Koen J Dechering

Universiteit Twente, Enschede, Provincie Overijssel, Netherlands

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Publications (32)175.29 Total impact

  • [Show abstract] [Hide abstract]
    ABSTRACT: The development of drugs and vaccines to reduce malaria transmission is an important part of eradication plans. The transmission reducing activity (TRA) of these agents is currently determined in the standard membrane feeding assay (SMFA) based on subjective microscopical read-outs and with limitations in up-scaling and throughput. Utilising a Plasmodium falciparum strain expressing the firefly luciferase protein, we present a luminescence based approach to SMFA evaluation that eliminates the requirement for mosquito dissections in favour of a simple approach where whole mosquitoes are homogenised and examined directly for luciferase activity. Analysis of 6860 Anopheles stephensi mosquitoes across 68 experimental feeds shows that the luminescence assay was as sensitive as microscopy for infection detection. The mean luminescence intensity of individual and pooled mosquitoes accurately quantifies mean oocyst intensity and generates comparable TRA estimates. The luminescence assay presented here could increase SMFA throughput so that 10-30 experimental feeds could be evaluated in a single 96-well plate. This new method of assessing Plasmodium infection and transmission intensity could expedite the screening of novel drug compounds, vaccine candidates and sera from malaria exposed individuals for TRA. Luminescence-based estimates of oocyst intensity in individual mosquitoes should be interpreted with caution.
    The Journal of Infectious Diseases 05/2014; · 5.85 Impact Factor
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    ABSTRACT: MicroRNAs (miRNAs) are a family of small, non-coding single-stranded RNA molecules involved in post-transcriptional regulation of gene expression. As such, they are believed to play a role in regulating the step-wise changes in gene expression patterns that occur during cell fate specification of multipotent stem cells. Here, we have studied whether terminal differentiation of C2C12 myoblasts is indeed controlled by lineage-specific changes in miRNA expression. Using a previously generated RNA polymerase II (Pol-II) ChIP-on-chip dataset, we show differential Pol-II occupancy at the promoter regions of six miRNAs during C2C12 myogenic versus BMP2-induced osteogenic differentiation. Overexpression of one of these miRNAs, miR-378, enhances Alp activity, calcium deposition and mRNA expression of osteogenic marker genes in the presence of BMP2. Our results demonstrate a previously unknown role for miR-378 in promoting BMP2-induced osteogenic differentiation.
    BMC Molecular Biology 01/2014; 15(1):1. · 2.80 Impact Factor
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    ABSTRACT: Achieving the goal of malaria elimination will depend on targeting Plasmodium pathways essential across all life stages. Here we identify a lipid kinase, phosphatidylinositol-4-OH kinase (PI(4)K), as the target of imidazopyrazines, a new antimalarial compound class that inhibits the intracellular development of multiple Plasmodium species at each stage of infection in the vertebrate host. Imidazopyrazines demonstrate potent preventive, therapeutic, and transmission-blocking activity in rodent malaria models, are active against blood-stage field isolates of the major human pathogens P. falciparum and P. vivax, and inhibit liver-stage hypnozoites in the simian parasite P. cynomolgi. We show that imidazopyrazines exert their effect through inhibitory interaction with the ATP-binding pocket of PI(4)K, altering the intracellular distribution of phosphatidylinositol-4-phosphate. Collectively, our data define PI(4)K as a key Plasmodium vulnerability, opening up new avenues of target-based discovery to identify drugs with an ideal activity profile for the prevention, treatment and elimination of malaria.
