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. 2018 May 11;41(5):615–622. doi: 10.1002/clc.22906

Absence of a primary role for TTN missense variants in arrhythmogenic cardiomyopathy: From a clinical and pathological perspective

Kai Chen 1, Jiangping Song 1,✉, Zhen Wang 1, Man Rao 1, Liang Chen 1, Shengshou Hu 1,✉
PMCID: PMC6490078  PMID: 29750433

Abstract

Background

Arrhythmogenic cardiomyopathy (ACM) is an inheritable heart disease characterized by fibro‐fatty replacement of the myocardium. TTN missense variants were previously reported as a pathogenic factor for ACM.

Hypothesis

TTN missense variants are commonly identified in ACM, but have limited effect on the phenotype of ACM.

Methods

We sequenced 15 ACM‐related genes in 35 patients who had a heart transplantation and quantified myocardium, and fibrous and adipose tissue in blocks of the explanted heart. Clinical and pathological characteristics were compared between patients with TTN variants and others. Pedigree analysis was performed in 3 families with TTN variants.

Results

TTN variants were detected in 11 patients (all missense, 9 heterozygous and 2 oligogenic form). The TTN truncating variant was absent in the cohort. Patients with TTN variants had late onset age of the disease (31 ±13 years vs 17 ±3 years, P = 0.049) and age of heart transplantation (41 ±14 years vs 24 ±9 years, P = 0.027), larger left ventricle end‐diastolic diameter (62 ±10 mm vs 45 ±10 mm, P = 0.019), smaller right ventricular outflow tract (34 ±14 mm vs 50 ±15 mm, P = 0.046), more myocardium (40.8% ±29.4% vs 13.8% ±11.0%, P = 0.017), and less adipose tissue (43.0% ±30.9% vs 66.9% ±18.5%, P = 0.036) in right ventricle than those with desmosomal variants. There was few difference between patients with TTN variants and those without variants. Pedigrees showed none of the family members with TTN missense variants had a disease phenotype, indicating a very low penetrance.

Conclusions

TTN missense variants was commonly identified in ACM patients in this cohort, but hardly played a primary role in ACM as causative variants.

Keywords: Arrhythmogenic Cardiomyopathy, Pathology, TTN Missense Variants

1. INTRODUCTION

Arrhythmogenic cardiomyopathy (ACM) is an inheritable heart disease characterized by fibro‐fatty replacement of the myocardium, which predisposes patients to ventricular arrhythmia and progressive ventricular dysfunction that may lead to heart transplantation.1, 2 More than 60% of ACM patients carry genetic variants that are inherited in an autosomal dominant trait in most cases and in an autosomal recessive trait in some cases.3, 4, 5 Desmosomal variants are most frequently detected, including PKP2,6 JUP,7 DSP,8 DSG2,9 and DSC2,10 and other genes have also been reported as associated with ACM, such as TGFβ‐3,11 RYR2,12 TMEM43,13 DES,14 TTN,15 LMNA,16 PLN,17 CTNNA3,18 CDH2 19 , and FLNC. 20 However, the interpretation of a rare variant should be cautious, for some of these variants were found in healthy people, especially missense variants.21

As the largest protein in heart tissue, titin encoded by TTN plays an important role for maintaining cardiac function. Truncating variants in TTN were extensively proven to be pathogenic in dilated cardiomyopathy (DCM).22 Missense variants in TTN, however, were commonly detected in DCM with uncertain pathogenicity.23 As for ACM, TTN variants were previously reported as pathogenic variants; however, cosegregation was detected in only 1 family.15 Brun et al24 found TTN variant carriers had a distinct clinical phenotype. Still, the pathogenicity of TTN missense variants is controversial.

To investigate TTN variants in ACM, we sequenced 15 ACM‐related genes in 35 ACM patients who had a heart transplantation. The clinical and pathological characteristics were compared between patients with TTN variants and others, and pedigrees of 3 families with TTN variants were analyzed.

