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Objectives: To investigate epidemiology, demography, and genetic and clinical characteristics of patients with familial Mediterranean fever (FMF) in Denmark. Method: In this population-based, cross-sectional cohort study, we identified FMF patients from discharge diagnoses using ICD-10 codes in the Danish National Patient Register, and linked data from the Danish Civil Registration System and laboratory databases for results of MEFV gene variant screening. Results: We identified 495 FMF patients (prevalence 1:11 680) with a median age of 29 years and a female ratio of 51%. The median age at diagnosis of FMF was 13 (IQR 7-22) years, with an estimated median diagnostic delay of 3 (IQR 0.7-6.9) years. The predominant ethnicities were Turkish (41.8%), Lebanese (15.8%), Syrian (6.5%), South-West Asian (7.9%), and South-East Asian (3.0%). The MEFV genotype distribution was 18.7% homozygous, 21.2% compound heterozygous, 32.0% heterozygous, 11.0% with complex alleles or unresolved zygosity, and 17.1% with no detected variants. M694V was the most prevalent variant in the overall cohort (32.5%). Homozygous or compound heterozygous MEFV exon 10 variants were associated with younger age at diagnosis (p < 0.001) and reduced number of hospital contacts before diagnosis (p = 0.008). The Charlson Comorbidity Index was ≥ 2 in 8.1% of patients. The prevalence of amyloidosis was 1.0%. Conclusions: FMF in Denmark is rare and patients are mainly of Eastern Mediterranean ethnicity. Diagnostic delay was long but patients with exon 10 MEFV variants were diagnosed at a younger age. Prolonged diagnostic delay is probably caused by lack of FMF awareness in the Danish healthcare system. [ABSTRACT FROM AUTHOR]

The continuing surge in commercial vessel sizes is putting increasing pressures on the world's port authorities to adopt effective expansion strategies to ensure that their asset is able to meet growing capacity demands. Among the key challenges is to assure that correct strategic planning and operational measures are adopted to guarantee safe and efficient traffic not only through its shipping channel, but also at the port berthing facilities. DHI and FORCE TECHNOLOGY have collaborated to develop a novel physicsbased vessel traffic management system named NCOS ONLINE. It is capable of taking into account of any relevant vessel constraints such as under-keel clearance (UKC), maneuverability and berth configuration that may constrict the movement of vessels through the channel or operability at berth, facilitating scenario planning and capacity assessment of proven unparalleled accuracy. The system incorporates the accuracy of high-end Full Mission Bridge Simulators with regards to vessel response under power and at berth. The underlying computational engines uses a powerful 3D panel method for vessel response calculations in combination with highly detailed environmental data such as wind, waves and hydrodynamics (water level and currents) simulated by use of MIKE Powered by DHI's recognized and scientific based computational models. The modular and integrated framework-based system has already been adopted by numerous port authorities, terminal operators and pilots worldwide for strategic port planning, design and 24/7 operational vessel traffic management. The paper focus on presenting the underlying equational framework and validation of the underlying physical response engines and provide a brief introduction to how they are integrated and operated through a series of user-friendly web dashboards. [ABSTRACT FROM AUTHOR]

Objective: The autoinflammatory disease familial Mediterranean fever (FMF), characterized by recurrent attacks of sterile fever, serosal, and/or synovial inflammation, is caused by variants in the Mediterranean fever gene, MEFV , coding for the pyrin inflammasome sensor. The diagnosis of FMF is mainly based on clinical symptoms and confirmed by detection of disease-associated MEFV variants. However, the diagnosis is challenging among patients carrying variants of uncertain clinical significance (VUS). In this study, we aimed to identify potential FMF discriminatory diagnostic markers in a cohort of clinically characterized FMF patients., Method: We established a cohort of clinically and MEFV genotype-characterized FMF patients by enrolling patients from major Danish hospitals (n = 91). The secretory profile of pyrin inflammasome-activated monocytes from healthy donors (HDs) and MEFV -characterized FMF patients (n = 28) was assessed by analysing cell supernatants for a custom-designed panel of 23 cytokines, chemokines, and soluble tumour necrosis factor receptors associated with monocyte and macrophage function., Results: MEFV genotypes in Danish FMF patients were associated with age at symptom onset (p < 0.05), FMF among relatives (p < 0.01), proportion of patients in colchicine treatment (p < 0.01), and treatment response (p < 0.05). Secretion of chemokines CCL1 and CXCL1 from pyrin-activated FMF monocytes was significantly decreased compared to HDs (p < 0.05), and could discriminate FMF patients with 'non-confirmatory' MEFV genotypes from HDs with 80.0% and 70.0% sensitivity for CCL1 and CXCL1, respectively (p < 0.05)., Conclusion: Our data suggest that a functional diagnostic assay based on CCL1 or CXCL1 levels in pyrin-activated patient monocytes may contribute to FMF diagnosis in patients with VUS.

