一般口演（Oral）

Oral Glia 一般口演グリア

7月26日（金）15:10～15:25　第9会場（朱鷺メッセ　3F　306+307） 2O-09a1-1 グリア浮腫は体温/機械刺激センサーTRPV4を介して神経細胞死を誘導する Koji Shibasaki（柴崎貢志）¹，Shouta Sugio（杉尾翔太）¹，Francois Seghers（Seghers Francois）²，David Krizaj（Krizaj David）³，Philippe Gailly（Gailly Phillipe）²，Yasuki Ishizaki（石崎泰樹）¹，Hidetaka Matsumoto（松本英孝）⁴ ¹群馬大院医分子細胞生物 ²Lab. Cell Physiol., Institute of Neurosci., Brussels, Belgium ³Dep. of Ophth, Univ Utah Sch Medicine ⁴群馬大医眼科 We successfully developed an acute retinal detachment model by region specific injection of hyaluronic acid in mice. By utilizing this specific model system, we investigated the molecular mechanisms how cell death of photoreceptors was triggered by retinal detachment. We focused on the TRPV4 ion channel, which is a thermosensor, osmosensor, mechanosensor or lipid sensor. We analyzed the retinal expression of TRPV4 by in situ hybridization, immunohistochemistry, a Ca ²⁺-imaging method or a whole cell patch clamp recording, and confirmed the TRPV4 was functionally expressed in Muller glia as previously shown (J. Neurosci. 2012). We previously reported that TRPV4 is expressed in ~30% subpopulation of astrocytes in brain, and the TRPV4 activation leads to gliotransmitter (ATP and glutamate) release and increases synaptic transmission (Shibasaki et al. JBC 2014). Because Muller glia performs many astrocyte-specific functions within the retina, we hypothesized that TRPV4 channels might contribute to the transduction of mechanical stress associated with retinal detachment. Retinal detachment in wild type mice induced apoptosis of rod photoreceptors but the numbers were significantly reduced (approximately 50%) in Muller glia specific TRPV4CKO mice. These results indicate that the TRPV4 activation was involved in photoreceptor cell death by the retinal detachment. Furthermore, we found that the TRPV4 in Muller glia can be activated by mechanical stimuli through their swelling, resulting in release of the inflammatory cytokine MCP-1. The MCP-1 recruited many macrophages, and those cells attacked and killed photoreceptors. We propose that retinal detachment adversely impacts the viability of photoreceptor neurons via glial swelling-induced TRPV4-activation and the MCP-1 release (J Neurosci 2018). Our study identified a novel molecular pathway that could exacerbate the effects of hypoxia on neuronal survival through glial-swelling following detachment of the retina.		7月26日（金）15:25～15:40　第9会場（朱鷺メッセ　3F　306+307） 2O-09a1-2 グリア細胞のABCA1機能異常は緑内障発症を誘導する Youichi Shinozaki（篠崎陽一）¹，Kazuhiko Namekata（行方和彦）²，Kenji Kashiwagi（柏木賢治）³，Nobuhiko Ohno（大野信彦）^4,5，Takahiro Segawa（瀬川高弘）⁶，Takayuki Harada（原田高幸）²，Schuichi Koizumi（小泉修一）¹ ¹山梨大院医薬理 ²東京都医学総合研視覚病態 ³山梨大院医眼科 ⁴生理研分子生理分子神経生理 ⁵自治医大医解剖 ⁶山梨大総合分析実験セ Glaucoma is progressive optic neuropathy which is characterized by degeneration of retinal ganglion cells (RGCs) and blindness. Although elevated intraocular pressure (IOP) has long been considered as primal cause of the pathology, it has become apparent that many risk factors other than IOP are involved in the etiology of glaucoma. Recent large scale genome wide association studies (GWAS) from three independent international cohorts have shown that single nucleotide polymorphism (SNP) of ABCA1 gene has the highest correlation for pathogenesis of glaucoma. The correlation between ABCA1 gene and glaucoma has clearly demonstrated, however, its causal relationship in glaucoma is still totally unrevealed. First, it is unclear whether or not ABCA1 affects intraocular pressure (IOP). Second, which of gain-of-neurotoxicity or loss-of-function of ABCA1 is involved in glaucoma. Third, which type of cells in the retina is dominantly expressing ABCA1. In IOP measurement, we did not find any differences between WT and conventional ABCA1 knockout (ABCA1KO) mice at 3 or 12 months old. We then asked the role of ABCA1 deficiency and found that the loss-of-function causes damages of RGCs. Next we investigated the cell type mainly expressing ABCA1. By magnetic cell sorting and qPCR, Abca1 mRNA was highly enriched in retinal macroglial cell types (i.e. astrocytes and Muller cells). Immunohistochemical analysis showed fiber-like patterns of ABCA1 in retinal ganglion cell layers which were co-localized with GFAP signals. ABCA1KO mice showed significant up-regulation of GFAP and Vimentin levels. Abca1KO mice also showed hypertrophic cell body and processes in astrocytes and Muller cells, indicating reactive phenotype of these cells. To clarify the role of astroglial ABCA1, we produced macroglia-selective ABCA1 knockout (Glia-ABCA1cKO) mice. Glia-ABCA1cKO mice also showed no IOP changes and dramatic increase in apoptotic RGC number at middle-age (12 months old). Associated with RGC damages, visual function estimated by multifocal electroretinogram showed clear reduction in middle-aged Glia-ABCA1cKO mice. Based on these findings, we concluded that (1) ABCA1 has no impact on IOP at least in the mice models; (2) loss-of-function of ABCA1 is involved in glaucoma; and (3) ABCA1 in glial cells has indispensable roles for pathogenesis of glaucoma.

