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Effects of ibuprofen on regeneration of the lateral olfactory tract
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Abstracts / Neuroscience Research 68S (2010) e223–e334
P2-e12 Paired immunoglobulin-like receptor B knockout does not enhance axonalregeneration or locomotor recovery after spinal cord injury Yuka Nakamura 1 , Yuki Fujita 1 , Masaki Ueno 1 , Toshiyuki Takai 2 , Toshihide Yamashita 1 1
Dept Molecular Neuroscience, Univ of Osaka, Osaka Immunol., IDAC, Univ of Tohoku, Miyagi
2
Dept Experimental
Myelin components that inhibit axonal regeneration are believed to significantly contribute tothe lack of axonal regeneration noted in the adult CNS. Three proteins found inmyelin-Nogo, myelin-associated glycoprotein, and oligodendrocyte-myelinglycoprotein-inhibit neurite outgrowth in vitro. All these proteins interact with the samereceptors, namely, the Nogo receptor (NgR) and paired immunoglobulin-like receptor B(PIR-B). As per previous reports, corticospinal tract (CST) regeneration is not enhanced inNgR-knockout mice after spinal cord injury. Therefore, we assessed CST regeneration inPIR-B-knockout mice. We found that hindlimb motor function, as assessed using the BassoMouse Scale, foot print test, inclined plane test, and beam walking test, did not differ betweenthe PIR-B-knockout and wild-type mice after dorsal hemisection of the spinal cord. Further, tracing of the CST fibers after injury did not reveal enhanced axonal regeneration or sproutingin the CST of the PIR-B-knockout mice. Systemic administration of NEP1-40, NgRantagonist, to PIR-B knockout mice did not enhance the regenerative response. These resultsindicate that PIR-B knockout is not sufficient to induce extensive axonal regeneration afterspinal cord injury. doi:10.1016/j.neures.2010.07.1125
P2-e13 Genetic deletion of paired immunoglobulin-like receptor B does not promote axonal plasticity or functional recovery after traumatic brain injury Shusaku Omoto 1,2 , Masaki Ueno 1 , Soichiro Mochio 2 , Toshiyuki Takai 3 , Toshihide Yamashita 1 1 3
Dept Mol Neurosci, Osaka Univ, Suita 2 Dept Neurology, Jikei Univ, Tokyo Dept Exp Immunol, Tohoku Univ, Sendai
The rewiring of neural networks is a fundamental step in recovering behavioral functions after brain injury. However, there is limited potential for axonal plasticity in the adult central nervous system. The myelin-associated proteins Nogo, myelin-associated glycoprotein (MAG), and oligodendrocyte myelin glycoprotein (OMgp) are known to inhibit axonal plasticity and thus targeting the inhibitory pathways they participate in is a potential means of promoting plasticity and functional recovery. Each of Nogo, MAG, and OMgp interacts with both the Nogo receptor (NgR) and paired immunoglobulinlike receptor B (PirB). Here, we determined whether blocking PirB or/and NgR activity enhances axonal reorganization and functional recovery after cortical injury. We found that axons of the contralesional corticospinal tract sprouted into the denervated side of the cervical spinal cord after unilateral injury of the motor cortex. The extent to which this axonal reorganization occurred was far greater in mice lesioned during early postnatal days than in mice lesioned at an age when myelin had begun to form. This suggests that myelin-associated proteins might limit axonal remodeling in vivo. However, the number of sprouting fibers within either the corticospinal or corticorubral tract was not enhanced in PirB−/− mice, even in those treated with the NgR antagonist NEP1-40. Blocking PirB and/or NgR signaling also failed to enhance functional recovery with three motor tests. Our results suggest that blocking the activity of PirB and NgR is not sufficient to promote axonal reorganization or functional recovery after cortical injury. doi:10.1016/j.neures.2010.07.1126
P2-e14 Protein transduction of p38 MAP kinase enhances adult neural stem cell migration. Makoto Hamanoue 1 , Taka-aki Mizuno 1 , Hirotaka James Okano 2 , Hideyuki Okano 2 , Ken Takamatsu 1 1
Department Physiol, Toho Univ Sch. of Med, Tokyo, Japan 2 Keio University Sch. of Med, Tokyo, Japan In the mice adult brain, neural stem cells (NSCs) proliferate in the subventricular zone (SVZ) of the cerebral cortex, and then migrate into olfactory bulb through rostral migratory stream to produce neurons. Though the migration of NSCs into the injured sites was detected, further enhancement of the migration of NSCs could be required for the regeneration of the lost brain function. In this study, immunohistochemical analysis of the adult mice brain
