BACKGROUND Dilated cardiomyopathy and hypertrophic cardiomyopathy arise from mutations in many genes. TTN, the gene encoding the sarcomere protein titin, has been insufficiently analyzed for cardiomyopathy mutations because of its enormous size. METHODS We analyzed TTN in 312 subjects with dilated cardiomyopathy, 231 subjects with hyper-trophic cardiomyopathy, and 249 controls by using next-generation or dideoxy sequencing. We evaluated deleterious variants for cosegregation in families and assessed clinical characteristics. RESULTS We identified 72 unique mutations (25 nonsense, 23 frameshift, 23 splicing, and 1 large tandem insertion) that altered full-length titin. Among subjects studied by means of next-generation sequencing, the frequency of TTN mutations was significantly higher among subjects with dilated cardiomyopathy (54 of 203 [27%]) than among subjects with hypertrophic cardiomyopathy (3 of 231 [1%], P = 3×10−16) or controls (7 of 249 [3%], P = 9×10−14). TTN mutations cosegregated with dilated cardiomyopathy in families (combined lod score, 11.1) with high (>95%) observed penetrance after the age of 40 years. Mutations associated with dilated cardiomyopathy were overrepresented in the titin A-band but were absent from the Z-disk and M-band regions of titin (P≤0.01 for all comparisons). Overall, the rates of cardiac outcomes were similar in subjects with and those without TTN mutations, but adverse events occurred earlier in male mutation carriers than in female carriers (P = 4×10−5). CONCLUSIONS TTN truncating mutations are a common cause of dilated cardiomyopathy, occurring in approximately 25% of familial cases of idiopathic dilated cardiomyopathy and in 18% of sporadic cases. Incorporation of sequencing approaches that detect TTN truncations into genetic testing for dilated cardiomyopathy should substantially increase test sensitivity, thereby allowing earlier diagnosis and therapeutic intervention for many patients with dilated cardiomyopathy. Defining the functional effects of TTN truncating mutations should improve our understanding of the pathophysiology of dilated cardiomyopathy. (Funded by the Howard Hughes Medical Institute and others.)

Mutations in sarcomere protein genes can cause hypertrophic cardiomyopathy (HCM), a disorder characterized by myocyte enlargement, fibrosis, and impaired ventricular relaxation. Here, we demonstrate that sarcomere protein gene mutations activate proliferative and profibrotic signals in non-myocyte cells to produce pathologic remodeling in HCM. Gene expression analyses of non-myocyte cells isolated from HCM mouse hearts showed increased levels of RNAs encoding cell-cycle proteins, Tgf-β, periostin, and other profibrotic proteins. Markedly increased BrdU labeling, Ki67 antigen expression, and periostin immunohistochemistry in the fibrotic regions of HCM hearts confirmed the transcriptional profiling data. Genetic ablation of periostin in HCM mice reduced but did not extinguish non-myocyte proliferation and fibrosis. In contrast, administration of Tgf-β-neutralizing antibodies abrogated non-myocyte proliferation and fibrosis. Chronic administration of the angiotensin II type 1 receptor antagonist losartan to mutation-positive, hypertrophynegative (prehypertrophic) mice prevented the emergence of hypertrophy, non-myocyte proliferation, and fibrosis. Losartan treatment did not reverse pathologic remodeling of established HCM but did reduce nonmyocyte proliferation. These data define non-myocyte activation of Tgf-β signaling as a pivotal mechanism for increased fibrosis in HCM and a potentially important factor contributing to diastolic dysfunction and heart failure. Preemptive pharmacologic inhibition of Tgf-β signals warrants study in human patients with sarcomere gene mutations.

The transcriptome is subject to multiple changes during pathogenesis, including the use of alternate 5′ startsites that can affect transcription levels and output. Current RNA sequencing techniques can assess mRNA levels, but do not robustly detect changes in 5′ start-site use. Here, we developed a transcriptome sequencing strategy that detects genome-wide changes in start-site usage (5′RNA-Seq) and applied this methodology to identify regulatory events that occur in hypertrophic cardiomyopathy (HCM). Compared with transcripts from WT mice, 92 genes had altered start-site usage in a mouse model of HCM, including four-and-a-half LIM domains protein 1 (Fhl1). HCM-induced altered transcriptional regulation of Fhl1 resulted in robust myocyte expression of a distinct protein isoform, a response that was conserved in humans with genetic or acquired cardiomyopathies. Genetic ablation of Fhl1 in HCM mice was deleterious, which suggests that Fhl1 transcriptional changes provide salutary effects on stressed myocytes in this disease. Because Fhl1 is a chromosome X-encoded gene, stress-induced changes in its transcription may contribute to gender differences in the clinical severity of HCM. Our findings indicate that 5′RNA-Seq has the potential to identify genomewide changes in 5′ start-site usage that are associated with pathogenic phenotypes.

