Martijn Dijkstra

Whole-genome sequence variation, population structure and demographic history of the Dutch population

Whole-genome sequencing enables complete characterization of genetic variation, but geographic clustering of rare alleles demands many diverse populations be studied. Here we describe the Genome of the Netherlands (GoNL) Project, in which we sequenced the whole genomes of 250 Dutch parent-offspring families and constructed a haplotype map of 20.4 million single-nucleotide variants and 1.2 million insertions and deletions. The intermediate coverage (∼13×) and trio design enabled extensive characterization of structural variation, including midsize events (30–500 bp) previously poorly catalogued and de novo mutations. We demonstrate that the quality of the haplotypes boosts imputation accuracy in independent samples, especially for lower frequency alleles. Population genetic analyses demonstrate fine-scale structure across the country and support multiple ancient migrations, consistent with historical changes in sea level and flooding. The GoNL Project illustrates how single-population whole-genome sequencing can provide detailed characterization of genetic variation and may guide the design of future population studies.

System-wide molecular evidence for phenotypic buffering in Arabidopsis

We profiled 162 lines of Arabidopsis for variation in transcript, protein and metabolite abundance using mRNA microarrays, two-dimensional polyacrylamide gel electrophoresis, gas chromatography time-of-flight mass spectrometry, liquid chromatography quadrupole time-of-flight mass spectrometry, and proton nuclear magnetic resonance. We added all publicly available phenotypic data from the same lines and mapped quantitative trait loci (QTL) for 40,580 molecular and 139 phenotypic traits. We found six QTL hot spots with major, system-wide effects, suggesting there are six breakpoints in a system otherwise buffered against many of the 500,000 SNPs.

Human Primary Adipocytes Exhibit Immune Cell Function: Adipocytes Prime Inflammation Independent of Macrophages

Background Obesity promotes inflammation in adipose tissue (AT) and this is implicated in pathophysiological complications such as insulin resistance, type 2 diabetes and cardiovascular disease. Although based on the classical hypothesis, necrotic AT adipocytes (ATA) in obese state activate AT macrophages (ATM) that then lead to a sustained chronic inflammation in AT, the link between human adipocytes and the source of inflammation in AT has not been in-depth and systematically studied. So we decided as a new hypothesis to investigate human primary adipocytes alone to see whether they are able to prime inflammation in AT. Methods and Results Using mRNA expression, human preadipocytes and adipocytes express the cytokines/chemokines and their receptors, MHC II molecule genes and 14 acute phase reactants including C-reactive protein. Using multiplex ELISA revealed the expression of 50 cytokine/chemokine proteins by human adipocytes. Upon lipopolysaccharide stimulation, most of these adipocyte-associated cytokines/chemokines and immune cell modulating receptors were up-regulated and a few down-regulated such as (ICAM-1, VCAM-1, MCP-1, IP-10, IL-6, IL-8, TNF-α and TNF-β highly up-regulated and IL-2, IL-7, IL-10, IL-13 and VEGF down-regulated. In migration assay, human adipocyte-derived chemokines attracted significantly more CD4+ T cells than controls and the number of migrated CD4+ cells was doubled after treating the adipocytes with LPS. Neutralizing MCP-1 effect produced by adipocytes reduced CD4+ migration by approximately 30%. Conclusion Human adipocytes express many cytokines/chemokines that are biologically functional. They are able to induce inflammation and activate CD4+ cells independent of macrophages. This suggests that the primary event in the sequence leading to chronic inflammation in AT is metabolic dysfunction in adipocytes, followed by production of immunological mediators by these adipocytes, which is then exacerbated by activated ATM, activation and recruitment of immune cells. This study provides novel knowledge about the prime of inflammation in human obese adipose tissue, opening a new avenue of investigations towards obesity-associated type 2 diabetes.

The Genome of the Netherlands: design, and project goals

Within the Netherlands a national network of biobanks has been established (Biobanking and Biomolecular Research Infrastructure-Netherlands (BBMRI-NL)) as a national node of the European BBMRI. One of the aims of BBMRI-NL is to enrich biobanks with different types of molecular and phenotype data. Here, we describe the Genome of the Netherlands (GoNL), one of the projects within BBMRI-NL. GoNL is a whole-genome-sequencing project in a representative sample consisting of 250 trio-families from all provinces in the Netherlands, which aims to characterize DNA sequence variation in the Dutch population. The parent–offspring trios include adult individuals ranging in age from 19 to 87 years (mean=53 years; SD=16 years) from birth cohorts 1910–1994. Sequencing was done on blood-derived DNA from uncultured cells and accomplished coverage was 14–15x. The family-based design represents a unique resource to assess the frequency of regional variants, accurately reconstruct haplotypes by family-based phasing, characterize short indels and complex structural variants, and establish the rate of de novo mutational events. GoNL will also serve as a reference panel for imputation in the available genome-wide association studies in Dutch and other cohorts to refine association signals and uncover population-specific variants. GoNL will create a catalog of human genetic variation in this sample that is uniquely characterized with respect to micro-geographic location and a wide range of phenotypes. The resource will be made available to the research and medical community to guide the interpretation of sequencing projects. The present paper summarizes the global characteristics of the project.

