Multi-omic profiling of cardiomyocyte ageing and functional decline

AIMS: Ageing is the strongest risk factor for heart failure, yet the molecular mechanisms underlying cardiomyocyte (CM) ageing remain unclear. We aimed to map the transcriptomic and epigenomic landscape of CM ageing and to test whether DNA hypermethylation is a causal driver of diastolic dysfunction. METHODS AND RESULTS: We performed single-nucleus multi-omics (concurrent snRNA-seq and snATAC-seq) on 4- and 28-month-old ventricular myocardium of C57BL/6J mice. Aged CMs showed widespread chromatin remodelling, with 28 324 regions having greater accessibility compared to only two with reduced accessibility. 1963 genes were differentially expressed in aged ventricular CMs, with 78.5% neighbouring differentially accessible regions. Promoter accessibility positively associated with expression. Reduced-representation bisulphite sequencing of ventricular CM nuclei identified 1422 regions associated with genes that were hypermethylated in aged CMs, compared to only 167 that were hypomethylated. CpG hypermethylation inversely correlated with differential gene expression. Multi-omic integration revealed ageing signatures shared across cell types and identified the long non-coding RNA Gm12381 as a CM-selective ageing marker. To test whether DNA hypermethylation is associated with ageing phenotypes, we used an engineered cardiotropic AAV (MyoAAV) to overexpress Dnmt3a in adult hearts. Dnmt3a overexpression induced CM hypermethylation, causing cardiac hypertrophy and diastolic dysfunction, key ageing phenotypes, and altering the transcriptome profile towards that of aged CMs. CONCLUSION: Gain of chromatin accessibility and CpG hypermethylation associated with transcriptomic reprogramming characterize CM ageing. Experimental elevation of DNA methylation is sufficient to induce diastolic dysfunction and hypertrophy, supporting DNMT3A-mediated hypermethylation as a mechanistic driver. Further work should test whether attenuating methylation prevents or reverses age-related cardiac dysfunction.