グリアの機能 Function of Glia
O3-6-5-1 神経炎症性病変における単球とミクログリアの機能解析 Functional analysis of monocytes and microglia in neuroinflammatory lesion ○山崎亮1,2, 大八木保政1, 吉良潤一3 ○Ryo Yamasaki1,2, Richard Ransohoff2, Haiyan Lu2, Nobuhiko Ohno2, Yasumasa Ohyagi1, Jun-ichi Kira3 九州大学大学院医学研究院　神経治療学1, クリーブランドクリニック　ラーナー研究所2, 九州大学大学院医学研究院　神経内科学3 Department of Neurological Therapeutics, Neurological Institute, Graduate School of Medical Sciences1, Cleveland Clinic, Lerner Research Institute2, Department of Neurology, Neurological Institute, Graduate School of Medical Sciences3 Microglia are the only myeloid cell populations in normal central nervous system (CNS) parenchyma. Once inflammation occurs in CNS, not only microglia but monocytes from peripheral blood that infiltrated into inflamed lesion activated and damaged CNS. They have common surface antigens and are resembles to each other when they are activated, so it has been controversial whether or not they have quite different roles in the pathogenesis of neuroinflammatory disorders. In this study, we have characterized different appearances (including ultra-fine structures) of monocytes and microglia, and we also have elucidated contrasting cytokine expression profiles and characteristics in phagocytosis between monocytes and microglia in inflamed mice CNS lesion. We observed inflamed lumbar CNS lesion of experimental autoimmune encephalomyelitis (EAE) model mice. We utilized CCR2-RFP/ CX3CR1-GFP mouse which have RFP in monocytes and GFP in microglia to distinguish each other in the lesion. At the onset stage, robust infiltration of monocytes was seen. At the peak stage, intrinsic microglia were activated and increased their number, while monocytes began to decrease. Immunohistochemical analysis showed that monocytes have complicated shape of nucleus and have small myelin debris, while microglia have simple shaped nucleus and have larger debris. Messenger RNA array assay revealed that monocytes expressed inflammatory cytokines at the onset stage while microglia expressed neuro-protective cytokines at the peak stage of EAE. The ultra-fine structures also differed; monocytes tended to have small mitochondria while microglia had needle-like mitochondria. Both of those cells attached myelinated axons and caused destructions of myelin, but monocytes tend to attach to the nodes of Ranvier, which were not observed for microglia. We concluded that monocytes from peripheral blood infiltrated first to cause demyelinations, and intrinsic microglia sweep myelin debris for remyelinations.	O3-6-5-2 P2Y1受容体によるグリア瘢痕形成調節 Astrocytic P2Y1 receptor regulates glial scar formation ○篠崎陽一1, 小泉修一1 ○Youichi Shinozaki1, Schuichi Koizumi1 山梨大学大学院 医学工学総合研究部 薬理学1 Dept Neuropharmacol, Univ of Yamanashi, Yamanashi1 ATP is released or leaked from injured cells. The released ATP dramatically changes glial phenotypes, and thus, affects pathogenesis of brain diseases. However, whether glial responses facilitate or rather inhibit the diseases is still the matter of debate. Here, we studied the neuroprotective role of purinergic signaling using in vivo stab injury model on mouse cerebral cortex. In the intact or contralateral side, fluoro-jade (FJ), a marker for neuronal death, and CD45+ signals were not observed, and were significantly increased in the ipsilateral side at 3 days after the stab injury. An ATP-degrading enzyme apyrase, a P2 receptor antagonist PPADS, and a P2Y1 receptor antagonist MRS2179 significantly reduced the FJ signals. In addition, P2Y1 receptor knockout (P2Y1KO) mice also exhibited reduced FJ and CD45+ signals. Immunohistochemical analysis revealed that reactive astrocytes formed glial scar, by which they enclosed CD45+ cells. P2Y1KO mice exhibited higher level of GFAP expression, more remarkable hypertrophy of astrocytes and tighter glial scar formation than wild type mice did. Although glial scar formation was accelerated, no enhanced proliferation was observed. We then analyzed the mechanism of enhanced glial scar formation and neuroprotection using in vitro scratch-wound model. In cultured astrocytes, the scratch induced a transient increase in extracellular ATP, which was followed by decrease in extracellular ATP mainly due to an increase in ectoATPase activity. Suppression of purinergic signaling by either apyrase, PPADS or MRS2179 enhanced scratch-evoked astrocytic migration rather than proliferation. Neither activation nor inhibition of P2Y1 receptors in cultured cortical neurons affected the scratch-induced damages. Our data suggest that the decrease in ATP/P2Y1 receptor-mediated signal should be a trigger that transforms astrocytes into migratory phenotype, which forms tighter glial scar structure thereby suppressing neuronal death.