    Nature 11/2013; · 38.60 Impact Factor
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    ABSTRACT: OBJECTIVES: Plasmodium gametocytes, responsible for malaria parasite transmission from humans to mosquitoes, represent a crucial target for new antimalarial drugs to achieve malaria elimination/eradication. We developed a novel colorimetric screening method for anti-gametocyte compounds based on the parasite lactate dehydrogenase (pLDH) assay, already standardized for asexual stages, to measure gametocyte viability and drug susceptibility. METHODS: Gametocytogenesis of 3D7 and NF54 Plasmodium falciparum strains was induced in vitro and asexual parasites were depleted with N-acetylglucosamine. Gametocytes were treated with dihydroartemisinin, epoxomicin, methylene blue, primaquine, puromycin or chloroquine in 96-well plates and the pLDH activity was evaluated using a modified Makler protocol. Mosquito infectivity was measured by the standard membrane feeding assay (SMFA). RESULTS: A linear correlation was found between gametocytaemia determined by Giemsa staining and pLDH activity. A concentration-dependent reduction in pLDH activity was observed after 72 h of drug treatment, whereas an additional 72 h of incubation without drugs was required to obtain complete inhibition of gametocyte viability. SMFA on treated and control gametocytes confirmed that a reduction in pLDH activity translates into reduced oocyst development in the mosquito vector. CONCLUSIONS: The gametocyte pLDH assay is fast, easy to perform, cheap and reproducible and is suitable for screening novel transmission-blocking compounds, which does not require parasite transgenic lines.
    Journal of Antimicrobial Chemotherapy 05/2013; · 5.34 Impact Factor
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    ABSTRACT: Mesenchymal stromal cells (hMSCs) are advancing into the clinic but the therapeutic efficacy of hMSCs faces the problem of donor variability. In bone tissue engineering, no reliable markers have been identified which are able to predict the bone-forming capacity of hMSCs prior to implantation. To this end, we isolated hMSCs from 62 donors and characterized systematically their in vitro lineage differentiation capacity, gene expression signature and in vivo capacity for ectopic bone formation. Our data confirms the large variability of in vitro differentiation capacity which did not correlate with in vivo ectopic bone formation. Using DNA microarray analysis of early passage hMSCs we identified a diagnostic bone-forming classifier. In fact, a single gene, CADM1, strongly correlated with the bone-forming capacity of hMSCs and could be used as a reliable in vitro diagnostic marker. Furthermore, data mining of genes expressed correlating with in vivo bone formation represented involvement in neurogenic processes and Wnt signaling. We will apply our data set to predict therapeutic efficacy of hMSCs and to gain novel insight in the process of bone regeneration. Our bio-informatics driven approach may be used in other fields of cell therapy to establish diagnostic markers for clinical efficacy.
    Biomaterials 03/2013; · 8.31 Impact Factor
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    ABSTRACT: DNA methylation is an important epigenetic regulator of gene expression. Abnormalities in DNA methylation patterns have been associated with various developmental and proliferative diseases, particularly cancer. Targeting DNA methyltransferases (DNMTs) represents a promising strategy for the treatment of such diseases. Current DNMT inhibitors suffer important drawbacks with respect to their efficacy, specificity, and toxicity. In this study, we have set up a robust in vitro bacterial M.SssI DNMT activity assay to systematically screen a collection of 26 240 compounds that were predicted to compete with the S-adenosyl-L-methionine (SAM) substrate of DNMT. This resulted in the identification of a novel set of structurally distinct inhibitors of M.SssI DNMT activity. Although molecular docking studies using an M.SssI homology model suggest that these compounds might compete with SAM binding, mode of activity (MoA) assays are still needed to confirm this hypothesis. Our set of novel M.SssI DNMT inhibitors, once confirmed in an orthogonal DNMT assay, may thus serve as a starting point to identify and characterize suitable lead candidates for further drug optimization.