2. METHODS

2.1. Study population

Thirty‐five index patients with ACM who had heart transplantation were included in our study between January 2004 and December 2014 from National Center of Cardiovascular Disease, Fuwai Hospital. The diagnosis of ACM was made retrospectively on the basis of the 2010 revised task force criteria for ACM.25 Pathological examination was performed on all of the explanted heart of the patients. Clinical data were carefully collected, including medical history, physical examination, family history, electrocardiogram, echocardiogram, Holter monitoring, and cardiac magnetic resonance whenever available. A major arrhythmic cardiovascular event (MACE) was defined as ventricular fibrillation, sustained ventricular tachycardia, or arrhythmic syncope. The composite endpoint included both heart transplantation and MACE.

All subjects or their guardians when younger than 18 years provided informed consent per individual institutional protocols. The study protocol was approved by the institutional review board at Fuwai Hospital, and all methods were performed in accordance with the Declaration of Helsinki.

2.2. Captured sequencing and data analysis

Genomic DNA was extracted from frozen heart tissue by liquid nitrogen using DNeasy Tissue kit (QIAGEN, Hilden, Germany). Genomic libraries with 300 to 500 base‐pair insertions were constructed by custom purified Tn5 transposase.26 Illumina HiSeq X Ten was used for sequencing. The ACM gene panel was designed to target all coding exons and their splice sites of 15 known genes associated with ACM, including PKP2, DSG2, DSC2, DSP, JUP, TTN, PLN, TMEM43, LMNA, DES, RYR2, TGFB3, CTNNA3, CDH2, and FLNC.

Sequencing reads were aligned to the human reference genome build hg19 with the Burrows‐Wheeler Aligner,27 and the downstream processing was performed with the Genome Analysis Toolkit.28 The variants were annotated with ANNOVAR (Annotate Variation).29 The following steps were applied to select candidate pathogenic variants. First, allele frequency of the candidate variants should be lower than 0.5% among the public databases, including the 1000 genome project, Exome Aggregation Consortium, CG46 databases, gnomAD_genome and gnomAD_exome. Second, for stop‐gain or stop‐loss variants, splice site variants, frameshift insertions or deletions, and non–frameshift insertions or deletions, all of them were defined as “candidate.” Finally, for missense variants, the pathogenicity should be annotated as “deleterious” or “highly pathogenic” by at least 3 software programs of SIFT, Polyphen2‐HDIV, Polyphen2‐HVAR, MutationTaster, or MutationAssessor. Meanwhile, the Genomic Evolutionary Rate Profiling score should be more than 4. All of the selected variants were confirmed by Sanger sequencing.

2.3. Pathological examination and quantification

Gross and microscopic examinations of explanted hearts were performed by 2 pathologists blinded to the clinical and genetic data. A total of 6 specimens were obtained from each explanted heart. Blocks of left ventricle (LV), including the anterior free wall of the left ventricle (ALV), lateral free wall of LV (LLV), and posterior free wall of the LV (PLV); blocks of the right ventricle (RV) including anterior free wall of RV (ARV) (right ventricular outflow tract), posterior free wall of the RV (PRV) (sub‐tricuspid area), and interventricular septum (IVS) were removed systematically for histological study. Samples were fixed in 10% formalin and were processed for histological examination. Tissue samples were stained with Masson's trichrome, and the slides were scanned into a digital image.

Subepicardial adipose tissue, including vessels and pericardium, were removed by Adobe Photoshop CS5 (Adobe Systems, San Jose, CA) before quantitative analysis. Endocardium was removed from fibrosis analysis when endocardial thickening was detected. The myocardium, fibrosis, and adipose tissue were evaluated using Image‐Pro Plus (version 4.0; Media Cybernetics, Rockville, MD) as previously reported.30, 31 The proportion of myocardium, fibrosis, and adipose tissue in the LV was the average of the values in the anterior, lateral, and posterior free wall of the LV, and the proportion in the RV was the average of values in the anterior and posterior free wall of the RV.