showed that p38 MAP kinase (p38) co-expressed in the cells which express doublecortin that is a marker for migrating NSCs. This result suggests that p38 could involve in NSCs migration. To introduce activated form of p38 into NSCs, the recombinant p38 protein fused to cell-penetrating peptide, Tat, at N-terminus, was purified from E. coli. Western blot analysis revealed that these purified-Tat-fused protein (Tat-p38) was phosphorylated, and has kinase activity against its substrate, ATF-2. To determine whether activation of p38 in NSCs could enhance migration, NSC purified from the adult mice cerebral hemisphere was cultured in the upper chamber of Transwell, and Tat-p38 was added to the lower chamber. After 16 h, cell number in the lower chamber was increased in the Tat-p38-treated wells compared to the well treated with p38 peptide alone or Tat-GFP. In addition, the wells treated with Tat-p38 protein that lost kinase activity by the replacement of amino acids showed no increase in the number of migrating cells, suggesting that Tat-p38 kinase protein could be potent inducer of NSCs migration. doi:10.1016/j.neures.2010.07.1127
P2-e16 Effects of ibuprofen on regeneration of the lateral olfactory tract Yasuyuki Sekiguchi 1 , Kumiko Yokouchi 2 , Nanae Fukushima 2 , Kyutaro Kawagishi 2 , Tetsuji Moriizumi 2 1 Department of Neurosurgery, Shinshu University School of Medicine, Matsumoto, Japan 2 Department of Anatomy, Shinshu University School of Medicine, Matsumoto, Japan
Ibuprofen, a nonsteroidal anti-inflammatory drug commonly used as a pain reliever, has recently been shown to stimulate neurite outgrowth in vitro and axonal regeneration in vivo by inhibiting the active RhoA signal induced by CNS myelin and chondroitin sulfate proteoglycans, both of which are known to be axon-inhibitory molecules, and thus has been highly expected to have beneficial effects to CNS injuries. The present study was undertaken to examine whether ibuprofen does have significant effects on regeneration of the lateral olfactory tract (LOT), the main fiber tract of the central olfactory system that connects the olfactory bulb to the olfactory cortex. After unilateral ablation of the olfactory bulb, the rats were trained to avoid cycloheximide solution by olfaction, since cycloheximide has a strong repellent action to rodents. These conditioned rats received transection of the LOT contralateral to the bulbectomy and subcutaneous continuous infusion of ibuprofen (60 mg/kg/day) via an osmotic mini-pump for 4 weeks. Olfactory function was estimated by the ability to discriminate between cycloheximide solution and water. An anterograde neuronal tracer, biotinylated dextran amine (BDA), was used to visualize axonal regeneration derived from the olfactory bulb. Morphological data on regeneration of the LOT will be presented along with functional analysis. doi:10.1016/j.neures.2010.07.1128
P2-e17 Fibrotic scar formed in the lesion site of the central nervous system. I. Impediment for axonal regeneration Hitoshi Kawano , Junko Kimura-Kuroda, Nozomu Yoshioka, Yukari Komuta, Kazunori Sango, Koki Kawamura Department of Developmental Morphology, Tokyo Metropolitan Institute for Neuroscience In the central nervous system (CNS) of adult mammals, transected axons display almost no regenerative capacity following traumatic injury. Various kinds of factors which occur around the lesion site, such as glial scar and chorndoitin sulfate proteoglycans, have been postulated to prevent the regrowth of severed axons. A fibrotic scar containing deposition of type IV collagen (Col IV) is also considered as an impediment for axonal regeneration. After traumatic injury, meningeal fibroblasts migrate in the lesion site, proliferate and secrete Col IV to form the fibrotic scar. We have demonstrated that suppression of the fibrotic scar formation is required for axonal regeneration in the damaged CNS in a variety of animal models, such as (1) suppression of Col IV synthesis (Kawano et al., 2005), (2) newborn mouse (Kawano et al., Ibid), (3) the mouse hypothalamic arcuate nucleus (Homma et al., 2006), (4) degradation of glycoaminoglycan side chains of chondroitin sulfate proteoglycans with chondroitinase ABC (Li et al., 2007) and (5) transplantation of olfactory ensheathing cells (Teng et al., 2008). In all of these cases, regenerated axons extended the fibrotic scar-eliminated lesion site and often elongated in the glial scar-occupied region. The fibrotic scar expresses various potent axonal growth-inhibitory molecules including NG2 proteoglycan, tenascin-C, semaphorin 3A and EphB2, which may act as chemical barriers for regenerating axons. We propose that the elimination of the fibrotic scar
