Rejuvenating the brain’s endogenous regenerative potential : focus on the role of MANF in brain development and ischemic brain injury

Stroke is one of the leading causes of death and a major cause of disabilities in adults. More than half of stroke victims suffer some type of disability, ranging from different levels of minor weak- ness in a limb to a complete loss of mobility. Currently, treatment of stroke requires a stringent re- habilitation programs. Nevertheless, two thirds of all patients will still have some type of difficulty with regular daily activities. Recent experimental findings raise the possibility that functional improvement after stroke may be achieved through neuronal replacement by endogenous neural stem cells (NSCs) residing in the adult brain. Therefore, additional understanding of the properties of NSCs will help to identify their optimal potential in cell-based therapy. Neurotrophic factors are a family of proteins that are important in neuronal development and function, and have been studied as possible drugs for ischemic brain injury. In addition to Brain-Derived Neurotrophic Factor (BDNF) and Glial cell line-Derived Neurotrophic Factor (GDNF), Mescenphalic Astrocyte-Derived Neurotrophic Factor (MANF) and Cerebral Dopamine Neurotrophic Factor (CDNF), that form a distinct family of evolutionary conserved proteins with neuroprotective effects, have potential in the treatment of stroke. While MANF has been shown to protect cortical neurons from death in a rodent model of ischemic brain injury, the effects of post-stroke MANF ad- ministration on cellular processes during the recovery phase are poorly understood. To shed light on the possible regenerative potential of MANF for the injured brain, we need to first investigate the roles of endogenous MANF in neural stem cells (NSC) in a normal or pathological condition. We developed and optimized a work platform for studying the regulation and effect of MANF on biological properties of NSCs and cortical development. Our findings reveal an important role of MANF in neurite outgrowth and neuronal migration in the developing cortex. In addition, we demonstrated that endogenous MANF has the potential to protect NSCs against oxygen and glucose-deprivation conditions. Next, using neurosphere and subventricular zone (SVZ) explant cultures, we further studied the effect of MANF administration on cell differentiation and migration. We presented the data that exogenously added MANF can induce neural/glial differentiation and promote cell migration out of SVZ explants. Also, utilizing the advantage of NSCs as a target for MANF, we discovered that exogenous MANF can induce the phosphorylation of STAT3 in NSCs. Finally, we used the rat model of ischemic stroke to compare the effects of MANF and GDNF in neurogenesis after stroke. While injection of GDNF into lateral ventricle has a strong mitogenic effect to increase neurogenesis in SVZ, it does not induce migration of neuroblasts towards the ischemic area. In contrast, MANF facilitates the migration of neuroblasts towards the lesioned cortex. Regarding long-term infusions in the peri-infarct zone, both GDNF and MANF recruited the neuroblasts in the infarct area. However, only MANF accelerated functional recovery after stroke. In summary, this work has extended the knowledge of MANF’s capacity for neuronal differentiation as well as migration, and the regenerative capacity for its therapeutic use in further studies.Havaitsemaan MANF: n mahdollista regeneratiivista potentiaalia loukkaantuneelle aivolle meidän on ensin tutkittava endogeenisen MANF: n roolit hermoston kantasoluissa (NSC) normaalissa tai patologisessa tilassa. Kehitimme ja optimoimme työympäristön MANF: n säätelyn ja vaikutuksen tutkimiseksi NSC: n ja kortikaalisen kehityksen biologisista ominaisuuksista. Tuloksemme paljastavat MANF: n tärkeän roolin neuriittikasvussa ja hermosolujen muuttumisessa kehitettävälle aivokuorelle. Lisäksi osoitimme, että endogeenisellä MANFilla on mahdollisuus suojata NSC: itä happea ja glukoosin heikkenemisolosuhteita vastaan. Seuraavaksi, käyttämällä neurosfääri- ja subventrikulaarisia vyöhykkeitä (SVZ) eksplantaatteja, tutkittiin edelleen MANF: n antamisen vaikutusta solujen erilaistumiseen ja migraatioon. Esitimme tiedot, jotka eksogeenisesti lisäsivät MANF: ia, voivat indusoida neuraalisen / gliaalisen erilaistumisen ja edistää solujen migraatiota SVZ-eksplantaateista. Lisäksi, käyttämällä NSC: ien etua MANF: n kohteena, havaitsimme, että eksogeeninen MANF voi indusoida STAT3: n fosforylaatiota NSC: issä. Lopuksi käytimme iskeemisen aivohalvausmallin vertaamaan MANF: n ja GDNF: n vaikutuksia neurogeneesiin aivohalvauksen jälkeen

