Autophagy is a fundamental degradative process, maintaining cellular homeostasis and functions in host defense against intracellular pathogens, including mycobacteria and Salmonella. The thesis... Show moreAutophagy is a fundamental degradative process, maintaining cellular homeostasis and functions in host defense against intracellular pathogens, including mycobacteria and Salmonella. The thesis investigated the function of an regulator of antibacterial autophagy, Damage Regulated Autophagy Modulator 1 (DRAM1) against infection and shows that DRAM1 restricts bacterial growth not only through canonical antibacterial autophagy (xenophagy) but also promotes an autophagy-related pathway, named LC3-associated phagocytosis (LAP). The function of DRAM1 in restricting bacterial proliferation is independent from the recognition of bacteria by xenophagy receptors. Mechanistically, DRAM1 promotes the infection-induced activation of autophagy and LAP as well as the maturation of bacteria-containing vesicles in both pathways. This maturation process, stimulated by DRAM1, involves multiple vesicle fusion steps directing bacteria to lysosomes. Through this maturation process, DRAM1 delivers the cytosolic protein Fau to bacteria-containing vesicles, where it serves as a precursor for antimicrobial peptides. The underlying mechanism may be explained by the discovery of an interaction between DRAM1 and the SNARE protein VTI1B. Overall, the work in this thesis contributes to ongoing research into the potential application of autophagy modulation as a host-directed therapy against infectious diseases. Show less
Replication stress, caused by Rev1 deficiency, is associated with mitochondrial dysfunction, and metabolic stress. However, the overall metabolic alterations and possible interventions to rescue... Show moreReplication stress, caused by Rev1 deficiency, is associated with mitochondrial dysfunction, and metabolic stress. However, the overall metabolic alterations and possible interventions to rescue the deficits due to Rev1 loss remain unclear. Here, we report that loss of Rev1 leads to intense changes in metabolites and that this can be manipulated by NAD + supplementation. Autophagy decreases in Rev1-/- mouse embryonic fibroblasts (MEFs) and can be restored by supplementing the NAD+ precursor nicotinamide riboside (NR). The abnormal mitochondrial morphology in Rev1-/-MEFs can be partially reversed by NR supplementation, which also protects the mito-chondrial cristae from rotenone-induced degeneration. In nematodes rev-1 deficiency causes sensitivity to oxidative stress but this cannot be rescued by NR supplementation. In conclusion, Rev1 deficiency leads to metabolic dysregulation of especially lipid and nucleotide metabolism, impaired autophagy, and mitochondrial anomalies, and all of these phenotypes can be improved by NR replenishment in MEFs. Show less
Exposure to cigarette smoke (CS) is the primary risk factor for developing chronic obstructive pulmonary disease. The impact of CS exposure on the molecular mechanisms involved in mitochondrial... Show moreExposure to cigarette smoke (CS) is the primary risk factor for developing chronic obstructive pulmonary disease. The impact of CS exposure on the molecular mechanisms involved in mitochondrial quality control in airway epithelial cells is incompletely understood. Undifferentiated or differentiated primary bronchial epithelial cells were acutely/chronically exposed to whole CS (WCS) or CS extract (CSE) in submerged or air-liquid interface conditions. Abundance of key regulators controlling mitochondrial biogenesis, mitophagy and mitochondrial dynamics was assessed. Acute exposure to WCS or CSE increased the abundance of components of autophagy and receptor-mediated mitophagy in all models. Although mitochondrial content and dynamics appeared to be unaltered in response to CS, changes in both the molecular control of mitochondrial biogenesis and a shift toward an increased glycolytic metabolism were observed in particular in differentiated cultures. These alterations persisted, at least in part, after chronic exposure to WCS during differentiation and upon subsequent discontinuation of WCS exposure. In conclusion, smoke exposure alters the regulation of mitochondrial metabolism in airway epithelial cells, but observed alterations may differ between various culture models used. Show less