Introduction: Diabetes mellitus is complicated by accelerated atherosclerosis, resulting in an increased risk of coronary artery disease (CAD) and thrombosis. Despite the proven benefits of aspirin, previous studies indicate a reduced cardiovascular protection from aspirin in diabetic patients. We aimed to investigate whether diabetes mellitus influenced the platelet response to aspirin in patients with CAD., Materials and Methods: Platelet aggregation and activation were evaluated during aspirin treatment in 85 diabetic and 92 non-diabetic patients with CAD. Adherence to aspirin was carefully controlled. All patients had CAD verified by coronary angiography and were taking 75 mg non-enteric coated aspirin daily., Results: Diabetic patients showed significantly higher levels of platelet aggregation compared to non-diabetic patients evaluated by VerifyNow® Aspirin (p=0.03) and Multiplate® aggregometry using arachidonic acid (AA) 0.5 mM (p=0.005) and 1.0 mM (p=0.009). In addition, platelet activation determined by soluble P-selectin was significantly higher in diabetics compared to non-diabetics (p=0.005). The higher AA-induced aggregation was associated with higher levels of HbA(1c). Compliance was confirmed by low levels of serum thromboxane B(2) (below 7.2 ng/mL). Diabetics had significantly higher levels of serum thromboxane B(2) (p<0.0001)., Conclusions: Diabetic patients with CAD had significantly higher levels of both platelet aggregation and activation compared to non-diabetic patients with CAD despite treatment with the same dosage of aspirin. These findings may partly explain the reduced cardiovascular protection from aspirin in diabetic patients., (Copyright © 2010 Elsevier Ltd. All rights reserved.)

Utilizing structure-based design, we have previously demonstrated that it is possible to obtain selective inhibitors of protein-tyrosine phosphatase 1B (PTP1B). A basic nitrogen was introduced into a general PTP inhibitor to form a salt bridge to Asp48 in PTP1B and simultaneously cause repulsion in PTPs containing an asparagine in the equivalent position [Iversen, L. F., et al. (2000) J. Biol. Chem. 275, 10300-10307]. Further, we have recently demonstrated that Gly259 in PTP1B forms the bottom of a gateway that allows easy access to the active site for a broad range of substrates, while bulky residues in the same position in other PTPs cause steric hindrance and reduced substrate recognition capacity [Peters, G. H., et al. (2000) J. Biol. Chem. 275, 18201-18209]. The current study was undertaken to investigate the feasibility of structure-based design, utilizing these differences in accessibility to the active site among various PTPs. We show that a general, low-molecular weight PTP inhibitor can be developed into a highly selective inhibitor for PTP1B and TC-PTP by introducing a substituent, which is designed to address the region around residues 258 and 259. Detailed enzyme kinetic analysis with a set of wild-type and mutant PTPs, X-ray protein crystallography, and molecular modeling studies confirmed that selectivity for PTP1B and TC-PTP was achieved due to steric hindrance imposed by bulky position 259 residues in other PTPs.

The accelerated formation of advanced glycation end-products (AGEs) due to elevated glycemia has repeatedly been reported as a central pathogenic factor in the development of diabetic microvascular complications. The effects of a novel inhibitor of AGE formation, NNC39-0028 (2,3-diaminophenazine), and a breaker of already formed AGE cross-links, N-phenacylthiazolium bromide (PTB), were investigated in streptozotocin-diabetic female Wistar rats. Diabetes for 24 weeks resulted in decreased tail collagen pepsin solubility, reflecting the formation of AGE cross-linking. Collagen solubility was significantly ameliorated by treatment with NNC39-0028, whereas PTB had no effect. Increased urinary albumin excretion (UAE) in diabetic rats was observed in serial measurements throughout the study period, and was not reduced by any treatment. Vascular dysfunction in the eye, measured as increased clearance of 125I-albumin, was induced by diabetes. NNC39-0028 did not affect this abnormality. This study demonstrated a pharmacological inhibition of collagen solubility alterations in diabetic rats without affecting diabetes-induced pathophysiology such as the increase in UAE or albumin clearance. Treatment with PTB, a specific breaker of AGE cross-links, had no effects in this study.