BN-embedded monolayer graphene with tunable electronic and topological properties

Finding an effective and controllable way to create a sizable energy gap in graphene-based systems has been a challenging topic of intensive research. We propose that the hybrid of boron nitride and graphene (h-BNC) at low BN doping serves as an ideal platform for band-gap engineering and valleytronic applications. We report a systematic first-principles study of the atomic configurations and band gap opening for energetically favorable BN patches embedded in graphene. Based on first-principles calculations, we construct a tight-binding model to simulate general doping configurations in large supercells. Unexpectedly, the calculations find a linear dependence of the band gap on the effective BN concentration at low doping, arising from an induced effective on-site energy difference at the two C sublattices as they are substituted by B and N dopants alternately. The significant and tunable band gap of a few hundred meVs, with preserved topological properties of graphene and feasible sample preparation in the laboratory, presents great opportunities to realize valley physics applications in graphene systems at room temperature

MANF Promotes Differentiation and Migration of Neural Progenitor Cells with Potential Neural Regenerative Effects in Stroke

Cerebral ischemia activates endogenous reparative processes, such as increased proliferation of neural stem cells (NSCs) in the subventricular zone (SVZ) and migration of neural progenitor cells (NPCs) toward the ischemic area. However, this reparative process is limited because most of the NPCs die shortly after injury or are unable to arrive at the infarct boundary. In this study, we demonstrate for the first time that endogenous mesencephalic astrocyte-derived neurotrophic factor (MANF) protects NSCs against oxygen-glucose-deprivation-induced injury and has a crucial role in regulating NPC migration. In NSC cultures, MANF protein administration did not affect growth of cells but triggered neuronal and glial differentiation, followed by activation of STAT3. In SVZ explants, MANF over expression facilitated cell migration and activated the STAT3 and ERK1/2 pathway. Using a rat model of cortical stroke, intracerebroventricular injections of MANF did not affect cell proliferation in the SVZ, but promoted migration of doublecortin (DCX)(+) cells toward the corpus callosum and infarct boundary on day 14 post-stroke. Long-term infusion of MANF into the per'-infarct zone increased the recruitment of DCX+ cells in the infarct area. In conclusion, our data demonstrate a neuroregenerative activity of MANF that facilitates differentiation and migration of NPCs, thereby increasing recruitment of neuroblasts in stroke cortex.Peer reviewe

Modulating Microglia/Macrophage Activation by CDNF Promotes Transplantation of Fetal Ventral Mesencephalic Graft Survival and Function in a Hemiparkinsonian Rat Model