Inherited cardiomyopathies include hypertrophic cardiomyopathy, dilated cardiomyopathy, arrhythmogenic right ventricular cardiomyopathy, left ventricular noncompaction, and restrictive cardiomyopathy. These diseases have a substantial genetic component and predispose to sudden cardiac death, which provides a high incentive to identify and sequence disease genes in affected individuals to identify pathogenic variants. Clinical genetic testing, which is now widely available, can be a powerful tool for identifying presymptomatic individuals. However, locus and allelic heterogeneity are the rule, as are clinical variability and reduced penetrance of disease in carriers of pathogenic variants. These factors, combined with genetic and phenotypic overlap between different cardiomyopathies, have made clinical genetic testing a lengthy and costly process. Next-generation sequencing technologies have removed many limitations such that comprehensive testing is now feasible, shortening diagnostic odysseys for clinically complex cases. Remaining challenges include the incomplete understanding of the spectrum of benign and pathogenic variants in the cardiomyopathy genes, which is a source of inconclusive results. This review provides an overview of inherited cardiomyopathies with a focus on their genetic etiology and diagnostic testing in the postgenomic era.

Objective: To explore the transcriptomic differences between patients with hypertrophic cardiomyopathy (HCM) and controls. Patients and Methods: RNA was extracted from cardiac tissue flash frozen at therapeutic surgical septal myectomy for 106 patients with HCM and 39 healthy donor hearts. Expression profiling of 37,846 genes was performed using the Illumina Human HT-12v3 Expression BeadChip. All patients with HCM were genotyped for pathogenic variants causing HCM. Technical validation was performed using quantitative real-time polymerase chain reaction (qRT-PCR) and Western blot. This study was started on January 1, 1999, and final analysis was completed on April 20, 2020. Results: Overall, 22% of the transcriptome (8443 of 37,846 genes) was expressed differentially between HCM and control tissues. Analysis by genotype revealed that gene expression changes were similar among genotypic subgroups of HCM, with only 4% (1502 of 37,846) to 6% (2336 of 37,846) of the transcriptome exhibiting differential expression between genotypic subgroups. The qRT-PCR confirmed differential expression in 92% (11 of 12 genes) of tested transcripts. Notably, in the context of coronavirus disease 2019 (COVID-19), the transcript for angiotensin I converting enzyme 2 (ACE2), a negative regulator of the angiotensin system, was the single most up-regulated gene in HCM (fold-change, 3.53; q-value ¼1.30Â10 À23 ), which was confirmed by qRT-PCR in triplicate (fold change, 3.78; P¼5.22Â10 À4 ), and Western blot confirmed greater than 5-fold overexpression of ACE2 protein (fold change, 5.34; P¼1.66Â10 À6 ). Conclusion: More than 20% of the transcriptome is expressed differentially between HCM and control tissues. Importantly, ACE2 was the most up-regulated gene in HCM, indicating perhaps the heart's compensatory effort to mount an antihypertrophic, antifibrotic response. However, given that the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) uses ACE2 for viral entry,

