Pathological Character Of Arrhythmogenic Right Ventricular Cardiomyopathy(ARVC) And The Correlationship Between ARVC And Desmosomal Proteins

Arrhythmogenic right ventricular cardiomyopathy （ARVC） is a rare heart muscle disease in which the right ventricle is "replaced" by fibrofatty tissue. ARVC is clinically characterized by ventricular tachyarrhythmias originating from the right ventricle, and by syncopal episodes and sudden death, which are often triggered by exercise. Pathologically ARVC is characterized by fibrofatty replacement of the myocardium, accompanied by vacuolated myocytes and interstitial fibrosis. In1995, the World Health Organization revised its classification of cardiomyopathy. ARVC was added to the previous classification. It is a major cause of sudden death in the young and in the athletes, the estimated prevalence of ARVC in the general population ranges from1in1,000to1in5,000, men are more frequently affected than women. There have been several genes identified which are linked to ARVC, most of them encode components of the cardiac desmosome. but the mechanisms are currently obscure.The desmosome, a specialized adhesive junction whose primary function is providing intercellular links in the desmosome-intermediate filament（IF） complex and lending great tensile strength to tissues, is crucial to tissues that experience mechanical stress, such as the skin, myocardium and gastrointestinal mucosa. It consists of five major component proteins encoded by three gene superfamilies:the desmosomal cadherins, desmoglein （DSG） and desmocollin（DSC）;the armadillo family, plakoglobin（JUP） and plakophilin （PKP）; and the plakin family, desmoplakin （DSP）. DSC and DSG are the desmosomal adhesion molecules, DP links the desmosomal plaque to the IF cytoskeleton, and JUP and PKP are adaptor proteins that link between the adhesion molecules and DSP.Since people found the first mutant desmosomal gene（JUP） in recessive form of ARVC（Naxos disease）, a lot of studies have been carried to aim to find out mutant desmosomal genes in ARVC, and so far it has been estimated that approximately50% of unrelated cases of ARVC have mutations in desmosomal genes. However, so many questions remain. First, what about the roughly50%of patients in whom the diagnosis of ARVC was firmly established on the basis of task force criteria but who were found not to have mutations in desmosomal protein genes? What is the cause of their disease? Do they even have the same disease as those with desmosomal mutations? Other candidate genes have been identified as potential causes of ARVC, including those encoding the cardiac ryanodine receptor （RyR2）, transforming growth factor-β_TGF-β, and transmembrane protein-43（TMEM43）, however, it seems unlikely that this would have accounted for a significant number of individuals with documented disease but no identifiable desmosomal mutation. Other mutations in as yet unidentified genes must play a significant role in causing ARVC. There may be nongenetic causes as well. A second important question is that it’s not clear that all or even most of these mutations are sufficient to cause disease. It remains a formal possibility that these patients may have additional mutations in nondesmosomal genes that could be critical in disease pathogenesis. Epigenetic factors may also play important roles. We simply do not know. Finally, cellular model showed that loss of desmosomal expression reduced Cx43at the gap junctions in ARVC that may be responsible for the conduction abnormalities and arrhythmogenicity, is there possibility that desmosomal mutations only cause arrhythmia rather than fibrofatty replacement of the myocardium, in other words, desmosomal mutations may be not specific for ARVC. One purpose of our research is to further study the pathological character of ARVC, especially the ventricular distribution of ARVC. another major object is to study the correlationship between ARVC and desmosomes.Part I The ventricular distribution of Arrhythmogenic Right Ventricular Cardiomyopathy:an autopsy studyBackground-Arrhythmogenic right ventricular cardiomyopathy （ARVC） is a rare cardiomyopathy characterized by fibrofatty replacement primarily of the right ventricular myocardium. It is a major cause of sudden death in the young and in the athletes. There are few autopsy studies of the