O3-6-5-3 脊髄損傷によりマイクログリア細胞に発現するタンパク質アルギニンメチル化酵素PRMT8の機能解明 Spinal cord injury (SCI)-mediated expression of protein arginine N-methyltransferase 8 (PRMT8)expression in activated microglia/macrophage ○森泰丈1, 小山佳久1, 宮田信吾2 ○Yasutake Mori1, Yoshihisa Koyama1, Shingo Miyata2 大阪大学大学院　医学系研究科　神経機能形態学1, 近畿大学　東洋医学研究所　分子脳科学研究部門2 Dept Anat and Neurosci, Univ of Osaka, Osaka1, Div of Mol Brain Sci, Res Inst of Traditional Asian Med, Kinki Univ, Osaka2 Protein arginine N-methyltransferase 8 (PRMT8) was originally reported as a neuron-specific type II PRMT with dominantly nuclear distribution. However, when we examined changes in PRMT8 expression level in the spinal cords injured by hemisection, unambiguous signals were observed in the cytoplasm of round-shaped cells that were densely packed around the lesion site. This group of cells were well-overlapped with CD11b-positive cells, not with neurons, demonstrating that PRMT8 is expressed in the activated microglia/macrophage cells. By Western blot analysis, PRMT8 from the injured spinal cord showed a slower mobility shift band with Mw of 60-62kDa than that from brain lysate with Mw of 42kDa. We demonstrated that the high molecular weight (HMW) PRMT8 is due to phosphorylation and the myristoylation might be caused by an additional N-terminal stretch harboring consensus myristoylation site that would stem from usage of another in-frame translation initiation site. These data raise the possibility that membrane bound type of PRMT8 can be a novel marker for activated microglia.	O3-6-5-4 アストロサイト微細突起とグルタミン酸トランスポーターEAAT2 Role of glutamate transporter EAAT2 in peripheral astrocyte process ○林真理子1, 安井正人1 ○Mariko Hayashi1, Masato Yasui1 慶應義塾大学　医学部　薬理学1 Dept. Pharmacol, Sch. Med,Keio Univ, Tokyo1 In brain, astrocytes have thousands of filopodia-like protrusions called peripheral astrocyte processes. But the mechanisms of how the functional membrane proteins, such as channels, receptors or transporters, are transported and localized to the processes are not known well.Excitatory amino acid transporter 2 (EAAT2 / GLT-1, SlcA2) is a glutamate transporter responsible for the recovery of the majority of glutamate released from excitatory neuronal synapses. mGFP-tagged EAAT2 strongly localizes to filopodia of both primary cultured astrocytes and COS-7 cells. In some cells, EAAT2 is localized to filopodia tips and shows rolling movements in the order of minutes. The movement of the filopodia was not affected by addition of glutamate or EAAT2 blocker, TBOA. When the mGFP-EAAT2 was expressed in astrocytes of hippocampus slice culture, the EAAT2 was localized to astrocyte plasma membrane of peripheral astrocyte processes, and occasionally localized to their tips with dynamic movement, while the majority of the processes are relatively stable. A MAGUK family PDZ domain protein PSD-95 and LIM domain protein Ajuba are known to interact with the EAAT2, and regulate its localization to plasma membrane. However, the filopodial localization of EAAT2 in COS-7 cells does not require the N- or C- terminal cytoplasmic regions, which are the major interaction sites of Ajuba and PSD-95, respectively. We also investigated the possibility of direct regulation of EAAT2 localization by some of the known actin-binding filopodia proteins. Coexpression of IRS-p53, Ezrin. EBP50/NHERF, VASP or coffiln do not have major impact on the localization of EAAT2 to the filopodia, and the localization of these proteins to the filopodia is weaker than that of EAAT2. These results suggests that EAAT2 plays some role in filopodia regulation in addition to turning off the glutamate signal and protecting neurons by glutamate re-uptake.

上部に戻る

前に戻る