During my research project we mainly focussed on studying the pathophysiology of venous and arterial thrombosis in mice. When we transiently lowered plasma protein levels of natural anticoagulants... Show moreDuring my research project we mainly focussed on studying the pathophysiology of venous and arterial thrombosis in mice. When we transiently lowered plasma protein levels of natural anticoagulants antithrombin and protein C using RNA interference, mice developed venous thrombosis in the head. In contrast to other mouse models for venous thrombosis where surgery is required for provoking the disease, mice injected with RNA interference against the mRNA of Serpinc1 and Proc (antithrombin and protein C, respectively) developed venous thrombosis without additional handlings. In this unique form of venous thrombosis, we studied the roles of platelets, neutrophils, and coagulation factor XII. These factors have been shown to be indispensable in experimental venous thrombosis in other mouse models, and they have been introduced as novel therapeutic targets. For the second part of my thesis we again used the RNA interference approach, to lower natural anticoagulation in atherosclerotic mice. When we lowered protein C in these mice, they developed atherothrombosis in the aortic root without any additional intervention. This unique form of atherothrombosis has been showed in multiple independent experiments, and we aimed to further characterize the process to learn more about prevention atherothrombosis in atherosclerotic mice and the role of protein C. Show less
Drug-induced organ toxicity is a major concern for pharmaceutical industry, due to removal of a high number of drugs from the market, as well as for public health, due to numerous hospitalizations... Show moreDrug-induced organ toxicity is a major concern for pharmaceutical industry, due to removal of a high number of drugs from the market, as well as for public health, due to numerous hospitalizations and patient death. The organs that are the primary target for such toxicities are the liver and the kidneys since both organs are continuously exposed to high concentrations of drugs and have high metabolic capacities. The immune system has been shown to be involved in the toxicity of several drugs- inducing liver and kidney injury. In particular, the cytokine tumor necrosis factor _ (TNF-_) has been shown to be the main constituent of the inflammatory processes responsible for the aggravation of drug-induced liver and kidney injuries. In this thesis, the role of TNF-_ in drug-induced organ injury was investigated. The main question was to assess the possible synergy between TNF-_ and druginduced cytotoxicities, and to unravel the underlying molecular mechanisms of such a synergy. Show less
Familial hypercholesterolemia (FH) is a genetic disorder characterized by high levels of low density lipoprotein cholesterol (LDL-C) and increasing the risk of cardio vascular diseases. FH and many... Show moreFamilial hypercholesterolemia (FH) is a genetic disorder characterized by high levels of low density lipoprotein cholesterol (LDL-C) and increasing the risk of cardio vascular diseases. FH and many other liver diseases can possibly be treated with RNA interference (RNAi). RNAi is a natural process of regulation of gene expression by binding of small RNA molecules to complementary sequences in the mRNA of a gene and hence inducing its degradation or translational repression. In this thesis, we aimed at developing a safe and robust RNAi-based therapy for FH by inhibiting Apolipoprotein B100 (ApoB). ApoB is a primary component of the low density lipoprotein (LDL), a lipoprotein that transports __bad__ cholesterol LDL-C. We have shown that RNAi therapy using AAV-delivered artificial miRNA is a promising approach for treatment of FH and possibly other liver diseases. Our mechanistic studies revealed differences in shRNA and miRNA processing and functioning in vivo. Finally, our findings have significant impact on understanding and overcoming toxicity and off-targeting related problems of RNAi based gene therapy in the liver using AAV vectors. Show less
Adverse drug reactions are problematic for both society and pharmaceutical industry. The costs are high in severe cases: for pharmaceutical companies due to the loss of income if a drug needs to be... Show moreAdverse drug reactions are problematic for both society and pharmaceutical industry. The costs are high in severe cases: for pharmaceutical companies due to the loss of income if a drug needs to be removed from the market; for society due to the extra healthcare that is required to treat the affected individuals. Liver damage upon drug intake is the most common type of adverse drug reaction and reason for drug withdrawal. What makes the liver such a vulnerable target is its role as the main drug-metabolizing organ and its large amount of stationary immune cells. This results in frequent exposure to drug metabolite-induced and inflammatory stress. In this thesis I have investigated the role of the signaling induced by the pro-inflammatory cytokine TNF_ in the exacerbation of drug-induced liver injury using an in vitro liver cell model. Using this model and methods such as high content imaging, gene array analysis and functional genomics we have gotten closer to understanding the molecular mechanisms of liver injury. This knowledge can be used to design novel tests for the drug-industry to assess the toxicity of novel drug candidates as well as to pin-point potentially susceptible individuals. Show less
