Epigenetics in Stroke Recovery

While the death rate from stroke has continually decreased due to interventions in the hyperacute stage of the disease, long-term disability and institutionalization have become common sequelae in the aftermath of stroke. Therefore, identification of new molecular pathways that could be targeted to improve neurological recovery among survivors of stroke is crucial. Epigenetic mechanisms such as post-translational modifications of histone proteins and microRNAs have recently emerged as key regulators of the enhanced plasticity observed during repair processes after stroke. In this review, we highlight the recent advancements in the evolving field of epigenetics in stroke recovery

MicroRNAs in the axon locally mediate the effects of chondroitin sulfate proteoglycans and cGMP on axonal growth

Axonal miRNAs locally regulate axonal growth by modulating local protein composition. Whether localized miRNAs in the axon mediate the inhibitory effect of Chondroitin sulfate proteoglycans (CSPGs) on the axon remains unknown. We showed that in cultured cortical neurons, axonal application of CSPGs inhibited axonal growth and altered axonal miRNA profiles, whereas elevation of axonal cyclic guanosine monophosphate (cGMP) levels by axonal application of sildenafil reversed the effect of CSPGs on inhibition of axonal growth and on miRNA profiles. Specifically, CSPGs elevated and reduced axonal levels of miR-29c and integrin β1 (ITGB1) proteins, respectively, while elevation of cGMP levels overcame these CSPG effects. Gain-of- and loss-of-function experiments demonstrated that miR-29c in the distal axon mediates axonal growth downstream of CSPGs and cGMP by regulating axonal protein levels of ITGB1, FAK, and RhoA. Together, our data demonstrate that axonal miRNAs play an important role in mediating the inhibitory action of CSPGs on axonal growth and that miR-29c at least partially mediates this process

Class IIa histone deacetylases affect neuronal remodeling and functional outcome after stroke

We have previously demonstrated that stroke induces nuclear shuttling of class IIa histone deacetylase 4 (HDAC4). Stroke-induced nuclear shuttling of HDAC4 is positively and significantly correlated with improved indices of neuronal remodeling in the peri-infarct cortex. In this study, using a rat model for middle cerebral artery occlusion (MCAO), we tested the effects of selective inhibition of class IIa HDACs on functional recovery and neuronal remodeling when administered 24hr after stroke. Adult male Wistar rats (n = 15-17/group) were subjected to 2 h MCAO and orally gavaged with MC1568 (a selective class IIa HDAC inhibitor), SAHA (a non-selective HDAC inhibitor), or vehicle-control for 7 days starting 24 h after MCAO. A battery of behavioral tests was performed. Lesion volume measurement and immunohistochemistry were performed 28 days after MCAO. We found that stroke increased total HDAC activity in the ipsilateral hemisphere compared to the contralateral hemisphere. Stroke-increased HDAC activity was significantly decreased by the administration of SAHA as well as by MC1568. However, SAHA significantly improved functional outcome compared to vehicle control, whereas selective class IIa inhibition with MC1568 increased mortality and lesion volume and did not improve functional outcome. In addition, MC1568 decreased microtubule associated protein 2 (MAP2, dendrites), phosphorylated neurofilament heavy chain (pNFH, axons) and myelin basic protein (MBP, myelination) immunoreactivity in the peri-infarct cortex. Quantitative RT-PCR of cortical neurons isolated by laser capture microdissection revealed that MC1568, but not SAHA, downregulated CREB and c-fos expression. Additionally, MC1568 decreased the expression of phosphorylated CREB (active) in neurons. Taken together, these findings demonstrate that selective inhibition of class IIa HDACs impairs neuronal remodeling and neurological outcome. Inactivation of CREB and c-fos by MC1568 likely contributes to this detrimental effect

MicroRNA-146a Promotes Oligodendrogenesis in Stroke

Stroke induces new myelinating oligodendrocytes that are involved in ischemic brain repair. Molecular mechanisms that regulate oligodendrogenesis have not been fully investigated. MicroRNAs (miRNAs) are small non-coding RNA molecules that post-transcriptionally regulate gene expression. MiR-146a has been reported to regulate immune response, but the role of miR-146a in oligodendrocyte progenitor cells (OPCs) remains unknown. Adult Wistar rats were subjected to the right middle cerebral artery occlusion (MCAo). In situ hybridization analysis with LNA probes against miR-146a revealed that stroke considerably increased miR-146a density in the corpus callosum and subventricular zone (SVZ) of the lateral ventricle of the ischemic hemisphere. In vitro, overexpression of miR-146a in neural progenitor cells (NPCs) significantly increased their differentiation into O4+ OPCs. Overexpression of miR-146a in primary OPCs increased their expression of myelin proteins, whereas attenuation of endogenous miR-146a suppressed generation of myelin proteins. MiR-146a also inversely regulated its target gene-IRAK1 expression in OPCs. Attenuation of IRAK1 in OPCs substantially increased myelin proteins and decreased OPC apoptosis. Collectively, our data suggest that miR-146a may mediate stroke-induced oligodendrogenesis

MicroRNA profiling in subventricular zone after stroke: MiR-124a regulates proliferation of neural progenitor cells through Notch signaling pathway.

The Notch signaling pathway regulates adult neurogenesis under physiological and pathophysiological conditions. MicroRNAs are small non-coding RNA molecules that regulate gene expression. The present study investigated the effect of miR-124a on the Notch signaling pathway in stroke-induced neurogenesis.We found that adult rats subjected to focal cerebral ischemia exhibited substantial reduction of miR-124a expression, a neuron specific miRNA, in the neural progenitor cells of the subventricular zone (SVZ) of the lateral ventricle, which was inversely associated with activation of Notch signals. In vitro, transfection of neural progenitor cells harvested from the SVZ of adult rat with miR-124a repressed Jagged-1 (JAG1), a ligand of Notch, in a luciferase construct containing the JAG1 target site. Introduction of miR-124a in neural progenitor cells significantly reduced JAG1 transcript and protein levels, leading to inactivation of Notch signals. Transfection of neural progenitor cells with miR-124a significantly reduced progenitor cell proliferation and promoted neuronal differentiation measured by an increase in the number of Doublecortin positive cells, a marker of neuroblasts. Furthermore, introduction of miR-124a significantly increased p27Kip1 mRNA and protein levels, a downstream target gene of the Notch signaling pathway.Collectively, our study demonstrated that in vivo, stroke alters miRNA expression in SVZ neural progenitor cells and that in vitro, miR-124a mediates stroke-induced neurogenesis by targeting the JAG-Notch signaling pathway