Overexpression of miR-124 Protects Against Neurological Dysfunction Induced by Neonatal Hypoxic-Ischemic Brain Injury.

Neonatal hypoxic-ischemic encephalopathy (HIE) is a major cause of lifelong disabilities worldwide, without effective therapies and clear regulatory mechanisms. MicroRNAs (miRNAs) act as a significant regulator in neuroregeneration and neuronal apoptosis, thus holding great potential as therapeutic targets in HIE. In this study, we established the hypoxia-ischemia (HI) model in vivo and oxygen-glucose deprivation (OGD) model in vitro. Zea-longa score and magnetic resonance imaging were applied to verify HI-induced neuronal dysfunction and brain infarction. Subsequently, a miRNA microarray analysis was employed to profile miRNA transcriptomes. Down-regulated miR-124 was found 24 h after HIE, which corresponded to the change in PC12, SHSY5Y, and neurons after OGD. To determine the function of miR-124, mimics and lentivirus-mediated overexpression were used to regulate miR-124 in vivo and in vitro, respectively. Our results showed that miR-124 overexpression obviously promoted cell survival and suppressed neuronal apoptosis. Further, the memory and neurological function of rats was also obviously improved at 1 and 2 months after HI, indicated by the neurological severity score, Y-maze test, open field test, and rotating rod test. Our findings showed that overexpression of miR-124 can be a promising new strategy for HIE therapy in future clinical practice.

A non-canonical rhodopsin-mediated insulin receptor signaling pathway in retinal photoreceptor neurons.

We previously reported a ligand-independent and rhodopsin-dependent insulin receptor (IR) neuroprotective signaling pathway in both rod and cone photoreceptor cells, which is activated through protein-protein interaction. Our previous studies were performed with either retina or isolated rod or cone outer segment preparations and the expression of IR signaling proteins were examined. The isolation of outer segments with large portions of the attached inner segments is a technical challenge. Optiprep™ density gradient medium has been used to isolate the cells and subcellular organelles, Optiprep™ is a non-ionic iodixanol-based medium with a density of 1.320 g/mL. We employed this method to examine the expression of IR and its signaling proteins, and activation of one of the downstream effectors of the IR in isolated photoreceptor cells. Identification of the signaling complexes will be helpful for therapeutic targeting in disease conditions.

Effect of the pituitary adenylate cyclase-activating polypeptide on the autophagic activation observed in in vitro and in vivo models of Parkinson's disease.

Parkinson's disease (PD) is a neurodegenerative disorder that leads to destruction of the midbrain dopaminergic (DA) neurons. This phenomenon is related to apoptosis and its activation can be blocked by the pituitary adenylate cyclase-activating polypeptide (PACAP). Growing evidence indicates that autophagy, a self-degradation activity that cleans up the cell, is induced during the course of neurodegenerative diseases. However, the role of autophagy in the pathogenesis of neuronal disorders is yet poorly understood and the potential ability of PACAP to modulate the related autophagic activation has never been significantly investigated. Hence, we explored the putative autophagy-modulating properties of PACAP in in vitro and in vivo models of PD, using the neurotoxic agents 1-methyl-4-phenylpyridinium (MPP(+)) and 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), respectively, to trigger alterations of DA neurons. In both models, following the toxin exposure, PACAP reduced the autophagic activity as evaluated by the production of LC3 II, the modulation of the p62 protein levels, and the formation of autophagic vacuoles. The ability of PACAP to inhibit autophagy was also observed in an in vitro cell assay by the blocking of the p62-sequestration activity produced with the autophagy inducer rapamycin. Thus, the results demonstrated that autophagy is induced in PD experimental models and that PACAP exhibits not only anti-apoptotic but also anti-autophagic properties.

Generating level-dependent models of cervical and thoracic spinal cord injury: Exploring the interplay of neuroanatomy, physiology, and function.