Parkinson's disease (PD) is characterized by the loss of dopaminergic neurons in substantia nigra pars compacta, which leads to the motor control deficits. Recently, cell transplantation is a cutting-edge technique for the therapy of PD. Nevertheless, one key bottleneck to realizing such potential is allogenic immune reaction of tissue grafts by recipients. Cerebral dopamine neurotrophic factor (CDNF) was shown to possess immune-modulatory properties that benefit neurodegenerative diseases. We hypothesized that co-administration of CDNF with fetal ventral mesencephalic (VM) tissue can improve the success of VM replacement therapies by attenuating immune responses. Hemiparkinsonian rats were generated by injecting 6-hydroxydopamine (6-OHDA) into the right medial forebrain bundle of Sprague Dawley (SD) rats. The rats were then intrastriatally transplanted with VM tissue from rats, with/without CDNF administration. Recovery of dopaminergic function and survival of the grafts were evaluated using the apomorphine-induced rotation test and smallanimal positron emission tomography (PET) coupled with [F-18] DOPA or [F-18] FE-PE2I, respectively. In addition, transplantation-related inflammatory response was determined by uptake of [F-18] FEPPA in the grafted side of striatum. Immunohistochemistry (IHC) examination was used to determine the survival of the grated dopaminergic neurons in the striatum and to investigate immune-modulatory effects of CDNF. The modulation of inflammatory responses caused by CDNF might involve enhancing M2 subset polarization and increasing fractal dimensions of 6-OHDA-treated BV2 microglial cell line. Analysis of CDNF-induced changes to gene expressions of 6-OHDA-stimulated BV2 cells implies that these alternations of the biomarkers and microglial morphology are implicated in the upregulation of protein kinase B signaling as well as regulation of catalytic, transferase, and protein serine/threonine kinase activity. The effects of CDNF on 6-OHDA-induced alternation of the canonical pathway in BV2 microglial cells is highly associated with PI3K-mediated phagosome formation. Our results are the first to show that CDNF administration enhances the survival of the grafted dopaminergic neurons and improves functional recovery in PD animal model. Modulation of the polarization, morphological characteristics, and transcriptional profiles of 6-OHDA-stimualted microglia by CDNF may possess these properties in transplantation-based regenerative therapies.Peer reviewe

UPR Responsive Genes Manf and Xbp1 in Stroke

Stroke is a devastating medical condition with no treatment to hasten recovery. Its abrupt nature results in cataclysmic changes in the affected tissues. Resident cells fail to cope with the cellular stress resulting in massive cell death, which cannot be endogenously repaired. A potential strategy to improve stroke outcomes is to boost endogenous pro-survival pathways. The unfolded protein response (UPR), an evolutionarily conserved stress response, provides a promising opportunity to ameliorate the survival of stressed cells. Recent studies from us and others have pointed toward mesencephalic astrocyte-derived neurotrophic factor (MANF) being a UPR responsive gene with an active role in maintaining proteostasis. Its pro-survival effects have been demonstrated in several disease models such as diabetes, neurodegeneration, and stroke. MANF has an ER-signal peptide and an ER-retention signal; it is secreted by ER calcium depletion and exits cells upon cell death. Although its functions remain elusive, conducted experiments suggest that the endogenous MANF in the ER lumen and exogenously administered MANF protein have different mechanisms of action. Here, we will revisit recent and older bodies of literature aiming to delineate the expression profile of MANF. We will focus on its neuroprotective roles in regulating neurogenesis and inflammation upon post-stroke administration. At the same time, we will investigate commonalities and differences with another UPR responsive gene, X-box binding protein 1 (XBP1), which has recently been associated with MANF's function. This will be the first systematic comparison of these two UPR responsive genes aiming at revealing previously uncovered associations between them. Overall, understanding the mode of action of these UPR responsive genes could provide novel approaches to promote cell survival.Peer reviewe

Modulating Microglia/Macrophage Activation by CDNF Promotes Transplantation of Fetal Ventral Mesencephalic Graft Survival and Function in a Hemiparkinsonian Rat Model

