Left ventricular non-compaction (LVNC) is the third most prevalent cardiomyopathy in children and its pathogenesis has been associated with the developmental defect of the embryonic myocardium. We show that patient-specific induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) generated from LVNC patients carrying a mutation in the cardiac transcription factor TBX20 recapitulate a key aspect of the pathological phenotype at the single-cell level and this was associated with perturbed transforming growth factor beta (TGF-β) signalling. LVNC iPSC-CMs have decreased proliferative capacity due to abnormal activation of TGF-β signalling. TBX20 regulates the expression of TGF-β signalling modifiers including one known to be a genetic cause of LVNC, PRDM16, and genome editing of PRDM16 caused proliferation defects in iPSC-CMs. Inhibition of TGF-β signalling and genome correction of the TBX20 mutation were sufficient to reverse the disease phenotype. Our study demonstrates that iPSC-CMs are a useful tool for the exploration of pathological mechanisms underlying poorly understood cardiomyopathies including LVNC.

Sex differences in the development of the normal heart and the prevalence of cardiomyopathies have been reported. The molecular basis of these differences remains unclear. Sex differences in the human heart might be related to patterns of gene expression. Recent studies have shown that sex specific differences in gene expression in tissues including the brain, kidney, skeletal muscle, and liver. Similar data is limited for the heart. Herein we address this issue by analyzing donor and post-mortem adult human heart samples originating from 46 control individuals to study whole-genome gene expression in the human left ventricle. Using data from the genotype tissue expression (GTEx) project, we compared the transcriptome expression profiles of male and female hearts. We found that genes located on sex chromosomes were the most abundant ones among the sexually dimorphic genes. The majority of differentially expressed autosomal genes were those involved in the regulation of inflammation, which has been found to be an important contributor to left ventricular remodeling. Specifically, genes on autosomal chromosomes encoding chemokines with inflammatory functions (e.g. CCL4, CX3CL1, TNFAIP3) and a gene that regulates adhesion of immune cells to the endothelium (e.g., VCAM1) were identified with sex-specific expression levels. This study underlines the relevance of sex as an important modifier of cardiac gene expression. These results have important implications in the understanding of the differences in the physiology of the male and female heart transcriptome and how they may lead to different sex specific difference in human cardiac health and its control.

Rationale:: Thousands of mutations across >50 genes have been implicated in inherited cardiomyopathies. However, options for sequencing this rapidly evolving gene set are limited because many sequencing services and off-the-shelf kits suffer from slow turnaround, inefficient capture of genomic DNA, and high cost. Furthermore, customization of these assays to cover emerging targets that suit individual needs is often expensive and time consuming. Objective:: We sought to develop a custom high throughput, clinical-grade next-generation sequencing assay for detecting cardiac disease gene mutations with improved accuracy, flexibility, turnaround, and cost. Methods and Results:: We used double-stranded probes (complementary long padlock probes), an inexpensive and customizable capture technology, to efficiently capture and amplify the entire coding region and flanking intronic and regulatory sequences of 88 genes and 40 microRNAs associated with inherited cardiomyopathies, congenital heart disease, and cardiac development. Multiplexing 11 samples per sequencing run resulted in a mean base pair coverage of 420, of which 97% had >20× coverage and >99% were concordant with known heterozygous single nucleotide polymorphisms. The assay correctly detected germline variants in 24 individuals and revealed several polymorphic regions in miR-499. Total run time was 3 days at an approximate cost of $100 per sample. Conclusions:: Accurate, high-throughput detection of mutations across numerous cardiac genes is achievable with complementary long padlock probe technology. Moreover, this format allows facile insertion of additional probes as more cardiomyopathy and congenital heart disease genes are discovered, giving researchers a powerful new tool for DNA mutation detection and discovery.

We aimed to identify the genetic cause of coronary artery disease (CAD) in an Iranian pedigree. Genetic linkage analysis identified three loci with an LOD score of 2.2. Twelve sequence variations identified by exome sequencing were tested for segregation with disease. A p.Val99Met causing mutation in ST6GALNAC5 was considered the likely cause of CAD. ST6GALNAC5 encodes sialyltransferase 7e. The variation affects a highly conserved amino acid, was absent in 800 controls, and was predicted to damage protein function. ST6GALNAC5 is positioned within loci previously linked to CAD-associated parameters. While hypercholesterolemia was a prominent feature in the family, clinical and genetic data suggest that this condition is not caused by the mutation in ST6GALNAC5. Sequencing of ST6GALNAC5 in 160 Iranian patients revealed a candidate causative stop-loss mutation in two other patients. The p.Val99Met and stop-loss mutations both caused increased sialyltransferase activity. Sequence data from combined Iranian and US controls and CAD affected individuals provided evidence consistent with potential role of ST6GALNAC5 in CAD. We conclude that ST6GALNAC5 mutations can cause CAD. There is substantial literature suggesting a relation between sialyltransferase and sialic acid levels and coronary disease. Our findings provide strong evidence for the existence of this relation.