The pulmonary veins in pigs and horses: research towards the development of a new treatment strategy of atrial fibrillation in human patients and horses Tim Vandecasteele Dissertation submitted in fulfillment of the requirements for the degree of Doctor of Philosophy (PhD) in Veterinary Sciences 2018 Promoters: Prof. dr. P. Cornillie Prof. dr. G. van Loon Prof. dr. W. Van den Broeck dr. G. Van Langenhove Department of Morphology Faculty of Veterinary Medicine Ghent University Tim Vandecasteele 2018 The pulmonary veins in pigs and horses: research towards the development of a new treatment strategy of atrial fibrillation in human patients and horses Front cover: heart image by Henry Vandyke Carter (Gray’s Anatomy) Printed by University Press, Zelzate, Belgium www.universitypress.be Success consists of going from failure to failure without loss of enthusiasm -Winston Churchill- Table of contents List of abbreviations Preface………………………………………………………………………………………………………………………………… 7 Chapter 1: General introduction………………………………………………………………………………………… 11 1.1 Heart and pulmonary veins (PVs)………………………………………….……………………………….. 13 1.2 Cardiac conduction………………………………………………………………………………………………… 20 1.3 Atrial fibrillation (AF)………………………………………………………………………………………….….. 25 1.4 PVs in horses……………………………………………………………………………………………………….…. 45 1.5 Pigs as cardiovascular model……………………………………………………………………….…………. 46 Chapter 2: Scientific aims………………………………………………………………………………………….…….... 73 Chapter 3: The pulmonary veins of the pig………………………………………………………………………… 77 Chapter 4: Presence of ganglia and telocytes in proximity to myocardial sleeve tissue in the porcine pulmonary veins wall…………………………………………………………………….……………………. 101 Chapter 5: A preclinical study of an implanted device in the pulmonary veins, intended for the treatment of atrial fibrillation in an ovine model……………………………………………………..… 123 Chapter 6: A preliminary study of pulmonary vein implant applicability and safety as a potential ablation platform in a follow-up study in pigs…………………………………………………… 139 Chapter 7: Immunohistochemical identification of stent-based ablation lesions in the superior vena cava and pulmonary veins……………………………………………………………………..…. 153 Chapter 8: Isolation of pulmonary veins using a thermo reactive implantable device with external energy transfer: Evaluation in a porcine model……………………………………………….... 169 Chapter 9: The pulmonary veins of the horse……………………………………………………..…………… 187 Chapter 10: 3D reconstruction of the porcine and equine pulmonary veins, supplemented with the identification of telocytes in the horse………………………………………………………………. 205 1 Chapter 11: Echocardiographic identification of atrial-related structures and vessels validated by CT images of equine hearts……………………………………………………………………....… 221 Chapter 12: General discussion…………………………………………………………………………………..…… 239 Summary…………………………………………………………………………………………………………………………. 263 Samenvatting………………………………………………………………………………………………………..……….. 269 Curriculum vitae……………………………………………………………………………………………………………… 275 Bibliography…………………………………………………………………………………………………………………….. 279 Dankwoord………………………………………………………………………………………………………………………. 285 2 List of abbreviations AccV accessory pulmonary vein ACT activated clotting time AEF atriooesophageal fistula AF atrial fibrillation ALL accessory lung lobe AN antrum Ao aorta AS atrial side AV aortic valve AVN atrioventricular node BT brachiocephalic trunk C temperature increase Ca caudal CaVC caudal vena cava CDLL caudal lung lobe CLL cranial lung lobe CLL(cp) cranial part cranial lung lobe CLL(cdp) caudal part cranial lung lobe CP cardiac plexus Cr cranial CrVC cranial vena cava CT computed tomography D dorsal DAB diaminobenzidine ECG electrocardiography EP electrophysiology H&E hematoxylin & eosin HPF high power field HR heat ring HRP horse radish peroxidase ICV inferior cava vein ILL intermediate lung lobe 3 IT intervenous tubercle L lumen LA left atrium LCLL left cranial lung lobe LCDLL left caudal lung lobe LiCd bronchus left caudal lung lobe LcdV1 left caudal pulmonary vein (cranial aspect) LcdV2 left caudal pulmonary vein (intermediate aspect) LcdV3 left caudal pulmonary vein (caudal aspect) LcrV left cranial pulmonary vein LiCrpCd bronchus left cranial lung lobe (caudal part) LiCrpCr bronchus left cranial lung lobe (cranial part) LIPV left inferior pulmonary vein LLL lingual lung lobe LPA left pulmonary artery LSPV left superior pulmonary vein LV left ventricle MRI magnetic resonance imaging MYBPC3 myosin binding protein C Myo myocardial sleeve NL nerves of left atrium O oblique OF oval fossa PA pulmonary arteries pAF paroxysmal atrial fibrillation PLLA-GP posterolateral left atrial ganglionated plexi PMLA-GP posteromedial left atrial ganglionated plexi Po podom PS pulmonary side PV pulmonary vein PVO pulmonary