The majority of spinal cord injuries (SCI) occur at the cervical level, which results in significant impairment. Neurologic level and severity of injury are primary endpoints in clinical trials; however, how level-specific damages relate to behavioural performance in cervical injury is incompletely understood. We hypothesized that ascending level of injury leads to worsening forelimb performance, and correlates with loss of neural tissue and muscle-specific neuron pools. A direct comparison of multiple models was made with injury realized at the C5, C6, C7 and T7 vertebral levels using clip compression with sham-operated controls. Animals were assessed for 10weeks post-injury with numerous (40) outcome measures, including: classic behavioural tests, CatWalk, non-invasive MRI, electrophysiology, histologic lesion morphometry, neuron counts, and motor compartment quantification, and multivariate statistics on the total dataset. Histologic staining and T1-weighted MR imaging revealed similar structural changes and distinct tissue loss with cystic cavitation across all injuries. Forelimb tests, including grip strength, F-WARP motor scale, Inclined Plane, and forelimb ladder walk, exhibited stratification between all groups and marked impairment with C5 and C6 injuries. Classic hindlimb tests including BBB, hindlimb ladder walk, bladder recovery, and mortality were not different between cervical and thoracic injuries. CatWalk multivariate gait analysis showed reciprocal and progressive changes forelimb and hindlimb function with ascending level of injury. Electrophysiology revealed poor forelimb axonal conduction in cervical C5 and C6 groups alone. The cervical enlargement (C5-T2) showed progressive ventral horn atrophy and loss of specific motor neuron populations with ascending injury. Multivariate statistics revealed a robust dataset, rank-order contribution of outcomes, and allowed prediction of injury level with single-level discrimination using forelimb performance and neuron counts. Level-dependent models were generated using clip-compression SCI, with marked and reliable differences in forelimb performance and specific neuron pool loss.

PI3K/Akt and ERK/MAPK Signaling Promote Different Aspects of Neuron Survival and Axonal Regrowth Following Rat Facial Nerve Axotomy.

The ERK/MAPK and PI3K/Akt signaling pathways play important role in neuronal survival and axonal regeneration after peripheral nerve injury. However, the relative importance and degree of functional overlap of the two pathways are still debated due to lack of in-vivo data. We used rats which underwent a facial nerve axotomy, and examined subsequent ERK/MAPK and PI3K/Akt signaling activity by quantifying phosphorylation of ERK and Akt. We also assessed the survival rate of facial neurons, number of regenerated axons, and the length of axonal regrowth in axotomized animals treated with an inhibitor of ERK/MAPK (U0126) or PI3K/Akt (LY294002) phosphorylation, or with vehicle. Axotomy increased phosphorylation of ERK and Akt in the facial nucleus 7 days after injury. The inhibition of ERK phosphorylation significantly reduced the length of regenerated axons, but not the other parameters. Inhibition of Akt phosphorylation significantly reduced the survival rate of facial neurons and the number of new axons, as well as the length of regenerated axons. The results indicate that facial nerve injury activates the ERK/MAPK and PI3K/Akt signaling pathways in the facial nerve nucleus and its axons. However, the pathways promoted aspects of regeneration with only slight overlap: PI3K/Akt signaling improved the survival of neurons, as well as axonal growth and branching, whereas ERK/MAPK signaling promoted only axonal extension.

The neurochemistry of peripheral nerve regeneration.

Peripheral nerve injuries (PNIs) can be most disabling, resulting in the loss of sensitivity, motor function and autonomic control in the involved anatomical segment. Although injured peripheral nerves are capable of regeneration, sub-optimal recovery of function is seen even with the best reconstruction. Distal axonal degeneration is an unavoidable consequence of PNI. There are currently few strategies aimed to maintain the distal pathway and/or target fidelity during regeneration across the zone of injury. The current state of the art approaches have been focussed on the site of nerve injury and not on their distal muscular targets or representative proximal cell bodies or central cortical regions. This is a comprehensive literature review of the neurochemistry of peripheral nerve regeneration and a state of the art analysis of experimental compounds (inorganic and organic agents) with demonstrated neurotherapeutic efficacy in improving cell body and neuron survival, reducing scar formation and maximising overall nerve regeneration.

Phosphorylation by PKA and Cdk5 Mediates the Early Effects of Synapsin III in Neuronal Morphological Maturation.

