miRNA-192-5p targets Dyrk1a to attenuate cerebral injury in MCAO mice by suppressing neuronal apoptosis and neuroinflammation.

Abstract

Introduction: Ischemic stroke (IS) is a leading cause of disability and mortality worldwide. Several studies have demonstrated the involvement of microRNAs (miRNAs) in brain diseases. miRNA-192-5p is a regulatory molecule in neurodegenerative diseases and its expression was found to be significantly downregulated in the whole blood samples of IS patients, but the specific role of miRNA-192-5p in IS not fully understood. Here, we investigated the role of miRNA-192-5p in a murine model of acute cerebral injury after IS.

Material and methods: Male C57BL/6J mice received an intracerebroventricular (i.c.v.) injection of agomir-192-5p or antagomir-192-5p 2 h before middle cerebral artery occlusion (MCAO). Infarct volume was assessed by 2,3,5 triphenyltetrazolium chloride (TTC) staining. Brain slices were subjected to Fluoro-Jade B, TUNEL, and immunofluorescence stainings. Contents of pro-inflammatory cytokines (TNF-α, IL-1β, and IL-6) were measured using enzyme-linked immunosorbent assay (ELISA) kits. In vitro, murine microglial BV-2 cells were subjected to oxygen-glucose deprivation (OGD), and the contents of pro-inflammatory cytokines were measured in cell lysates.

Results: miRNA-192-5p was downregulated in the ischemic penumbra of the cerebral cortex. Pretreatment with agomir-192-5p attenuated neurological deficits and reduced cerebral edema and infarct volume in MCAO mice. Agomir-192-5p-treated animals had fewer degenerating and apoptotic neurons in the ischemic penumbra. Additionally, agomir-192-5p significantly suppressed neuroinflammation as evidenced by decreased immunostaining for GFAP and Iba1 and decreased levels of pro-inflammatory cytokines. Antagomir-192-5p pretreatment showed the opposite effect. Furthermore, dual specificity tyrosine phosphorylation regulated kinase 1A (Dyrk1a) was identified as a target gene of miRNA-192-5p, and the elevated Dyrk1a expression in the ischemic penumbra was markedly reduced by agomir-192-5p. Dyrk1a overexpression in BV-2 microglial cells impaired miRNA-192-5p-mediated inhibition of OGD-induced activation of BV-2 microglial cells. Opposite results were obtained using miRNA-192-5p inhibitor and Dyrk1a siRNA.

Conclusions: We found that intracerebroventricular administration of miRNA-192-5p before MCAO attenuatedacute cerebral injury by suppressing neuronal apoptosis and neuroinflammation in mice, and these protective effects might be mediated by downregulation of Dyrk1a. This study would help identify novel therapeutic targets for IS.