2.4. Statistical analysis

Statistics of clinical and pathological variables were expressed as mean ±standard deviation for continuous variables, and counts and frequencies (%) for categorical variables. Comparisons between groups were made by the analysis of variance (ANOVA) test on continuous variables using the Brown‐Forsythe statistic; when the assumption of equal variances did not hold, post hoc was also applied. Categorical variables of the groups were compared with a χ2 or Fisher exact analysis when necessary. The cumulative freedom from birth to clinical outcome (MACE, heart transplantation, and composite endpoint) was determined by the Kaplan–Meier method, and differences in survival between groups were evaluated with a log‐rank test. Reported probability values were 2 sided, and values <0.05 were considered statistically significant. SPSS statistical software (version 23, IBM Corp., Armonk, NY) was used for the analyses.

3. RESULTS

3.1. Clinical data

Our study cohort (all Han Chinese) showed a male dominance with 23 males (66%) and 12 females (34%). All of the patients fulfilled the 2010 revised Task Force Criteria (see Supporting Table S1 in the online version of this article). The clinical characteristics are shown in Table 1. The average age of the disease onset and heart transplantation was 30 ±12 years and 37 ±13 years, respectively. Eleven of the patients (31%) had a family history of cardiomyopathy or suspected cardiomyopathy.

Table 1.

Summary of clinical features of the subjects

Characteristics Total, N = 35 TTN, N = 9 DS, N = 6 NV, N = 16 P Value
Gender, male, n (%) 23 (66) 7 (78) 4 (67) 11 (69) 0.863
Age of disease onset, y 30 ±12 31 ±13 17 ±3 32 ±11 0.021a , b
Age of heart transplantation, y 37 ±13 41 ±14 24 ±9 38 ±10 0.022a , b
Duration from onset to heart transplantation, mo 86 ±79 117 ±85 84 ±82 71 ±71 0.363
Presyncope, n (%) 4 (11) 1 (11) 1 (17) 1 (6) 0.751
Syncope, n (%) 9 (26) 2 (22) 2 (33) 3 (19) 0.767
Family history, n (%) 11 (31) 3 (33) 3 (50) 4 (25) 0.534
MACE, n (%) 12 (34) 4 (44) 2 (33) 4 (25) 0.606
LA, mm 40 ± 11 44 ± 6 25 ± 6 44 ±11 <0.001a , b
LVEDD, mm 59 ±12 62 ±10 45 ±10 62 ±11 0.006a , b
LVEF, % 30 ±13 32 ±16 42 ±17 26 ±8 0.051
RVOT, mm 35 ± 13 34 ±14 50 ±15 31 ±8 0.005a , b
Therapy, n (%)
Amiodarone 8 (23) 2 (22) 1 (17) 3 (19) 0.961
β‐blocker 15 (43) 2 (22) 3 (50) 8 (50) 0.363
Sotalol 2 (6) 1 (11) 1 (17) 0 (0) 0.292
Radiofrequency ablation 1 (3) 0 (0) 0 (0) 1 (6) 0.616
ICD 3 (9) 0 (0) 2 (33) 1 (6) 0.081
CRT or CRT‐D 4 (11) 1 (11) 0 (0) 3 (19) 0.496
Pacemaker 1 (3) 0 (0) 0 (0) 1 (6) 0.616
Electrical conversion or CPR 5 (14) 1 (11) 2 (33) 1 (6) 0.236
Diagnosis, n(%)
RV dysfunction and structural alterations, major 31 (89) 7 (78) 6 (100) 14 (88) 0.452
RV dysfunction and structural alterations, minor 4 (11) 2 (22) 0 (0) 2 (13) 0.452
TWI in V1–3 without CRBBB, major 6 (17) 0 (0) 2 (33) 3 (19) 0.210
TWI in V1–2 without CRBBB or in V4, V5 or V6, minor 8 (23) 2 (22) 1 (17) 4 (25) 0.917
TWI in V1–4 with CRBBB, minor 7 (20) 3 (33) 1 (17) 3 (19) 0.654
Epsilon wave in V1–3, major/depolarization, major 16 (46) 2 (22) 4 (67) 9 (56) 0.160
VT of LBBB with superior axis, major 15 (43) 5 (56) 2 (33) 7 (44) 0.689
>500 VES/24 hours, minor 14 (40) 5 (56) 3 (50) 4 (25) 0.264
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Abbreviations: CPR, cardiopulmonary resuscitation; CRBBB, complete right bundle branch block; CRT, cardiac resynchronization therapy; CRT‐D, cardiac resynchronization therapy with defibrillator; DS, patients with desmosomal variants; HT, heart transplantation; ICD, implantable cardioverter defibrillator; LA, left atrial; LBBB, left bundle branch block; LVEDD, left ventricle end diastolic dimension; LVEF, left ventricle ejection fraction; MACE, major arrhythmia cardiovascular event; NV, patients with no variants; RV, right ventricle; RVOT, right ventricle outflow tract; TTN, patients with TTN variants; TWI, T‐wave inversion; VES, ventricular extrasystole; VT, ventricular tachycardia.

