In this thesis a novel technology is described to target adenovirus vectors. Adenovirus vectors are powerful tools to modulate gene expression. The use of these vectors however, is hampered by the... Show moreIn this thesis a novel technology is described to target adenovirus vectors. Adenovirus vectors are powerful tools to modulate gene expression. The use of these vectors however, is hampered by the fact that many for gene therapy interesting cell types do not, or only at low levels express the CAR receptor, necessary for infection. We developed a linker protein consisting of the virus-binding moiety of CAR genetically fused to the chicken protein avidin. Biotinylated ligands for cell specific receptors are bound to the linker protein via the avidin-biotin interaction. This now targeting protein is used to redirect adenovirus vectors to previously refractory cell types. Using this technology endothelial cell lines as well as primary endothelial cells can by infected at low MOI__s using an biotinylated cyclic RGD peptide. Primary bone marrow derived macrophages and macrophage cell lines are easily infected using a biotinylated dA6dG10 oligo nucleotide ligand. In vivo experiments showed a marked reduction of adenovirus mediated transgene expression by the liver, the organ responsible for virus uptake when unmodified adenovirus vectors are administered, after addition of several different ligands to the virus via the linker protein. Show less
Various modalities are available in the diagnostic and prognostic evaluation of patients presenting with known or suspected coronary artery disease (CAD). A rapidly expanding technique is... Show moreVarious modalities are available in the diagnostic and prognostic evaluation of patients presenting with known or suspected coronary artery disease (CAD). A rapidly expanding technique is noninvasive coronary angiography with Multi-Slice Computed Tomography (MSCT), which allows accurate detection of significant stenoses. The main value of the technique lies in the noninvasive exclusion of CAD in patients with intermediate pre-test likelihood. Although imaging in populations such as patients with previous stent placement appears to be more challenging, promising results have been obtained in these populations as well. However, it remains important to realize that the presence of coronary atherosclerosis with luminal obstruction does not invariably imply the presence of ischemia. Accordingly, a noninvasive angiographic imaging technique as MSCT cannot be used to predict the hemodynamical importance of lesions. In patients with borderline stenosis, therefore, functional testing (which can be performed by nuclear imaging, stress echocardiography or MRI) will remain necessary to determine management. Nonetheless, detection of CAD at a far earlier stage than functional imaging is an important advantage of MSCT. Initial investigations suggest that MSCT may distinguish different plaque characteristics between various presentations. Potentially, this information could be useful for risk stratification. Finally, additional non-coronary information can be derived as well. LV function can be evaluated with high accuracy while also information on the cardiac venous system can be obtained. Show less
One of the most important characteristics of atherosclerosis is the chronic inflammatory response in which T cells and NKT cells are very important. In this thesis several methods to modulate the... Show moreOne of the most important characteristics of atherosclerosis is the chronic inflammatory response in which T cells and NKT cells are very important. In this thesis several methods to modulate the activity of these T and NKT cells in atherosclerosis are described. The induction of regulatory T cells (as mentioned in chapters 2 and 3) positively influences the immune response in atherosclerosis. In addition, treatment with dendritic cells (DCs) loaded with oxidized LDL (oxLDL) resulted in the activation of oxLDL-specific T cells and in an increase in oxLDL-specific antibodies. These IgG antibodies have a protecting effect on atherosclerosis (chapter 4). Furthermore, the complex role of NKT cells in the immunology of atherosclerosis is described (chapters 5, 6 and 7). NKT cells can play a protective role in atherosclerosis when they are 1) activated directly by _-GalCer or when they are 2) activated by OCH loaded on DCs. The studies described in this thesis led to a better insight in the role of the immune system in high-fat diet induced atherosclerosis, and they also create a possible therapeutic protocol which can probably be applied to reduce the incidence of cardiovascular diseases in humans. Show less
With the use of combinatorial phage display, solid phase peptide synthesis and a multidiscipline of molecular and cellular assays in vascular biology, the research described in this thesis has... Show moreWith the use of combinatorial phage display, solid phase peptide synthesis and a multidiscipline of molecular and cellular assays in vascular biology, the research described in this thesis has resulted in the identification of two novel peptides targeting to SR-AI and CD40 respectively which hold promise as targeted contrast agents for the diagnosis of atherosclerosis symptom. In addition, a peptide named VIVIT and its derivatives had been discovered and synthesized which constitute a more selective and less toxic drug candidate than currently used immunosuppressant cyclosporine A or FK506, leading to new generation immunosuppressants and therapeutics for autoimmune diseases such as rheumatoid arthritis or allograft transplantation and cardiovascular disorders including atherosclerosis, restenosis and cardiac hypertrophy. Show less
