Abstract T P209: SIRT3 is Upregulated During Stroke and Contributes to Ischemic Damage by Depressing the Transcription of Nuclear Genes
Introduction: The impairment of energy metabolism during stroke is a major contributor to ensuing tissue damage and death. The NAD+-dependent histone deacetylase SIRT3 has been identified as a key regulator of mitochondrial energy metabolism. It has also been found to repress the transcription of nuclear genes via histone deacetylation. This study aimed to elucidate the function of nuclear SIRT3 in ischemic stroke.
Methods: Focal cerebral ischemia was induced by transient right middle cerebral artery occlusion (MCAO) for 90min followed by 72hrs of reperfusion. Infarct area was quantified with Sigma ScanPro5 on 30μm coronal sections stained with Cresyl Violet. Expression of SIRT3 and acetylated lysine proteins was examined by immunohistochemistry (IHC) and western blotting with anti-SIRT3 and anti-acetylated lysine antibodies. NAD+ levels were measured with a colorimetric kit.
Results: SIRT3 expression was increased in the ischemic hemisphere of wild-type (WT) mice at 72hrs. Higher magnification of the IHC image demonstrated localization of SIRT3 to both the nucleus and mitochondria of labeled cells. Subcellular fractionations confirmed this localization, and revealed a dramatic increase in nuclear translocation of SIRT3 with ischemia. With the same duration of MCAO and reperfusion, SIRT3 knockout (KO) mice displayed a reduction in striatal infarct from 60.6±7.2% to 24.9±9.7% (P<0.05), as well as reduced cortical and hemispheric infarct compared to WT mice. Western blotting of whole cell lysate from WT and KO mice following stroke revealed increased cellular protein acetylation in KO mice. KO mice displayed higher intra-ischemic NAD+ levels than WT mice.
Conclusions: Our findings suggest that SIRT3 contributes to brain injury during stroke via ischemia-induced upregulation and translocation to the nucleus, where it deacetylates both histone and non-histone proteins and represses nuclear gene transcription. Consistent with the neuroprotection afforded by the protein’s absence in KO mice, the inhibition of SIRT3 may serve as a potential therapeutic strategy for minimizing stroke damage.
Author Disclosures: J. Crapser: None. R. Verma: None. L. McCullough: None.
This research has received full or partial funding support from the American Heart Association, Founders - Connecticut, Maine, Massachusetts, New Hampshire, New Jersey, New York, Rhode Island, Vermont.
- © 2014 by American Heart Association, Inc.