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153 Relationship between micro RNA-145 and vascular smooth muscle cell senescence in diabetes
  1. Karen Hemmings1,
  2. Kirsten Riches-Suman2,
  3. Neil Turner2,
  4. Porter Karen2,
  5. David O’Regan3
  1. 1University of Leeds, LIGHT Laboratories, Clarendon Way, Leeds, UK
  2. 2University of Leeds
  3. 3Leeds Teaching Hospitals

Abstract

Introduction Saphenous vein (SV) graft failure after coronary bypass surgery is a significant problem in Type 2 diabetic (T2DM) patients, characterised by aberrant smooth muscle cell (SMC) remodelling. SV-SMC cultured from diabetic patients express elevated levels of microRNA-145, exhibit distinct morphology, impaired proliferation and DNA damage. This study explored a potential relationship between miR-145, DNA damage response signalling (DDR) and SV-SMC senescence.

Methods Using native human non-diabetic (ND) and T2DM-SMC, DDR signalling proteins (ATM, ATR, p21) were measured (real-time PCR) and subsequently quantified in ND cells overexpressing miR-145 (premiR-145 transfection). Conditioned medium (CM) from miR-145 overexpressing cells was analysed by ELISA for indicators of a senescence associated secretory phenotype (SASP) and to explore intracellular signalling (immunoblotting) in naïve ND-SMC. The effect of siRNA knockdown or pharmacological inhibition of ATM/ATR on cell proliferation was also explored (cell counting).

Results Increased expression of ATM, ATR and p21 was observed in T2DM-SMC relative to ND-SMC (n=8, p<0.05). MiR-145 overexpression increased ATR (n=7, p<0.01), and a trend to increased p21 was observed. Secretion of IL-6, IL-8 and MCP-1 was increased by 1.5, 2.0 and 5.5-fold, respectively (n=8, p<0.01) in miR-145-overexpressing cells; this CM induced persistent (≥96h) p38α phosphorylation in naïve cells. Pharmacological inhibition or gene-silencing of ATR, but not ATM significantly inhibited cell proliferation (n=4, p<0.05).

Conclusion Aberrant expression of DDR proteins in T2DM-SMC can be mimicked, at least partially by miR-145 overexpression. MiR-145 drives a SASP, resulting in chronic p-38 phosphorylation. MiR-145 holds potential as a therapeutic target to ameliorate SMC senescence.

  • vascular smooth muscle cell
  • diabetes
  • senescence

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