論文 - 木村　俊介

Airway M cells arise in the lower airway due to RANKL signaling and reside in the bronchiolar epithelium associated with iBALT in murine models of respiratory disease

Shunsuke Kimura*, Mami Mutoh, Meri Hisamoto, Hikaru Saito, Shun Takahashi, Takanori Asakura, Makoto Ishii, Yutaka Nakamura, Junichiro Iida, Koji Hase and Toshihiko Iwanaga

Frontiers in Immunology 10   1323 - 1323 2019年06月

研究論文（学術雑誌）, 共著, 査読有り

　概要を見る

Microfold (M) cells residing in the follicle-associated epithelium of mucosa-associated lymphoid tissues are specialized for sampling luminal antigens to initiate mucosal immune responses. In the past decade, glycoprotein 2 (GP2) and Tnfaip2 were identified as reliable markers for M cells in the Peyer's patches of the intestine. Furthermore, RANKL-RANK signaling, as well as the canonical and non-canonical NFκB pathways downstream, is essential for M-cell differentiation from the intestinal stem cells. However, the molecular characterization and differentiation mechanisms of M cells in the lower respiratory tract, where organized lymphoid tissues exist rarely, remain to be fully elucidated. Therefore, this study aimed to explore M cells in the lower respiratory tract in terms of their specific molecular markers, differentiation mechanism, and functions. Immunofluorescence analysis revealed a small number of M cells expressing GP2, Tnfaip2, and RANK is present in the lower respiratory tract of healthy mice. The intraperitoneal administration of RANKL in mice effectively induced M cells, which have a high capacity to take up luminal substrates, in the lower respiratory epithelium. The airway M cells associated with lymphoid follicles were frequently detected in the pathologically induced bronchus-associated lymphoid tissue (iBALT) in the murine models of autoimmune disease as well as pulmonary emphysema. These findings demonstrate that RANKL is a common inducer of M cells in the airway and digestive tracts and that M cells are associated with the respiratory disease. We also established a two-dimensional culture method for airway M cells from the tracheal epithelium in the presence of RANKL successfully. This model may be useful for functional studies of M cells in the sampling of antigens at airway mucosal surfaces.

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Sox8 is essential for M cell maturation to accelerate IgA response at the early stage after weaning in mice.

Kimura S, Kobayashi N, Nakamura Y, Kanaya T, Takahashi D, Fujiki R, Mutoh M, Obata Y, Iwanaga T, Nakagawa T, Kato N, Sato S, Kaisho T, Ohno H, Hase K

The Journal of experimental medicine 216 （ 4 ） 831 - 846 2019年03月

研究論文（学術雑誌）, 共著, 査読有り,  ISSN  0022-1007

　概要を見る

Microfold (M) cells residing in the follicle-associated epithelium (FAE) of the gut-associated lymphoid tissue are specialized for antigen uptake to initiate mucosal immune responses. The molecular machinery and biological significance of M cell differentiation, however, remain to be fully elucidated. Here, we demonstrate that Sox8, a member of the SRY-related HMG box transcription factor family, is specifically expressed by M cells in the intestinal epithelium. The expression of Sox8 requires activation of RANKL-RelB signaling. Chromatin immunoprecipitation and luciferase assays revealed that Sox8 directly binds the promoter region of Gp2 to increase Gp2 expression, which is the hallmark of functionally mature M cells. Furthermore, genetic deletion of Sox8 causes a marked decrease in the number of mature M cells, resulting in reduced antigen uptake in Peyer's patches. Consequently, juvenile Sox8-deficient mice showed attenuated germinal center reactions and antigen-specific IgA responses. These findings indicate that Sox8 plays an essential role in the development of M cells to establish mucosal immune responses.

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Osteoprotegerin-dependent M cell self-regulation balances the gut immunity and infection

Y Nakamura, S Kimura, N Kobayashi, K Hase

EUROPEAN JOURNAL OF IMMUNOLOGY 49, 1379-1379 2019年

Intestinal Lymphatic Endothelial Cells Produce R-Spondin3.