    Journal of Biomolecular Screening 10/2012; · 2.21 Impact Factor
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    Malaria Journal 10/2012; 11(1). · 3.49 Impact Factor
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    ABSTRACT: We present here the x-ray structures of the progesterone receptor (PR) in complex with two mixed profile PR modulators whose functional activity results from two differing molecular mechanisms. The structure of Asoprisnil bound to the agonist state of PR demonstrates the contribution of the ligand to increasing stability of the agonist conformation of helix-12 via a specific hydrogen-bond network including Glu(723). This interaction is absent when the full antagonist, RU486, binds to PR. Combined with a previously reported structure of Asoprisnil bound to the antagonist state of the receptor, this structure extends our understanding of the complex molecular interactions underlying the mixed agonist/antagonist profile of the compound. In addition, we present the structure of PR in its agonist conformation bound to the mixed profile compound Org3H whose reduced antagonistic activity and increased agonistic activity compared with reference antagonists is due to an induced fit around Trp(755), resulting in a decreased steric clash with Met(909) but inducing a new internal clash with Val(912) in helix-12. This structure also explains the previously published observation that 16α attachments to RU486 analogs induce mixed profiles by altering the binding of 11β substituents. Together these structures further our understanding of the steric and electrostatic factors that contribute to the function of steroid receptor modulators, providing valuable insight for future compound design.
    Journal of Biological Chemistry 04/2012; 287(24):20333-43. · 4.65 Impact Factor
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    ABSTRACT: The progesterone receptor is able to bind to a large number and variety of ligands that elicit a broad range of transcriptional responses ranging from full agonism to full antagonism and numerous mixed profiles inbetween. We describe here two new progesterone receptor ligand binding domain x-ray structures bound to compounds from a structurally related but functionally divergent series, which show different binding modes corresponding to their agonistic or antagonistic nature. In addition, we present a third progesterone receptor ligand binding domain dimer bound to an agonist in monomer A and an antagonist in monomer B, which display binding modes in agreement with the earlier observation that agonists and antagonists from this series adopt different binding modes.
    Journal of Biological Chemistry 10/2011; 286(40):35079-35086. · 4.65 Impact Factor
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    ABSTRACT: The progesterone receptor is able to bind to a large number and variety of ligands that elicit a broad range of transcriptional responses ranging from full agonism to full antagonism and numerous mixed profiles inbetween. We describe here two new progesterone receptor ligand binding domain x-ray structures bound to compounds from a structurally related but functionally divergent series, which show different binding modes corresponding to their agonistic or antagonistic nature. In addition, we present a third progesterone receptor ligand binding domain dimer bound to an agonist in monomer A and an antagonist in monomer B, which display binding modes in agreement with the earlier observation that agonists and antagonists from this series adopt different binding modes.
    Journal of Biological Chemistry 08/2011; 286(40):35079-86. · 4.65 Impact Factor
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    ABSTRACT: Mesenchymal progenitor cells can be differentiated in vitro into myotubes that exhibit many characteristic features of primary mammalian skeletal muscle fibers. However, in general, they do not show the functional excitation-contraction coupling or the striated sarcomere arrangement typical of mature myofibers. Epigenetic modifications have been shown to play a key role in regulating the progressional changes in transcription necessary for muscle differentiation. In this study, we demonstrate that treatment of murine C2C12 mesenchymal progenitor cells with 10 μM of the DNA methylation inhibitor 5-azacytidine (5AC) promotes myogenesis, resulting in myotubes with enhanced maturity as compared to untreated myotubes. Specifically, 5AC treatment resulted in the up-regulation of muscle genes at the myoblast stage, while at later stages nearly 50% of the 5AC-treated myotubes displayed a mature, well-defined sarcomere organization, as well as spontaneous contractions that coincided with action potentials and intracellular calcium transients. Both the percentage of striated myotubes and their contractile activity could be inhibited by 20 nM TTX, 10 μM ryanodine, and 100 μM nifedipine, suggesting that action potential-induced calcium transients are responsible for these characteristics. Our data suggest that genomic demethylation induced by 5AC overcomes an epigenetic barrier that prevents untreated C2C12 myotubes from reaching full maturity.