The main cause of cardiovascular disease (CVD) is atherosclerosis. Several genes that affect atherosclerosis development have been identified by the use of genetically modified mice (i.e.... Show moreThe main cause of cardiovascular disease (CVD) is atherosclerosis. Several genes that affect atherosclerosis development have been identified by the use of genetically modified mice (i.e. transgenic and knock-out mouse models). Many of these genes exert their role in atherosclerosis development as a result of effects on lipoprotein metabolism and inflammation. Transgenic mouse models have also been proven to be suitable for evaluating the mechanisms underlying the anti-atherosclerotic action of experimental drugs aimed to reduce atherogenic lipoprotein levels. However, thus far no suitable animal model was present to evaluate the mechanism of action of anti-atherosclerotic effect of HDL-raising therapeutic strategies. In this thesis, we further explored the role of apolipoprotein CI (apoCI) and cholesteryl ester transfer protein (CETP) in lipoprotein metabolism, inflammation, and atherosclerosis. Furthermore, we developed a mouse model that will be suitable for testing potential high-density-lipoprotein (HDL) raising therapies as a novel strategy to treat CVD. Show less
Accurate visualization and quantification of atherosclerosis in a non-invasive manner by means of MR is, nowadays, of high importance, not only regarding morphology but also composition of the... Show moreAccurate visualization and quantification of atherosclerosis in a non-invasive manner by means of MR is, nowadays, of high importance, not only regarding morphology but also composition of the atherosclerotic plaques. MR has proven to be capable of detecting early, subclinical vulnerable plaque. However, when the analysis of atherosclerosis is based on visual interpretation of the images or manually delineated structures, the outcome is often not reliable. The main objective of this thesis was to investigate novel image processing techniques to automatically quantify markers of the severity of atherosclerosis in a reproducible manner, by automatically outlining the boundaries of the blood vessel wall, lumen and plaque burden. Different algorithms have been developed and applied to different vascular beds (aorta and carotid arteries) proving to be versatile and powerful tools, that provide quantitative reproducible parameters. These new techniques have been validated in limited populations, proving to be accurate and reproducible. They are, therefore, suitable to be further adapted and to be employed in the clinical vascular research. This assists the physician in making a diagnosis and identifying high-risk patients, who may benefit from treatment. Show less
Restenosis is the main drawback of percutaneous coronary interventions (PCI). Recently, drug-eluting stents (DES) were introduced successfully in clinical practice leading to a drop in the post... Show moreRestenosis is the main drawback of percutaneous coronary interventions (PCI). Recently, drug-eluting stents (DES) were introduced successfully in clinical practice leading to a drop in the post-angioplasty restenosis rate. Nevertheless, safety and efficacy concerns related to the active compounds of such stents still exist. This thesis aimed at the development of a local delivery system, in combination with the cuff-induced mouse model of neointima formation, in order to closely mimic the DES mode-of-action and to further answer to such remaining questions related to this novel technology. Altogether, these comprehensive studies on efficacy and safety of anti-restenotic compounds using a novel developed mouse model of neointimal hyperplasia have resulted in a thorough understanding of the mode-of-action of these anti-restenotic drugs on neointimal hyperplasia inhibition as well as to further assess the potential adverse effects on vessel wall biology and on the (treated) pre-existing underlying atherosclerotic lesions. Especially the discovery of novel anti-restenotic strategies to be used in combination with the DES technology is of specially importance for further preclinical studies in more advance animal models of restenosis (e.g. porcine coronary artery model) as well as directly in the field of interventional cardiology per se. Show less
In dit promotieonderzoek is zijn de effecten van vetstapeling en ontstekingsreactie tijdens het proces van atherosclerose. We hebben aangetoond dat het ontstekingsremmende eiwit interleukine-9, een... Show moreIn dit promotieonderzoek is zijn de effecten van vetstapeling en ontstekingsreactie tijdens het proces van atherosclerose. We hebben aangetoond dat het ontstekingsremmende eiwit interleukine-9, een stof die door bepaalde immunologische cellen geproduceerd wordt, een remmende werking heeft op het ontstaan van atherosclerose in het algemeen en van vetstapeling in macrofagen in het bijzonder. Aan de andere kant blijkt uit mijn promotieonderzoek dat vetstapeling van macrofagen de gevoeligheid van deze cellen voor ontstekingen beïnvloedt. LPS is in staat om een zeer sterke ontstekingsreactie te stimuleren en om de expressie van verschillende genen die betrokken zijn bij vetstapeling te beïnvloeden. Door gebruik te maken van muizen die geen scavenger receptor BI (SR-BI) tot expressie brengen, hebben we aangetoond dat SR-BI beschermd tegen de door LPS gestimuleerde ontstekingsreactie. Tevens blijkt dat een dieet met een hoog cholesterol gehalte een grote invloed heeft op parenchymcellen in de lever. Voornamelijk FABP5 en vier nieuwe vetzuurbindende eiwitten lijken een belangrijke rol te spelen in de reactie van deze cellen op het dieet. Ook het ontstekingremmende interleukine 10, waarvan bekend is dat het atherosclerose kan remmen en een verlaging van cholesterol in het bloed kan veroorzaken, beïnvloedt vele genen betrokken bij vethuishouding in parenchymcellen van de lever. Show less