Synapsin III (SynIII) is a neuron-specific phosphoprotein that plays a unique role in neuronal development. SynIII is phosphorylated by cAMP-dependent protein kinase (PKA) at a highly conserved phosphorylation site and by cyclin-dependent kinase-5 (Cdk5) at a newly described site. Although SynIII is known to be involved in axon elongation in vitro, the role of its phosphorylation by PKA and Cdk5 in the modulation of this process is unknown. We expressed either wild-type (WT) or phosphorylation-site mutants of SynIII in primary SynIII knock-out (KO) mouse neurons at early stages of in vitro development. Whereas the neurite elongation phenotype of SynIII KO neurons was fully rescued by the expression of WT SynIII, the expression of nonphosphorylatable and pseudo-phosphorylated PKA mutants was ineffective. Also, the nonphosphorylatable Cdk5 mutant was unable to rescue the neurite elongation phenotype of SynIII KO neurons. By contrast, the pseudo-phosphorylated mutant rescued the delay in neuronal maturation and axonal elongation, revealing a Cdk5-dependent regulation of SynIII function. Interestingly, SynIII KO neurons also exhibited decreased survival that was fully rescued by the expression of WT SynIII, but not by its phosphorylation mutants, and was associated with increased activated caspase3 and altered tropomyosin receptor kinase B isoform expression. These results indicate that PKA and Cdk5 phosphorylation is required for the physiological action of SynIII on axon specification and neurite outgrowth and that the expression of a functional SynIII is crucial for cell survival. Significance statement: Synapsin III is an atypical member of the synapsin family of synaptic vesicle-associated phosphoproteins that is precociously expressed in neurons and is downregulated afterward. Although experimental evidence suggests a specific role for Synapsin III in neuronal development, the molecular mechanisms are still largely unknown. We found that Synapsin III plays a central role in early stages of neuronal development involving neuronal survival, polarization, and neuritic growth and that these effects are dependent on phosphorylation by cAMP-dependent protein kinase and cyclin-dependent protein kinase-5. These results explain the recently described neurodevelopmental defects in the migration and orientation of Synapsin III-depleted cortical neurons and support the potential association of Synapsin III with neurodevelopmental disorders such as schizophrenia.

Peroxisome proliferator-activated receptor-γ coactivator-1α mediates neuroprotection against excitotoxic brain injury in transgenic mice: role of mitochondria and X-linked inhibitor of apoptosis protein.

Peroxisome proliferator-activated receptor gamma coactivator-1α (PGC-1α) is a transcriptional coactivator involved in the regulation of mitochondrial biogenesis and cell defense. The functions of PGC-1α in physiology of brain mitochondria are, however, not fully understood. To address this we have studied wild-type and transgenic mice with a two-fold overexpression of PGC-1α in brain neurons. Data showed that the relative number and basal respiration of brain mitochondria were increased in PGC-1α transgenic mice compared with wild-type mitochondria. These changes occurred concomitantly with altered levels of proteins involved in oxidative phosphorylation (OXPHOS) as studied by proteomic analyses and immunoblottings. Cultured hippocampal neurons from PGC-1α transgenic mice were more resistant to cell degeneration induced by the glutamate receptor agonist kainic acid. In vivo kainic acid induced excitotoxic cell death in the hippocampus at 48 h in wild-type mice but significantly less so in PGC-1α transgenic mice. However, at later time points cell degeneration was also evident in the transgenic mouse hippocampus, indicating that PGC-1α overexpression can induce a delay in cell death. Immunoblotting showed that X-linked inhibitor of apoptosis protein (XIAP) was increased in PGC-1α transgenic hippocampus with no significant changes in Bcl-2 or Bcl-X. Collectively, these results show that PGC-1α overexpression contributes to enhanced neuronal viability by stimulating mitochondria number and respiration and increasing levels of OXPHOS proteins and the anti-apoptotic protein XIAP.

Safranal acts as a neurorestorative agent in rats with cerebral ischemic stroke via upregulating SIRT1.