a

TTN vs DS P < 0.05.

b

DS vs NT‐ND P < 0.05.

3.2. Genetic analysis revealed high variants burden in TTN

We detected 24 rare variants in 19 of the 35 patients (54%): 11 in TTN, 5 in DSG2, 2 in PKP2, DSC2, and LMNA, respectively, and 1 in DSP and PLN (Table 2). All of the rare variants in TTN were missense variants, and truncating variants in TTN were not detected in our cohort. Of the 11 TTN missense variants, 8 are located in high percentage spliced in (PSI) exons, 2 in median PSI exons, and 1 in a low PSI exon. Monogenic variants were detected in 17 patients, among whom 9 (26%) patients had a TTN missense variant, and 6 (17%) patients had desmosomal variants. Interestingly, all patients with desmosomal variants had 2 mutated sites: 3 DSG2 compound variants, 1 DSG2 homozygous variants, and 1 DSC2 and 1 PKP2 compound variants. Oligogenic variants were detected in 2 patients: 1 had DSP and TTN variants, and 1 had DSG2, LMNA, and TTN variants. Among the patients with TTN missense variants, all harbored TTN heterozygous variants.

Table 2.

Summary of variants identified in the arrhythmogenic cardiomyopathy study population

Gene Transcript DNA Change Protein Change Type Patient No. MAF SIFT Polyphen2‐HDIV Polyphen2‐HVAR MutationTaster MutationAssessor PSI ACMG
TTN NM_133378 c.5419C > T p.P1807S Missense HT‐19 0.0001 D D P D N High PSI, Z‐disc border VUS
c.22412G > A p.C7471Y Missense HT‐2 0.001 D D D D NA Median PSI, I‐band VUS
c.24956 T > G p.L8319 W Missense HT‐14a 0.0003 D D D D NA Median PSI, I‐band VUS
c.33516C > G p.D11172E Missense HT‐34 NA D D D D N Low PSI, I‐band VUS
c.44968C > T p.P14990S Missense HT‐3 0.0003 T D D D M High PSI, A‐band VUS
c.61804G > A p.E20602K Missense HT‐32 3.238*10−5 D D P D L High PSI, A‐band VUS
c.67264G > A p.G22422S Missense HT‐29 NA D D D D H High PSI, A‐band VUS
c.68137G > T p.A22713S Missense HT‐16 0.0001 D D P D M High PSI, A‐band VUS
c.82024 T > C p.Y27342H Missense HT‐1a NA D D D D H High PSI, A‐band VUS
c.89303C > T p.P29768L Missense HT‐20 0.0001 D D D D M High PSI, A‐band VUS
c.96320G > A p.R32107K Missense HT‐39 0.0002 D D P D N High PSI, M‐band VUS
DSG2 NM_001943 c.136C > T p.R46W Missense HT‐22a 0.0001 D D D N H NA LP
c.593A > G p.Y198C Missense HT‐36a, HT‐22a 0.0001 D D D D H NA VUS
c.991G > A p.E331K Missense HT‐6a 0.0001 D D D A M NA VUS
c.1592 T > G p.F531C Missense HT‐36a, HT‐6a, HT‐21b 0.0012 D D D D H NA VUS
c.2390 T > A p.L797Q Missense HT‐1a NA D D D D M NA VUS
PKP2 NM_004572 c.746G > C p.S249 T Missense HT‐4a NA D D D D M NA VUS
c.2554delG p.E852fs Frameshift deletion HT‐4a NA NA NA NA NA NA NA VUS
DSC2 NM_004949 c.395G > A p.R132H Missense HT‐23a 0.0001 D D D D H NA VUS
c.905G > A p.G302D Missense HT‐23a NA D D D D H NA VUS
DSP NM_001008844 c.943C > T p.R315C Missense HT‐14a 0.001 D D P D N NA VUS
PLN NM_002667 c.36_38del p.12_13del Non–frameshift deletion HT‐27 NA NA NA NA NA NA NA P
LMNA NM_005572 c.797C > T p.T266I Missense HT‐12 NA D D D D M NA LP
c.922C > T p.Q308* Stopgain HT‐1a NA NA NA NA NA NA NA LP
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Abbreviations

