arrhythmogenic rv cardiomyopathy/dysplasia...preface this book covers all the recent research...

Arrhythmogenic RV Cardiomyopathy/DysplasiaRecent Advances

FRANK I. MARKUS

ANDREA NAVA

GAETANO THIENE

FRANK I. MARCUS • ANDREA NAVA • GAETANO THIENE (Eds)

Arrhythmogenic RVCardiomyopathy/Dysplasia

Recent Advances

Indice III

FRANK I. MARCUS

Section of CardiologyDepartment of MedicineSarver Heart CenterUniversity of ArizonaTucson (AZ), USA

ANDREA NAVA

Department of Cardiological, Thoracicand Vascular SciencesUniversity of Padua, Italy

GAETANO THIENE

Department of Medical Diagnostic SciencesUniversity of Padua, Italy

Library of Congress Control Number: 2007922495

ISBN 978-88-470-0489-4 Springer Milan Berlin Heidelberg New York

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IV Indice

PREFACE

This book covers all the recent research highlights of arrhythmogenic right ventric-ular cardiomyopathy/dysplasia (ARVC/D), a recently discovered heart muscle dis-ease which is a major threat to the life of affected young people. It summarizes near-ly 25 years of investigation on the etiology, genetics, pathology, clinical features, di-agnosis, and treatment of ARVC/D. In particular, a 5-year research programsupported by grants from both the European Community and the National Heart,Lung and Blood Institutes has contributed to the discovery of seven disease-causinggenes, thus opening new avenues for the early identification of affected patients andprevention of sudden death.

A Workshop was held in Venice, Italy, October 3, 2005, as part of the Venice Ar-rhythmia Meeting, where the European and American investigators presented anddiscussed several major achievements which are now reported in this book. As a re-sult of these coordinated efforts, great advances have been made in the recognitionand understanding of the disease, which are summarized in this book.

Molecular genetics has established this cardiomyopathy as a familial disordercaused by mutation of the genes that encode cell junction proteins, resulting in de-fective cellular adhesion with specific immunohistochemical and ultrastructural al-terations. Remodelling at the intercalated disk may trigger a cascade of events in-cluding apoptotic cell death, fibrofatty replacement, and electrical instability. Theleft ventricle may be involved early in the course of some genetic types of this dis-ease. This finding alters the traditional concept of a disease confined to the rightventricle.

Genetic screening can detect symptomatic carriers in the early stage of the dis-ease. Magnetic resonance imaging with gadolinium permits in vivo identification offibrous tissue. Electroanatomic mapping can reveal areas of fibrofatty replacementof the right ventricular myocardium. The implantable cardioverter defibrillator hasbeen shown to prevent sudden cardiac death. Whether it should be implanted forsecondary as well as for primary prevention is still controversial.

Screening prior to participation in competitive sports has been found to be effec-tive for the identification of subjects at risk and is life-saving by disqualifying af-fected individuals and avoiding competitive type effort. Sudden death of young ath-letes declined fivefold after implementation of preparticipation screening, mainlydue to identification of ARVC/D.

A panel of experts have contributed to writing this monograph, which will be anessential reference for clinicians and scholars in human genetics, pathology, cardiol-ogy, and radiology as well as in forensic and sports medicine.

We would like to express our gratitude to the European Commission, Brussels,CARIPARO Foundation, Padua, and Veneto Region, Venice, for their financial sup-port, which made this publication possible. A special thanks is given to Chiara Car-turan and Kathy Gear for their continuous, outstanding collaboration over the yearsof these investigations.

The Editors

Indice V

TABLE OF CONTENTS

LIST OF CONTRIBUTORS . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . IX

ABOUT THE EDITORS . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . XIII

Introduction: ARRHYTHMOGENIC RIGHT VENTRICULAR CARDIOMYOPATHY/DYSPLASIA CLARIFIED . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1Gaetano Thiene, Andrea Nava, Frank I. Marcus

CHAPTER 1 ADVANCES IN GENETICS: DOMINANT FORMS . . . . . . . . . . . . . . . . . . . . . 7Alessandra Rampazzo, Gian Antonio Danieli

CHAPTER 2 ADVANCES IN GENETICS: RECESSIVE FORMS . . . . . . . . . . . . . . . . . . . . . 15Nikos Protonotarios, Adalena Tsatsopoulou

CHAPTER 3 GENOTYPE-PHENOTYPE CORRELATIONS . . . . . . . . . . . . . . . . . . . . . . . . . . 21Barbara Bauce, Andrea Nava

CHAPTER 4 AUTOPSY AND ENDOMYOCARDIAL BIOPSY FINDINGS . . . . . . . . . . . 29Cristina Basso, Gaetano Thiene

CHAPTER 5 CELL ADHESION PATHOLOGY . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 45Jeffrey E. Saffitz

CHAPTER 6 ULTRASTRUCTURAL SUBSTRATES . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 53Elzbieta Czarnowska, Mila Della Barbera, Gaetano Thiene, Marialuisa Valente, Cristina Basso

CHAPTER 7 TRANSGENIC ANIMAL MODELS . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 61Matteo Vatta, Zhao Yang, Jeffrey A.Towbin

CHAPTER 8 SPONTANEOUS ANIMAL MODELS . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 69Philip R. Fox, Cristina Basso, Gaetano Thiene, Barry J. Maron

CHAPTER 9 POSSIBLE CAUSATIVE OR CONTRIBUTING ROLE OF VIRUSES . . . 79Fiorella Calabrese, Elisa Carturan, Gaetano Thiene, Jeffrey A.Towbin

CHAPTER 10 DIAGNOSIS: TASK FORCE CRITERIA INCLUDING MODIFICATIONSFOR FAMILY MEMBERS . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 87Deirdre Ward, Petros Syrris, Srijita Sen-Chowdhry,William J. McKenna

Indice VII

CHAPTER 11 STRENGTHS AND WEAKNESSES OF THE TASK FORCE CRITERIA –PROPOSED MODIFICATIONS . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 97Frank I. Marcus, Duane Sherrill

CHAPTER 12 IDIOPATHIC RIGHT VENTRICULAR OUTFLOW TRACT TACHYCARDIA . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 105Gianfranco Buja, Barbara Ignatiuk,Thomas Wichter, Domenico Corrado

CHAPTER 13 ELECTROCARDIOGRAPHIC MANIFESTATIONS . . . . . . . . . . . . . . . . . . . . 121Wojciech Zareba, Katarzyna Piotrowicz, Pietro Turrini

