Abstract 18846: The Dual Effects of Shear Stress on Vascular Endothelial Autophagy and Mitochondrial DNA Damage
Vascular oxidative stress preferentially develops at arterial curvatures and branching points where oscillatory shear stress (OSS) occurs. Autophagy, a tightly regulated intracellular bulk degradation/recycling system, is implicated in endothelial homeostasis. In this study, we assessed the effects of OSS on autophagy and mitochondrial DNA (mtDNA) damage. Human aortic endothelial cells (HAEC) exposed to OSS (τavg= 0±3 dyne/cm2 for 4 hours) increased relative LC3-II/LC3-I ratios (control: 1.0±0.02, OSS: 2.4±0.51, p<0.01, n=4), as supported by an increase in autophagosome punctures by LC3-green fluorescence protein (LC3-GFP). OSS-induced increase in LC3-II/LC3-I ratios were significantly attenuated in response to N-acetyl cysteine (NAC), MnSOD over-expression, and JNK siRNA (siJNK) knockdown (p<0.05, n=4). Inhibition of autophagy by ATG5 siRNA increased OSS-induced MitoSox intensity for mitochondrial superoxide (mtO2•-) by 2.03±0.25-fold (p< 0.05, n=4), whereas Rapamycin-induced autophagy reduced OSS induced MitoSox intensity by 78.4% (p<0.05, n=4). OSS-induced reduction of mitochondrial complex II activity was also attenuated in response to Rapamycin (OSS: 62.1±11.2%; Rapamycin+OSS: 130.1±40.4% (relative to static control), p<0.05, n=4). In comparison with static and pulsatile flow (PSS=23 dyne/cm2) conditions, OSS also significantly increased mtDNA damage by 4.5±1.3-fold (p<0.01, n=4), which was significantly reversed in response to NAC, MnSOD over-expression, and siJNK knockdown (p<0.05, n=4). In the fat-fed New Zealand White rabbits, quantification for immuno-staining revealed prominent endothelial JNK phosphorylation and DNA damage in the OSS-exposed aortic arch in contrast with nearly absent staining in the PSS-exposed descending aorta (p< 0.05, n=4). Furthermore, analysis of caspase-3 activity was negative for OSS-exposed HAEC and OSS-exposed aorta area from normal diet-fed animals. Despite atherogenic nature of OSS, our findings suggest that OSS imparts dual effects in endothelial autophagy and mtDNA damage with an implication in maintaining homeostasis.
- © 2013 by American Heart Association, Inc.