheless, <em>2</em>4% of patients with normal <em>D</em>-<em>dimer</em>s had additional imaging tests for PE.

CONCLUSIONS

The D-dimer ELISA has a high negative predictive value for excluding PE. By paying more attention to normal D-dimer results, fewer chest CT scans and lung scans will be required, and improvements may be realized in diagnostic efficiency and cost reduction.

The Escherichia coli cy<em>d</em>AB operon enco<em>d</em>es the high-affinity terminal oxi<em>d</em>ase of the oxygen respiratory chain, cytochrome <em>d</em> oxi<em>d</em>ase. The sensor-regulator pair, ArcB-ArcA, is responsible for the microaerobic activation of the cy<em>d</em>AB operon, whereas the anaerobic regulator Fnr represses its expression in the absence of oxygen. Fnr bin<em>d</em>s in vitro at two sites within the cy<em>d</em>AB promoter element. To <em>d</em>iscern whether these two regions have an in vivo function in the anaerobic regulation of cy<em>d</em>AB, the Fnr-bin<em>d</em>ing motifs were mutagenize<em>d</em> in<em>d</em>ivi<em>d</em>ually an<em>d</em> in combination. The effects of these mutations on in vivo gene expression were <em>d</em>etermine<em>d</em> by lac fusion an<em>d</em> primer extension analysis. Our results show that the Fnr-<em>2</em> site is critical for Fnr-me<em>d</em>iate<em>d</em> anaerobic repression of the two main cy<em>d</em>AB promoters, P1 an<em>d</em> P<em>2</em>. In contrast, the Fnr-1 site has an auxiliary role in the anaerobic repression of P1, but not of P<em>2</em>. Transcription from P1 <em>d</em>i<em>d</em> not affect ArcA-me<em>d</em>iate<em>d</em> activation or Fnr-me<em>d</em>iate<em>d</em> repression of P<em>2</em>, in<em>d</em>icating that oxygen regulation is exerte<em>d</em> on both promoters in an in<em>d</em>epen<em>d</em>ent fashion. In a<em>d</em><em>d</em>ition, three new promoters were i<em>d</em>entifie<em>d</em> in the cy<em>d</em>AB control region, an<em>d</em> the 5' en<em>d</em>s of the correspon<em>d</em>ing transcripts were mappe<em>d</em>. Two of these promoters, <em>d</em>esignate<em>d</em> P3 an<em>d</em> P4, are co-or<em>d</em>inately regulate<em>d</em> with P1 an<em>d</em> P<em>2</em> in response to oxygen, ArcA an<em>d</em> Fnr. The P5 promoter is not Fnr regulate<em>d</em> an<em>d</em> is only weakly activate<em>d</em> by ArcA. The contribution of these three a<em>d</em><em>d</em>itional promoters to the overall cy<em>d</em>AB expression is most relevant un<em>d</em>er aerobic con<em>d</em>itions. Our results suggest a unique repression mo<em>d</em>el, in which one Fnr <em>dimer</em> boun<em>d</em> to one single site (Fnr-<em>2</em>) is sufficient to <em>d</em>ownregulate transcription from four cy<em>d</em>AB promoters. In conclusion, transcription of the cy<em>d</em>AB operon is <em>d</em>riven by a complex regulatory element containing at least five promoters that act in unison to provi<em>d</em>e a<em>d</em>equate oxygen control of gene expression.