Chromatin Accessibility of Human Mitral Valves and Functional Assessment of MVP Risk Loci.

RATIONALE: Mitral valve prolapse (MVP) is a common valvopathy that leads to mitral insufficiency, heart failure, and sudden death. Functional genomic studies in mitral valves are needed to better characterize MVP-associated variants and target genes. OBJECTIVE: To establish the chromatin accessibility profiles and assess functionality of variants and narrow down target genes at MVP loci. METHODS AND RESULTS: We mapped the open chromatin regions in nuclei from 11 human pathogenic and 7 nonpathogenic mitral valves by an assay for transposase-accessible chromatin with high-throughput sequencing. Open chromatin peaks were globally similar between pathogenic and nonpathogenic valves. Compared with the heart tissue and cardiac fibroblasts, we found that MV-specific assay for transposase-accessible chromatin with high-throughput sequencing peaks are enriched near genes involved in extracellular matrix organization, chondrocyte differentiation, and connective tissue development. One of the most enriched motifs in MV-specific open chromatin peaks was for the nuclear factor of activated T cells family of TFs (transcription factors) involved in valve endocardial and interstitial cell formation. We also found that MVP-associated variants were significantly enriched (P<0.05) in mitral valve open chromatin peaks. Integration of the assay for transposase-accessible chromatin with high-throughput sequencing data with risk loci, extensive functional annotation, and gene reporter assay suggest plausible causal variants for rs2641440 at the SMG6/SRR locus and rs6723013 at the IGFBP2/IGFBP5/TNS1 locus. CRISPR-Cas9 deletion of the sequence including rs6723013 in human fibroblasts correlated with increased expression only for TNS1. Circular chromatin conformation capture followed by high-throughput sequencing experiments provided evidence for several target genes, including SRR, HIC1, and DPH1 at the SMG6/SRR locus and further supported TNS1 as the most likely target gene on chromosome 2. CONCLUSIONS: Here, we describe unprecedented genome-wide open chromatin profiles from human pathogenic and nonpathogenic MVs and report specific gene regulation profiles, compared with the heart. We also report in vitro functional evidence for potential causal variants and target genes at MVP risk loci involving established and new biological mechanisms. Graphic Abstract: A graphic abstract is available for this article.

Normal distribution of melanocytes in the mouse heart.

We report the consistent distribution of a population of pigmented trp-1-positive cells in several important septal and valvular structures of the normal mouse (C57BL/6) heart. The pigmented cell population was first apparent by E16.5 p.c. in the right atrial wall and extended into the atrium along the interatrial septum. By E17.5, these cells were found along the apical membranous interventricular septum near or below the surface of the endocardium. The most striking distribution of dark pigmented cells was found in the tricuspid and mitral valvular leaflets and chordae tendineae. The normal distribution of pigmented cells in the valvuloseptal apparatus of C57BL/6 adult heart suggests that a premelanocytic lineage may participate in the earlier morphogenesis of the valve leaflets and chordae tendineae. The origin of the premelanocyte lineage is currently unknown. The most likely candidate populations include the neural crest and the epicardially derived cells. The only cell type in the heart previously shown to form melanocytes is the neural crest. The presence of neural crest cells, but not melanocytes, in some of the regions we describe has been reported by others. However, previous reports have not shown a contribution of melanocytes or neural crest derivatives to the atrioventricular valve leaflets or chordae tendineae in mouse hearts. If these cells are of neural crest origin, it would suggest a possibly greater contribution and persistence of neural crest cells to the valvuloseptal apparatus than has been previously understood.