MAF column: MAF, minor allele frequency; NA, data not applicable.

SIFT column: D, deleterious; T, tolerate.

Polyphen2 HDIV and HVAR columns: B, benign; D, probably damage; P, possible damage.

MutationTaster column: A, disease causing automatic; D, disease causing; N, polymorphism; P, polymorphism automatic.

MutationAssessor column: H, high pathogenic; M, medium pathogenic; L, low pathogenic; N, no pathogenic.

PSI column: NA, data not applicable; PSA, percentage spliced in.

ACMG column: ACMG, American College of Medical Genetics; LP, likely pathogenic; P, pathogenic; VUS, variant of uncertain significance.

a

Patient with compound heterozygous variants.

b

Patient with homozygous variants.

3.3. Similar phenotype in patients with TTN variants to those without variants

Compared to patients with TTN missense variants and those without variants, patients with desmosomal variants demonstrated a younger age of onset of the disease (17 ±3 years vs 31 ±13 years and 32 ±11 years, P = 0.049 and P = 0.024, respectively) and a younger age for heart transplantation (24 ±9 years vs 41 ±14 and 38 ±10 years, P = 0.027 and P = 0.049, respectively) (Table 1). Kaplan–Meier analysis showed lower composite endpoint‐free survival rate in patients with desmosomal variants than those without variants (P = 0.004) (Figure 1). Regarding ventricular alteration, patients with desmosomal variants had a significantly larger right ventricular out flow tract (50 ±15 mm vs 34 ±14 mm and 31 ±8 mm, P = 0.046 and P = 0.004, respectively), and a smaller left ventricle end‐diastolic dimension (45 ±10 mm vs 62 ±10 mm and 62 ±11 mm, P = 0.019 and P = 0.007, respectively) and left atrial dimension (25 ±6 mm vs 44 ±16 mm and 44 ±11 mm, P = 0.002 and P = 0.001, respectively) than patients with TTN missense variants and those without variants.

Figure 1.

Figure 1

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Survival analysis. Follow‐up from birth to endpoint. (A) Survival‐free from composite endpoint. (B) Survival‐free from heart transplantation. (C) Survival‐free from major arrhythmic cardiovascular event. Abbreviations: DS, patients with single desmosomal variants; NV, patients with no detectable variants; TTN, patients with TTN variants only; HT, heart transplantation; MACE, major arrhythmic cardiovascular event

3.4. Digital quantitative evaluation fibro‐fatty infiltration

The characteristic histology feature of ACM is loss of myocardium and fibro‐fatty infiltration in gross pathology and microscopic pathology (Figure 2A,B). Myocardium decrease was most obviously detected in RV, where the percentage of myocardium was significantly lower than that in LV and IVS (37.0% ±26.7% vs 56.3% ±20.3% and 64.9% ±17.7%, P < 0.001 and P < 0.001, respectively) (Figure 2C). Interestingly, we found more fibrous tissue infiltration in LV than RV (27.5% ±14.8% vs 21.1% ±11.6%, P = 0.011) and more adipose tissue in RV than that in LV and IVS (41.9% ±26.8% vs 16.2% ±14.0% and 7.1% ±8.1%, P < 0.001 and P < 0.001) (Figure 2C).