Safranal is an active ingredient of saffron (Crocus sativus L.). Its neuroprotective role in ischemic stroke (IS) through reducing oxidative stress damage has been widely reported. However, the neurorestorative mechanisms of safranal are still in the preliminary stage of exploration. the present study is aimed to discuss the effects of safranal on the recovery of neural function after IS. A middle cerebral artery occlusion/reperfusion (MCAO/R) rat model and an oxygen-glucose deprivation/reoxygenation (OGD/R) model in rat brain microvascular endothelial cells (RBMEC) were established to explore the effects of safranal on IS in vivo and in vitro. It was found that safranal dramatically reduced infarct size and Nissl's body loss in rats subjected to MCAO/R. Safranal also promoted neuron survival, stimulated neurogenesis, induced angiogenesis and increased SIRT1 expression in vivo and in vitro. Silencing of SIRT1 reversed the above effects of safranal on OGD/R-induced RBMEC. The present study indicated that safranal was a promising compound to exert neurorestorative effect in IS via upregulating SIRT1 expression. These results offer insight into developing new mechanisms in the recovery of neural function after safranal treatment of IS.

Pharmacological inhibition of the integrated stress response accelerates disease progression in an amyotrophic lateral sclerosis mouse model.

BACKGROUND AND PURPOSE: The integrated stress response (ISR) regulates translation in response to diverse stresses. ISR activation has been documented in amyotrophic lateral sclerosis (ALS) patients and ALS experimental models. In experimental models, both ISR stimulation and inhibition prevented ALS neurodegeneration; however, which mode of ISR regulation would work in patients is still debated. We previously demonstrated that the ISR modulator ISRIB (Integrated Stress Response InhiBitor, an eIF2B activator) enhances survival of neurons expressing the ALS neurotoxic allele SOD1 G93A. Here, we tested the effect of two ISRIB-like eIF2B activators (2BAct and PRXS571) in the disease progression of transgenic SOD1G93A mice. EXPERIMENTAL APPROACH: After biochemical characterization in primary neurons, SOD1G93A mice were treated with 2BAct and PRXS571. Muscle denervation of vulnerable motor units was monitored with a longitudinal electromyographic test. We used a clinical score to document disease onset and progression; force loss was determined with the hanging wire motor test. Motor neuronal survival was assessed by immunohistochemistry. KEY RESULTS: In primary neurons, 2BAct and PRXS571 relieve the ISR-imposed translational inhibition while maintaining high ATF4 levels. Electromyographic recordings evidenced an earlier and more dramatic muscle denervation in treated SOD1G93A mice that correlated with a decrease in motor neuron survival. Both compounds anticipated disease onset and shortened survival time. CONCLUSION AND IMPLICATIONS: 2BAct and PRXS571 anticipate disease onset, aggravating muscle denervation and motor neuronal death of SOD1G93A mice. This study reveals that the ISR works as a neuroprotective pathway in ALS motor neurons and reveals the toxicity that eIF2B activators may display in ALS patients.

Reduced Cytotoxicity by Repetitive mRNA Transfection in Differentiated Neurons.

Background and Objectives: mRNA-based protein expression technology has been used to express functional proteins. We have previously generated dopamine neurons from rat-embryo derived neural precursor cells (NPCs) through repeated transfection of synthetic transcription factor mRNA encoding dopamine-inducible genes. However, NPCs began to die approximately 10 d post-transfection. In this study, we examined a long-term transfection protocol that did not affect cell viability. Methods and Results: Experiments were performed in eight groups sorted according to the start date of mRNA transfection. mRNA was transfected into NPCs daily for 21 d and live cell images of each group were recorded. NPCs which were differentiated for more than five days showed sustained gene expression and appreciable viability despite daily mRNA transfection for 21 d. Conclusions: Repeated mRNA transfection requires cells with a sufficient differentiation period.

Cell-specific alterations in autophagy-lysosomal activity near the chronically implanted microelectrodes.

Intracortical microelectrodes that can record and stimulate brain activity have become a valuable technique for basic science research and clinical applications. However, long-term implantation of these microelectrodes can lead to progressive neurodegeneration in the surrounding microenvironment, characterized by elevation in disease-associated markers. Dysregulation of autophagy-lysosomal degradation, a major intracellular waste removal process, is considered a key factor in the onset and progression of neurodegenerative diseases. It is plausible that similar dysfunctions in autophagy-lysosomal degradation contribute to tissue degeneration following implantation-induced focal brain injury, ultimately impacting recording performance. To understand how the focal, persistent brain injury caused by long-term microelectrode implantation impairs autophagy-lysosomal pathway, we employed two-photon microscopy and immunohistology. This investigation focused on the spatiotemporal characterization of autophagy-lysosomal activity near the chronically implanted microelectrode. We observed an aberrant accumulation of immature autophagy vesicles near the microelectrode over the chronic implantation period. Additionally, we found deficits in autophagy-lysosomal clearance proximal to the chronic implant, which was associated with an accumulation of autophagy cargo and a reduction in lysosomal protease level during the chronic period. Furthermore, our evidence demonstrates reactive astrocytes have myelin-containing lysosomes near the microelectrode, suggesting its role of myelin engulfment during acute implantation period. Together, this study sheds light on the process of brain tissue degeneration caused by long-term microelectrode implantation, with a specific focus on impaired intracellular waste degradation.