Ogasawara R, Hashimoto D, Kimura S, Hayase E, Ara T, Takahashi S, Ohigashi H, Yoshioka K, Tateno T, Yokoyama E, Ebata K, Kondo T, Sugita J, Onozawa M, Iwanaga T, Teshima T

Scientific reports 8 （ 1 ） 10719 - 10719 2018年07月

共著, 査読有り

　概要を見る

The R-Spondin (R-Spo) family regulates WNT signaling and stimulates the proliferation and differentiation of intestinal stem cells (ISCs). R-Spo plays a critical role in maintaining intestinal homeostasis, but endogenous producers of R-Spo in the intestine remain to be investigated. We found that R-Spo3 was the major R-Spo family member produced in the intestine and it was predominantly produced by CD45-CD90+CD31+ lymphatic endothelial cells (LECs) in the lamina propria of the intestinal mucosa. Transcriptome analysis demonstrated that LECs highly expressed R-Spo receptor, Lgr5, suggesting an autocrine stimulatory loop in LECs. LECs were significantly reduced in number, and their R-Spo3 production was impaired in intestinal graft-versus-host disease (GVHD) after allogeneic hematopoietic stem cell transplantation. The impaired production of R-Spo3 in the intestine may be a novel mechanism of delayed tissue repair and defective mucosal defense in intestinal GVHD. We demonstrate a novel role of intestinal LECs in producing R-Spondin3 to maintain intestinal homeostasis.

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Histochemical characteristics of regressing vessels in the hyaloid vascular system of neonatal mice: Novel implication for vascular atrophy.

Kishimoto A, Kimura S, Nio-Kobayashi J, Takahashi-Iwanaga H, Park AM, Iwanaga T

Experimental eye research （Academic Press）  172   1 - 9 2018年03月

研究論文（学術雑誌）, 共著, 査読有り,  ISSN  0014-4835

　概要を見る

The hyaloid vasculature constitutes a transitory system nourishing the internal structures of the developing eye, but the mechanism of vascular regression and its cell biological characteristics are not fully understood. The present study aimed to reveal the specificity of the hyaloid vessels by a systematic immunohistochemical approach for marker substances of myeloid cells and the extracellular matrix (ECM) in neonatal mice. Macrophages immunoreactive for F4/80, cathepsin D, and LYVE-1 gathered around the vasa hyaloidea propria (VHP), while small round cells in vascular lumen of VHP were selectively immunoreactive for galectin-3
their segmented nuclei and immunoreactivities for Ly-6G, CD11b, and myeloperoxidase indicated their neutrophilic origin. VHP possessed thick ECM and a dense pericyte envelope as demonstrated by immunostaining for laminin, type IV collagen, integrin β1, and NG2. The galectin-3+ cells loosely aggregated with numerous erythrocytes in the lumen of hyaloid vessels in a manner reminiscent of vascular congestion. Galectin-3 is known to polymerize and form a complex with ECM and NG2 as well as recruit leukocytes on the endothelium. Observation of galectin-3 KO mice implicated the involvement of galectin-3 in the regression of hyaloid vasculature. Since macrophages may play central roles including blocking of the blood flow and the induction of apoptosis in the regression, galectin-3+ neutrophils may play a supportive role in the macrophage-mediated involution of the hyaloid vascular system.

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Molecular insights into the mechanisms of M-cell differentiation and transcytosis in the mucosa-associated lymphoid tissues.

Kimura S

Anatomical science international （Springer Tokyo）  93 （ 1 ） 23 - 34 2018年01月

単著, 査読有り,  ISSN  1447-6959

　概要を見る

Microfold cells (M cells), which are located in the follicle-associated epithelium (FAE) covering mucosal lymphoid follicles, are specialized epithelial cells that initiate mucosal immune responses. These cells take luminal antigens and transport them via transcytosis across the FAE to the antigen-presenting cells underneath. Several intestinal pathogens exploit M cells as their portal for entry to invade the host and cause disease conditions. Recent studies have revealed that the uptake of antigens by M cells is essential for efficient antigen-specific IgA production and that this process likely maintains the homeostasis of mucosal tissues. The present article reviews recent advances in understanding the molecular mechanism of M-cell differentiation and describes the molecules expressed by M cells that are associated with antigen uptake and/or the transcytosis process. Current efforts to augment M-cell-mediated uptake for use in the development of effective mucosal vaccines are also discussed.