ventricular distribution of the disease.Methods and Results-Fifty cases of sudden cardiac death with fibrofatty replacement in either ventricle from a single medical examiner’s office were studied. Distribution of disease as determined grossly and microscopically was correlated with activity at time of death, race, and presence of inflammation. Extent of disease was right ventricular in6cases （12%, age25±5years）, biventricular in25（50%, age36±3years）, and left ventricular in19（38%, age37±3years）（p=0.13）. Inflammation was present in44%of biventricular ARVC, vs.74%of left ventricular ARVC, and83%of right ventricular ARVC （p=0.06）.Conclusions-Arrhythmogenic right ventricular cardiomyopathy, when presenting with sudden death, is usually biventricular. There is a trend that right ventricular involvement occurs at an earlier age, and that right ventricular involvement shows more inflammation, suggesting different stages of disease.Part Ⅱ Allograft pathology in patients transplanted for idiopathic dilated cardiomyopathyBackground:There are few morphologic studies of idiopathic dilated cardiomyopathy （CM） treated with transplant.Methods:We prospectively correlated gross, histologic and clinical findings of hearts explanted in a5-year period from patients with a clinical diagnosis of non-ischemic CM。Results:Of64patients with a clinical diagnosis of dilated cardiomyopathy, there were42men （age51±13years） and22women （age42±18years）. The pathologic diagnosis was idiopathic （dilated） cardiomyopathy （DC） in55（86%）, and the left diagnosis were fibrofatty change consistent with arrhythmogenic right ventricular cardiomyopathy （ARVC）（n=6）, amyloidosis （n=2）, and sarcoidosis （n=1）, none of which were suspected clinically. The55hearts with idiopathic DC had a mean heart weight of508grams （range220-980grams）. Pathologic subsets of the DC group included4hearts without enlargement, cavity dilatation, or significant histologic findings （minimal DC）;3hearts with histologic evidence of healed myocarditis; and5hearts with mildly non-compacted left ventricle with hypertrabeculation. Five patients had prior mitral or tricuspid valve replacement/repairs to manage heart failure. There were7post-partum DC,1with a histologic pattern of healed myocarditis, and1alcoholism-associated DC. Familial DC comprised16%（9of55patients）.Conclusion:when the extent of ARVC is left ventricular or biventricular, it’s easily misdiagnosed as Dilated cardiomyopathy. Dilated cardiomyopathy is a heterogeneous group morphologically. Features of non-compaction are not uncommon.Part Ⅲ Quantitative Immunohistochemistry of Desmosomal Proteins （Plakoglobin, Desmoplakin and Plakophilin） in Arrhythmogenic Cardiomyopathy:an Autopsy StudyBackground-Arrhythmogenic right ventricular cardiomyopathy （ARVC） is a genetic disorder related to mutations in desmosomal proteins. There have been few histologic studies quantitating expression of desmosomal proteins in autopsy samples of ARVC.Methods-We studied23hearts （16males,7females） dying suddenly with ARVC. Their age at death was33-16and33-16years, respectively. Control subject tissues were21hearts （17men and4women, ages34-12and32-12years, respectively） from people dying from non-cardiac causes （n=15）, dilated cardiomyopathy （n=3） and coronary artery disease （n=3）. Sections of heart were taken from short axis cuts:28sections from controls, including14from the left ventricle （LV）, and14from the right ventricle （RV） and50sections from ARVC （24LV,26RV）.Areas free of fibrofatty change or scarring were assessed. Immunohistochemical stains against plakoglobin （using two different antibodies）, plakophilin, desmoplakin were applied and area expression analyzed by computerized morphometry. Desmin was stained as a control for fixation and similarly analyzed.Results-The mean area of desmin expression was similar in controls and ARVC （85%vs.86%, p=0.75）. Plakoglobin expression was4.5%±1.4.