Since the adult heart has minimal capacity to repair itself, myocardial infarction often leads to pathological remodeling and ultimately to the development of fatal heart failure. Upon ischemic... Show moreSince the adult heart has minimal capacity to repair itself, myocardial infarction often leads to pathological remodeling and ultimately to the development of fatal heart failure. Upon ischemic injury, the epicardium, the outer layer of the heart which is essential for cardiac development, becomes re-activated and displays reparative potential. In this process, epicardial epithelial-to-mesenchymal transition (epiMT) is an essential step. We hypothesize that the reparative capacity of the heart can be improved by enhancing the participation of the epicardium to cardiac repair, particularly by stimulating the occurrence of epiMT. Therefore, the aim of my thesis is to find ways to boost epiMT in the injured heart. In this thesis, we describe a cell culture model which allows us to study epiMT. Using this model, we identify novel epiMT regulators. Because EMT is also involved in pathological remodeling, application of an epiMT stimulator should be transient and local. Therefore, we describe a method to locally administer these factors to the injured mouse heart. Show less
In the current thesis the embryological origin of the developing atrioventricular (AV) canal and atrioventricular node (AVN) was investigated, using different techniques, including... Show moreIn the current thesis the embryological origin of the developing atrioventricular (AV) canal and atrioventricular node (AVN) was investigated, using different techniques, including immunohistochemical staining, lineage tracing and genetic characterization. In addition, the role of the RHOA-ROCK signaling pathway on development of key components of the cardiac conduction system (CCS) was studied. Finally, the potential role of the epicardium in autonomic modulation of the heartbeat was investigated. Show less
This thesis describes the use of zebrafish to study Noonan-(NS) and LEOPARD syndromes (LS), two autosomal dominant disorders with overlapping symptoms, caused by mutations in protein-tyrosine... Show moreThis thesis describes the use of zebrafish to study Noonan-(NS) and LEOPARD syndromes (LS), two autosomal dominant disorders with overlapping symptoms, caused by mutations in protein-tyrosine phosphatase, non-receptor type 11 (PTPN11). Intriguingly, while NS mutations result in a more __active__ state of Shp2, LS mutations give rise to a PTP defective protein. First, we studied the role of ptpn11 in zebrafish embryonic development. Whereas ptpn11a is crucial for development ptpn11b seems dispensible. Moreover, using phoshoproteomics we show hypo- and hypertyrosyl phosphorylation of Fer kinase and PZR (Protein zero related), respectively and study their roles as (potential) interacting proteins in the etiology of both NS and LS. Defective heart development is a prominent symptom of NS and LS. Embryos expressing NS and LS Shp2 showed impaired expression of laterality markers, impaired cilia function in Kupffer__s vesicle and at later stages, heart function defects. Finally we identified several mutations in Alpha-2-Macroglobulin-Like-1 (A2ML1) in patients diagnosed with NS. Functional characterization of these mutations in zebrafish showed NS-like developmental defects. This is the first example of an extracellular factor in a disorder clinically related to NS and LS, providing potential new leads for better understanding of the molecular basis of these developmental syndromes. Show less
This thesis is separated in two parts (Part I and Part II) in which normal and abnormal heart development are studied and related to congenital heart disease, in particular to the etiology of... Show moreThis thesis is separated in two parts (Part I and Part II) in which normal and abnormal heart development are studied and related to congenital heart disease, in particular to the etiology of supraventricular arrhythmias in fetuses and neonates. Part I describes the development of the posterior heart field derived venous pole of the heart specifically in correlation to the role of Shox2 and podoplanin in that particular area. Furthermore, the developmental processes in this region seem to have an important role in the anlage of the cardiac conduction system and epicardial lineage development of the heart. In the second part of this thesis (Part II) the aetiology of a specific subtype of supraventricular tachycardias i.e. atrioventricular reentry tachycardias (AVRTs) are related to normal heart development in human and mouse. AVRTs are one of the most common types of tachyarrhythmias at the perinatal period of development. We demonstrate that perinatal AVRTs might be related to incomplete formation of the isolating annulus fibrosus resulting in the temporary persistence of accessory myocardial connections between the atria and ventricles. We furthermore, demonstrate the late outcome of fetal brady- and tachyarrhythmia cases. Show less
Clinical mapping studies demonstrate that cardiac arrhythmias are often encountered at specific anatomical sites. The anatomical development of the heart and the cardiac conduction system are... Show moreClinical mapping studies demonstrate that cardiac arrhythmias are often encountered at specific anatomical sites. The anatomical development of the heart and the cardiac conduction system are narrowly related. The thesis starts with a Chapter 1 that provides a general overview of the basics of cardiac development, development of the cardiac conduction system, markers for the developing cardiac conduction system and anatomical predilection sites for the occurrence of clinical arrhythmias. Next, attention is focused on clinical arrhythmias in adults in relation to cardiac anatomy, the treatment of these arrhythmias and imaging techniques used to visualise the substrate. Subsequently, in part I (Chapter 2-4) of the thesis, a developmental origin of clinical arrhythmias is hypothesized. The developing cardiac conduction system is studied and a developmental origin of clinical arrhythmias is hypothesized based on the spatial expression pattern of the conduction system marker CCS-lacZ in murine embryos. Furthermore, incorporation of the primitive pulmonary vein in the left atrium is studied in sequential developmental stages in humans. In part II (Chapter 5-10), the treatment of clinical arrhythmias that are initiated and/or perpetuated at specific anatomical locations in the heart (particularly the pulmonary veins) guided by imaging techniques for visualisation of the substrate, is described. Show less
Ligation of the right lateral vitelline vein in chicken embryos (venous clip) results in changes in the intracardial blood flow patterns, and in functional and morphological cardiovascular defects.... Show moreLigation of the right lateral vitelline vein in chicken embryos (venous clip) results in changes in the intracardial blood flow patterns, and in functional and morphological cardiovascular defects. This demonstrates that blood flow, of which shear stress is a derivative, plays an important role in cardiovascular development. A general mechanism of shear stress sensing by endothelial cells remained to be elucidated. We postulate that the cytoskeleton functions as a central shear stress transducer, which uses a primary cilium for detection of low shear forces. During normal cardiogenesis we demonstrated that shear stress responsive genes lung Kruppel-like factor (KLF2) and endothelial nitric oxide synthase (NOS-3) are expressed in the endothelium and endocardium of structural lumen constrictions, where endothelin-1 (ET-1), a growth factor and vasoconstrictor, is absent and shear stress is high. After vitelline ligation KLF2 and NOS-3 were locally increased in the heart whereas ET-1 was down-regulated. Infusion of ET-1 or antagonists of the endothelin-A (ETA) and/or endothelin-B (ETB) receptor in the chicken embryonic circulation resulted in similar but less severe defects in cardiac function and morphology, demonstrating that components of the ET-1 pathway play a role in the development of cardiovascular defects in the venous clip model. Show less
