Notch: een moleculaire schakelaar die instaat voor de metabole aanpassingen nodig voor redox homeostase in rustende endotheelcellen

Blood vessels ensure nutrient and oxygen supply to all cells via transport through the endothelium. This endothelial cell (EC) layer lining the luminal side of blood vessels forms a selective interface between the circulating blood and the surrounding tissue. ECs display high plasticity: i.e. in health, ECs in established blood vessels are quiescent and rarely divide, but upon stimulation in pathological conditions (ischemic, inflammatory or malignant diseases), they switch to a proliferative state and can migrate to form new sprouts. In relation to their involvement in pathological conditions, these activated ECs are subject of intense research in the cardiovascular field. Recent findings demonstrate the existence of a metabolic switch coinciding with the angiogenic switch. While most studies focus on targeting the angiogenic switch and preventing activation and sprouting, we sought to study the switch to quiescence with the aim to obtain valuable insight into the metabolic adaptation associated with this switch. Given their distinct environmental conditions, functional role and physiology, quiescent and angiogenic ECs are expected to have different metabolic needs. Previous data of the host lab showed that angiogenic ECs use the majority of metabolic energy for biosynthesis reactions to synthesize DNA, proteins and lipids, and to facilitate migration. Conversely, since quiescent cells are exempted from these laborious tasks, we hypothesized that reprogramming of basal endothelial metabolism is necessary to facilitate this ‘pro-quiescent switch’ from rapid growth to quiescence. In this thesis work, diverse metabolic assays and measurements combined with biochemical and molecular techniques were applied to characterize the nature and mechanisms of this pro-quiescent switch. This revealed that quiescent ECs are not generally hypometabolic, but display an activated oxidative pentose phosphate pathway (oxPPP) as well as increased fatty acid b-oxidation (FAO). These findings contradict the commonly held opinion that decreased metabolism is a hallmark of quiescence. This metabolic switch does not result in increased ATP or biomass synthesis, but instead promotes NADPH production, providing the necessary reducing power needed to lower the oxidative status of the cell. This level of redox control is most likely needed to prevent high levels of reactive oxygen species (ROS) since EC reside in an oxygen rich and oxidative environment. This prevents ROS sensitive activation of ECs and to avert prolonged oxidative stress associated with EC dysfunction (as is observed in various cardiovascular diseases). Furthermore, the data indicates that FAO, which in contrast to the oxPPP is not able to directly produce NADPH, likely stimulates NADPH formation in ECs via the mitochondrial NADPH producing malic enzyme 3 (ME3), isocitrate dehydrogenase 1 (IDH1) and nicotinamide nucleotide transhydrogenase (NNT). Functionally, I demonstrate that FAO in quiescent ECs is essential to constrain ROS sensitive induction of endothelial permeability and counteracts permeability defects and EC dysfunction as observed in cardiovascular diseases. I further show that Dll4-Notch signaling serves as a molecular hub controlling the metabolic adaptations associated with the induction of EC quiescence. Interestingly, Dll4-Notch directly modulates the FAO flux through binding of the Notch intracellular domain (NICD) in complex with the transcription factor CSL to the promoter of carnitine palmitoyl transferase 1a (CPT1a), a key enzyme of FAO, and subsequent transcriptional activation. Taken together, I provide valuable information regarding the metabolic profile of quiescent ECs and present a molecular mechanism responsible for the metabolic switch associated with the reverse angiogenic switch.status: publishe