Histone Deacetylase 3 Inhibition Decreases Cerebral Edema and Protects the Blood-Brain Barrier after Stroke

Abstract Introduction: We have previously shown that selective inhibition of histone deacetylase 3 (HDAC3) decreases infarct volume and improves long-term functional outcomes after stroke. In this study, we examined the effects of HDAC3 inhibition on cerebral edema and blood-brain barrier (BBB) leakage and explored its underlying mechanisms.Methods: Adult male Wistar rats were subjected to 2-h middle cerebral artery occlusion (MCAO) and randomly treated i.p. with either vehicle or a selective HDAC3 inhibitor (RGFP966) at 2 and 24 hours after stroke. Modified neurological severity scores (mNSS) were calculated at 2 hours, 1 day, and 3 days. H&E and Evans blue dye (EBD) assay were employed to assess cerebral edema and BBB leakage, respectively. Western Blot for matrix metalloproteinase-9 (MMP9) and immunostaining for HDAC3, GFAP, Iba-1, albumin, aquaporin-4, claudin-5, ZO-1, and NF-kappa were performed. Results: Early RGFP966 administration decreased cerebral edema (p=0.002) and BBB leakage, as measured by EBD assay and albumin extravasation (p=0.001). RGFP966 significantly increased the expression of tight junction proteins (claudin-5 and ZO-1) in the peri-infarct area. RGFP966 also significantly decreased HDAC3 expression in GFAP+ astrocytes, which correlated with better mNSS (r=0.67, p=0.03) and decreased cerebral edema (r=0.64, p=0.04). RGFP966 decreased the expression of aquaporin-4 in GFAP+ astrocytes (p=0.002), as well as, the inflammatory markers Iba-1, NF-kappa, and MMP9 in the ischemic brain (p<0.05).Conclusions: Early HDAC3 inhibition decreases cerebral edema and BBB leakage. BBB protection by RGFP966 is mediated in part by the upregulation of tight junction proteins, downregulation of aquaporin-4 and HDAC3 in astrocytes, and decreased neuroinflammation.