Attenuation of histone H4 lysine 16 acetylation (H4K16ac) elicits a neuroprotection against ischemic stroke by alleviating the autophagic/lysosomal dysfunction in neurons at the penumbra.

A modest autophagy benefits neuroprotection while an excessive autophagy leads to neuronal death after cerebral ischemia, but what governs an appropriate autophagy remains to be understood. Studies indicated that acetylation of histone H4 at lysine16 (H4K16ac) strongly modulated autophagic/lysosomal signaling pathway. Thus, this study was to investigate whether the autophagic neuronal injury could be alleviated by amending H4K16ac level after ischemic stroke. A rat model of middle cerebral artery occlusion (MCAO)/reperfusion was prepared to investigate dynamic variations between H4K16ac and autophagy at the penumbra. The results illustrated that the significantly elevated H4K16ac was coupled with dramatically promoted autophagic activity at 4 h after the insult, suggesting H4K16ac tightly controlled autophagic signaling. After that, H4K16ac level was altered by pretreatment with trichostatin A (TSA, a H4K16ac facilitator) and MG149 (a H4K16ac inhibitor), respectively. Four hours after MCAO/reperfusion, the penumbral tissues were obtained to detect the key proteins in autophagic/lysosomal pathway by western blot and immunofluorescence, respectively. Meanwhile, the infarct volume, neurological deficits, and neuron survival were assessed to evaluate the neurological outcomes. The results showed that TSA-promoted H4K16ac led to an excessively up-regulated autophagy resulting in autophagic/lysosomal dysfunction, as indicated by the accumulated autophagic substrates and exacerbated lysosomal inefficiency in neurons. By contrast, MG149-depressed H4K16ac significantly down-regulated autophagic activity and thereby restored the impaired autophagic flux. Consequently, the neurological injury was markedly alleviated in MCAO + MG149 group, compared with that in MCAO group. Our study suggests that the H4K16ac attenuation elicits neuroprotection against ischemic stroke by ameliorating autophagic/lysosomal dysfunction in neurons.

Corrigendum: Vi4-miR-185-5p-Igfbp3 Network Protects the Brain From Neonatal Hypoxic Ischemic Injury via Promoting Neuron Survival and Suppressing the Cell Apoptosis.

[This corrects the article DOI: 10.3389/fcell.2020.529544.].

Agonism of the α7-acetylcholine receptor/PI3K/Akt pathway promotes neuronal survival after subarachnoid hemorrhage in mice.

Subarachnoid hemorrhage (SAH) results in severe neuronal dysfunction and degeneration. Since the nicotinic acetylcholine α7 receptors (α7-AChR) are involved in neuronal function and survival, we investigated if stimulation of α7-AChR would promote neuronal survival and improve behavioral outcome following SAH in mice. Male mice subjected to SAH were treated with either galantamine (α7-AChR agonist) or vehicle. Neurobehavioral testing was performed 24 h after SAH, and mice were euthanized for analysis of neuronal cell death or a cell survival (PI3K/Akt) signaling pathway. Neuron cell cultures were subjected to hemoglobin toxicity to assess the direct effects of α7-AChR agonism independent of other cells. Treatment with the α7-AChR agonist promoted neuronal survival and improved functional outcomes 24 h post-SAH. The improved outcomes corresponded with increased PI3K/Akt activity. Antagonism of α7-AChR or PI3K effectively reversed galantamine's beneficial effects. Tissue from α7-AChR knockout mice confirmed α7-AChR's role in neuronal survival after SAH. Data from the neuronal cell culture experiment supported a direct effect of α7-AChR agonism in promoting cell survival. Our findings indicate that α7-AChR is a therapeutic target following SAH which can promote neuronal survival, thereby improving neurobehavioral outcome. Thus, the clinically relevant α7-AChR agonist, galantamine, might be a potential candidate for human use to improve outcome after SAH.

