The p70-S6K inhibitor LY2584702 reverses oxidative stress-induced senescence in small airway epithelial cells.

Background: Overactivation of the mammalian target of rapamycin (mTOR) pathway induced by oxidative stress leads to downregulation of the anti-ageing protein sirtuin-1 (SIRT1), resulting in cellular senescence similar to that seen in COPD. P70 S6 kinase (p70-S6K) acts downstream of mTOR to regulate the cell cycle and DNA damage response. Inhibiting p70-S6K may be able to reverse cell cycle arrest and senescence and identify a therapeutic target for diseases of lung ageing. Aim: To examine the effect of inhibiting p70-S6K on SIRT1 and markers of senescence in H₂O₂-treated small airway epithelial cells (SAECs). Methods: SAECs isolated from non-smokers were cultured with or without 300µM H₂O₂ for 48h, followed by treatment with 10^-12-10^-5M LY2584702 (p70-S6K inhibitor) or 0.1% DMSO for 24h. Protein expression of SIRT1 was quantified using Western blot. Senescence associated-β-Galactosidase staining was used to determine senescent cells and ELISA for CXCL8 release. Results: H₂O₂ treatment significantly reduced SIRT1 protein (65%, p<0.01, n=3), increased CXCL8 (75%, p<0.01, n=5) and SA-β-Gal staining (51%, p<0.05, n=3). LY2584702 exhibited a concentration dependent effect, 10^-5M treatment increased SIRT1 protein expression (70%, p<0.01, n=3), reduced CXCL8 protein release (57%, p<0.01, n=3) and SA-β-Gal staining (30%, p<0.01, n=3). Conclusion: Inhibition of p70-S6K reverses oxidative stress-induced senescence in SAECs. Further investigation into the mechanism of p70-S6K in regulating SIRT1 may help identify further drug targets to combat cellular senescence in COPD.