CHAPTER 14 ECHOCARDIOGRAPHY . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 129Danita Yoerger-Sanborn, Michael H. Picard, Barbara Bauce

CHAPTER 15 MR AND CT IMAGING . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 135Harikrishna Tandri, Hugh Calkins, David A. Bluemke

CHAPTER 16 DIAGNOSTIC ROLE OF ANGIOGRAPHY . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 147Thomas Wichter, Julia Indik, Luciano Daliento

CHAPTER 17 ELECTROPHYSIOLOGIC STUDY INCLUDING ELECTROANATOMICMAPPING . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 159Domenico Corrado, James Daubert, Cristina Basso, Gianfranco Buja, Gaetano Thiene

CHAPTER 18 RISK STRATIFICATION AND ANTIARRHYTHMIC DRUG THERAPY 171Thomas Wichter, Domenico Corrado, Matthias Paul

CHAPTER 19 CATHETER ABLATION OF VENTRICULAR TACHYCARDIA . . . . . . . . . 181Guy Fontaine, Jérôme Lacotte, Françoise Hidden-Lucet, Robert Frank

CHAPTER 20 THE ROLE OF THE IMPLANTABLE CARDIAC DEFIBRILLATOR IN THE MANAGEMENT . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 189Olaf Hedrich, Gianfranco Buja, Domenico Corrado, Mark S. Link,Thomas Wichter,N.A. Mark Estes III

CHAPTER 21 MANAGEMENT OF HEART FAILURE . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 199Elzbieta Katarzyna Wlodarska, Marek Konka

CHAPTER 22 SUDDEN DEATH IN YOUNG ATHLETES . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 205Domenico Corrado, Cristina Basso, Gaetano Thiene

SUBJECT INDEX . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 213

VIII Table of Contents

CRISTINA BASSO, MD, PhDDepartment of Medical Diagnostic SciencesUniversity of Padua, Italy

BARBARA BAUCE, MD, PhDDepartment of Cardiological, Thoracicand Vascular SciencesUniversity of Padua, Italy

DAVID A. BLUEMKE, MD, PhDRussel H. Morgan Department of Radiology and Radiological ScienceJohns Hopkins Medical InstitutionsBaltimore (MD), USA

GIANFRANCO BUJA, MDDepartment of Cardiological, Thoracicand Vascular SciencesUniversity of Padua, Italy

FIORELLA CALABRESE, MDDepartment of Medical Diagnostic SciencesUniversity of Padua, Italy

HUGH CALKINS, MDDivision of Cardiology Johns Hopkins Medical InstitutionsBaltimore (MD), USA

ELISA CARTURAN


DOMENICO CORRADO, MD, PhDDepartment of Cardiological, Thoracicand Vascular SciencesUniversity of Padua, Italy

ELZBIETA CZARNOWSKA, PhDDepartment of PathologyThe Children’s Memorial Health InstituteWarzaw, Poland

LUCIANO DALIENTO, MDDepartment of Cardiological, Thoracicand Vascular SciencesUniversity of Padua, Italy

GIAN ANTONIO DANIELI, BScDepartment of BiologyUniversity of Padua, Italy

JAMES DAUBERT, MDDivision of CardiologyDepartment of Internal MedicineStrong Memorial HospitalUniversity of RochesterRochester (NY), USA

MILA DELLA BARBERA


N.A. MARK III ESTES, MDTufts University School of MedicineNew England Cardiac Arrhythmia CenterTufts-New England Medical CenterBoston (MA), USA

GUY FONTAINE, MD, PhDDepartment of Cardiac ArrhythmiasInstitute of CardiologyUniversity Hospital and School of Medicine Pitié-SalpêtrièreParis, France

PHILIP R. FOX, DVMCaspary Institute The Animal Medical CenterNew York (NY), USA

ROBERT FRANK, MDDepartment of Cardiac ArrhythmiasInstitute of CardiologyUniversity Hospital and School of Medicine Pitié-SalpêtrièreParis, France

OLAF HEDRICH, MDTufts-New England Medical CenterCardiac Arrhythmia ServiceBoston (MA), USA

Elenco autori IX

LIST OF CONTRIBUTORS

FRANÇOISE HIDDEN-LUCET, MDDepartment of Cardiac ArrhythmiasInstitute of CardiologyUniversity Hospital and School of Medicine Pitié-SalpêtrièreParis, France

BARBARA IGNATIUK, MD, PhDDepartment of Cardiological, Thoracicand Vascular SciencesUniversity of Padua, Italy

JULIA INDIK, MDDepartment of CardiologySarver Heart CenterUniversity of Arizona College of MedicineTucson (AZ), USA

MAREK KONKA, MD, PhDDepartment of Congenital Heart DiseaseInstitute of CardiologyWarsaw, Poland

JÉRÔME LACOTTE, MDDepartment of Cardiac ArrhythmiasInstitute of CardiologyUniversity Hospital and School ofMedicine Pitié-SalpêtrièreParis, France

MARK S. LINK, MDTufts-New England Medical CenterCardiac Arrhythmia ServiceBoston (MA), USA

FRANK I. MARCUS, MDSection of CardiologyDepartment of MedicineSarver Heart CenterUniversity of ArizonaTucson (AZ), USA

BARRY J. MARON, MDHypertrophic Cardiomyopathy CenterMinneapolis Heart Institute FoundationMinneapolis (MN), USA

WILLIAM J. MCKENNA, MD, DSc, FRCPDepartment of MedicineUniversity College London and University College London Hospitals TrustLondon, UK

ANDREA NAVA, MDDepartment of Cardiological, Thoracicand Vascular SciencesUniversity of Padua, Italy

MATTHIAS PAUL, MDDepartment of Cardiology and AngiologyUniversity Hospital of MünsterMünster, Germany

MICHAEL H. PICARD, MDCardiac Ultrasound LaboratoryMassachusetts General HospitalBoston (MA), USA

KATARZYNA PIOTROWICZ, MDHeart Research Follow-up ProgramDivision of CardiologyUniversity of Rochester Medical CenterRochester (NY), USA

NIKOS PROTONOTARIOS, MDYannis Protonotarios Medical CentreHora Naxos, Naxos, Greece

ALESSANDRA RAMPAZZO, BSc, PhDDepartment of BiologyUniversity of Padua, Italy

JEFFREY SAFFITZ, MD, PhDHarvard Medical School and Department of PathologyBeth Israel Deaconess Medical CenterBoston (MA), USA