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Development of intestinal M cells and follicle-associated epithelium is regulated by TRAF6-mediated NF-κB signaling.

Kanaya T, Sakakibara S, Jinnohara T, Hachisuka M, Tachibana N, Hidano S, Kobayashi T, Kimura S, Iwanaga T, Nakagawa T, Katsuno T, Kato N, Akiyama T, Sato T, Williams IR, Ohno H

The Journal of experimental medicine 215 （ 2 ） 501 - 519 2018年01月

研究論文（学術雑誌）, 共著, 査読有り,  ISSN  0022-1007

　概要を見る

M cells are located in the follicle-associated epithelium (FAE) that covers Peyer's patches (PPs) and are responsible for the uptake of intestinal antigens. The differentiation of M cells is initiated by receptor activator of NF-κB. However, the intracellular pathways involved in M cell differentiation are still elusive. In this study, we demonstrate that the NF-κB pathway activated by RANK is essential for M cell differentiation using in vitro organoid culture. Overexpression of NF-κB transcription factors enhances the expression of M cell-associated molecules but is not sufficient to complete M cell differentiation. Furthermore, we evaluated the requirement for tumor necrosis factor receptor-associated factor 6 (TRAF6). Conditional deletion of TRAF6 in the intestinal epithelium causes a complete loss of M cells in PPs, resulting in impaired antigen uptake into PPs. In addition, the expression of FAE-associated genes is almost silenced in TRAF6-deficient mice. This study thus demonstrates the crucial role of TRAF6-mediated NF-κB signaling in the development of M cells and FAE.

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M cell-dependent antigen uptake on follicle-associated epithelium for mucosal immune surveillance.

Nakamura Y, Kimura S, Hase K

Inflammation and regeneration 38   15 - 15 2018年

共著, 査読有り,  ISSN  1880-9693

　概要を見る

The follicle-associated epithelium (FAE) covering mucosa-associated lymphoid tissue is distinct from the villous epithelium in cellular composition and functions. Interleukin-22 binding protein (IL-22BP), provided by dendritic cells at the sub-epithelial dome region, inhibits the IL-22-mediated secretion of antimicrobial peptides by the FAE. The Notch signal from stromal cells underneath the FAE diminishes goblet cell differentiation. These events dampen the mucosal barrier functions to allow luminal microorganisms to readily gain access to the luminal surface of the FAE. Furthermore, receptor activator of nucleic factor-kappa B ligand (RANKL) from a certain stromal cell type induces differentiation into microfold (M) cells that specialize in antigen uptake in the mucosa. Microfold (M) cells play a key role in mucosal immune surveillance by actively transporting external antigens from the gut lumen to the lymphoid follicle. The molecular basis of antigen uptake by M cells has been gradually identified in the last decade. For example, GPI-anchored molecules (e.g., glycoprotein 2 (GP2) and cellular prion protein (PrPC)) and β1-integrin facilitate the transport of specific types of xenobiotics. The antigen transport by M cells initiates antigen-specific mucosal immune responses represented by the induction of secretory immunoglobulin A (S-IgA). Meanwhile, several invasive pathogens exploit M cells as a portal to establish a systemic infection. Recent findings have uncovered the molecular machinery of differentiation and functions of M cells.

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Enhanced gap junction intercellular communication inhibits catabolic and pro-inflammatory responses in tenocytes against heat stress.

