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Ephrins and Eph receptor(s) have recently been implicated in regulating neurogenesis in the adult subventricular zone (SVZ) and rostral migratory stream. Here, we examined the role of ephrinB3-EphB3 signaling in mediating the SVZ response to traumatic brain injury (TBI). Analysis of EphB3 expression showed colocalization with glial fibrillary acidic protein-positive neural stem progenitor cells (NSPCs) and doublecortin-positive neuroblasts, whereas ephrinB3 was expressed outside the neurogenic region. TBI resulted in a significant reduction in EphB3 expression, which coincided with enhanced NSPC survival and proliferation at 3 and 7 days postinjury. Analysis of mice lacking either ephrinB3 (ephrinB3 2/2 ) or EphB3 (EphB3 2/2 ) showed a significant increase in bromodeoxyuridine (BrdU) incorporation and Ki67 immunoreactivity in the SVZ. Interestingly, cell death was dissimilar between knockout mice, where cell death was reduced in EphB3 2/2 but increased in ephrinB3 2/2 mice. Lateral ventricle infusion of soluble preclustered ephrinB3-Fc reversed the proliferative and cell death defects in ephrinB3 2/2 but not EphB3 2/2 mice and prevented TBIinduced proliferation in wild-type NSPCs. Coincidently, tumor suppressor p53 expression was increased following EphB3 stimulation and is reduced in the absence of either EphB3 or ephrinB3. Furthermore, pharmacological inhibition and siRNA knockdown of p53-attenuated ephrinB3-Fcmediated growth suppression while having no effect on cell death in cultured NSPCs. These data demonstrate that EphB3 signaling suppresses NSPC proliferation in a p53-dependent manner, induces cell death in the absence of ligand stimulation and is transiently reduced in the SVZ to initiate the expansion and survival of endogenous adult NSPCs following TBI.

Carbonyl sulfide (COS) is a gas that may play important roles in mammalian and bacterial biology, but its study is limited by a lack of suitable donor molecules. We report here the use of N-thiocarboxyanhydrides (NTAs) as COS donors that release the gas in a sustained manner under biologically relevant conditions with innocuous peptide byproducts. Carbonic anhydrase converts COS into H2S, allowing NTAs to serve as either COS or H2S donors, depending on the availability of the enzyme. Analysis of the pseudo-first-order H2S release rate under biologically relevant conditions revealed a release half-life of 75 min for the small molecule NTA under investigation. A polynorbornene bearing pendant NTAs made by ring-opening metathesis polymerization was also synthesized to generate a polymeric COS/H2S donor. A half-life of 280 min was measured for the polymeric donor. Endothelial cell proliferation studies revealed an enhanced rate of proliferation for cells treated with the NTA over untreated controls.

Embryonic stem (ES) cells represent an ideal source for cell engraftment in the damaged central nervous system (CNS). Understanding key signals that control ES cell differentiation may improve cell type-specific differentiation that is suitable for transplantation therapy. We tested the hypothesis that extracellular signalregulated kinase (ERK) 1/2 phosphorylation is an early signaling event required for the neuronal differentiation of ES cells. Cultured mouse ES cells were treated with an all-trans-retinoic-acid (RA) protocol to generate neurally induced progenitor cells. Western blot analysis showed a dramatic increase in ERK 1/2 phosphorylation (p-ERK 1/2) 1-5 days after RA induction, which was attenuated in the presence of the p-ERK 1/2-specific inhibitor UO126. Phospho-ERK 1/2 inhibition significantly reduced the number of NeuN-positive cells and the expression of associated cytoskeletal proteins. In differentiating ES cells, there was increased nuclear translocation of STAT3 and decreased protein expression levels of GDNF, BDNF and NGF. STAT3 translocation was attenuated by UO126. Finally, caspase-3 activation was observed in the presence of UO126, suggesting that the ERK pathway also contributes to the survival of differentiating ES cells. These data indicate that ERK 1/2 phosphorylation is a key event required for early neuronal differentiation and survival of ES cells.