SRIJITA SEN-CHOWDHRY, MA, MBBS, MRCPDepartment of MedicineUniversity College London and University College London Hospitals TrustLondon, UK

DUANE SHERRILL, PhDMel and Enid Zuckerman College ofPublic HealthUniversity of ArizonaTucson (AZ), USA

PETROS SYRRIS, PhDDepartment of MedicineUniversity College London and University College London Hospitals TrustLondon, UK

HARIKRISHNA TANDRI, MDDivision of CardiologyJohns Hopkins Medical InstitutionsBaltimore (MD), USA

GAETANO THIENE, MDDepartment of Medical Diagnostic SciencesUniversity of Padua, Italy

JEFFREY A.TOWBIN, MDPediatric CardiologyTexas Children’s HospitalBaylor College of MedicineHouston (TX), USA

ADALENA TSATSOPOULOU, MDYannis Protonotarios Medical CentreHora Naxos, Naxos, Greece

X List of Contributors

List of Contributors XI

PIETRO TURRINI, MD, PhDDepartment of CardiologyCivil Hospital of CamposampieroCamposampiero (PD), Italy

MARIALUISA VALENTE, MDDepartment of Medical Diagnostic SciencesUniversity of Padua, Italy

MATTEO VATTA, PhDPediatric CardiologyTexas Children’s HospitalBaylor College of MedicineHouston (TX), USA

DEIRDRE WARD, MRCPIDepartment of MedicineUniversity College London and University College London Hospitals TrustLondon, UK

THOMAS WICHTER, MD, FESCDepartment of Cardiology and AngiologyUniversity Hospital of MünsterMünster, Germany

ELZBIETA KATARZYNA WLODARSKA, MD, PhDDepartment of Congenital Heart DiseaseInstitute of CardiologyWarsaw, Poland

ZHAO YANG, MDPediatric CardiologyTexas Children’s HospitalBaylor College of MedicineHouston (TX), USA

DANITA YOERGER-SANBORN, MD, MMScCardiac Ultrasound LaboratoryCardiology DivisionMassachusetts General HospitalHarvard Medical SchoolBoston (MA), USA

WOJCIECH ZAREBA, MD, PhDHeart Research Follow-up Programand Clinical Cardiology ResearchDivision of Cardiology University of Rochester Medical CenterRochester (NY), USA

Elenco degli autori XIII

FRANK I. MARCUS, MDFrank Marcus is Professor of Medicine at the Sarver Heart Center,University of Arizona, Tucson, USA. He reported the first clinicalseries of ARVC/D patients and continues to contribute to the un-derstanding of the diagnosis and treatment of this disease. He isthe principal investigator of the North American Multidiscipli-nary study of ARVC/D supported by the NHLBI, that consists ofa registry, a data coordination center, a genetic laboratory andseveral international core laboratories.

ANDREA NAVA, MDAndrea Nava is Professor of Cardiology at the University of Pad-ua, Italy. He first recognized the familial occurrence of ARVC/Din the Veneto Region and has made major contributions to theearly identification, discovery of defective genes and phenotypicexpression of the disease. He is in charge of the family screeningprogram that consists of clinical and genetic investigations.

GAETANO THIENE, MDGaetano Thiene is Professor of Cardiovascular Pathology and Di-rector of the Institute of Pathological Anatomy at the Universityof Padua, Italy. He was trained both in Cardiology and Pathology.He first reported ARVC/D as a major cause of sudden, unexpect-ed death in the young and in athletes and established diagnosticcriteria for endomyocardial biopsy. He coordinates the Europeanstudy and has implemented the European Registry of ARVC/Dsupported by the European Commission.

ABOUT THE EDITORS

The first monograph on arrhythmogenic right ven-tricular cardiomyopathy/dysplasia (ARVC/D) waspublished 10 years ago [1]. Since then, there havebeen major advancements in the basic knowledge ofthe disease as well as a better understanding of the di-agnosis and treatment. A workshop was held inVenice, Italy on October 3, 2005, where research onvarious aspects of this disease, both biological andclinical was presented.

This book has assembled contributions in the formof a monograph rather than as the publication of Pro-ceedings. In addition, some topics were added in a sim-ilar format to that of the first monograph [1], whichfollowed a meeting on ARVC/D held in Paris in 1996.

In the last 10 years our understanding of this dis-ease has been impressive. This is the logical conse-quence of a research strategy with clear goals.

At the turn of the millennium, following a series ofmeetings of experts from both sides of the Atlantic, itbecame evident that we had to merge the expertise ofscientists and clinicians attracted by the mystery ofARVC/D and its devastating physical and social conse-quences into an “army”for the fight against the disease.

An International Registry was considered manda-tory in order to collect study material and concen-trate efforts on this rare disease [2].

It was then decided to apply for grants to the Eu-ropean Commission (EC) and to the National Instituteof Health (NIH). Two teams were created, one in Eu-rope coordinated by Gaetano Thiene [3] and one inNorth America coordinated by Frank Marcus [4]. Uti-lizing a similar database and having some Core Labo-ratories in common, the two projects were initiated.The structure was somewhat different: the EuropeanRegistry enrolled patients who were both previously di-agnosed as well as those with the recent onset of symp-toms, whereas the North American Registry enrolledonly newly diagnosed patients. Guidelines for diag-nostic criteria and protocols were implemented. Ge-netic investigation was an integral part of both studies.Fortunately, the two projects were approved and fund-ed for 5 years, thus allowing the start of a major inter-disciplinary study of ARVC/D. The results exceeded our

best expectations, resulting in numerous importantpublications in well-recognized cardiovascular jour-nals. In this monograph the advances in our knowledgewill be summarized in didactic presentations.

A brief overview of the major advances is as follows:1. The genetic background of this hereditary-famil-

ial, monogenic disease has been clarified. Despitegenetic heterogeneity with rare variants, it hasbeen demonstrated that both autosomal and re-cessive forms are due to defects of genes encod-ing desmosomal proteins of the intercalated disc:plakoglobin [5], desmoplakin, [6] plakophilin[7], desmoglein [8], and desmocollin [9]. This ex-plains why the disease is now called a desmoso-mal cardiomyopathy [10, 11]. To date, seven dis-ease genes have been identified during the courseof the EC and NIH research projects – an unbe-lievable achievement. Genetic screening is nowfeasible for the detection of gene carriers and ear-ly clinical diagnosis [12].

