title: Targeted acetylation of NF-kappaB/RelA and histones by epigenetic drugs reduces post-ischemic brain injury in mice with an extended therapeutic window.
abstract: Nuclear factor-kappaB (NF-κB) p50/RelA is a key molecule with a dual effect in the progression of ischemic stroke. In harmful ischemia, but not in preconditioning insult, neurotoxic activation of p50/RelA is characterized by RelA-specific acetylation at Lys310 (K310) and deacetylation at other Lys residues. The derangement of RelA acetylation is associated with activation of Bim promoter.
  
  
  OBJECTIVE
  With the aim of producing neuroprotection by correcting altered acetylation of RelA in brain ischemia, we combined the pharmacological inhibition of histone deacetylase (HDAC) 1-3, the enzymes known to reduce global RelA acetylation, and the activation of sirtuin 1, endowed with a specific deacetylase activity on the K310 residue of RelA. To afford this aim, we tested the clinically used HDAC 1-3 inhibitor entinostat (MS-275) and the sirtuin 1 activator resveratrol.
  
  
  METHODS
  We used the mouse model of transient middle cerebral artery occlusion (MCAO) and primary cortical neurons exposed to oxygen glucose deprivation (OGD).
  
  
  RESULTS
  The combined use of MS-275 and resveratrol, by restoring normal RelA acetylation, elicited a synergistic neuroprotection in neurons exposed to OGD. This effect correlated with MS-275 capability to increase total RelA acetylation and resveratrol capability to reduce RelA K310 acetylation through the activation of an AMP-activated protein kinase-sirtuin 1 pathway. The synergistic treatment reproduced the acetylation state of RelA peculiar of preconditioning ischemia. Neurons exposed to the combined drugs totally recovered the optimal histone H3 acetylation. Neuroprotection was reproduced in mice subjected to MCAO and treated with MS-275 (20μg/kg and 200μg/kg) or resveratrol (6800μg/kg) individually. However, the administration of lowest doses of MS-275 (2μg/kg) and resveratrol (68μg/kg) synergistically reduced infarct volume and neurological deficits. Importantly, the treatment was effective even when administered 7h after the stroke onset. Chromatin immunoprecipitation analysis of cortices harvested from treated mice showed that the RelA binding and histone acetylation increased at the Bcl-xL promoter and decreased at the Bim promoter.
  
  
  CONCLUSIONS
  Our study reveals that epigenetic therapy shaping acetylation of both RelA and histones may be a promising strategy to limit post-ischemic injury with an extended therapeutic window.
authors.affiliation: Division of Pharmacology and Experimental Therapeutics, Department of Biomedical Sciences & Biotechnologies and National Institute of Neuroscience, School of Medicine, University of Brescia, Italy.
authors.forename: Annamaria
authors.initials: A
authors.lastname: Lanzillotta
authors.affiliation: Division of Pharmacology, Department of Neuroscience and National Institute of Neuroscience, School of Medicine, Federico II University of Naples, Italy.
authors.forename: Giuseppe
authors.initials: G
authors.lastname: Pignataro
authors.affiliation: Division of Pharmacology and Experimental Therapeutics, Department of Biomedical Sciences & Biotechnologies and National Institute of Neuroscience, School of Medicine, University of Brescia, Italy.
authors.forename: Caterina
authors.initials: C
authors.lastname: Branca
authors.affiliation: Division of Pharmacology, Department of Neuroscience and National Institute of Neuroscience, School of Medicine, Federico II University of Naples, Italy.
authors.forename: Ornella
authors.initials: O
authors.lastname: Cuomo
authors.affiliation: Division of Pharmacology and Experimental Therapeutics, Department of Biomedical Sciences & Biotechnologies and National Institute of Neuroscience, School of Medicine, University of Brescia, Italy.
authors.forename: Ilenia
authors.initials: I
authors.lastname: Sarnico
authors.affiliation: Division of Pharmacology and Experimental Therapeutics, Department of Biomedical Sciences & Biotechnologies and National Institute of Neuroscience, School of Medicine, University of Brescia, Italy.
authors.forename: Marina
authors.initials: M
authors.lastname: Benarese
authors.affiliation: Division of Pharmacology, Department of Neuroscience and National Institute of Neuroscience, School of Medicine, Federico II University of Naples, Italy; IRCCS SDN, Naples, Italy.
authors.forename: Lucio
authors.initials: L
authors.lastname: Annunziato
authors.affiliation: Division of Pharmacology and Experimental Therapeutics, Department of Biomedical Sciences & Biotechnologies and National Institute of Neuroscience, School of Medicine, University of Brescia, Italy; IRCCS, S. Camillo Hospital, Venice, Italy.
authors.forename: PierFranco
authors.initials: P
authors.lastname: Spano
authors.affiliation: Division of Pharmacology and Experimental Therapeutics, Department of Biomedical Sciences & Biotechnologies and National Institute of Neuroscience, School of Medicine, University of Brescia, Italy; IRCCS, S. Camillo Hospital, Venice, Italy. Electronic address: pizzi@med.unibs.it.
authors.forename: Marina
authors.initials: M
authors.lastname: Pizzi
mesh_terms: D000107:Acetylation / Q000187:drug effects; D000818:Animals; D001549:Benzamides / Q000494:pharmacology; D001921:Brain / Q000187:drug effects / Q000378:metabolism / Q000473:pathology; D002545:Brain Ischemia / Q000188:drug therapy* / Q000378:metabolism / Q000473:pathology; D015687:Cell Hypoxia / Q000187:drug effects / Q000502:physiology; D004195:Disease Models, Animal; D044127:Epigenesis, Genetic / Q000187:drug effects* / Q000502:physiology; D005947:Glucose / Q000172:deficiency; D056572:Histone Deacetylase Inhibitors / Q000494:pharmacology; D006657:Histones / Q000378:metabolism*; D020244:Infarction, Middle Cerebral Artery; D008297:Male; D008810:Mice, Inbred C57BL; D016328:NF-kappa B / Q000378:metabolism*; D009474:Neurons / Q000378:metabolism / Q000473:pathology; D018696:Neuroprotective Agents / Q000494:pharmacology*; D011725:Pyridines / Q000494:pharmacology; D000077185:Resveratrol; D056564:Sirtuin 1 / Q000378:metabolism; D013267:Stilbenes / Q000494:pharmacology; D051996:Transcription Factor RelA / Q000378:metabolism*
chemical_list: D001549:Benzamides; D056572:Histone Deacetylase Inhibitors; D006657:Histones; D016328:NF-kappa B; D018696:Neuroprotective Agents; D011725:Pyridines; C498144:Rela protein, mouse; D013267:Stilbenes; D051996:Transcription Factor RelA; C118739:entinostat; C470545:Sirt1 protein, mouse; D056564:Sirtuin 1; D005947:Glucose; D000077185:Resveratrol