Laboratory of Clinical Chemistry and Haematology

Venlo, Netherlands

Laboratory of Clinical Chemistry and Haematology

Venlo, Netherlands
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Haitjema S.,University Utrecht | Meddens C.A.,University Utrecht | Van Der Laan S.W.,Wilhelmina Childrens Hospital | Kofink D.,Regenerative Medicine Center Utrecht | And 26 more authors.
Circulation: Cardiovascular Genetics | Year: 2017

Background - As genome-wide association efforts, such as CARDIoGRAM and METASTROKE, are ongoing to reveal susceptibility loci for their underlying disease - atherosclerotic disease - identification of candidate genes explaining the associations of these loci has proven the main challenge. Many disease susceptibility loci colocalize with DNA regulatory elements, which influence gene expression through chromatin interactions. Therefore, the target genes of these regulatory elements can be considered candidate genes. Applying these biological principles, we used an alternative approach to annotate susceptibility loci and identify candidate genes for human atherosclerotic disease based on circular chromosome conformation capture followed by sequencing. Methods and Results - In human monocytes and coronary endothelial cells, we generated 63 chromatin interaction data sets for 37 active DNA regulatory elements that colocalize with known susceptibility loci for coronary artery disease (CARDIoGRAMplusC4D) and large artery stroke (METASTROKE). By circular chromosome conformation capture followed by sequencing, we identified a physical 3-dimensional interaction with 326 candidate genes expressed in at least 1 of these cell types, of which 294 have not been reported before. We highlight 16 genes based on expression quantitative trait loci. Conclusions - Our findings provide additional candidate-gene annotation for 37 disease susceptibility loci for human atherosclerotic disease that are of potential interest to better understand the complex pathophysiology of cardiovascular diseases. © 2017 American Heart Association, Inc.

Bonadonna P.,Allergy Unit | Zanotti R.,University of Verona | Melioli G.,Instituto Giannina Gaslini | Antonini F.,Instituto Giannina Gaslini | And 4 more authors.
Allergy: European Journal of Allergy and Clinical Immunology | Year: 2012

Background: Systemic mastocytosis (SM) may be associated with hymenoptera allergy. In such cases, immunotherapy is a life-saving treatment, but a circumstantiated diagnosis is needed for its prescription. Patients with SM and previous reactions to stings, but with negative tests represent a diagnostic dilemma. The basophil activation test (BAT) may be helpful in refining the diagnosis. Objective: We assessed the usefulness of BAT in subpopulations of mastocytosis patients, including those with negative tests for insect allergy. Methods: Within a population of patients with mastocytosis and previous stings, we studied by BAT and augmented intradermal test (IDT) (10 μg/ml) two groups: (1) with reactions to stings and negative tests; (2) without reactions and negative tests. Basophil activation test was performed with different venoms, assessing at flow cytometry basophils' activation. Results: Sixty-three patients had mastocytosis and 52 had reactions to previous hymenoptera stings. Of them, seven proved negative to diagnostic tests. In six of seven of those patients, BAT was negative with all venoms, and in one, basophils resulted activated also with the negative control. In six patients without previous reactions and negative tests, BAT was totally negative in five of six patients and weakly positive to Hornet in one. Finally, the IDT at 10 μg/ml venom produced nonspecific positive results in most cases. Conclusion: In patients with mastocytosis, the negative results of standard tests are reliable, because BAT and IDT at higher concentration do not add useful information. © 2012 John Wiley & Sons A/S.

Shab-Bidar S.,Tehran University of Medical Sciences | Bours S.P.G.,Maastricht University | Geusens P.P.M.M.,Maastricht University | Geusens P.P.M.M.,Hasselt University | And 4 more authors.
European Journal of Endocrinology | Year: 2013