2. The pathological substrate of the disease has beenclarified at the ultrastructural level with evidenceof remodeling of intercalated disc (widening ofintercellular space with abnormal desmosomes)[13]. These structural abnormalities can poten-tially trigger a cascade of events following parietalstretch (apoptotic cell death, fibrofatty replace-ment, electrical instability). There is now evi-dence that the left ventricle is also involved. Insome variants of the disease it has been shownthat the left ventricle is primarily affected, thus ex-panding the previous concept that the disease isconfined to the right ventricle [14-16]. The diag-nostic role of endomyocardial biopsy has beenimproved by updating morphometric parameters.

3. Both the advantages and limitations of imagingmodalities have been clarified and are beginningto be subjected to quantitative analysis. Magnet-ic resonance imaging is being expanded in scopenot only to study the morphology and dysfunc-tion of the ventricles, but also to identify tissuecomposition, particularly fibrosis utilizing gadol-inium late enhancement.

Introduction: ARRHYTHMOGENIC RIGHT VENTRICULAR CARDIOMYOPATHY/DYSPLASIA CLARIFIEDGaetano Thiene, Andrea Nava, Frank I. Marcus

4. With regard to advances in the invasive diagnos-tic techniques, electroanatomic mapping is beingevaluated to detect areas of decreased electricalactivity, which has been found to correspond todiffuse segmental fibrofatty atrophy [17]. Thismay be important not only for the diagnosis ofthe disease, but also for the identification of areasthat may be the target for catheter ablation. Nev-ertheless, the precise diagnostic role of elec-troanatomic mapping needs further clarification.Also the role of ablation for treatment of ventric-ular arrhythmias needs to be reinvestigated uti-lizing the technique of electroanatomic mapping.

5. It is indisputable that the implantable cardiovert-er defibrillator (ICD) has been lifesaving in pa-tients with ARVC/D who have malignant ventric-ular arrhythmias including hemodynamically un-stable ventricular tachycardia [18, 19]. The efficacyof the ICD in this setting is astonishing and recallsthe miracle of the resuscitation of Lazarus, friendof Jesus Christ, from the tomb, painted by Giottoin the Scrovegni Chapel in Padua, where Jesus said“veni foras, Lazare” (John’s Gospel chapter 11, line43-44) (Fig. 1). Whether the ICD should be em-ployed as primary as well as for secondary pre-vention is still controversial.

6. Primary prevention of sudden death in the youngand in athletes from ARVC/D may be possible bylifestyle changes, particularly avoiding participa-tion in vigorous and certainly in competitivesports. Preparticipation screening for those whoengage in competitive sports has been shown tobe highly effective for identification of the indi-

viduals at risk, including those with ARVC/D. InItaly, sudden death of young athletes declined fivefold after the implementation of preparticipationscreening primarily due to identification of car-diomyopathies [20]. The recognition of ARVC/Das a disease entity, as well as the utilization of strictdiagnostic criteria [21], accounts for this impor-tant achievement.

7. Recent developments from in vitro and in vivoanalyses of mutated proteins in transgenic miceare providing mechanistic explanations, with tar-geted therapies on the horizon for affected pa-tients [22-25].These studies suggest that sudden cardiac death in

patients with ARVC/D may be prevented by differentapproaches (Fig. 2):

2 Gaetano Thiene, Andrea Nava, Frank I. Marcus

CardiacARREST

SUBSTRATE TRIGGER

ArrhythmicMECHANISM

Curativetherapy

Sportsdisqualification,

lifestyle

Implant ofdefibrillator

Drug therapy,ablation

Fig. 2 • Diagram illustrating the various levels of interven-tions for sudden death prevention in ARVC/D

Fig. 1 • The resuscitation ofLazarus, painted by Giottoin the Scrovegni Chapel inPadua (C. 1304), is com-pared to the rescue fromcardiac arrest by ICD; ecgtracing,courtesy of Dr.Moss

Introduction: Arrhythmogenic Right Ventricular Cardiomyopathy/Dysplasia Clarified 3

1. Avoiding the trigger, such as strenuous exercise inpatients who are identified as having the diseaseby clinical or genetic screening;

2. Preventing life-threatening arrhythmias usingdrug therapy or ablation;

3. ICD implantation, an extremely effective therapyto treat life-threatening ventricular arrhythmiasthat can result in cardiac arrest.The selection of appropriate therapy for the indi-

vidual patient awaits further investigation.All the above-mentioned therapeutic and preven-

tive measures are palliative, not curative. The defini-tive cure of the disease is still elusive. Cardiac trans-plantation is employed to treat end-stage cardiac fail-ure or for refractory electrical instability, but thistherapy is not without problems, particularly theneed to prevent acute rejection as well as allograft vas-culopathy. Prevention of myocyte apoptotic death, in-flammation, and fibrofatty replacement, the basicmechanisms of myocardial injury and repair, will re-quire understanding the pathogenetic mechanisms ofARVC/D. At present, replacement of the defectivegenes (gene therapy) is theoretically possible only bydisease identification at the early embryonic stage, withpreimplantation genetic diagnosis, an issue that raisesmajor ethical questions [26].

Thus, although the genetic basis of ARVC/D hasbeen largely clarified, there is much work to be doneto better understand the cell biology of the disease,to know how to slow disease progression, and ulti-mately to prevent disease transmission.

Finally, some historical notes. It was in 1961 thatProfessor Sergio Dalla Volta from the University ofPadua reported cases with “auricularization of theright ventricular pressure” with an amazing fibrofat-ty, nonischemic pathology of the right ventricle [27].One of those patients survived until 1995, and un-derwent transplantation due to end-stage cardiacfailure. Interestingly, the heart specimen showed se-vere right ventricular enlargement with a nearly nor-mal left ventricle (Fig. 3).

Attention was focused on the disease following theclinical description of ARVC/D by Marcus et al. in1982 [28]. In 1988 Nava et al. elucidated the patternof transmission in family members [29], and Thieneet al. discovered the disease as a previously unrecog-nized and important cause of sudden death in theyoung [30]. However, the first description of the dis-ease can be traced to the book De Motu Cordis etAneurismatibus by Giovanni Maria Lancisi, publishedin 1736. (Dr. Arnold Katz, personal communication)(Fig. 4). In the 5th chapter of this book, paragraph 47,Lancisi reported a family with disease recurrence infour generations. Signs and symptoms were palpita-tions, heart failure, dilatation and aneurysms of theright ventricle, and sudden death, all features consis-tent with the current diagnostic criteria of the disease.Thus, we know that the disease is not new, only new-ly investigated. Nevertheless, tremendous strides havebeen made in recognition and understanding of thedisease which are summarized in this monograph.