Parkinson’s disease (PD) is characterized by the loss of dopaminergic neurons in substantia nigra pars compacta, which leads to the motor control deficits. Recently, cell transplantation is a cutting-edge technique for the therapy of PD. Nevertheless, one key bottleneck to realizing such potential is allogenic immune reaction of tissue grafts by recipients. Cerebral dopamine neurotrophic factor (CDNF) was shown to possess immune-modulatory properties that benefit neurodegenerative diseases. We hypothesized that co-administration of CDNF with fetal ventral mesencephalic (VM) tissue can improve the success of VM replacement therapies by attenuating immune responses. Hemiparkinsonian rats were generated by injecting 6-hydroxydopamine (6-OHDA) into the right medial forebrain bundle of Sprague Dawley (SD) rats. The rats were then intrastriatally transplanted with VM tissue from rats, with/without CDNF administration. Recovery of dopaminergic function and survival of the grafts were evaluated using the apomorphine-induced rotation test and small-animal positron emission tomography (PET) coupled with [18F] DOPA or [18F] FE-PE2I, respectively. In addition, transplantation-related inflammatory response was determined by uptake of [18F] FEPPA in the grafted side of striatum. Immunohistochemistry (IHC) examination was used to determine the survival of the grated dopaminergic neurons in the striatum and to investigate immune-modulatory effects of CDNF. The modulation of inflammatory responses caused by CDNF might involve enhancing M2 subset polarization and increasing fractal dimensions of 6-OHDA-treated BV2 microglial cell line. Analysis of CDNF-induced changes to gene expressions of 6-OHDA-stimulated BV2 cells implies that these alternations of the biomarkers and microglial morphology are implicated in the upregulation of protein kinase B signaling as well as regulation of catalytic, transferase, and protein serine/threonine kinase activity. The effects of CDNF on 6-OHDA-induced alternation of the canonical pathway in BV2 microglial cells is highly associated with PI3K-mediated phagosome formation. Our results are the first to show that CDNF administration enhances the survival of the grafted dopaminergic neurons and improves functional recovery in PD animal model. Modulation of the polarization, morphological characteristics, and transcriptional profiles of 6-OHDA-stimualted microglia by CDNF may possess these properties in transplantation-based regenerative therapies

Local Administration of AAV-BDNF to Subventricular Zone Induces Functional Recovery in Stroke Rats

Peer reviewe

Augmenting hematoma-scavenging capacity of innate immune cells by CDNF reduces brain injury and promotes functional recovery after intracerebral hemorrhage

During intracerebral hemorrhage (ICH), hematoma formation at the site of blood vessel damage results in local mechanical injury. Subsequently, erythrocytes lyse to release hemoglobin and heme, which act as neurotoxins and induce inflammation and secondary brain injury, resulting in severe neurological deficits. Accelerating hematoma resorption and mitigating hematoma-induced brain edema by modulating immune cells has potential as a novel therapeutic strategy for functional recovery after ICH. Here, we show that intracerebroventricular administration of recombinant human cerebral dopamine neurotrophic factor (rhCDNF) accelerates hemorrhagic lesion resolution, reduces peri-focal edema, and improves neurological outcomes in an animal model of collagenase-induced ICH. We demonstrate that CDNF acts on microglia/macrophages in the hemorrhagic striatum by promoting scavenger receptor expression, enhancing erythrophagocytosis and increasing anti-inflammatory mediators while suppressing the production of pro-inflammatory cytokines. Administration of rhCDNF results in upregulation of the Nrf2-HO-1 pathway, but alleviation of oxidative stress and unfolded protein responses in the perihematomal area. Finally, we demonstrate that intravenous delivery of rhCDNF has beneficial effects in an animal model of ICH and that systemic application promotes scavenging by the brain's myeloid cells for the treatment of ICH.Peer reviewe

Search for dark matter produced in association with bottom or top quarks in √s = 13 TeV pp collisions with the ATLAS detector

A search for weakly interacting massive particle dark matter produced in association with bottom or top quarks is presented. Final states containing third-generation quarks and miss- ing transverse momentum are considered. The analysis uses 36.1 fb−1 of proton–proton collision data recorded by the ATLAS experiment at √s = 13 TeV in 2015 and 2016. No significant excess of events above the estimated backgrounds is observed. The results are in- terpreted in the framework of simplified models of spin-0 dark-matter mediators. For colour- neutral spin-0 mediators produced in association with top quarks and decaying into a pair of dark-matter particles, mediator masses below 50 GeV are excluded assuming a dark-matter candidate mass of 1 GeV and unitary couplings. For scalar and pseudoscalar mediators produced in association with bottom quarks, the search sets limits on the production cross- section of 300 times the predicted rate for mediators with masses between 10 and 50 GeV and assuming a dark-matter mass of 1 GeV and unitary coupling. Constraints on colour- charged scalar simplified models are also presented. Assuming a dark-matter particle mass of 35 GeV, mediator particles with mass below 1.1 TeV are excluded for couplings yielding a dark-matter relic density consistent with measurements