Objective: Guidelines on the need for dose adaptation of vitamin D3 supplementation according to baseline serum 25(OH)D are inconclusive. The effects of increasing doses of vitamin D3 at lower baseline serum 25(OH)D values on the serum 25(OH)D after 4.2 and 11 months were determined in an observational study. Design: A prospective observational study. Methods: Out of 1481 consecutive women and men with a recent clinical fracture, 707 had a baseline 25(OH)D level <50 nmol/I and were supplemented with increasing doses of vitamin D3 (400, 800, 1700, and ≥3500 IU/day) according to the lower baseline 25(OH)D. Final analysis was restricted to the 221 participants who had full follow-up data available for 11 months. Results: Serum 25(OH)D ≥50 nmol/I was achieved in 5 7-76% of patients after 4.2 months and in 73-79% after 11 months. These percentages were similar for all doses (P = 0.06 and P = 0.91 respectively). The mean achieved 25(OH)D was similar for all dose groups (56.1-64.0 nmol/I after 4.2 months and 60.2-76.3 nmol/I after 11 months). With multivariate analysis, the increase in 25(OH)D (17 ± 32.0 after 4.2 months and 24.3 ± 34.0 nmol/I after 11 months) was dependent on the baseline 25(OH)D (P<0.001), not on suppIementation dose, season, age, BMI, or gender. Conclusions: The increase in serum 25(OH)D was significantIy Iarger with higher vitamin D3 suppIementation doses. However, this dose-effect response was mainly explained by the baseline 25(OH)D, not the supplementation dose, with a greater magnitude of response at lower baseline 25(OH)D concentrations. In 21-2 7% of patients, serum 25(OH)D3 levels did not reach 50 nmol/l after 11 months, at any dose. Further studies are needed to identify possibIe causes of suboptimal response such as non-compliance, undiagnosed malabsorption syndromes, or variability in cholecalciferol content of the vitamin D supplements. © 2013 European Society of Endocrinology.

Tersteeg C.,Laboratory of Clinical Chemistry and Haematology | Tersteeg C.,Laboratory of Experimental Cardiology | Heijnen H.F.,Laboratory of Clinical Chemistry and Haematology | Heijnen H.F.,Cell Microscopy Center | And 10 more authors.
Circulation Research | Year: 2014

RATIONALE:: Platelets are the most important cells in the primary prevention of blood loss after injury. In addition, platelets are at the interface between circulating leukocytes and the (sub)endothelium regulating inflammatory responses. OBJECTIVE:: Our aim was to study the dynamic process that leads to the formation of procoagulant and proinflammatory platelets under physiological flow. METHODS AND RESULTS:: In the present study, we describe the formation of extremely long, negatively charged membrane strands that emerge from platelets adhered under flow. These flow-induced protrusions (FLIPRs) are formed in vitro on different physiological substrates and are also detected in vivo in a mouse carotid injury model. FLIPRs are formed downstream the adherent and activated platelets and reach lengths of 250 μm. FLIPR formation is shear-dependent and requires cyclophilin D, calpain, and Rac1 activation. It is accompanied by a disassembly of the F-actin and microtubule organization. Monocytes and neutrophils roll over FLIPRs in a P-selectin/P-selectin glycoprotein ligand-1-dependent manner, retrieving fragments of FLIPRs as microparticles on their surface. Consequently, monocytes and neutrophils become activated, as demonstrated by increased CD11b expression and L-selectin shedding. CONCLUSIONS:: The formation of long platelet membrane extensions, such as the ones presented in our flow model, may pave the way to generate an increased membrane surface for interaction with monocytes and neutrophils. Our study provides a mechanistic model for platelet membrane transfer and the generation of monocyte/neutrophil-microparticle complexes. We propose that the formation of FLIPRs in vivo contributes to the well-established proinflammatory function of platelets and platelet-derived microparticles. © 2014 American Heart Association, Inc.

Marks M.S.,University of Pennsylvania | Heijnen H.F.G.,Laboratory of Clinical Chemistry and Haematology | Raposo G.,University Pierre and Marie Curie | Raposo G.,French National Center for Scientific Research
Current Opinion in Cell Biology | Year: 2013

Lysosome-related organelles (LROs) comprise a group of cell type-specific subcellular compartments with unique composition, morphology and structure that share some features with endosomes and lysosomes and that function in varied processes such as pigmentation, hemostasis, lung plasticity and immunity. In recent years, studies of genetic diseases in which LRO functions are compromised have provided new insights into the mechanisms of LRO biogenesis and the regulated secretion of LRO contents. These insights have revealed previously unappreciated specialized endosomal sorting processes in all cell types, and are expanding our views of the plasticity of the endosomal and secretory systems in adapting to cell type-specific needs. © 2013 Elsevier Ltd.