Regulation of adipokine production in human adipose tissue by propionic acid

Eur J Clin Invest 2010; 40 (5): 401–407

The MOLGENIS toolkit: rapid prototyping of biosoftware at the push of a button

BackgroundThere is a huge demand on bioinformaticians to provide their biologists with user friendly and scalable software infrastructures to capture, exchange, and exploit the unprecedented amounts of new *omics data. We here present MOLGENIS, a generic, open source, software toolkit to quickly produce the bespoke MOLecular GENetics Information Systems needed.MethodsThe MOLGENIS toolkit provides bioinformaticians with a simple language to model biological data structures and user interfaces. At the push of a button, MOLGENIS’ generator suite automatically translates these models into a feature-rich, ready-to-use web application including database, user interfaces, exchange formats, and scriptable interfaces. Each generator is a template of SQL, JAVA, R, or HTML code that would require much effort to write by hand. This ‘model-driven’ method ensures reuse of best practices and improves quality because the modeling language and generators are shared between all MOLGENIS applications, so that errors are found quickly and improvements are shared easily by a re-generation. A plug-in mechanism ensures that both the generator suite and generated product can be customized just as much as hand-written software.ResultsIn recent years we have successfully evaluated the MOLGENIS toolkit for the rapid prototyping of many types of biomedical applications, including next-generation sequencing, GWAS, QTL, proteomics and biobanking. Writing 500 lines of model XML typically replaces 15,000 lines of hand-written programming code, which allows for quick adaptation if the information system is not yet to the biologist’s satisfaction. Each application generated with MOLGENIS comes with an optimized database back-end, user interfaces for biologists to manage and exploit their data, programming interfaces for bioinformaticians to script analysis tools in R, Java, SOAP, REST/JSON and RDF, a tab-delimited file format to ease upload and exchange of data, and detailed technical documentation. Existing databases can be quickly enhanced with MOLGENIS generated interfaces using the ‘ExtractModel’ procedure.ConclusionsThe MOLGENIS toolkit provides bioinformaticians with a simple model to quickly generate flexible web platforms for all possible genomic, molecular and phenotypic experiments with a richness of interfaces not provided by other tools. All the software and manuals are available free as LGPLv3 open source at http://www.molgenis.org.

Improved imputation quality of low-frequency and rare variants in European samples using the 'Genome of The Netherlands'

Although genome-wide association studies (GWAS) have identified many common variants associated with complex traits, low-frequency and rare variants have not been interrogated in a comprehensive manner. Imputation from dense reference panels, such as the 1000 Genomes Project (1000G), enables testing of ungenotyped variants for association. Here we present the results of imputation using a large, new population-specific panel: the Genome of The Netherlands (GoNL). We benchmarked the performance of the 1000G and GoNL reference sets by comparing imputation genotypes with ‘true’ genotypes typed on ImmunoChip in three European populations (Dutch, British, and Italian). GoNL showed significant improvement in the imputation quality for rare variants (MAF 0.05–0.5%) compared with 1000G. In Dutch samples, the mean observed Pearson correlation, r2, increased from 0.61 to 0.71. We also saw improved imputation accuracy for other European populations (in the British samples, r2 improved from 0.58 to 0.65, and in the Italians from 0.43 to 0.47). A combined reference set comprising 1000G and GoNL improved the imputation of rare variants even further. The Italian samples benefitted the most from this combined reference (the mean r2 increased from 0.47 to 0.50). We conclude that the creation of a large population-specific reference is advantageous for imputing rare variants and that a combined reference panel across multiple populations yields the best imputation results.