Figure 2.

Figure 2

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Pathological features of arrhythmogenic cardiomyopathy. (A) gross feature of right ventricle of a patient with TTN missense variant (top). Gross feature of right ventricle of a patient with DSG2 variant (bottom). (B) Masson's trichrome stain of specimens from patients with TTN missense variant, desmosomal variants, and without variants. (C) Quantification of myocardium, fibrous tissue, and adipose tissue in the left ventricle, right ventricle, and interventricular septum of ACM patients. *P < 0.05; **P < 0.01; ***P < 0.001. Abbreviations: DS, patients with single desmosomal variants; IVS, interventricular septum; LV, left ventricle; NV, patients with no detectable variants; RV, right ventricle; TTN, patients with TTN variants only

We then compared the percentage of myocardium, fibrous tissue, and adipose tissue. There was no difference between groups in LV (including ALV, LLV and PLV) and IVS (Figure 3A,C; and Supporting Figure S1 in the online version of this article). In RV, compared to patients with TTN missense variants and those without variants, patients with desmosomal variants had much less myocardium (13.8% ±11.0% vs 41.0% ±26.4% and 40.8% ±29.4%, P = 0.006 and P = 0.017, respectively) and more adipose tissue (66.9% ±18.5% vs 34.3% ±23.0% and 43.0% ±30.9%, P = 0.001 and P = 0.036, respectively) (Figure 3B). Patients with TTN missense variants had less fibrous tissue than those without variants in RV (16.2% ±7.2% vs 24.7% ±12.9%, P = 0.035) (Figure 3B). In ARV, there was a trend that patients with desmosomal variants had less myocardium than patients with TTN missense variants and without variants, but no statistical significance was detected. Patients with desmosomal variants had much more adipose tissue than those without variants (70.5% ±20.8% vs 31.3% ±20.5%, P = 0.004), and patients without variants had more fibrous tissue than those with TTN missense variants (29.5% ±14.3% vs 14.4% ±7.1%, P = 0.015) (see Supporting Figure S2A in the online version of this article). In PRV, although no significance was found, there was a trend that patients with desmosomal variants had less myocardium and more adipose tissue than those with TTN missense variants and those without variants (see Supporting Figure S2B in the online version of this article).

Figure 3.

Figure 3

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Quantification of myocardium, fibrous tissue, and adipose tissue in subgroups of patients. Abbreviations: DS, patients with desmosomal variants; IVS, interventricular septum; LV, left ventricle; NV, patients with no detectable variants; RV, right ventricle; TTN, patients with TTN missense variants. *P < 0.05; **P < 0.01; ***P < 0.001

3.5. Pedigree analysis

We performed specific TTN variants testing in 3 available families, and TTN missense variants were detected in 11 family members (see Supporting Figure S3 in the online version of this article). Clinical evaluation of the family members showed no abnormity, indicating TTN missense variants had either a very low penetrance or negligible pathogenicity on the disease.

4. DISCUSSION

In this study, we reported a large heart transplant cohort of ACM with well‐characterized pathological features. In our cohort, 26% of the patients had TTN missense variants, and 17% had desmosomal variants. The genetic background of our cohort was distinctive, with less desmosomal variants than that of the cohorts from previous studies.32, 33 Our results demonstrated the genetic features of a subgroup in Chinese ACM patients who had heart transplantation.