Fig. 4 • The first description of ARVC/D was reported in thisbook of Giovanni Maria Lancisi, Professor of Anatomy inRome, 1736

Fig. 3 • The heart specimen with ARVC/D of the original pa-tient reported by Professor Dalla Volta in 1961. The patientunderwent cardiac transplantation 35 years later because ofright ventricular failure: note the massive dilatation of theright ventricle and the small, normal left ventricle

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11. Maron BJ, Towbin JA, Thiene G et al (2006) Contem-porary definitions and classification of the cardiomy-opathies: An American Heart Association ScientificStatement from the Council on Clinical Cardiology,Heart Failure and Transplantation Committee; Quali-ty of Care and Outcomes Research and Functional Ge-nomics and Translational Biology InterdisciplinaryWorking Groups; and Council on Epidemiology andPrevention. Circulation 113:1807-1816

12. Corrado D, Thiene G (2006) Arrhythmogenic rightventricular cardiomyopathy/dysplasia: Clinical impact ofmolecular genetic studies. Circulation 113:1634-1637

13. Basso C, Czarnowska E, Della Barbera M et al (2006)Ultrastructural evidence of intercalated disc remodel-ling in arrhythmogenic right ventricular cardiomy-opathy: An electron microscopy investigation on en-domyocardial biopsies. Eur Heart J 27:1847-1854

14. Bauce B, Basso C, Rampazzo A et al (2005) Clinicalprofile of four families with arrhythmogenic right ven-tricular cardiomyopathy caused by dominant desmo-plakin mutations. Eur Heart J 26:1666-1675

15. Norman M, Simpson M, Mogensen J et al (2005) Nov-el mutation in desmoplakin causes arrhythmogenic leftventricular cardiomyopathy. Circulation 112:636-642

16. Sen-Chowdry S, Prasad SK, Syrris P et al (2006) Car-diovascular magnetic resonance in arrhythmogenicright ventricular cardiomyopathy revisited: Compari-son with task force criteria and genotype. J Am CollCardiol 48:2132-2140

17. Corrado D, Basso C, Leoni L et al (2005) Three-dimensional electroanatomic voltage mapping increasesaccuracy of diagnosing arrhythmogenic right ventricu-lar cardiomyopathy/dysplasia. Circulation 111:3042-3050

18. Corrado D, Leoni L, Link MS et al (2003) Implantablecardioverter-defibrillator therapy for prevention ofsudden death in patients with arrhythmogenic rightventricular cardiomyopathy/dysplasia. Circulation108:3084-3091

19. Wichter T, Paul M, Wollmann C et al (2004) Im-plantable cardioverter/defibrillator therapy in ar-rhythmogenic right ventricular cardiomyopathy. Sin-gle-center experience of long-term follow-up and com-plications in 60 patients. Circulation 109:1503-1508

20. Corrado D, Basso C, Pavei A et al (2006) Trends in sud-den cardiovascular death in young competitive athletesafter implementation of a preparticipation screeningprogram. JAMA 296:1593-1601

21. McKenna WJ, Thiene G, Nava A et al (1994) Diagno-sis of arrhythmogenic right ventricular dysplasia/car-diomyopathy. Task Force of the Working Group My-ocardial and Pericardial Disease of the European Soci-ety of Cardiology and of the Scientific Council onCardiomyopathies of the International Society andFederation of Cardiology. Br Heart J 71:215-218

22. Garcia-Gras E, Lombardi R, Giocondo MJ et al (2006)Suppression of canonical Wnt/beta-catenin signalingby nuclear plakoglobin recapitulates phenotype of ar-rhythmogenic right ventricular cardiomyopathy. J ClinInvest 116:1825-1828

23. Yang Z, Bowles NE, Scherer SE et al (2006) Desmoso-mal dysfunction due to mutations in desmoplakincauses arrhythmogenic right ventricular dysplasia/car-diomyopathy. Circ Res 99:646-655

24. Kirchhof P, Fabritz L, Zwiener M et al (2006) Age andtraining dependent development of arrhythmogenicright ventricular cardiomyopathy in heterozygousplakoglobin deficient mice. Circulation 114:1799-1806

4 Gaetano Thiene, Andrea Nava, Frank I. Marcus

Introduction: Arrhythmogenic Right Ventricular Cardiomyopathy/Dysplasia Clarified 5

25. Marcus F, Towbin JA (2006) The mystery of arrhyth-mogenic right ventricular dysplasia/cardiomyopathy.From observation to mechanistic explanation. Circu-lation 114:1794-1795

26. Sermon K, Van Steirteghem A, Liebaers I (2004) Preim-plantation genetic diagnosis. Lancet 363:1633-1641

27. Dalla Volta S, Battaglia G, Zerbini E (1961) “Auricu-larization” of right ventricular pressure curve. AmHeart J 61:25-33

28. Marcus FI, Fontaine GH, Guiraudon G et al (1982)Right ventricular dysplasia: A report of 24 adult cases.Circulation 65:384-398

29. Nava A, Thiene G, Canciani B et al (1988) Familial oc-currence of right ventricular dysplasia: A study involv-ing nine families. J Am Coll Cardiol 12:1222-1228

30. Thiene G, Nava A, Corrado D et al (1988) Right ven-tricular cardiomyopathy and sudden death in youngpeople. N Engl J Med 318:129-133

Introduction

Arrhythmogenic right ventricular cardiomyopathy/dysplasia (ARVC/D) is a progressive cardiomyopathywith different clinical-pathological patterns: (a)“silent” cardiomyopathic abnormalities localized inthe right ventricle in asymptomatic victims of sud-den death; (b) “overt” disease characterized by seg-mental or global right ventricular structural changes,often associated with histological evidence of left ven-tricular involvement and underlying symptomaticventricular arrhythmias; and (c) “end-stage” biven-tricular cardiomyopathy mimicking dilated car-diomyopathy, leading to progressive heart failure andeventually requiring heart transplantation [1]. Ascoring system to establish the diagnosis of ARVC/Dhas been developed on the basis of the presence ofmajor and minor criteria encompassing structural,histological, electrocardiographic, arrhythmic, andgenetic features of the disease [2].

The clinical manifestations of the disease mostlyoccur between the second and fourth decade of life;they include electrocardiographic depolarization/re-polarization changes, arrhythmias of right ventricu-lar origin, and structural and functional abnormali-ties of the right ventricle. In ARVC/D, the myocardi-um of right ventricular free wall is partially or almostentirely replaced by fibro-fatty tissue [3-5], and in-volves the epicardium, midmyocardium, and usual-ly spares the subendocardium. The anterior rightventricular outflow tract, the apex, and the inferior-posterior wall are primarily involved [6]. Ventriculartachycardias are thought to be due to re-entrantmechanism, due to slow conduction within the my-ocardiocytes embedded in fibrous tissue and fat.