Bours S.P.G.,VieCuri Medisch Centrum Noord Limburg | Bours S.P.G.,Maastricht University | Van Geel T.A.C.M.,Maastricht University | Geusens P.P.M.M.,Maastricht University | And 7 more authors.
Journal of Clinical Endocrinology and Metabolism | Year: 2011

Background: Previously undetected contributors to secondary osteoporosis and metabolic bone diseases (SECOB) are frequently found in patients with osteoporosis, but the prevalence in patients at the time they present with a clinical fracture is unknown. Methods: All consecutive patients with a recent clinical vertebral or nonvertebral fracture, who were able and willing to be investigated (n = 626: 482 women, 144 men, age range 50-97 yr) had bone mineral density and laboratory investigations (serum calcium, inorganic phosphate, 25-hydroxyvitamin D, creatinine, intact PTH, TSH, free T4, serum and urine protein electrophoresis, and in men also serum testosterone). Results: Known SECOB contributors were present in 23.0% of patients and newly diagnosed SECOB contributors in 26.5%: monoclonal proteinemia (14 of 626), renal insufficiency grade III or greater (54 of 626), primary (17 of 626) and secondary (64 of 626) hyperparathyroidism, hyperthyroidism (39 of 626), and hypogonadism in men (12 of 144). Newly diagnosed SECOBs, serum 25-hydroxyvitamin D less than 50 nmol/liter (in 63.9%), and dietary calcium intake less than 1200 mg/d (in 90.6%) were found at any age, in both sexes, after any fracture (except SECOB in men with finger and toe fractures) and at any level of bone mineral density. Conclusion: At presentation with a fracture, 26.5% of patients have previously unknown contributors to SECOB, which are treatable or need follow-up, and more than 90% of patients have an inadequate vitamin D status and/or calcium intake. Systematic screening of patients with a recent fracture identifies those in whom potentially reversible contributors to SECOB and calcium and vitamin D deficiency are present. Copyright © 2011 by The Endocrine Society.

Tersteeg C.,Laboratory of Clinical Chemistry and Haematology | Tersteeg C.,Laboratory of Experimental Cardiology | De Maat S.,Laboratory of Clinical Chemistry and Haematology | De Meyer S.F.,Catholic University of Leuven | And 8 more authors.
Circulation | Year: 2014

BACKGROUND - : Von Willebrand factor (VWF) multimer size is controlled through continuous proteolysis by ADAMTS13 (a disintegrin and metalloproteinase with a thrombospondin type I motif, member 13). This prevents spontaneous platelet agglutination and microvascular obstructions. ADAMTS13 deficiency is associated with thrombotic thrombocytopenic purpura, in which life-threatening episodes of microangiopathy damage kidneys, heart, and brain. Enigmatically, a complete ADAMTS13 deficiency does not lead to continuous microangiopathy. We hypothesized that plasmin, the key enzyme of the fibrinolytic system, serves as a physiological backup enzyme for ADAMTS13 in the degradation of pathological platelet-VWF complexes. METHODS AND RESULTS - : Using real-time microscopy, we determined that plasmin rapidly degrades platelet-VWF complexes on endothelial cells in absence of ADAMTS13, after activation by urokinase-type plasminogen activator or the thrombolytic agent streptokinase. Similarly, plasmin degrades platelet-VWF complexes in platelet agglutination studies. Plasminogen directly binds to VWF and its A1 domain in a lysine-dependent manner, as determined by enzyme-linked immunosorbent assay. Plasma levels of plasmin-α2-antiplasmin complexes increase with the extent of thrombocytopenia in patients with acute episodes of thrombotic thrombocytopenic purpura, independent of ADAMTS13 activity. This indicates that plasminogen activation takes place during microangiopathy. Finally, we show that the thrombolytic agent streptokinase has therapeutic value for Adamts13 mice in a model of thrombotic thrombocytopenic purpura. CONCLUSIONS - : We propose that plasminogen activation on endothelial cells acts as a natural backup for ADAMTS13 to degrade obstructive platelet-VWF complexes. Our findings indicate that thrombolytic agents may have therapeutic value in the treatment of microangiopathies and may be useful to bypass inhibitory antibodies against ADAMTS13 that cause thrombotic thrombocytopenic purpura. © 2014 American Heart Association, Inc.

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