7月26日（金）15:40～15:55　第9会場（朱鷺メッセ　3F　306+307） 2O-09a1-3 小胞体膜貫通型転写因子OASISのアストロサイト細胞老化における役割 Koji Matsuhisa（松久幸司）¹，Atsushi Saito（齋藤敦）²，Rie Asada（浅田梨絵）¹，Masayuki Kaneko（金子雅幸）¹，Kazunori Imaizumi（今泉和則）¹ ¹広島大学大学院医歯薬保健学研究科分子細胞情報学 ²広島大学大学院医歯薬保健学研究科ストレス分子動態学 Old Astrocyte Specifically Induced Substance (OASIS) was originally identified as a gene induced in long-term cultured astrocytes. OASIS protein is an Endoplasmic reticulum (ER)-resident type-II transmembrane transcriptional factor containing transcription activation- and basic leucine zipper domains in its N-terminal cytoplasmic region. We demonstrated that OASIS protein is cleaved in its transmembrane region in response to ER stress, and the released N-terminal fragments translocate into the nucleus followed by the induction of its target genes. However, the roles of OASIS in cellular senescence remain to be elucidated. DNA damage is known as one of the major factors leading to cellular senescence. In this study, we investigated the roles of OASIS in cellular senescence induced by DNA damage. First, we examined the mRNA and protein levels of OASIS in mouse astrocytes treated with 1 μg/ml doxorubicin or irradiated with 5 Gy of X-ray, both of which induce DNA double strand break (DSB). Both mRNA and proteins of OASIS increased after the DSB. Interestingly, active forms of OASIS (cleaved N-terminal fragment of OASIS) also elevated. The levels of ER stress marker BiP and spliced form of Xbp1 mRNA were not changed, indicating that OASIS is induced and activated in a DNA damage-dependent manner, not depend on ER stress. p21 is known as a negative modulator of cell cycle progression and is required for cellular senescence. p21 is known to be one of the targets of OASIS. Induction of p21 by treatment with 1 μg/ml doxorubicin was canceled in OASIS KO astrocytes. These results indicate that OASIS promotes transcription of p21 in cellular senescence of astrocytes. We treated wild-type or OASIS KO astrocytes with 1 μg/ml doxorubicin for 3 days, and stained those cells by Senescence Associated (SA) β-gal. OASIS KO astrocytes showed no SA β-gal signals although wild-type astrocytes were intensely stained. Furthermore, the number of BrdU-labeled OASIS KO astrocytes did not decrease by the treatment with doxorubicin, indicating that OASIS plays important roles in the inhibition of cell growth caused by DNA damage. Collectively, we demonstrated that OASIS induces cellular senescence of astrocytes via promotion of transcription of p21.		7月26日（金）15:55～16:10　第9会場（朱鷺メッセ　3F　306+307） 2O-09a1-4 神経細胞はアストロサイトのself-avoidanceを誘導する Mariko Hayashi（林真理子）¹，Yuko Sekino（関野祐子）²，Kaoru Sato（佐藤薫）³ ¹国際医療福祉大 ²東京大院薬 ³国立医薬品食品衛生研薬理 An astrocyte makes thousands of finely branched processes to approach neurons. Each astrocyte expands processes to form an exclusive territory with limited penetration of processes of neighboring astrocytes. This is a phenomenon called tiling. Whether their extensive branching sterically interferes entry of other astrocyte processes, or astrocyte processes actively avoid each other to form exclusive territories was not known. Here we show that neurons induce multi-order branches of astrocytes tiled in two-dimensional in vitro culture. Astrocyte processes did not overlap with other processes, both of their own and of neighboring astrocytes. As a result, exclusive territories of astrocytes were formed. In some neurons, tiling is established by pruning of overlapped processes, as well as self-avoidance of expanding processes. Tracking morphological changes of astrocytes during process formation showed that they tile by expanding processes avoiding the direction of other processes, and not by pruning once formed processes overlapped with others. Astrocyte process in the neighborhood of other astrocyte processes tend to be stabilized and do not further expand. Astrocytes settled at DIV4 and started to expand process already showed process self-avoidance, at which stage neurons are also expanding dendrites and mature synapses are not formed yet. These astrocytes do not avoid neurons, indicating the avoidance mechanism utilizes astrocyte specific proteins induced by neurons. Our results indicate that premature neurons before synapse formation were capable of inducing astrocyte process formation and self-avoidance.