Purpose Non-medullary thyroid cancer (NMTC) treatment is based on the ability of thyroid follicular cells to accumulate radioactive iodide (RAI). However, in a subset of NMTC patients tumor... Show morePurpose Non-medullary thyroid cancer (NMTC) treatment is based on the ability of thyroid follicular cells to accumulate radioactive iodide (RAI). However, in a subset of NMTC patients tumor dedifferentiation occurs, leading to RAI resistance. Digoxin has been demonstrated to restore iodide uptake capacity in vitro in poorly differentiated and anaplastic NMTC cells, termed redifferentiation. The aim of the present study was to investigate the in vivo effects of digoxin in TPO-Cre/LSL-Braf(V600E) mice and digoxin-treated NMTC patients. Methods Mice with thyroid cancer were subjected to 3D ultrasound for monitoring tumor growth and I-124 PET/CT for measurement of intratumoral iodide uptake. Post-mortem analyses on tumor tissues comprised gene expression profiling and measurement of intratumoral autophagy activity. Through PALGA (Dutch Pathology Registry), archived tumor material was obtained from 11 non-anaplastic NMTC patients who were using digoxin. Clinical characteristics and tumor material of these patients were compared to 11 matched control NMTC patients never treated with digoxin. Results We found that in mice, tumor growth was inhibited and I-124 accumulation was sustainably increased after short-course digoxin treatment. Post-mortem analyses revealed that digoxin treatment increased autophagy activity and enhanced expression of thyroid-specific genes in mouse tumors compared to vehicle-treated mice. Digoxin-treated NMTC patients exhibited significantly higher autophagy activity and a higher differentiation status as compared to matched control NMTC patients, and were associated with favourable clinical outcome. Conclusions These in vivo data support the hypothesis that digoxin may represent a repositioned adjunctive treatment modality that suppresses tumor growth and improves RAI sensitivity in patients with RAI-refractory NMTC. Show less
Oncolytic reovirus has the natural preference to kill transformed cells while sparing normal cells. Its administration has been shown to be safe in clinical trials, but the anti-cancer efficacy... Show moreOncolytic reovirus has the natural preference to kill transformed cells while sparing normal cells. Its administration has been shown to be safe in clinical trials, but the anti-cancer efficacy remains to be improved. In this thesis, we discuss several aspects that are important for the design of a potent anti-cancer therapeutic strategy using reovirus. We explored which cellular factors and pathways are important for efficient reovirus replication. Furthermore, we genetically modified the reovirus genome to encode potentially therapeutic transgenes. We tested the potency of these recombinant reoviruses, and describe what we believe is the most promising strategy to move forward. Moreover, we discuss the stability issues that we encountered during the generation of recombinant reoviruses. Finally, we discuss the various challenges and opportunities in how to proceed. Show less
This review discusses the relevant metabolic pathways and their regulators which show potential for T cell metabolism-based immunotherapy in diseases hallmarked by both metabolic disease and... Show moreThis review discusses the relevant metabolic pathways and their regulators which show potential for T cell metabolism-based immunotherapy in diseases hallmarked by both metabolic disease and autoimmunity. Multiple therapeutic approaches using existing pharmaceuticals are possible from a rationale in which T cell metabolism forms the hub in dampening the T cell component of autoimmunity in metabolic diseases. Future research into the effects of a metabolically aberrant micro-environment on T cell metabolism and its potential as a therapeutic target for immunomodulation could lead to novel treatment strategies for metabolic disease-associated autoimmunity. Show less