LncRNA TCONS_00041002 improves neurological outcomes in neonatal rats with hypoxic-ischemic encephalopathy by inhibiting apoptosis and promoting neuron survival.

It has been reported that Neonatal hypoxic-ischemic encephalopathy (HIE) could induce apoptosis in neonates and result in cognitive and sensory impairments, which are associated with poor developmental outcomes. Despite the improvement in neonatology, there is still no clinically effective treatment for HIE presently. Long non-coding RNAs (lncRNAs) play important roles in cellular homeostasis. Nevertheless, their effects in developing rat brains with HI is little known. Here, we established HIE model in neonate rats and explored the expression and function of lncRNAs in HI, and found the expression of 19 lncRNAs was remarkably changed in the brains of HI rats, compared to the sham group. Among them, three lncRNAs (TCONS_00041002, TCONS_00070547, TCONS_00045572) were enriched in the apoptotic process via gene ontology (GO) and pathway analysis, which were selected for the further qRT-PCR verification. Through lentivirus-mediated overexpression of these three lncRNAs, we found that overexpression of TCONS_00041002 attenuated the cell apoptosis, and increased the vitality of neurons after oxygen-glucose deprivation (OGD), therefore reduced the brain infarction and further promoted the neuron survival as well as improved the neurological disorders in the rats subjected to HIE. What's more, ceRNA network prediction and co-expression verification showed that the expression of TCONS_00041002 was positively associated with Foxe1, Pawr and Nfkbiz. Altogether, this study has exhibited that lncRNA TCONS_00041002 participates in the cell apoptosis and neuronal survival of HIE and represents a potential new target for the treatment of HIE.

Fully Characterized Mature Human iPS- and NMP-Derived Motor Neurons Thrive Without Neuroprotection in the Spinal Contusion Cavity.

Neural cell interventions in spinal cord injury (SCI) have focused predominantly on transplanted multipotent neural stem/progenitor cells (NSPCs) for animal research and clinical use due to limited information on survival of spinal neurons. However, transplanted NSPC fate is unpredictable and largely governed by injury-derived matrix and cytokine factors that are often gliogenic and inflammatory. Here, using a rat cervical hemicontusion model, we evaluate the survival and integration of hiPSC-derived spinal motor neurons (SMNs) and oligodendrocyte progenitor cells (OPCs). SMNs and OPCs were differentiated in vitro through a neuromesodermal progenitor stage to mimic the natural origin of the spinal cord. We demonstrate robust survival and engraftment without additional injury site modifiers or neuroprotective biomaterials. Ex vivo differentiated neurons achieve cervical spinal cord matched transcriptomic and proteomic profiles, meeting functional electrophysiology parameters prior to transplantation. These data establish an approach for ex vivo developmentally accurate neuronal fate specification and subsequent transplantation for a more streamlined and predictable outcome in neural cell-based therapies of SCI.

Novel pharmacotherapy: NNI-362, an allosteric p70S6 kinase stimulator, reverses cognitive and neural regenerative deficits in models of aging and disease.