Familial occurrence of ARVC/D is rather com-mon. Evidence has been found for autosomal dom-inant inheritance with variable penetrance in about50% of cases [7].

ARVC/D has been reported in different humanpopulations [8-10], although it is not known if the dis-ease is equally prevalent in different geographical areas.

Ten years ago, we estimated that prevalence rateof ARVC/D in the Veneto region (northeast Italy) isabout 6:10000 [11]. This figure is probably low, be-cause many cases escape diagnosis. In Italy, 12.5%-25% of sudden deaths in athletes under the age of 35are due to undiagnosed ARVC/D [12].

Two international registries have been established;one in North America and one in Europe, to deter-mine the clinical, pathological, and genetic featuresof ARVC/D, to validate diagnostic criteria, and to de-fine strategies for disease management and suddendeath prevention [13-15].

Since identification of the first ARVC/D locus in1994 by Rampazzo et al. [11], ten loci have been de-tected [11, 16-24], but only five disease genes havebeen identified [22-26] (Table 1.1).

Disease Genes

The first identified ARVC/D gene in a dominantform was ryanodine receptor-2, involved in ARVD2[25]. In ARVD2, there is fibro-fatty substitution ofthe myocardial tissue, though much less pronouncedthan in the typical ARVC/D. The distinctive featureof this form is the presence of polymorphic, effort-induced arrhythmias. RYR2 is one of the largest hu-man genes (105 exons), encoding a 565Kda proteinlocated in the membrane of smooth sarcoplasmicreticulum. The homo-tetrameric structure known ascardiac ryanodine receptor plays a pivotal role in in-tracellular calcium homeostasis and excitation-con-traction coupling in cardiomyocytes [27, 28]. AllRYR2 mutations detected in ARVD2 patients weremissense resulting in substitutions involving aminoacids highly conserved through evolution in criticaldomains of the protein [25, 29].

Mutations in the human RYR2 gene have alsobeen associated with catecholaminergic polymor-phic ventricular tachycardia (CPVT; OMIM 604772)[30, 31] and familial polymorphic ventricular tachy-cardia (FPVT); OMIM 604772) [32, 33]. Putativepathogenic mutations in RYR2 have been reported in

ADVANCES IN GENETICS: DOMINANTFORMSAlessandra Rampazzo, Gian Antonio Danieli

CHAPTER

1

20 out 240 patients referred for long-QT syndromegenetic testing [34].

All RYR2 mutations described to date cluster inthree specific domains: the N-terminal amino-acidresidues 176-433, the centrally located residues 2246-2504, and the C-terminal residues 3778-4959. De-tection of RYR2 mutations in both ARVD2 andCPVT patients raises the question of the possible ex-istence of a single genetic defect, different pheno-types of which might be simply due to variable ex-pression and incomplete penetrance. Both ARVD2and CPVT-RyR2 missense mutations would alter theability of the calcium channel to remain closed. In-tense adrenergic stimulation due to emotional orphysical stress can lead to calcium overload, thustriggering severe arrhythmias. The functional role ofmutations R176Q, L433P, N2386I, and T2504M,previously detected in ARVD2 patients [25], was re-cently investigated [35]. RyR2 mutants N2386I andR176Q/T2504M exhibited enhanced sensitivity tocaffeine activation and increased Ca2+ release, inagreement with the current hypothesis that defectiveRyR2 causes Ca2+ leak. In contrast, RyR2 L433P mu-tation showed reduced response to caffeine activa-tion. This mutation might be interpreted as a “loss-of-function.” Therefore, RyR2 mutations might beeither “gain-of-function” or “loss-of-function,” thussuggesting heterogeneity in functional consequencesof RyR2 mutations. Even with this additional infor-mation, the question of whether ARVD2 and CPVTare different diseases due to different mutations ofthe RYR2 gene still remains unsettled.

The first disease gene linked to autosomal dominantARVC/D showing typical right ventricular phenotypewas Desmoplakin (DSP) [22]. In 2002, genome scan ina family with ARVC/D indicated a linkage with a regionof chromosome 6 short arm including DSP gene. DNAsequencing of all DSP exons in the affected persons ofthis family revealed a missense mutation in exon 7(C1176G; AGC→AGG) (Fig. 1.1). The involved aminoacid (Ser299Arg) is at the center of a coiled, charged re-gion, separating the two short helices of DSP subdo-main Z. The amino acid substitution suppresses a pu-tative phosphorylation site, which, on the other hand,is fully conserved in related proteins belonging to thesame family. This mutation is thought to disrupt a pro-tein kinase C phosphorylation site which is involvedin plakoglobin binding and in clustering of desmoso-mal cadherin-plakoglobin complexes.

Desmoplakin, together with plakoglobin, anchorsto desmosomal cadherins, forming an ordered arrayof nontransmembrane proteins, which bind to keratinintermediate filaments (Fig. 1.2) [36]. The primarystructure of desmoplakin contains three functionaldomains: the N-terminal, which binds to the desmo-some via connection with plakoglobin andplakophilin; the rod segment, which is predicted toform a dimeric coil; and the C-terminal domain,which binds intermediate filaments [37]. Alternativesplicing of the protein produces two isoforms, desmo-plakin I and desmoplakin II. The cDNAs encodingthese two highly related proteins differ in a 1.8 Kbasesequence that is missing in DSPII, most likely due todifferential splicing of a longer transcript [38].

8 Alessandra Rampazzo, Gian Antonio Danieli

Locus Chromosome Gene Function Mutations References

ARVD1 14q24.3 TGFb3 Cytokine stimulating Regulatory 11, 26fibrosis and modulating mutations in 5’cell adhesion and 3’ UTRs

ARVD2 1q42-q43 RYR2 Calcium homeostasis Missense mutations 16, 25

ARVD3 14q12-q22 unknown – – 17

ARVD4 2q32.1-q32.2 unknown – – 18

ARVD5 3p23 unknown – – 19

ARVD6 10p12-p14 unknown – – 20

ARVD7 10q22.3 unknown – – 21

ARVD8 6p24 DSP Cell-cell adhesion Missense, nonsense 22and splice site mutations

ARVD9 12p11.2 PKP2 Cell-cell adhesion Missense, nonsense, 23insertion/deletionand splice site mutations

ARVD10 18q12.1 DSG2 Cell-cell adhesion Missense, nonsense,insertion/deletion and splice site mutations 24

Table 1.1 • Known ARVC/D loci and disease-genes.

