Vascular diseases pose a significant burden on society, mainly due to the lack of effective treatment methods. A major reason for this is the shortcomings of current preclinical model systems. In... Show moreVascular diseases pose a significant burden on society, mainly due to the lack of effective treatment methods. A major reason for this is the shortcomings of current preclinical model systems. In this study, we have taken steps toward developing a more complex and relevant model system for (brain) blood vessels to address this issue. We used human induced pluripotent stem cells (hiPSCs) in the lab as a cellular source to generate the different types of cells needed for stable blood vessels. These cells were then combined in 3D microscopic culture environments (so-called vessel-on-chip systems) to closely mimic physiological conditions. Using this model, we were able to demonstrate specific abnormalities in a hereditary vascular disease, which was not possible with more 'traditional' culture methods. Additionally, we included brain cells to better investigate brain-specific disorders in the future. This work lays the essential foundation for a better understanding and treatment of complex vascular diseases, while potentially reducing the number of animal models needed. Show less
Manukjan, N.; Majcher, D.; Leenders, P.; Caiment, F.; Herwijnen, M. van; Smeets, H.J.; ... ; Foulquier, S. 2023
Cerebral small vessel disease is characterised by decreased cerebral blood flow and blood–brain barrier impairments which play a key role in the development of white matter lesions. We... Show moreCerebral small vessel disease is characterised by decreased cerebral blood flow and blood–brain barrier impairments which play a key role in the development of white matter lesions. We hypothesised that cerebral hypoperfusion causes local hypoxia, affecting oligodendrocyte precursor cell—endothelial cell signalling leading to blood–brain barrier dysfunction as an early mechanism for the development of white matter lesions. Bilateral carotid artery stenosis was used as a mouse model for cerebral hypoperfusion. Pimonidazole, a hypoxic cell marker, was injected prior to humane sacrifice at day 7. Myelin content, vascular density, blood–brain barrier leakages, and hypoxic cell density were quantified. Primary mouse oligodendrocyte precursor cells were exposed to hypoxia and RNA sequencing was performed. Vegfa gene expression and protein secretion was examined in an oligodendrocyte precursor cell line exposed to hypoxia. Additionally, human blood plasma VEGFA levels were measured and correlated to blood–brain barrier permeability in normal-appearing white matter and white matter lesions of cerebral small vessel disease patients and controls. Cerebral blood flow was reduced in the stenosis mice, with an increase in hypoxic cell number and blood–brain barrier leakages in the cortical areas but no changes in myelin content or vascular density. Vegfa upregulation was identified in hypoxic oligodendrocyte precursor cells, which was mediated via Hif1α and Epas1. In humans, VEGFA plasma levels were increased in patients versus controls. VEGFA plasma levels were associated with increased blood–brain barrier permeability in normal appearing white matter of patients. Cerebral hypoperfusion mediates hypoxia induced VEGFA expression in oligodendrocyte precursor cells through Hif1α/Epas1 signalling. VEGFA could in turn increase BBB permeability. In humans, increased VEGFA plasma levels in cerebral small vessel disease patients were associated with increased blood–brain barrier permeability in the normal appearing white matter. Our results support a role of VEGFA expression in cerebral hypoperfusion as seen in cerebral small vessel disease. Show less
Clinical development of drugs for central nervous system (CNS) disorders has been particularly challenging and still suffers from high attrition rates. This high attrition is mainly due to lack of... Show moreClinical development of drugs for central nervous system (CNS) disorders has been particularly challenging and still suffers from high attrition rates. This high attrition is mainly due to lack of efficacy during clinical development. To improve the prediction of CNS drug effects, knowledge of the drug concentration at the CNS target-site is indispensable. Unfortunately, measuring drug concentrations in the human CNS has major practical and ethical constraints. Therefore, alternative approaches to predict the drug pharmacokinetics (PK) at the target-site(s) in the human CNS should be searched for.In this research, a comprehensive CNS physiologically based PK (PBPK) model for prediction of drug concentration-time profiles in multiple CNS compartments was developed for both rats and humans. The CNS PBPK model only requires knowledge of physicochemical properties of the drugs, with the influence of the net active transporters on the drug exchange across the BBB and the BCSFB that can be obtained from in silico predictions, literature information and in vitro studies (if needed). Because of this, the developed CNS PBPK model is a powerful tool to predict drug PK in the CNS in the early stage of the drug development. Show less
Alzheimer's Disease (AD) is the main contributor to the syndrome dementia, one for which no cure has yet been developed. One of the key issues with developing a cure is the presence of the blood... Show moreAlzheimer's Disease (AD) is the main contributor to the syndrome dementia, one for which no cure has yet been developed. One of the key issues with developing a cure is the presence of the blood-brain barrier (BBB), a restrictive network involving the cerebral vasculature which prevents passage of therapeutics into the brain. Currently many if not all of the clinically most advanced therapeutic avenues involve the use of antibodies to target the production, oligomerization and aggregation of the peptide Amyloid beta. In this thesis, an antibody variant derived from camelids, such as llamas, is examined. The llama antibody is devoid of the light chain, and therefore significantly smaller and easier to manipulate than conventional antibodies. Various strategies have been explored, including the use ofdirect intrathecal injections, liposomes and adeno associated virusses to allow the llama antibody fragment to enter and remain in the brain of mouse models of AD. Various imaging modalities are used to determine the success of BBB passage, including micro-SPECT, intravital multiphoton microscopy, in vivo whole body fluorescence imaging and ex vivo IHC and IF. The thesis supplies a foundation for delivery of llama antibody fragments into the brain. Show less
