Prelamin A drives vascular calcification by reorganization of the epigenetic landscape to promote inflammaging

Background: Vascular calcification is a ubiquitous ageing pathology markedly accelerated in patients with metabolic disorders including diabetes and renal failure. Prelamin A is a biomarker of vascular ageing which accelerates vascular smooth muscle cell (VSMC) senescence and calcification. Using prelamin A as a model we explored the earliest events in VSMC ageing, focussing on the epigenetic landscape as an environmentally modifiable target. Methods: Adenoviral expression of prelamin A was used to induce VSMC ageing and epigenetic modifications were monitored using immunofluorescence and targeted PCR-arrays. Epigenetic findings were verified in vitro using drugs targeting epigenetic modifiers and in vivo using immunohistochemistry in human vessels from young, aged and calcified patients. Transcriptomics, ChIP-seq and antibody arrays were used to identify and verify gene targets impacted by changes in the epigenetic landscape and to monitor effects on inflammation, ageing indices and calcification. A novel mouse model of VSMC-specific inducible prelamin A expression was used to study epigenetic ageing in vivo and calcification was induced with vitamin D. Results: Prelamin A accumulation in VSMCs caused rapid global alterations in the repressive heterochromatin marks, H3K9me3 and H3K27me3 which coincided with downregulation of the epigenetic writers EZH2 and SUV39H1. These same epigenetic changes were recapitulated in primary human VSMCs induced to calcify in response to mineral dysregulation and in the calcified arteries of adults, and young children on dialysis. Global analysis of H3K9me3 and H3K27me3 marks in response to prelamin A expression showed deregulation of cross-talking NF-κB and HIPK2 stress response inflammatory pathways consistent with robust activation of a senescence associated secretory phenotype (SASP). Treatment of VSMCs with EZH2 and SUV39H1 inhibitors amplified the SASP and accelerated senescence and mineral deposition. Induction of prelamin A expression in VSMCs in mice induced rapid loss of heterochromatin which preceded DNA damage and senescence and coincided with early activation of SASP factors creating an environment permissive to calcification. Conclusions: Loss of heterochromatin is accelerated by metabolic disease and acts to promote VSMC inflammaging and a tissue microenvironment prone to calcification. Pre-senescent cells with an active SASP may be an early target for tackling vascular ageing pathologies. tackling vascular ageing pathologies.