CHAPTER 1 • Advances In Genetics: Dominant Forms 9

Mutations in the desmoplakin gene have beenshown to be responsible for some cases of an auto-somal dominant skin disorder (striate palmoplantarkeratoderma) without cardiac involvement [39-41];a dominant form of ARVC/D without skin disease

[22]; an autosomal recessive condition characterizedby dilated cardiomyopathy, woolly hair, and kerato-derma (so-called Carvajal syndrome) [42], an auto-somal recessive condition characterized by ARVC/D,woolly hair, and keratoderma [43] and a left-sidedARVC/D named arrhythmogenic left ventricular car-diomyopathy (ALVC) [44].

Mutations in DSP gene were detected in differentfamilies: they include twelve missense, two nonsense,and two splice-site mutations. In our experience, DSPmutations may account for a considerable number ofARVC/D cases.

In 2004, Gerull et al. [23] selected plakophilin-2 (PKP2) as candidate gene because a homozygousdeletion caused a lethal cardiac defect in mice [45].PKP2 gene encodes plakophilin-2, an essential pro-tein of the cardiac desmosome. By sequencing all 14exons of the PKP2 gene, including flanking intron-ic splice sequences, the authors identified 25 differ-ent heterozygous mutations (twelve insertion-dele-tion, six nonsense, four missense, and three splicesite mutations) in 32 of 120 unrelated ARVC/Dprobands [23]. Plakophilin-2 is an armadillo-relat-ed protein, located in the outer dense plaque ofdesmosomes. It links desmosomal cadherins todesmoplakin and the intermediate filament system(Fig. 1.2). Plakophilins are also present in the nu-cleus, where they may play a role in transcription-al regulation [46]. Gerull et al. [23] speculated thatlack of plakophilin-2 or incorporation of mutantplakophilin-2 in the cardiac desmosomes mightimpair cell-cell contacts and, as a consequence,

Fig. 1.1 • Family pedigree of the ARVC/D index case carrying the S299R DSP mutation and sequence electropherogramshowing the heterozygous missense mutation

Fig. 1.2 • Schematic representation of relationships betweendesmosomal proteins in myocardiocytes.DSC2:desmocollin-2;DSG2: desmoglein-2; DSP: desmoplakin; PKP2: plakophilin-2;PKP4: plakophilin-4; JUP: plakoglobin; DES: desmin

might disrupt association between adjacent car-diomyocytes.

The frequency of PKP2 mutations amongARVC/D cases ranged from 11% to 43% in differentstudies [47-49]. These differences might be attributedto different geographical origin of cases or simply toselection bias.

Recently, we decided to shift from linkage studiesin ARVC/D families to a candidate gene approach.Thus, we screened different genes encoding desmo-somal proteins. When analyzing DSG2 gene(Desmoglein-2, the only isoform expressed in cardiacmyocytes), we detected nine heterozygous mutationsin eight of 50 unrelated individuals with ARVC/Dwhich proved negative for mutations of DSP, PKP2,and TGFβ3 genes [24]. Among these, five were mis-sense mutations, two were insertion-deletions, onewas a nonsense and one was a splice site mutation;one patient had two different DSG2 mutations (com-pound heterozygote). Endomyocardial biopsy, ob-tained from five patients, showed extensive loss ofmyocytes with fibro-fatty tissue replacement. In threepatients, electron microscopy showed intercalateddisc paleness, decreased desmosome number, and in-tercellular gap widening [24]. Mutations in DSG2gene were also detected in an independent study [50].It is interesting to note that, in this study, there wasone patient with compound-heterozygous mutationsin DSG2 (Fig. 1.3).

In 2005, our group identified the gene involved inARVD1 [26]. The large critical interval for ARVD1 in-cluded 40 known genes; five of them (POMT2,KIAA0759, KIAA1036, C14orf4, and TAIL1) wereunsuccessfully screened for pathogenic ARVC/D mu-tations [51, 52]. Among genes mapped to the ARVD1critical region and expressed in myocardium, trans-forming growth factor-beta3 (TGFβ3) appeared to be

a good candidate, since it encodes a cytokine stimu-lating fibrosis and modulating cell adhesion. Afterprevious analyses failed to detect any mutation in thecoding region of this gene, mutation screening was ex-tended to the promoter and untranslated regions(UTRs). A nucleotide substitution (c.-36G>A) in5′UTR of TGFβ3 gene was detected in all affected sub-jects belonging to a large ARVD1 family. After the in-vestigation was extended to 30 unrelated ARVC/D in-dex patients, an additional mutation (c.1723C>T) wasidentified in the 3’ UTR of one proband. In vitro ex-pression assays of constructs containing the mutationsshowed that mutated UTRs were twofold more activethan wild type [26].

TGFβ3 is a member of the transforming growthfactor superfamily, which includes a diverse range ofproteins regulating many different physiologicalprocesses. TGFβ1, -β2, and -β3 are the prototype ofthe TGFβ superfamily. They inhibit proliferation inmost types of cells and induce apoptosis of epithe-lial cells. Conversely, they stimulate mesenchymalcells to proliferate and produce extracellular matrixand they induce a fibrotic response in various tis-sues in vivo.

Finding TGFβ3 mutations associated with ARVC/Dis very interesting, since it is well established thatTGFβs stimulate mesenchymal cells to proliferate andto produce extracellular matrix components. Sincemutations in UTRs of the TGFβ3 gene, detected inARVC/D, showed enhanced gene expression in vitro,it is likely that they could promote myocardial fibro-sis in vivo. Myocardial fibrosis may disrupt electricaland mechanical behavior of myocardium and extra-cellular matrix abnormalities may predispose to re-entrant ventricular arrhythmias. In agreement withthis hypothesis, endomyocardial biopsy in the twoprobands in which TGFβ3 UTR mutations were de-tected showed extensive replacement-type fibrosis.Moreover, it has been shown that TGFβs modulate ex-pression of genes encoding desmosomal proteins indifferent cell types. cDNA microarray analysis, per-formed on RNA from cardiac fibroblasts incubated inthe presence or in the absence of exogenous TGFβs,revealed increased expression of different genes, in-cluding plakoglobin [53]. Yoshida et al. [54] reportedthat TGFβ1 exposure of cultured airway epithelialcells increases the content of desmoplakins I and II.This suggests that regulation of cell-cell junctionalcomplexes may be an important effect of TGFβs.Therefore, overexpression of TGFβ3, caused by UTRsmutations, might affect cell-to-cell junction stability,thus leading to disease expression similar to that ob-served in ARVC/D due to mutations of genes encod-ing desmosomal proteins.