, respectively) develop significant hypertrophic cardiomyopathy (HCM) compared with male and female wild-type mice (M-␣MHC ϩ/ϩ and F-␣MHC ϩ/ϩ , respectively) after ϳ30 wk of age. We tested the hypothesis that myofilament mechanical performance differs between M-␣MHC R403Q/ϩ and F-␣MHC R403Q/ϩ at younger ages (10 -20 wk) and could account for sex differences in HCM development. The sensitivity of chemically skinned myocardial strips to Ca 2ϩ activation (pCa50) was significantly (P Ͻ 0.05) enhanced in male mice independent of genotype (M-␣MHC Ϫ3 , F-␣MHC ϩ/ϩ 25 Ϯ 3 10 Ϫ3 muscle lengths/s ϫ normalized tension). We did not find a statistically significant sex ϫ mutation interaction for any measure of myofilament performance. Therefore, sarcomeric incorporation of the R403Q myosin similarly enhanced left ventricular myofilament mechanical performance in both male and female mice. The sex-dependent development of HCM due to the R403Q myosin may then be inhibited by female sex hormones, which may additionally underlie the observed sex differences for pCa50 and unloaded shortening velocity. hypertrophic cardiomyopathy; myosin heavy chain; isometric tension; calcium sensitivity; force-velocity HYPERTROPHIC CARDIOMYOPATHY (HCM) in the absence of recognizable hemodynamic risk factors can arise naturally as a consequence of a mutant allele for myosin heavy chain (MHC) (41). Assessing the functional consequences of HCM-associated mutations, such as the R403Q point mutation in cardiac ␣-MHC or ␤-MHC, has played an important role in the identification of modifications in myofilament performance that may lead to the HCM phenotype (9,19,36,51). However, functional data from some models of the R403Q-mutated MHC have been indefinite. For example, myosin isolated from myocardium of HCM patients carrying an allele for the R403Q mutation reportedly reduce (5) or enhance (33) actin filament velocity (V actin ) in the myosin motility assay. Additionally, actin-activated myosin ATPase and V actin of R403Q mutant MHC expressed by baculovirus/insect cell systems are reportedly reduced for rat cardiac ␣-MHC (44) and yet enhanced in chicken gizzard smooth muscle MHC (53).Data from mouse models of the R403Q mutation in ␣-MHC, on the other hand, have provided more consistent findings. Myosin isolated from myocardium of ␣-MHC R403Q/R403Q homozygous mice demonstrate a marked enhancement of actinactivated ATPase, V actin , and stall force of V actin by ␣-actinin (7, 47). Similarly, cardiac myosin isolated from ␣MHC R403Q/ϩ heterozygous mice shows enhanced ATPase (2, 31, 47). Data from ATPase and V actin assays generally correspond to each other and reflect actomyosin cross-bridge kinetics (48), whose enhancement in the R403Q myosin has been corroborated by a shorter cross-bridge time-on measured in the laser trap (47).Left ventricular (LV) function in the ␣MHC R403Q/ϩ heterozygous mouse, which mimics the human genotype leading to HCM, is characterized in part by an earlier and higher peak LV pressure and an elevated free wall fractional shorteni...

Background-The geographic isolation and homogeneous population of Iceland are ideally suited to ascertain clinical and genetic characteristics of hypertrophic cardiomyopathy (HCM) at the population level. Methods and Results-Medical records and cardiac imaging studies obtained between 1997 and 2010 were reviewed to identify Icelandic patients with HCM. Surviving patients were recruited for clinical and genetic studies. A previously identified Icelandic mutation, MYBPC3 c.927-2A>G, was genotyped, and mutation-negative samples were sequenced for HCM genes and other hypertrophic genes. Record review identified 180 patients with HCM. Genetic analyses of 151 patients defined pathogenic mutations in 101 (67%), including MYBPC3 c.927-2A>G (88 patients, 58%), 4 other MYBPC3 or MYH7 mutations (5 patients, 3.3%), and 2 GLA mutations (8 patients, 5.3%). Haplotype and genetic genealogical data defined MYBPC3 c.927-2A>G as a founder mutation, introduced into the Icelandic population in the 15th century, with a current population prevalence of 0.36%. MYBPC3 c.927-2A>G mutation carriers exhibited phenotypic diversity but were younger at diagnosis (42 versus 49 years; P=0.001) and sustained more adverse events (15% versus 2%; P=0.02) than mutation-negative patients. All-cause mortality for patients with HCM was similar to that of an age-matched Icelandic population (hazard ratio, 0.98; P=0.9). HCM-related mortality (0.78%/y) occurred at a mean age of 68 compared with 81 years for non-HCM-related mortality (P=0.02). Conclusions-A founder MYBPC3 mutation that arose >550 years ago is the predominant cause of HCM in Iceland. The MYBPC3 c.927-2A>G mutation is associated with low adverse event rates but earlier cardiovascular mortality, illustrating the impact of genotype on outcomes in HCM.

Hypertrophic cardiomyopathy (HCM) is a genetically heterogeneous cardiac muscle disorder with a diverse natural history, characterized by unexplained left ventricular hypertrophy (LVH), with histopathological hallmarks including myocyte enlargement, myocyte disarray and myocardial fibrosis. Although these features can cause significant cardiac symptoms, many young individuals with HCM are asymptomatic or mildly symptomatic. Sudden cardiac death (SCD) may occur as the initial clinical manifestation. Over the past few decades, HCM has been considered a disease of sarcomere, and typically as an autosomal dominant disease with variable expressivity and incomplete penetrance. Important insights into the genetic landscape of HCM have enhanced our understanding of the molecular pathogenesis, empowered gene-based diagnostic testing to identify at-risk individuals, and offered potential targets for the development of therapeutic agents. This article reviews the current knowledge on the clinical genetics and management of HCM.