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1 Prelamin A promotes DNA damage, calcification and senescence in human vascular smooth muscle cells
  1. Y Liu,
  2. C M Shanahan
  1. Cardiovascular Division, James Black Centre, King's College London, London, UK

Abstract

Vascular calcification is prominent in the ageing population yet little is understood about the mechanisms involved in senescence-associated phenotypic change in VSMCs. Our previous data showed that VSMCs acquire nuclear defects due to the accumulation of the nuclear lamina protein-prelamin A as they approach senescence both in vitro and in vivo. Over-expression of prelamin A accelerated VSMC senescence and DNA damage1, and prelamin A accumulated at sites of calcification in diseased arteries. In this study, we investigated whether prelamin A accumulation has an effect on VSMC osteogenic differentiation and the mechanisms underlying this process. Using immunostaining and Western blot we show that there was increased prelamin A accumulation in calcifying VSMCs. Adenoviral over-expression of prelamin A significantly accelerated VSMC mineralisation, shown by alizarin red staining, increased 45Calcium incorporation (6.97±0.58-fold [p<0.01]) and alkaline phosphatase (ALP) activity (2.11±0.16-fold [p<0.05]) compared to controls. Moreover, prelamin A over-expression also significantly elevated mRNA levels of Runx2, osteocalcin, BMP2 and ALP in VSMCs. Coculture of VSMC over-expressing prelamin A with C2C12 cells significantly enhanced osteogenic differentiation of C2C12€s due to increased BMP2 secretion. Prelamin A over-expression also induced persistent DNA damage in VSMCs, shown by 53BP1 immunostaining and western blotting for p-ATM/ATR, and p-H2AX. More importantly, the DNA damage response inhibitor, CGK733, significantly blocked prelamin A over-expression induced VSMC osteogenic differentiation. This is the first study to show that prelamin A links DNA damage to VSMC mineralisation and suggests that age-related vascular calcification shares similar mechanisms to the accelerated VSMC ageing observed in patients with progeria syndromes.

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