%in controls, vs.4.3%±1.3%in ARVC （p=0.5）. Plakophilin staining was4.5%±1.4%in controls vs.4.3%±1.3%in ARVC （p=0.5）. Desmoplakin staining was3.2±1.0%in controls vs.3.2±0.9%in ARVC （p=0.6）. There were no significant differences when staining was compared between right and left ventricles （all p>0.1）.Conclusion-there is a small but insignificant decrease in desmosomal protein expression in ARVC vs. controls, suggesting that routine immunohistochemical staining of uninvolved myocardium is not a useful tool for the diagnosis of ARVCPart IV PKP2mutations in sudden death due to arrhythmogenic right ventricular cardiomyopathy （ARVC） and sudden unexpected death with negative autopsyBackground:Plakophilin2（PKP2） is a desmosome-related protein with numerous armadillo repeats, which has been linked to arrhythmogenic right ventricular cardiomyopathy （ARVC）. Fatal arrhythmias resulting in sudden death also occur in the absence of morphologic cardiac abnormalities at autopsy, and have been linked to ion channel mutations in a subset of cases, but so far not to PKP2.Methods:We sequenced all14exons of PKP2in DNA extracted from post-mortem heart tissues of25patients dying with ARVC and25with sudden unexpected death with negative autopsy. The primers were designed using the Primer Express3.0software. Direct sequencing for both sense and antisense strands was performed with a BigDye Terminator DNA sequencing kit on a3130x1Genetic Analyzer. Mutation damage prediction was made using Mutation Taster, Polyphen and SIFT software.Results:In6of25ARVC samples,6PKP2mutations were identified,4of which were likely significant, and3of which were novel （p.N641del, p.L64PfsX22, p.G269R）. In6of25cases of SUDNA,6PKP2mutations were identified,3of which were likely significant, and4of which not previous described （p.P665S, p.Y217TfsX45,p.E540, p.S615T）.Conclusions:PKP2mutations are not specific for ARVC and may result in sudden death with negative autopsy. The link between ARVC and desmosomal mutations may not be causal but related to an association between defective desmosomal proteins and arrhythmias.Conclusions1. Arrhythmogenic right ventricular cardiomyopathy, when presenting with sudden death, is usually biventricular.2. The most common locations of left ventricular involvement are the posterolateral walls in a subepicardial distribution.3. There is a trend that right ventricular involvement occurs at an earlier age, and that right ventricular involvement shows more inflammation, suggesting different stages of disease.4. When the extent of ARVC is left ventricular or biventricular, it’s easily misdiagnosed as Dilated cardiomyopathy.5. There is a small but insignificant decrease in desmosomal protein expression in ARVC vs. controls.6. Routine immunohistochemical staining of uninvolved myocardium is not a useful tool for the diagnosis of ARVC..7. PKP2mutations are not specific for ARVC and may result in sudden death with negative autopsy.8Desmosomal mutations may not in themselves cause fibrofatty change and the syndrome of ARVC. More studies of desmosomal genotype are necessary in populations other than ARVC, in order to definitively establish a link between ARVC and the desmosome.Originalities of this work1. Arrhythmogenic right ventricular cardiomyopathy （ARVC） is a rare cardiomyopathy characterized by fibrofatty replacement primarily of the right ventricular myocardium. Although there are reports of single or a few autopsy cases found that fibrofatty replacement could also be found in the left ventricular myocardium. There is no autopsy study of the ventricular distribution of the disease with so large scale. The result will helpful for medical examiner and autopsy pathologists.2. There are few morphologic studies of idiopathic dilated cardiomyopathy （CM） treated with transplant.3Arrhythmogenic right ventricular cardiomyopathy （ARVC） is a genetic disorder related to mutations in desmosomal proteins. There have been some studies found that the expresson of desmosomal proteins changed in samples of ARVC. However, there have been few histologic studies quantitating expression of desmosomal proteins in autopsy samples of ARVC compared to control group.4Recent evidence indicates that mutations in several desmosomal components play a major role in the pathogenesis of arrhythmogenic right ventricular cardiomyopathy （ARVC）, among the genes in which mutations have been identified associated with ARVC, plakophilin-2accounts for a large proportion. Almost all the studies sequenced the DNA extracted from the blood samples from ARVC patients to detect the desmosomal mutation, few study totally use the heart tissue samples which may reflect mutations more directly. Moreover, there has been no PKP2mutation study in samples of sudden death with negative autopsy.</sub>