This review discusses the relevant metabolic pathways and their regulators which show potential for T cell metabolism-based immunotherapy in diseases hallmarked by both metabolic disease and... Show moreThis review discusses the relevant metabolic pathways and their regulators which show potential for T cell metabolism-based immunotherapy in diseases hallmarked by both metabolic disease and autoimmunity. Multiple therapeutic approaches using existing pharmaceuticals are possible from a rationale in which T cell metabolism forms the hub in dampening the T cell component of autoimmunity in metabolic diseases. Future research into the effects of a metabolically aberrant micro-environment on T cell metabolism and its potential as a therapeutic target for immunomodulation could lead to novel treatment strategies for metabolic disease-associated autoimmunity. Show less
This review discusses the relevant metabolic pathways and their regulators which show potential for T cell metabolism-based immunotherapy in diseases hallmarked by both metabolic disease and... Show moreThis review discusses the relevant metabolic pathways and their regulators which show potential for T cell metabolism-based immunotherapy in diseases hallmarked by both metabolic disease and autoimmunity. Multiple therapeutic approaches using existing pharmaceuticals are possible from a rationale in which T cell metabolism forms the hub in dampening the T cell component of autoimmunity in metabolic diseases. Future research into the effects of a metabolically aberrant micro-environment on T cell metabolism and its potential as a therapeutic target for immunomodulation could lead to novel treatment strategies for metabolic disease-associated autoimmunity. Show less
This review discusses the relevant metabolic pathways and their regulators which show potential for T cell metabolism-based immunotherapy in diseases hallmarked by both metabolic disease and... Show moreThis review discusses the relevant metabolic pathways and their regulators which show potential for T cell metabolism-based immunotherapy in diseases hallmarked by both metabolic disease and autoimmunity. Multiple therapeutic approaches using existing pharmaceuticals are possible from a rationale in which T cell metabolism forms the hub in dampening the T cell component of autoimmunity in metabolic diseases. Future research into the effects of a metabolically aberrant micro-environment on T cell metabolism and its potential as a therapeutic target for immunomodulation could lead to novel treatment strategies for metabolic disease-associated autoimmunity. Show less
De resultaten beschreven in dit proefschrift verschaffen nieuwe inzichten in verschillende aspecten van de behandeling van inflammatoire darmziekten. Uit grote studies is gebleken dat mutaties in... Show moreDe resultaten beschreven in dit proefschrift verschaffen nieuwe inzichten in verschillende aspecten van de behandeling van inflammatoire darmziekten. Uit grote studies is gebleken dat mutaties in genen die betrokken zijn bij het autofagie proces (autophagos: __zelf-eten__), geassocieerd zijn met de ziekte van Crohn. In dit proefschrift wordt beschreven hoe deze mutatie in dendritische cellen leidt tot hyperactivatie van het immuunsysteem. Dit mechanisme kan mogelijk een nieuw aangrijpingspunt verschaffen voor nieuwe therapie voor pati_nten die een mutatie hebben in dit gen. Anti-tumor-necrosis-factor-alpha (anti-TNF_) therapie neemt een steeds belangrijkere plaats in in de behandeling van IBD; het werkingsmechanisme is echter niet geheel duidelijk. In dit proefschrift wordt beschreven hoe regulatoire macrophagen met immuunsuppressieve en wondgenezende eigenschappen ge_nduceerd worden door anti-TNF_ therapie. Dit werkingsmechanisme kan nieuwe aangrijpingspunten verschaffen voor toekomstige therapie_n. Tevens wordt in dit proefschrift een experimentele behandeling van Crohn met mesenchymale stamcellen (MSCs) beschreven. Deze behandeling is haalbaar en veilig gebleken, hetgeen aanleiding geeft tot verder onderzoek naar de effectiviteit van MSCs in de behandeling van Crohn. Tot slot werd het v__rkomen van kwaadaardige lymfomen onderzocht in IBD pati_nten in Nederland. Deze lymfomen bleken niet vaker voor te komen in de totale groep IBD pati_nten vergeleken met gezonde mensen, wel werd een verhoogde incidentie gezien in bepaalde leeftijdsgroepen. Daarnaast bleek het ontstaan van een EBV positief lymfoom (epstein Barr virus) sterk geassocieerd te zijn met het gebruik van azathioprine, een ander immuunsuppressieve therapie. Samenvattend beschrijft dit proefschrift verschillende aspecten van IBD therapie, en worden nieuwe inzichten en aangrijpinspunten verschaft Show less
