一般口演

一般口演8

2021/10/1　11:00～12:00　オンデマンド D会場 O8-1 グリア細胞からの腸管神経新生におけるLRRK2の役割の解明 Leucine-rich repeat kinase 2 regulates the acquisition of bi-phenotype in enteric neurons. ^○前川達則, Rei Kawashima, Fumitaka Kawakami, Daichi Moriya, Takafumi Ichikawa 北里大学大学院　医療系研究科　生体制御生化学 ^○Tatsunori Maekawa, Rei Kawashima, Fumitaka Kawakami, Daichi Moriya, Takafumi Ichikawa Department of Regulation Biochemistry, Graduate School of Medical Sciences, Kitasato University [Background and aim] The enteric nervous system (ENS) had been thought that no newborn neurons are generated in mature ENS. However, recent findings reported new evidence in which enteric glial cells have the potential to differentiate into neurons through bi-phenotypic status that express both glial specific protein and neuronal specific protein under certain conditions. Among these backgrounds, we have revealed that leucine-rich repeat kinase 2 (LRRK2) is expressed in the ENS and our previous analysis showed that LRRK2 knockout (KO) mice have more bi-phenotypic cells than control mice. In this study, to elucidate the role of LRRK2 in the differentiation of bi-phenotypic cells in the ENS, we performed 1) validation of the factors initiating the differentiation of bi-phenotypic cells in vivo and 2) identification of LRRK2-expressing cells in the ENS. [Methods] Immunofluorescence staining was performed to identify subtypes of enteric neuron and glial cell. 5-HT4 agonist and EdU were administrated into mice followed by colon preparations were subjected to immunofluorescence staining. LRRK2 reporter mice were generated and analyzed by immunofluorescence staining. [Results] KO mice didn’t show that the increment of bi-phenotypic cells after the treatment of 5-HT4 agonist, as shown in WT mice. EdU positive cells were identified in glial cells but neurons. The expression of LRRK2 in both enteric neurons and glial cells. [Conclusion] Given that EdU positive cells were observed only in glial cells in LRRK2 KO mice, proliferated cells could be a compensatory effect following the increment of differentiated bi-phenotypic cells in KO mice. LRRK2-expression in glial cells indicates that LRRK2 has a role under 5-HT4 signaling pathway in the glial cell.		2021/10/1　11:00～12:00　オンデマンド D会場 O8-2 サブタイプ特異的なヒト神経細胞をカスタム作成する Platform for robust and custom generation of various subtype-specific human neurons ^○石川充, Emi Sen, Hideyuki Okano Department of Physiology, Keio University School of Medicine ^○Mitsuru Ishikawa, Emi Sen, Hideyuki Okano Department of Physiology, Keio University School of Medicine A large number of reports has been already accumulated on methods to induce differentiation of human ES/iPS cells into various types of neurons. However, due to technical issues, not all of these methods can be easily reproduced in any research or facility. The Dual-SMAD inhibition method, which is often used for neuroectoderm differentiation, is an enormously powerful tool, but the state of intracellular SMAD signaling and brain region patterning in Petri dishes is ever-changing and difficult to control properly. Another issue is that human iPS cells tend to have high clonal variation. To solve these issues, we have focused on and improved a method to drive neural differentiation using transient gene expression by introducing pro-neural factors NEUROG2 or ASCL1 into human ES/iPS cells. We successfully introduced these TetO-driven genes into hES/iPS cells with PiggyBac vector and induced neuronal differentiation by treating doxycycline. In particular, the PiggyBac vector can be used to increase the number of copies of GOI inserted into the genome. The vector in this study also contains drug resistance genes, allowing for drug selection at the iPS cell stage. Therefore, the selected iPS cells can be differentiated into neurons with a high efficiency of almost 100%, and at the same time, they can be grown, maintained, and cryopreserved at a pluripotent stem cell state. Having achieved such a robust induction of neuronal differentiation, our next step is to investigate the production of subtype-specific neurons in a semi-comprehensive manner. In this study, we will introduce our attempt to produce more diverse neurons individually according to the experimental purpose while utilizing scRNA-Seq results.

2021/10/1　11:00～12:00　オンデマンド D会場 O8-3 真皮マクロファージはNGF発現レベルを調節して痛覚感受性を決定する Dermal macrophages set pain sensitivity by modulating tissue NGF levels ^○田中達英¹, Hiroaki Okuda², Ayami Isonishi¹, Kouko Tatsumi¹, Akio Wanaka¹ 1.奈良県立医科大学医学部解剖学第2講座, 2.Department of Functional Anatomy, Graduate School of Medical Science, Kanazawa University ^○Tatsuhide Tanaka¹, Hiroaki Okuda², Ayami Isonishi¹, Kouko Tatsumi¹, Akio Wanaka¹ 1.Department of Anatomy and Neuroscience, Nara Medical University, 2.Department of Functional Anatomy, Graduate School of Medical Science, Kanazawa University Crosstalk between peripheral neurons and immune cells plays important roles in pain sensation. We identified sorting nexin 25 (Snx25) as a pain-modulating gene in a transgenic mouse line with reduced pain behavior. Snx25 conditional-KO (cKO) in monocyte/macrophage-lineage cells but not in the peripheral sensory neurons reduced pain responses in both normal and neuropathic conditions. Cross transplantation experiments of bone marrows between cKO and wild type (WT) mice revealed that cKO macrophages caused dull phenotype in WT mice and WT macrophages in turn increased pain behavior in cKO mice. SNX25 in dermal macrophages enhances NGF production. We conclude that dermal macrophages set pain sensitivity by producing and secreting NGF into the dermis in addition to their host defense functions.		2021/10/1　11:00～12:00　オンデマンド D会場 O8-4 【演題取消】