Abstracts / Neuroscience Research 68S (2010) e223–e334
P2-e12 Paired immunoglobulin-like receptor B knockout does not enhance axonalregeneration or locomotor recovery after spinal cord injury Yuka Nakamura 1 , Yuki Fujita 1 , Masaki Ueno 1 , Toshiyuki Takai 2 , Toshihide Yamashita 1 1
Dept Molecular Neuroscience, Univ of Osaka, Osaka Immunol., IDAC, Univ of Tohoku, Miyagi
2
Dept Experimental
Myelin components that inhibit axonal regeneration are believed to significantly contribute tothe lack of axonal regeneration noted in the adult CNS. Three proteins found inmyelin-Nogo, myelin-associated glycoprotein, and oligodendrocyte-myelinglycoprotein-inhibit neurite outgrowth in vitro. All these proteins interact with the samereceptors, namely, the Nogo receptor (NgR) and paired immunoglobulin-like receptor B(PIR-B). As per previous reports, corticospinal tract (CST) regeneration is not enhanced inNgR-knockout mice after spinal cord injury. Therefore, we assessed CST regeneration inPIR-B-knockout mice. We found that hindlimb motor function, as assessed using the BassoMouse Scale, foot print test, inclined plane test, and beam walking test, did not differ betweenthe PIR-B-knockout and wild-type mice after dorsal hemisection of the spinal cord. Further, tracing of the CST fibers after injury did not reveal enhanced axonal regeneration or sproutingin the CST of the PIR-B-knockout mice. Systemic administration of NEP1-40, NgRantagonist, to PIR-B knockout mice did not enhance the regenerative response. These resultsindicate that PIR-B knockout is not sufficient to induce extensive axonal regeneration afterspinal cord injury. doi:10.1016/j.neures.2010.07.1125
P2-e13 Genetic deletion of paired immunoglobulin-like receptor B does not promote axonal plasticity or functional recovery after traumatic brain injury Shusaku Omoto 1,2 , Masaki Ueno 1 , Soichiro Mochio 2 , Toshiyuki Takai 3 , Toshihide Yamashita 1 1 3
Dept Mol Neurosci, Osaka Univ, Suita 2 Dept Neurology, Jikei Univ, Tokyo Dept Exp Immunol, Tohoku Univ, Sendai
The rewiring of neural networks is a fundamental step in recovering behavioral functions after brain injury. However, there is limited potential for axonal plasticity in the adult central nervous system. The myelin-associated proteins Nogo, myelin-associated glycoprotein (MAG), and oligodendrocyte myelin glycoprotein (OMgp) are known to inhibit axonal plasticity and thus targeting the inhibitory pathways they participate in is a potential means of promoting plasticity and functional recovery. Each of Nogo, MAG, and OMgp interacts with both the Nogo receptor (NgR) and paired immunoglobulinlike receptor B (PirB). Here, we determined whether blocking PirB or/and NgR activity enhances axonal reorganization and functional recovery after cortical injury. We found that axons of the contralesional corticospinal tract sprouted into the denervated side of the cervical spinal cord after unilateral injury of the motor cortex. The extent to which this axonal reorganization occurred was far greater in mice lesioned during early postnatal days than in mice lesioned at an age when myelin had begun to form. This suggests that myelin-associated proteins might limit axonal remodeling in vivo. However, the number of sprouting fibers within either the corticospinal or corticorubral tract was not enhanced in PirB−/− mice, even in those treated with the NgR antagonist NEP1-40. Blocking PirB and/or NgR signaling also failed to enhance functional recovery with three motor tests. Our results suggest that blocking the activity of PirB and NgR is not sufficient to promote axonal reorganization or functional recovery after cortical injury. doi:10.1016/j.neures.2010.07.1126
P2-e14 Protein transduction of p38 MAP kinase enhances adult neural stem cell migration. Makoto Hamanoue 1 , Taka-aki Mizuno 1 , Hirotaka James Okano 2 , Hideyuki Okano 2 , Ken Takamatsu 1 1
Department Physiol, Toho Univ Sch. of Med, Tokyo, Japan 2 Keio University Sch. of Med, Tokyo, Japan In the mice adult brain, neural stem cells (NSCs) proliferate in the subventricular zone (SVZ) of the cerebral cortex, and then migrate into olfactory bulb through rostral migratory stream to produce neurons. Though the migration of NSCs into the injured sites was detected, further enhancement of the migration of NSCs could be required for the regeneration of the lost brain function. In this study, immunohistochemical analysis of the adult mice brain
