Posterior heart field and epicardium in cardiac development : PDGFRα and EMT

The processes of heart development, in which a primitive heart tube transforms into a specialized organ with two atria and ventricles divided by septa, are not only important for the understanding of congenital heart defects. These processes also give more insight for the development of heart regeneration therapies. The aim of this thesis was two fold. In the first part, we described the expression pattern of PDGF-A, -C and their receptor PDGFR-_ in the avian heart. Additionally, we studied mouse embryos in which the Pdgfr_ gene was mutated. These data show new insights into the role of PDGFR in a gene regulatory network which supports the development of cardiac structures. The second part of this thesis focused on the process of EMT in human adult EPDC and the role of the EPDC in the differentiation of cardiomyocytes. We gained more insight into the factors involved in EMT of the adult epicardium and showed that EPDCs are indispensible for proper integration of cardiomyocytes into a synchronously beating syncythium. These results might be beneficial for endogenous regulated cell-based cardiac repair.Nederlandse Hartstichting, Jurriaanse Stichting, Lead Pharma Holding BVUBL - phd migration 201

Myocardial perfusion MRI shows impaired perfusion of the mouse hypertrophic left ventricle

There is growing consensus that myocardial perfusion deficits play a pivotal role in the transition from compensated to overt decompensated hypertrophy. The purpose of this study was to systematically study myocardial perfusion deficits in the highly relevant model of pressure overload induced hypertrophy and heart failur

Epicardium-derived cells are important for correct development of the Purkinje fibers in the avian heart

During embryonic development, the proepicardial organ (PEO) grows out over the heart surface to form the epicardium. Following epithelial-mesenchymal transformation, epicardium-derived cells (EPDCs) migrate into the heart and contribute to the developing coronary arteries, to the valves, and to the myocardium. The peripheral Purkinje fiber network develops from differentiating cardiomyocytes in the ventricular myocardium. Intrigued by the close spatial relationship between the final destinations of migrating EPDCs and Purkinje fiber differentiation in the avian heart, that is, surrounding the coronary arteries and at subendocardial sites, we investigated whether inhibition of epicardial outgrowth would disturb cardiomyocyte differentiation into Purkinje fibers. To this end, epicardial development was inhibited mechanically with a membrane, or genetically, by suppressing epicardial epithelial-to-mesenchymal transformation with antisense retroviral vectors affecting Ets transcription factor levels (n = 4, HH39-41). In both epicardial inhibition models, we evaluated Purkinje fiber development by EAP-300 immunohistochemistry and found that restraints on EPDC development resulted in morphologically aberrant differentiation of Purkinje fibers. Purkinje fiber hypoplasia was observed both periarterially and at subendocardial positions. Furthermore, the cells were morphologically abnormal and not aligned in orderly Purkinje fibers. We conclude that EPDCs are instrumental in Purkinje fiber differentiation, and we hypothesize that they coo

Fibrosis has no detrimental effect on beating frequency in native-based cardiac microtissue models

Introduction Fibrosis is a hallmark of cardiac disease leading to changes in myocardial architecture that may hamper ventricular function. However, the influence of changes in the cardiac microenvironment on cardiomyocyte contractility remains unclear. Here, we describe the characterization of the cardiac microenvironment related to fibrosis in mouse hearts suffering from dystrophinopathies. The in vivo characteristics were compared to matrix organization in novel engineered in vitro cardiac microtissues. This high throughput cardiac model system allows investigation of the effect of fibrosis on tissue contraction in healthy and fibrotic microenvironments. Materials and Methods Mdx mice are surrogates for Duchenne muscular dystrophy and suffer from heart disease at 10 months of age. Changes in structure and composition of cardiac ECM mdx hearts were investigated and compared to matrix organization in age-matched controls. Microfabricated tissue gauses (µTUGs) with flexible microposts with uniaxial or biaxial constraints to manipulate matrix organization were fabricated using soft lithography. Neonatal mouse cardiomyocytes (CMs) and fibroblasts (cFBs) were resuspended in collagen/matrigel and seeded in the µTUGs to form microtissues to recapitulate the in vivo composition. Dynamic contractile behavior of the tissues was monitored for 7 days using deflection of the microposts. ECM distribution was determined at culture day 7 with immunofluorescence. Results Mdx mouse hearts were characterized by induction of patchy fibrosis in the left ventricle which caused disorganization of the ECM. The fibrotic areas were composed of collagen I, III and fibronectin and decreased the left ventricular myocardial stiffness. Using the µTUG system we obtained aligned (anisotropic) and disorganized (isotropic) ECM organization. In vitro distribution of collagens and fibronectin was homogenous, and not patchy as in the native tissue. Although CMs in anisotropic microtissues were more aligned, no higher contractile forces were generated. However, strain analysis showed that contraction was much more homogenous in anisotropic tissue compared to isotropic. Beating frequency was not affected by ECM organization but only by the percentage of cFBs in the microtissues. Discussion and Conclusion In this study characteristics of the healthy vs diseased cellular microenvironment were implemented in a novel in vitro cardiac tissue model. Our data indicate that ECM organization does not disturb the frequency of contraction but the number of cFBs does. Furthermore, we showed that these models are suitable to study contractility in cellular and pathophysiological processes that occur during heart disease and may facilitate in the optimization of new therapies for cardiac disease