The TTN missense variants have been reported as a causal gene variant of ACM in 2011.15 In our cohort, TTN missense variants were commonly identified in ACM patients. Although the TTN truncating variant could be detected in 10% to 25% of patients with dilated cardiomyopathy,34 it was not detected in ACM patients. Therefore, TTN truncating variants might not be the genetic causes of ACM in our cohort. In accordance with previous study,24 individual patients with desmosomal variants had early onset of the disease; therefore, they had a poorer prognosis than those without variants. Additionally, patients with desmosomal variants had a more classic ACM phenotype, with a severely enlarged RV but a relatively better LV morphology. We did not detect any difference in inverted T wave or pacemaker implantation between groups. The difference between our study and previous studies might be caused by either ethnic difference, as we included Chinese ACM patients, or end‐stage characteristics of ACM, as our cohort was comprised of end‐stage ACM patients only. In addition, the differences between patients with desmosomal variants and those without variants were detected in both our cohort and other cohorts, indicating desmosomal variants had a certain effect on ACM phenotype. However, the characteristics of patients with TTN missense variants were similar to those without a variant, indicating TTN missense variants could not induce a specific clinical phenotype in ACM patients in our cohort. Quantitative evaluation of myocardium, fibrous tissue, and adipose tissue showed more infiltration of fat tissue and loss of myocardium in the RV of the patients with desmosomal variants, which explained why these patients had a worse enlarged RV condition. Histology analysis indicated desmosomal variants remarkably impaired RV, which led to a subgroup of classic ACM. Similar histological characteristic of patients with TTN missense variants and those without variants suggested TTN missense variants might not lead a subgroup of ACM.

As titin is the largest protein and ranks as the third abundant in heart muscle,35 the frequency of TTN missense variants in cardiomyopathies is far above what would be expected according to the prevalence.36 As a result, the pathogenesis of TTN missense variants is controversial. Pedigree analysis demonstrated a very low penetrance of TTN missense variants, indicating TTN missense variants had little pathogenicity on the disease. Li et al37 found oligogenic etiology in hypertrophic cardiomyopathy, suggesting TTN missense variants could contribute additive effect to the pathogenicity of cardiomyopathy. Herein we assumed TTN missense variants might superimpose genetic predisposition for ACM, but not the primary factor that led to ACM.

Our study has some limitations. First, the total sample size of our cohort is limited. However, this is the largest heart transplantation cohort with ACM. Second, we only focused on proven ACM‐related genes. Whole genome sequencing or whole exon sequencing should be performed on patients without known variants. Third, the pathologic features reflected only end‐stage ACM. To study ACM patients in an early stage who died suddenly might also be of great importance.

5. CONCLUSION

TTN missense variants were commonly identified in our ACM cohort, but TTN truncating variants were absent. Patients with TTN missense variants had similar clinical and pathological features to patients without variants and showed very low penetrance, indicating absence of primary pathogenesis in ACM patients in our cohort.

Conflicts of interest

The authors declare no potential conflicts of interest.

Supporting information

Figure S1. Pathological ingredient of left ventricle.

Figure S2. Pathological ingredient of right ventricle.

Figure S3. Pedigree of patients with TTN variants.

Click here for additional data file. (14MB, docx)

Table S1 Diagnosis of the patients in the cohort.

Click here for additional data file. (27.2KB, docx)

Chen K, Song J, Wang Z, Rao M, Chen L, Hu S. Absence of a primary role for TTN missense variants in arrhythmogenic cardiomyopathy: From a clinical and pathological perspective. Clin Cardiol. 2018;41:615–622. 10.1002/clc.22906

Funding information This study was supported by the CAMS Innovation Fund for Medical Sciences (CIFMS, 2016‐I2M‐1‐015), the National Natural Science Foundation of China (grant no. 81670376), and the PUMC Youth Fund and the Fundamental Research Funds for the Central Universities.

Contributor Information

Jiangping Song, Email: fwsongjiangping@163.com.

Shengshou Hu, Email: fwhushengshou@163.com.

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Associated Data

This section collects any data citations, data availability statements, or supplementary materials included in this article.

Supplementary Materials

Figure S1. Pathological ingredient of left ventricle.

Figure S2. Pathological ingredient of right ventricle.

Figure S3. Pedigree of patients with TTN variants.

Click here for additional data file. (14MB, docx)

Table S1 Diagnosis of the patients in the cohort.

Click here for additional data file. (27.2KB, docx)

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