showed that p38 MAP kinase (p38) co-expressed in the cells which express doublecortin that is a marker for migrating NSCs. This result suggests that p38 could involve in NSCs migration. To introduce activated form of p38 into NSCs, the recombinant p38 protein fused to cell-penetrating peptide, Tat, at N-terminus, was purified from E. coli. Western blot analysis revealed that these purified-Tat-fused protein (Tat-p38) was phosphorylated, and has kinase activity against its substrate, ATF-2. To determine whether activation of p38 in NSCs could enhance migration, NSC purified from the adult mice cerebral hemisphere was cultured in the upper chamber of Transwell, and Tat-p38 was added to the lower chamber. After 16 h, cell number in the lower chamber was increased in the Tat-p38-treated wells compared to the well treated with p38 peptide alone or Tat-GFP. In addition, the wells treated with Tat-p38 protein that lost kinase activity by the replacement of amino acids showed no increase in the number of migrating cells, suggesting that Tat-p38 kinase protein could be potent inducer of NSCs migration. doi:10.1016/j.neures.2010.07.1127
P2-e16 Effects of ibuprofen on regeneration of the lateral olfactory tract Yasuyuki Sekiguchi 1 , Kumiko Yokouchi 2 , Nanae Fukushima 2 , Kyutaro Kawagishi 2 , Tetsuji Moriizumi 2 1 Department of Neurosurgery, Shinshu University School of Medicine, Matsumoto, Japan 2 Department of Anatomy, Shinshu University School of Medicine, Matsumoto, Japan
Ibuprofen, a nonsteroidal anti-inflammatory drug commonly used as a pain reliever, has recently been shown to stimulate neurite outgrowth in vitro and axonal regeneration in vivo by inhibiting the active RhoA signal induced by CNS myelin and chondroitin sulfate proteoglycans, both of which are known to be axon-inhibitory molecules, and thus has been highly expected to have beneficial effects to CNS injuries. The present study was undertaken to examine whether ibuprofen does have significant effects on regeneration of the lateral olfactory tract (LOT), the main fiber tract of the central olfactory system that connects the olfactory bulb to the olfactory cortex. After unilateral ablation of the olfactory bulb, the rats were trained to avoid cycloheximide solution by olfaction, since cycloheximide has a strong repellent action to rodents. These conditioned rats received transection of the LOT contralateral to the bulbectomy and subcutaneous continuous infusion of ibuprofen (60 mg/kg/day) via an osmotic mini-pump for 4 weeks. Olfactory function was estimated by the ability to discriminate between cycloheximide solution and water. An anterograde neuronal tracer, biotinylated dextran amine (BDA), was used to visualize axonal regeneration derived from the olfactory bulb. Morphological data on regeneration of the LOT will be presented along with functional analysis. doi:10.1016/j.neures.2010.07.1128
P2-e17 Fibrotic scar formed in the lesion site of the central nervous system. I. Impediment for axonal regeneration Hitoshi Kawano , Junko Kimura-Kuroda, Nozomu Yoshioka, Yukari Komuta, Kazunori Sango, Koki Kawamura Department of Developmental Morphology, Tokyo Metropolitan Institute for Neuroscience In the central nervous system (CNS) of adult mammals, transected axons display almost no regenerative capacity following traumatic injury. Various kinds of factors which occur around the lesion site, such as glial scar and chorndoitin sulfate proteoglycans, have been postulated to prevent the regrowth of severed axons. A fibrotic scar containing deposition of type IV collagen (Col IV) is also considered as an impediment for axonal regeneration. After traumatic injury, meningeal fibroblasts migrate in the lesion site, proliferate and secrete Col IV to form the fibrotic scar. We have demonstrated that suppression of the fibrotic scar formation is required for axonal regeneration in the damaged CNS in a variety of animal models, such as (1) suppression of Col IV synthesis (Kawano et al., 2005), (2) newborn mouse (Kawano et al., Ibid), (3) the mouse hypothalamic arcuate nucleus (Homma et al., 2006), (4) degradation of glycoaminoglycan side chains of chondroitin sulfate proteoglycans with chondroitinase ABC (Li et al., 2007) and (5) transplantation of olfactory ensheathing cells (Teng et al., 2008). In all of these cases, regenerated axons extended the fibrotic scar-eliminated lesion site and often elongated in the glial scar-occupied region. The fibrotic scar expresses various potent axonal growth-inhibitory molecules including NG2 proteoglycan, tenascin-C, semaphorin 3A and EphB2, which may act as chemical barriers for regenerating axons. We propose that the elimination of the fibrotic scar