    The FASEB Journal 07/2011; 25(11):3861-72. · 5.70 Impact Factor
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    ABSTRACT: The progressive restriction of differentiation potential from pluripotent embryonic stem cells, via multipotent progenitor cells to terminally differentiated, mature somatic cells, involves step-wise changes in transcription patterns that are tightly controlled by the coordinated action of key transcription factors and changes in epigenetic modifications. While previous studies have demonstrated tissue-specific differences in DNA methylation patterns that might function in lineage restriction, it is unclear at what exact developmental stage these differences arise. Here, we have studied whether terminal, multi-lineage differentiation of C2C12 myoblasts is accompanied by lineage-specific changes in DNA methylation patterns. Using bisulfite sequencing and genome-wide methylated DNA- and chromatin immunoprecipitation-on-chip techniques we show that in these cells, in general, myogenic genes are enriched for RNA polymerase II and hypomethylated, whereas osteogenic genes show lower polymerase occupancy and are hypermethylated. Removal of DNA methylation marks by 5-azacytidine (5AC) treatment alters the myogenic lineage commitment of these cells and induces spontaneous osteogenic and adipogenic differentiation. This is accompanied by upregulation of key lineage-specific transcription factors. We subsequently analyzed genome-wide changes in DNA methylation and polymerase II occupancy during BMP2-induced osteogenesis. Our data indicate that BMP2 is able to induce the transcriptional program underlying osteogenesis without changing the methylation status of the genome. We conclude that DNA methylation primes C2C12 cells for myogenesis and prevents spontaneous osteogenesis, but still permits induction of the osteogenic transcriptional program upon BMP2 stimulation. Based on these results, we propose that cell type-specific DNA methylation patterns are established prior to terminal differentiation of adult progenitor cells. This article is part of a Special Issue entitled: 11th European Symposium on Calcium.
    Biochimica et Biophysica Acta 05/2011; 1813(5):839-49. · 4.66 Impact Factor
  • Arthritis & Rheumatology 03/2011; 63(3):849. · 7.48 Impact Factor
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    ABSTRACT: Human mesenchymal stromal cells are regarded as the golden standard for cell-based therapies. They present multilineage differentiation potential and trophic and immunosuppressive abilities, making them the best candidate for clinical applications. Several molecules have been described to increase bone formation and were mainly discovered by candidate approaches towards known signaling pathways controlling osteogenesis. However, their bone forming potential is still limited, making the search for novel molecules a necessity. High-throughput screening (HTS) not only allows the screening of a large number of diverse chemical compounds, but also allows the discovery of unexpected signaling pathways and molecular mechanisms for a certain application, even without the prior knowledge of the full molecular pathway. Typically HTS is performed in cell lines, however, in this manuscript we have performed a phenotypical screen on more clinically relevant human mesenchymal stromal cells, as a proof of principle that HTS can be performed in those cells and can be used to find small molecules that impact stem cell fate. From a library of pharmacologically active small molecules, we were able to identify novel compounds with increased osteogenic activity. These compounds allowed achieving levels of bone-specific alkaline phosphatase higher than any other combination previously known. By combining biochemical techniques, we were able to demonstrate that a medium to high-throughput phenotypic assay can be performed in academic research laboratories allowing the discovery of novel molecules able to enhance stem cell differentiation.
    PLoS ONE 01/2011; 6(10):e26678. · 3.53 Impact Factor
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    ABSTRACT: Human mesenchymal stem cells (hMSCs) are being considered for several areas of clinical therapy, due to their multipotent nature. For instance, osteogenic hMSCs are applied in bone tissue engineering, but current differentiation protocols need further optimization before they can be clinically applied. Protein kinase C (PKC) family members have been implicated in bone metabolism, which prompted us to use a pharmaceutical approach to manipulate PKC signalling in hMSCs. Inhibition of PKC resulted in a dose-dependent inhibition of dexamethasone-induced osteogenic differentiation. Surprisingly, PKC activation using phorbol 12-myristate 13-acetate (PMA) also resulted in inhibition of osteogenesis, although we observed that inhibition was more pronounced at low than at high concentrations of PMA. Furthermore, we observed that inhibition of PKCdelta blocked alkaline phosphatase (ALP, an early marker of osteogenic differentiation) expression, whereas inhibition of the conventional PKC subfamily and PKCmicro using Gö6976 resulted in an induction of ALP activity, collagen (I) expression and mineralization. In conclusion, inhibition of the conventional PKCs/PKCmicro and activation of PKCdelta could further benefit osteogenic differentiation of hMSCs in vitro and in vivo, which is currently under investigation.