The aim of this study was to define the structural elements that determine the differences in substrate recognition capacity of two protein-tyrosine phosphatases (PTPs), PTP1B and PTPalpha, both suggested to be negative regulators of insulin signaling. Since the Ac-DADE(pY)L-NH(2) peptide is well recognized by PTP1B, but less efficiently by PTPalpha, it was chosen as a tool for these analyses. Calpha regiovariation analyses and primary sequence alignments indicate that residues 47, 48, 258, and 259 (PTP1B numbering) define a selectivity-determining region. By analyzing a set of DADE(pY)L analogs with a series of PTP mutants in which these four residues were exchanged between PTP1B and PTPalpha, either in combination or alone, we here demonstrate that the key selectivity-determining residue is 259. In PTPalpha, this residue is a glutamine causing steric hindrance and in PTP1B a glycine allowing broad substrate recognition. Significantly, replacing Gln(259) with a glycine almost turns PTPalpha into a PTP1B-like enzyme. By using a novel set of PTP inhibitors and x-ray crystallography, we further provide evidence that Gln(259) in PTPalpha plays a dual role leading to restricted substrate recognition (directly via steric hindrance) and reduced catalytic activity (indirectly via Gln(262)). Both effects may indicate that PTPalpha regulates highly selective signal transduction processes.

Dicarbonyl compounds are supposed to be reactive intermediates in the non-enzymatic glycation of proteins. A process that eventually could lead to the development of late diabetic complications. Glyoxal lysine dimer (GOLD) and methylglyoxal lysine dimer (MOLD) have previously been described as such reactive dicarbonyls. Here a new compound 3-deoxyglucosone lysine dimer (DOLD), a cross-link resulting from the reaction between hippuryl-lysine and 3-deoxyglucosone, has been isolated by HPLC and the structure determined by mass spectrometry and NMR.

Human alpha 2-macroglobulin (alpha 2M) is a unique 720-kDa proteinase inhibitor with a broad specificity. Unlike most other proteinase inhibitors, it does not inhibit proteolytic activity by blocking the active site of the proteinase. During complex formation with a proteinase, alpha 2M entraps the proteinase molecule in a reaction that involves large conformational changes in alpha 2M. We describe the molecular cloning of alpha 2M cDNA from the human hepatoblastoma cell line HepG2. The cDNA was subcloned under control of the adenovirus major late promoter in a mammalian expression vector and introduced into the baby hamster kidney (BHK) cell line. Transformed clones were isolated and tested for production of human alpha 2M with a specific enzyme-linked immunosorbent assay. Human recombinant alpha 2M (r alpha 2M), secreted and purified from isolated transfected BHK cell lines, was structurally and functionally compared to alpha 2M purified from human serum. The results show that r alpha 2M was secreted from the BHK cells as an active proteinase-binding tetramer with functional thiol esters. Cleavage reactions of r alpha 2M with methylamine and trypsin showed that the recombinant product, which was correctly processed at the N-terminus, exhibited molecular characteristics similar to those of the human serum derived reference. Moreover, r alpha 2M-trypsin complex bound to purified human placental alpha 2M receptor with an affinity indistinguishable from that of a complex formed from serum-derived alpha 2M and trypsin.

A rapid assay to detect and quantitate immunoglobulin A1 (IgA1) protease activity was developed by the use of a high-performance gel-permeation chromatography column. The assay measured the disappearance of intact substrate and the emergence of cleavage fragments and the results could be expressed in absolute units. The utility of the assay was demonstrated in the partial purification of an IgA1 protease from a strain of Haemophilus influenzae.

The secretory IgA system plays an important role in protecting the mucous membranes of the respiratory tract from attacks by microorganisms and potential allergens. We present evidence of in vivo cleavage of S-IgA1 in nasopharyngeal secretions by IgA1 proteases excreted by certain bacteria colonizing the upper respiratory tract. A procedure in two stages, which includes separation of secretion constituents by HPLC and subsequent immunochemical analysis of the fractions by two ELISA systems, identified the S-IgA fragments observed in some nasopharyngeal secretions as intact (FC alpha)2 . SC and Fab alpha, respectively. It is conceivable that colonization of areas of the respiratory tract by increased numbers of IgA1 protease-producing bacteria might cause a local impairment of the mucosal immune barrier. It is hypothesized that such bacterium-induced changes may be a primary event in the pathogenesis of certain inflammatory respiratory diseases and some forms of atopy.