Fig. 1.3 • Pedigree of the proband carrying two DSG2 mu-tations (988G>A, 1881-2A>G). Hatched symbol representsan individual of unknown disease status.Presence (+) or ab-sence (-) of the DSG2 mutation is indicated


Desmosomes are important cell-cell adhesionjunctions, predominantly found in the epidermisand heart. They couple cytoskeletal elements to plas-ma membrane at cell-cell or cell-substrate adhesions.Whereas adherens junctions are linked with micro-filaments at cell-cell interfaces, desmosomes anchorstress-bearing intermediate filaments at sites ofstrong intercellular adhesion. The resulting scaffoldplays a key role in providing mechanical integrity totissues such as epidermis and heart, which experiencemechanical stress. Desmosomes include proteinsfrom at least three distinct gene families: cadherins,armadillo proteins, and plakins (Fig. 1.2). Desmoso-mal cadherins include desmogleins and desmo-collins; members of both subfamilies are single-passtransmembrane glycoproteins, mediating Ca2+-de-pendent cell-cell adhesion. Armadillo proteins in-clude plakoglobin and plakophilins (PKP1-3). Theplakin family proteins include desmoplakin, plectin,and the cell envelope proteins envoplakin andperiplakin. Desmoplakin (involved in ARVD8),plakophilin-2 (involved in ARVD9), desmoglein-2(involved in ARVD10), and plakoglobin (involved inNaxos syndrome, the autosomal recessive form ofARVC/D) are desmosomal proteins. Based on presentevidence we may conclude that different defects inproteins of desmosomal complex lead to ARVC/D.Therefore, additional components of the desmoso-mal complex may be targets for pathogenic muta-tions leading to ARVC/D.

Molecular Pathogenesis

The reported involvement of different desmosomalproteins in ARVC/D and the discovery that someRYR2 mutations may produce ARVD2 leads us topropose a comprehensive hypothesis on the molec-ular pathogenesis of ARVC/D [22]. According to thishypothesis, the predilitation of involvement of theright ventricle in ARVC/D might be due to greater di-latation and thinning of its wall, in comparison withthe left ventricular free wall. Possibly, defective pro-teins in cardiac desmosomes might impair cell-to-cellcontacts and, hence, might affect the response of ven-tricular myocardium to mechanical stretch. This al-teration would occur preferably in myocardial areassubjected to high strain, like the right ventricular out-flow tract, the apex, and subtricuspid areas.

According to present knowledge, mechanicalforces applied to adherens junctions activate stretch-sensitive calcium channels via cadherins’ mechanicalintracellular signaling [55]. Data on stretch-activat-

ed channels in ventricular cardiomyocytes point tothe relevance of these channels in transduction ofmechanical forces into a cellular electrochemical sig-nal, via increase of intracellular calcium concentra-tion [56-58].

Volume overload of the right ventricle in a patientwith genetically defective intercellular junctions (asin case of mutant plakoglobin, desmoplakin,plakophilin, desmoglein, or TGFβ3) would produceunusual stretching resulting in excessive calciumload. Stretching of cardiomyocytes is known to mod-ulate the elementary calcium release process fromryanodine receptor release channels [59]. Therefore,a genetically impaired response to mechanical stressmight adversely affect intracellular calcium concen-tration and excitation-contraction coupling, thusproducing arrhythmias. On the other hand, volumeoverload of the right ventricle in carriers of RYR2mutations would cause calcium overload, because ofdefective Ca++ homeostasis. The existence of a dom-inant form of ARVC/D due to RyR2 mutations sup-ports the hypothesis of a key pathogenic role of in-tracellular calcium overload in the molecular patho-genesis of the disease.

Mutation Screening

We performed mutation screening in 90 unrelatedprobands fulfilling the International ARVC/D TaskForce criteria; the screening by DHPLC and subse-quent DNA sequencing involved coding sequences ofknown ARVC/D genes. Plakophilin-2 was involved in21% of cases, desmoplakin-2 in 20%, desmoglein-2in 11%, and TGFβ3 in 2% (unpublished results). In46% of cases no mutation was detected. This is notsurprising, since in 50% of reported ARVC/D loci(ARVD3, ARVD4, ARVD5, ARVD6, ARVD7) the in-volved gene has not been identified.

In our series of patients screened for mutations,eight compound heterozygotes were detected, sug-gesting that this condition may be more frequentthan expected among ARVC/D patients. It is difficultto establish whether all of these cases are compoundheterozygotes for pathogenic mutations, since it is al-most impossible to discriminate between a rare vari-ant with pathogenic effect and a rare DNA poly-morphism.

Present knowledge on the molecular genetics of thedominant forms of ARVC/D may permit detection ofasymptomatic carriers in families with ARVC/D. How-ever, it must be noted that genotype-phenotype cor-relations may be established only for clearly patho-genic mutations (i.e., nonsense, frameshifts, splice-site

mutations with evidence of modified RNA length, etc.)and once all genes reportedly involved in ARVC/Dwould have been screened.

Mutation screening is time- and effort-consum-ing. Routine methods (direct sequencing of codingsegments or DHPLC followed by DNA sequencing)reach about 98% detection rate due to undetectablemutations in intronic sequences or in regulatory el-ements, or unexpected large deletions. Moreover, thepresence of compound heterozygotes carrying onemutation in a known ARVD gene and one mutationin a gene still unknown might produce misleading re-sults. Therefore, genetic assessment of asymptomaticrelatives of ARVC/D patients still poses several tech-nical, clinical, and ethical problems. However, iden-tification of additional genes involved in dominantforms of ARVC/D and collection of data regardingpathogenic mutations in known genes will provideinformation to establish safe protocols for genetic in-vestigation in families with ARVC/D, genetic coun-seling, and risk assessment.

Acknowledgements

The financial support of European Commission (Pro-ject QLG1-CT-2000-01091, UE) and of NIH, USA(Grant U04HL65652) is gratefully acknowledged.

P.S. Following submission of this manuscript, muta-tions in desmocollin-2 gene, encoding a desmosomalcadherin, have been reported to be associated withARVC/D [60-62].

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arrhythmogenic rv cardiomyopathy/dysplasia...preface this book covers all the recent research...

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