Aging is known to slow the neurogenic capacity of the hippocampus, one of only two mammalian adult neurogenic niches. The reduction of adult-born neurons with age may initiate cognitive decline progression which is exacerbated in chronic neurodegenerative disorders, e.g., Alzheimer's disease (AD). With physiologic neurogenesis diminished, but still viable in aging, non-invasive therapeutic modulation of this neuron regeneration process remains possible. The discovery of truly novel neuron regenerative therapies could be identified through phenotypic screening of small molecules that promote adult-born neurons from human neural progenitor cells (hNPCs). By identifying neuron-generating therapeutics and potentially novel mechanism of actions, therapeutic benefit could be confirmed through in vivo proof-of-concept studies. The key aging and longevity mTOR/p70S6 kinase axis, a commonly targeted pathway, is substrate for potential selective kinase modulators to promote new hippocampal neurons from NPCs. The highly regulated downstream substrate of mTOR, p70S6 kinase, directly controls pleiotropic cellular activities, including translation and cell growth. Stimulating this kinase, selectively in an adult neurogenic niche, should promote NPC proliferation, and cell growth and survival in the hippocampus. Studies of kinase profiling and immunocytochemistry of human progenitor neurogenesis suggest that the novel small molecule NNI-362 stimulates p70S6 kinase phosphorylation, which, in turn, promotes proliferation and differentiation of NPCs to neurons. NNI-362 promoted the associative reversal of age- and disease-related cognitive deficits in aged mice and Down syndrome-modeled mice. This oral, allosteric modulator may ultimately be beneficial for age-related neurodegenerative disorders involving hippocampal-dependent cognitive impairment, specifically AD, by promoting endogenous hippocampal regeneration.

Vi4-miR-185-5p-Igfbp3 Network Protects the Brain From Neonatal Hypoxic Ischemic Injury via Promoting Neuron Survival and Suppressing the Cell Apoptosis.

Neonatal hypoxic ischemic encephalopathy (HIE) due to birth asphyxia is common and causes severe neurological deficits, without any effective therapies currently available. Neuronal death is an important driving factors of neurological disorders after HIE, but the regulatory mechanisms are still uncertain. Long non-coding RNA (lncRNA) or ceRNA network act as a significant regulator in neuroregeneration and neuronal apoptosis, thus owning a great potential as therapeutic targets in HIE. Here, we found a new lncRNA, is the most functional in targeting the Igfbp3 gene in HIE, which enriched in the cell growth and cell apoptosis processes. In addition, luciferase reporter assay showed competitive regulatory binding sites to the target gene Igfbp3 between TCONS00044054 (Vi4) and miR-185-5p. The change in blood miR-185-5p and Igfbp3 expression is further confirmed in patients with brain ischemia. Moreover, Vi4 overexpression and miR-185-5p knock-out promote the neuron survival and neurite growth, and suppress the cell apoptosis, then further improve the motor and cognitive deficits in rats with HIE, while Igfbp3 interfering got the opposite results. Together, Vi4-miR-185-5p-Igfbp3 regulatory network plays an important role in neuron survival and cell apoptosis and further promote the neuro-functional recovery from HIE, therefore is a likely a drug target for HIE therapy.

Adenosine 2A receptor inhibition protects phrenic motor neurons from cell death induced by protein synthesis inhibition.

Respiratory motor neuron survival is critical for maintenance of adequate ventilation and airway clearance, preventing dependence to mechanical ventilation and respiratory tract infections. Phrenic motor neurons are highly vulnerable in rodent models of motor neuron disease versus accessory inspiratory motor pools (e.g. intercostals, scalenus). Thus, strategies that promote phrenic motor neuron survival when faced with disease and/or toxic insults are needed to help preserve breathing ability, airway defense and ventilator independence. Adenosine 2A receptors (A2A) are emerging as a potential target to promote neuroprotection, although their activation can have both beneficial and pathogenic effects. Since the role of A2A receptors in the phrenic motor neuron survival/death is not known, we tested the hypothesis that A2A receptor antagonism promotes phrenic motor neuron survival and preserves diaphragm function when faced with toxic, neurodegenerative insults that lead to phrenic motor neuron death. We utilized a novel neurotoxic model of respiratory motor neuron death recently developed in our laboratory: intrapleural injections of cholera toxin B subunit (CtB) conjugated to the ribosomal toxin, saporin (CtB-Saporin). We demonstrate that intrapleural CtB-Saporin causes: 1) profound phrenic motor neuron death (~5% survival); 2) ~7-fold increase in phrenic motor neuron A2A receptor expression prior to cell death; and 3) diaphragm muscle paralysis (inactive in most rats; ~7% residual diaphragm EMG amplitude during room air breathing). The A2A receptor antagonist istradefylline given after CtB-Saporin: 1) reduced phrenic motor neuron death (~20% survival) and 2) preserved diaphragm EMG activity (~46%). Thus, A2A receptors contribute to neurotoxic phrenic motor neuron death, an effect mitigated by A2A receptor antagonism.