    Journal of Tissue Engineering and Regenerative Medicine 07/2010; 4(5):329-39. · 4.43 Impact Factor
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    ABSTRACT: Oligonucleotide arrays have become one of the most widely used high-throughput tools in biology. Due to their sensitivity to experimental conditions, normalization is a crucial step when comparing measurements from these arrays. Normalization is, however, far from a solved problem. Frequently, we encounter datasets with significant technical effects that currently available methods are not able to correct. We show that by a careful decomposition of probe specific amplification, hybridization and array location effects, a normalization can be performed that allows for a much improved analysis of these data. Identification of the technical sources of variation between arrays has allowed us to build statistical models that are used to estimate how the signal of individual probes is affected, based on their properties. This enables a model-based normalization that is probe-specific, in contrast with the signal intensity distribution normalization performed by many current methods. Next to this, we propose a novel way of handling background correction, enabling the use of background information to weight probes during summarization. Testing of the proposed method shows a much improved detection of differentially expressed genes over earlier proposed methods, even when tested on (experimentally tightly controlled and replicated) spike-in datasets. When a limited number of arrays are available, or when arrays are run in different batches, technical effects have a large influence on the measured expression of genes. We show that a detailed modelling and correction of these technical effects allows for an improved analysis in these situations.
    BMC Bioinformatics 03/2010; 11:156. · 3.02 Impact Factor
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    ABSTRACT: Since the discovery that bone morphogenetic proteins (BMPs) are able to induce ectopic bone formation, considerable effort has been devoted to apply it for bone regeneration. BMP activity needs to be temporally and spatially controlled and the organism has devised ways to achieve it. Here we show that the BMP inhibitor chordin-like 1 can interfere with BMP2 signalling thereby affecting the osteogenic differentiation of MC3T3-E1 cells. Besides its function as a BMP antagonist, chordin-like 1 enhanced the proliferation of human mesenchymal stem cells (hMSCs) in a BMP2-independent manner. When MC3T3-E1 cells were exposed to recombinant chordin-like 1 there was an inhibition of alkaline phosphatase (ALP) expression, whereas in the case of hMSCs no effect was observed. However, chordin-like 1 dose-dependently increased the proliferation of hMSCs. This effect is probably BMP2 independent because the chordin-like 1 concentration that stimulates proliferation does not interfere with BMP signalling monitored by a Smad-dependent reporter gene. Our data point towards a novel, BMP-independent role of chordin-like 1 in hMSC proliferation.
    Cells Tissues Organs 01/2010; 191(6):443-52. · 1.96 Impact Factor
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    ABSTRACT: Bone marrow-derived human mesenchymal stem cells (hMSCs) have the in vitro capacity to differentiate into osteoblasts, chondrocytes or adipocytes, depending on the applied stimulus. In order to identify novel regulators of osteogenesis in hMSCs, osteo-transcriptomics was performed whereby differentiation induced by dexamethasone (DEX), DEX+ bone morphogenetic protein 2 (BMP2), and DEX+ Vitamin D(3) (1,25(OH)(2)D(3)) was studied over a course of 12 days. Microarray analysis revealed that 2095 genes were significantly regulated by DEX+ 1,25(OH)(2)D(3), of which 961 showed accelerated expression kinetics compared to treatment by DEX alone. The majority of these genes were accelerated 24-48 h after onset of osteogenic treatment. Gene ontology (GO) analysis of these 1,25(OH)(2)D(3)-accelerated genes indicated their involvement in biological processes related to cellular differentiation and cell cycle regulation. When compared to cells treated with DEX or DEX+BMP2, treatment with DEX+ 1,25(OH)(2)D(3) clearly accelerated osteoprogenitor commitment and osteoblast maturation, as measured by alkaline phosphatase (ALP) activity and calcification of the matrix. Cell cycle progression, as observed after initial growth arrest, was not significantly accelerated by 1,25(OH)(2)D(3) and was not required for onset and progression of osteogenesis. However, expression of c-Myc was accelerated by 1,25(OH)(2)D(3), and binding sites for c-MYC were enriched in promoters of genes accelerated by 1,25(OH)(2)D(3). Lentiviral overexpression of c-MYC strongly promoted DEX+ BMP2-induced osteoblast differentiation and matrix maturation. In conclusion, our studies show for the first time that 1,25(OH)(2)D(3) strongly accelerates expression of genes involved in differentiation of hMSCs and, moreover, identify c-MYC as a novel regulator of osteogenesis.