Life experience can leave lasting marks, such as epigenetic changes, in the brain. How life experience is translated into storable epigenetic information remains largely unknown. With unbiased data-driven approaches, we predicted that Egr1 , a transcription factor important for memory formation, plays an essential role in brain epigenetic programming. We performed EGR1 ChIP-seq and validated thousands of EGR1 binding sites with methylation patterns established during postnatal brain development. More specifically, these EGR1 binding sites become hypomethylated in mature neurons but remain heavily methylated in glia. We further demonstrated that EGR1 recruits a DNA demethylase TET1 to remove the methylation marks and activate downstream genes. The frontal cortices from the knockout mice lacking Egr1 or Tet1 share strikingly similar profiles in both gene expression and DNA methylation. In summary, our study reveals EGR1 programs the brain methylome together with TET1 providing new insight into how life experience may shape the brain methylome.

Traumatic brain injuries (TBIs) account for the majority of injury-related deaths in the United States with roughly two million TBIs occurring annually. Due to the spectrum of severity and heterogeneity in TBIs, investigation into the secondary injury is necessary in order to formulate an effective treatment. A mechanical consequence of trauma involves dysregulation of the blood–brain barrier (BBB) which contributes to secondary injury and exposure of peripheral components to the brain parenchyma. Recent studies have shed light on the mechanisms of BBB breakdown in TBI including novel intracellular signaling and cell–cell interactions within the BBB niche. The current review provides an overview of the BBB, novel detection methods for disruption, and the cellular and molecular mechanisms implicated in regulating its stability following TBI.

Eph receptor tyrosine kinases and their membrane-bound ligands, ephrins, have a variety of roles in the developing and adult central nervous system that require direct cell–cell interactions; including regulating axon path finding, cell proliferation, migration and synaptic plasticity. Recently, we identified a novel pro-survival role for ephrins in the adult subventricular zone, where ephrinB3 blocks Eph-mediated cell death during adult neurogenesis. Here, we examined whether EphB3 mediates cell death in the adult forebrain following traumatic brain injury and whether ephrinB3 infusion could limit this effect. We show that EphB3 co-labels with microtubule-associated protein 2-positive neurons in the adult cortex and is closely associated with ephrinB3 ligand, which is reduced following controlled cortical impact (CCI) injury. In the complete absence of EphB3 (EphB3−/−), we observed reduced terminal deoxynucleotidyl transferase-dUTP nick end labeling (TUNEL), and functional improvements in motor deficits after CCI injury as compared with wild-type and ephrinB3−/− mice. We also demonstrated that EphB3 exhibits dependence receptor characteristics as it is cleaved by caspases and induces cell death, which is not observed in the presence of ephrinB3. Following trauma, infusion of pre-clustered ephrinB3-Fc molecules (eB3-Fc) into the contralateral ventricle reduced cortical infarct volume and TUNEL staining in the cortex, dentate gyrus and CA3 hippocampus of wild-type and ephrinB3−/− mice, but not EphB3−/− mice. Similarly, application of eB3-Fc improved motor functions after CCI injury. We conclude that EphB3 mediates cell death in the adult cortex through a novel dependence receptor-mediated cell death mechanism in the injured adult cortex and is attenuated following ephrinB3 stimulation.

BackgroundThe continuum of pro- and anti-inflammatory response elicited by traumatic brain injury (TBI) is suggested to play a key role in the outcome of TBI; however, the underlying mechanisms remain ill -defined.MethodsHere, we demonstrate that using bone marrow chimeric mice and systemic inhibition of EphA4 receptor shifts the pro-inflammatory milieu to pro-resolving following acute TBI.ResultsEphA4 expression is increased in the injured cortex as early as 2 h post-TBI and on CX3CR1gfp-positive cells in the peri-lesion. Systemic inhibition or genetic deletion of EphA4 significantly reduced cortical lesion volume and shifted the inflammatory profile of peripheral-derived immune cells to pro-resolving in the damaged cortex. These findings were consistent with in vitro studies showing EphA4 inhibition or deletion altered the inflammatory state of LPS-stimulated monocyte/macrophages towards anti-inflammatory. Phosphoarray analysis revealed that EphA4 may regulate pro-inflammatory gene expression by suppressing the mTOR, Akt, and NF-κB pathways. Our human metadata analysis further demonstrates increased EPHA4 and pro-inflammatory gene expression, which correlates with reduced AKT concurrent with increased brain injury severity in patients.ConclusionsOverall, these findings implicate EphA4 as a novel mediator of cortical tissue damage and neuroinflammation following TBI.