vein ostia PVs pulmonary veins PVI pulmonary vein isolation 4 PVS pulmonary vein stenosis PT pulmonary trunk QCA quantitative coronary angiography RA right atrium RAA right atrial appendage RCd bronchus right caudal lung lobe RCDLL right caudal lung lobe RcdV1 right caudal pulmonary vein (cranial aspect) RcdV2 right caudal pulmonary vein (intermediate dorsal aspect) RcdV3 right caudal pulmonary vein (intermediate ventral aspect) RcdV4 right caudal pulmonary vein (caudal aspect) RCLL right cranial lung lobe RCr bronchus trachealis (right cranial lung lobe) RcrV right cranial pulmonary vein Rim bronchus right middle lung lobe RLa bronchus accessory lung lobe RPA right pulmonary artery Rt right RV right ventricle SCV superior caval vein SG stellate ganglion SLA-GP superior left atrial ganglionated plexi SN sinus node TC terminal crest Tcb telocyte cell body TEM transmission electron microscopy Tpr telocyte prolongation V ventral VN vagal nerves VWT-peak peak temperature during ablation VWT-pre vessel wall temperature prior to ablation 5 6 Preface 7 8 Preface The research presented in this manuscript was originally initiated from a rather simple, straightforward question: “is the pig a suitable animal model to test a newly developed implantable device as a novel treatment option for atrial fibrillation in human patients?” Atrial fibrillation is in fact the most important cardiac arrhythmia in man, affecting an increasing number of people, especially at higher age. Although various treatment options have been developed, the method of the last resort, i.e. catheter ablation, has an unsatisfyingly high recurrence rate of at least 20-40% (Darby, 2016), requiring the patient to undergo this time-consuming procedure multiple times under general anesthesia. The intended procedure involving the newly developed ablation device is both aiming for a significantly lower recurrence rate as well as a much shorter intervention time. However, it still needed to be tested in an animal model. The pig was selected as cardiovascular model as pigs are frequently used as an animal model in different research domains and the porcine heart is comparable in size with that of humans. As the device was to be inplanted in the pulmonary veins, the main focus of the investigation resided in the comprehensive elucidation of the anatomical organization of these vessels draining into the left atrium of the heart. Although the cause and development of atrial fibrillation has still not been fully elucidated, in man, the role of the pulmonary vein walls as sites of ectopic foci triggering the arrhythmia have long been recognized. As such, many in-depth morphological and electrophysiological studies of the pulmonary vein walls in man are available in literature, which is in huge contrast with the availability of similar data for the pig. The first histological literature on the human pulmonary veins dates back to the 19th century, yet it is only since the 2000’s that the scientific interest in these structures re-emerged (Stieda, 1877; Nathan and Eliakim, 1966; Moubarak et al., 2000; Hassink et al., 2003). Therefore, as a first step of this investigation, the complete unravelment of the topographic organization of the porcine pulmonary veins was pursued. Secondly, the walls of these veins were histologically investigated to obtain an overview of all present structures and cell types that either might initiate or facilitate the propagation of ectopic pulses and/or might be affected by the intended ablation (heat scarring) in this region. These data made it possible to proceed towards the invasive procedures in anesthetized pigs. A prototype of the new device for atrial fibrillation treatment was developed, applied and tested on pigs and adjusted during different study phases. Simultaneously with the examination of the porcine pulmonary veins, this research was extended towards the equine heart. In fact, also horses are quite prone to develop atrial fibrillation. However, in contrast to the situation in man, similar advances in the understanding of the underlying mechanisms of atrial fibrillation and the development of new treatment options in the equine patient are still too far out of reach. This is mainly because of our current inability to unambiguously visualize and identify the individual pulmonary veins in the living animal and consequently determine 9 Preface their involvement in impaired cardiac function. Echocardiography is the most important, if not the only feasible visualisation technique for the equine heart. However, due to the far more complex topography of drainage area of the equine pulmonary veins into the left atrium and the size of horses, most structures in this region are too difficult to identify on two-dimensional cross sectionial