    Bone 10/2009; 46(3):613-27. · 4.46 Impact Factor
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    ABSTRACT: Collagen is the main protein component of the extracellular matrix of bone, and it has structural and instructive properties. Collagen undergoes many post-translational modifications, including extensive crosslinking. Although defective crosslinking has been implicated in human syndromes (e.g., osteogenesis imperfecta or Ehlers-Danlos syndrome), it is not clear to what extent crosslinking is necessary for collagen's instructive properties during bone formation. Here we report that inhibition of collagen crosslinking in the mouse pre-osteoblast cell line MC3T3-E1 impairs the osteogenic program. Genome-wide expression profiling of beta-aminopropionitrile-treated and control cells revealed that matrix deposition by MC3T3-E1 cells provides a feed back signal, driving cells through the differentiation process, that is strongly impaired when crosslinking is inhibited. Inhibition of crosslinking did not affect osteogenic differentiation of human mesenchymal stem cells (hMSCs), shown by the expression of alkaline phosphatase and genome-wide gene expression analysis, although it enhances matrix mineralization. In conclusion, collagen crosslinking harbors instructive properties in MC3T3-E1 differentiation but plays a more-passive role in differentiation of bone marrow-derived hMSCs.
    Tissue Engineering Part A 09/2009; 15(12):3857-67. · 4.64 Impact Factor
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    ABSTRACT: The development and maintenance of a healthy skeleton depends on the migration of cells to areas of new bone formation. Osteoblasts, the bone forming cells of the body, mature from mesenchymal stem cells under the influence of bone morphogenetic protein. It is unclear at what developmental stage the osteoblasts start to migrate to their functional location. We have studied migration of immature pre-osteoblasts and of mature osteoblasts in response to Platelet-derived growth factor (PDGF) and sphingosine-1-phosphate (S1P). PDGF is a growth factor involved in bone remodeling and fracture healing whereas S1P is a circulating sphingolipid known to control cell trafficking. Our data indicate that PDGF acts as a chemotactic cue for pre-osteoblasts. In contrast, S1P is a chemorepellent to these cells. Upon Bone Morphogenetic Protein 2-induced conversion to the osteoblast phenotype, the chemotaxis response to PDGF is retained whereas the sensitivity to S1P is lost. By RNA interference and overexpression experiments we showed that the expression level of the S1P2 receptor is the sole determinant controlling responsiveness to S1P. The combined data indicate that migration of osteoblasts is controlled by the balance between PDGF, S1P and the differentiation state of the cells. We propose that this mechanism preserves the osteoprogenitor pool in the bone marrow, only allowing the more differentiated cell to travel to sites of bone formation.
    Journal of Cellular Biochemistry 10/2008; 105(4):1128-38. · 3.06 Impact Factor

Publication Stats

494 Citations
175.29 Total Impact Points

Institutions

  • 2009–2013
    • Universiteit Twente
      • Department of Tissue Regeneration (TR)
      Enschede, Provincie Overijssel, Netherlands
  • 2012
    • Netherlands eScience Center
      Amsterdamo, North Holland, Netherlands
  • 2003–2012
    • Radboud University Nijmegen
      • Department of Cell and Applied Biology
      Nymegen, Gelderland, Netherlands
  • 2011
    • Merck
      Whitehouse Station, New Jersey, United States
  • 2005
    • Erasmus MC
      • Department of Pathology
      Rotterdam, South Holland, Netherlands