Endovascular Aneurysm Sealing for Juxtarenal Aneurysm Using the Nellix Device and Chimney Covered Stents

Purpose: To show the feasibility of the Nellix device in conjunction with a chimney technique for treating juxtarenal aneurysms in two patients who were deemed unsuitable for fenestrated endovascular aneurysm repair or open surgery. Case Reports: Two men aged 83 and 81 years were referred with a juxtarenal abdominal aortic aneurysm (66 and 69 mm, respectively). Both were considered for open surgery as well as custom-made fenestrated stent-graft but deemed unsuitable for both options. They were both treated using the Nellix endoprosthesis in combination with chimney grafts to preserve the renal arteries. Technical success was achieved in both cases, with successful aneurysm exclusion and target vessel preservation (the right renal artery in the first case and both renal arteries in the second). At 6 months, duplex ultrasound and computed tomographic angiography of the first patient showed no signs of endoleak and patent renal arteries. The second patient developed a right retroperitoneal hematoma with minor extravasation near the lower pole of the right kidney for which coil embolization was necessary. The subsequent clinical sequelae led to respiratory insufficiency and ultimately death. Conclusion: The use of the Nellix endoprosthesis combined with chimney grafts is technically feasible. The addition of chimney grafts can increase the applicability of endovascular aneurysm sealing to treat short-neck and juxtarenal aneurysms. Further studies are needed to confirm these findings and establish longer term outcome.

Comparison of isotope-labeled amino acid incorporation rates (CILAIR) provides a quantitative method to study tissue secretomes

Adipose tissue is an endocrine organ involved in regulation of whole-body energy metabolism via storage of lipids and secretion of various peptide hormones (adipokines). We previously characterized the adipose tissue secretome and showed that [13C]lysine incorporation into secreted proteins can be used to determine the origin of identified proteins. In the present study we determined the effect of insulin on the secretome by comparing incorporation rates of 13C-labeled lysine in the presence and absence of insulin. Human visceral adipose tissue from one patient was divided over six dishes. After subsequent washes to remove serum proteins, [13C]lysine-containing medium was added. Three dishes also received 60 nm insulin. The other three were controls. After 72 h of culture, media were collected and processed separately, involving concentration by ultrafiltration and fractionation by SDS-PAGE followed by in-gel digestion of excised bands and LC-MS/MS analyses. The obtained spectra were used for database searching and calculation of heavy/light ratios. The three control data sets shared 342 proteins of which 156 were potentially secreted and contained label. The three insulin-derived data sets shared 361 proteins of which 141 were potentially secreted and contained label. After discarding secreted proteins with very low label incorporation, 121 and 113 proteins remained for control and insulin data sets, respectively. The average coefficient of variation for control triplicates was 10.0% and for insulin triplicates was 18.3%. By comparing heavy/light ratios in the absence and presence of insulin we found 24 up-regulated proteins and four down-regulated proteins, and 58 proteins showed no change. Proteins involved in the endoplasmic reticulum stress response and in extracellular matrix remodeling were up-regulated by insulin. In conclusion, comparison of isotope-labeled amino acid incorporation rates (CILAIR) allows quantitative assessment of changes in protein secretion without the need for 100% label incorporation, which cannot be reached in differentiated tissues or cells.

Dutch experience with the fenestrated Anaconda endograft for short-neck infrarenal and juxtarenal abdominal aortic aneurysm repair

OBJECTIVE In the past decennium, the management of short-neck infrarenal and juxtarenal aortic aneurysms with fenestrated endovascular aneurysm repair (FEVAR) has been shown to be successful, with good early and midterm results. Recently, a new fenestrated device, the fenestrated Anaconda (Vascutek, Renfrewshire, Scotland), was introduced. The aim of this study was to present the current Dutch experience with this device. METHODS A prospectively held database of patients treated with the fenestrated Anaconda endograft was analyzed. Decision to treat was based on current international guidelines. Indications for FEVAR included an abdominal aortic aneurysm (AAA) with unsuitable neck anatomy for EVAR. Planning was performed on computed tomography angiography images using a three-dimensional workstation. RESULTS Between May 2011 and September 2013, 25 patients were treated in eight institutions for juxtarenal (n = 23) and short-neck AAA (n = 2). Median AAA size was 61 mm (59-68.5 mm). All procedures except one were performed with bifurcated devices. A total of 56 fenestrations were incorporated, and 53 (94.6%) were successfully cannulated and stented. One patient died of bowel ischemia caused by occlusion of the superior mesenteric artery. On completion angiography, three type I endoleaks and seven type II endoleaks were observed. At 1 month of follow-up, all endoleaks had spontaneously resolved. Median follow-up was 11 months (range, 1-29 months). There were no aneurysm ruptures or aneurysm-related deaths and no reinterventions to date. Primary patency at 1 month of cannulated and stented target vessels was 96%. CONCLUSIONS Initial and short-term results of FEVAR using the fenestrated Anaconda endograft are promising, with acceptable technical success and short-term complication rates. Growing experience and long-term results are needed to support these findings.