Maeda E, Kimura S, Yamada M, Tashiro M, Ohashi T

Journal of cell communication and signaling （Springer Netherlands）  11 （ 4 ） 369 - 380 2017年06月

研究論文（学術雑誌）, 共著, 査読有り,  ISSN  1873-9601

　概要を見る

Elevation of tendon core temperature during severe activity is well known. However, its effects on tenocyte function have not been studied in detail. The present study tested a hypothesis that heat stimulation upregulates tenocyte catabolism, which can be modulated by the inhibition or the enhancement of gap junction intercellular communication (GJIC). Tenocytes isolated from rabbit Achilles tendons were subjected to heat stimulation at 37 °C, 41 °C or 43 °C for 30 min, and changes in cell viability, gene expressions and GJIC were examined. It was found that GJIC exhibited no changes by the stimulation even at 43 °C, but cell viability was decreased and catabolic and proinflammatory gene expressions were upregulated. Inhibition of GJIC demonstrated further upregulated catabolic and proinflammatory gene expressions. In contrast, enhanced GJIC, resulting from forced upregulation of connexin 43 gene, counteracted the heat-induced upregulation of catabolic and proinflammatory genes. These findings suggest that the temperature rise in tendon core could upregulate catabolic and proinflammatory activities, potentially leading to the onset of tendinopathy, and such upregulations could be suppressed by the enhancement of GJIC. Therefore, to prevent tendon injury at an early stage from becoming chronic injury, tendon core temperature and GJIC could be targets for post-activity treatments.

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Intrarenal signaling mediated by CCK plays a role in salt intake-induced natriuresis.

Takahashi-Iwanaga H, Kimura S, Konno K, Watanabe M, Iwanaga T

American journal of physiology. Renal physiology （AMER PHYSIOLOGICAL SOC）  313 （ 1 ） ajprenal.00539.2016 - F29 2017年03月

研究論文（学術雑誌）, 共著, 査読有り,  ISSN  1931-857X

　概要を見る

The natriuretic hormone CCK exhibits its gene transcripts in total kidney extracts. To test the possibility of CCK acting as an intrarenal mediator of sodium excretion, we examined mouse kidneys by 1) an in situ hybridization technique for CCK mRNA in animals fed a normal- or a high-sodium diet; 2) immuno-electron microscopy for the CCK peptide, 3) an in situ hybridization method and immunohistochemistry for the CCK-specific receptor CCKAR; 4) confocal image analysis of receptor-mediated Ca2+ responses in isolated renal tubules; and 5) metabolic cage experiments for the measurement of urinary sodium excretion in high-salt-fed mice either treated or untreated with the CCKAR antagonist lorglumide. Results showed the CCK gene to be expressed intensely in the inner medulla and moderately in the inner stripe of the outer medulla, with the expression in the latter being enhanced by high sodium intake. Immunoreactivity for the CCK peptide was localized to the rough endoplasmic reticulum of the medullary interstitial cells in corresponding renal regions, confirming it to be a secretory protein. Gene transcripts, protein products, and the functional activity for CCKAR were consistently localized to the late proximal tubule segments (S2 and S3) in the medullary rays, and the outer stripe of the outer medulla. Lorglumide significantly diminished natriuretic responses of mice to a dietary sodium load without altering the glomerular filtration rate. These findings suggest that the medullary interstitial cells respond to body fluid expansion by CCK release for feedback regulation of the late proximal tubular reabsorption.

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Macrophage ubiquitin-specific protease 2 modifies insulin sensitivity in obese mice

Natsuko Saito, Shunsuke Kimura, Tomomi Miyamoto, Sanae Fukushima, Misato Amagasa, Yoshinori Shimamoto, Chieko Nishioka, Shiki Okamoto, Chitoku Toda, Kohei Washio, Atsushi Asano, Ichiro Miyoshi, Eiki Takahashi, Hiroshi Kitamura

Biochemistry and Biophysics Reports （Elsevier B.V.）  9   322 - 329 2017年03月

研究論文（学術雑誌）, 共著, 査読有り,  ISSN  2405-5808

　概要を見る

We previously reported that ubiquitin-specific protease (USP) 2 in macrophages down-regulates genes associated with metabolic diseases, suggesting a putative anti-diabetic role for USP2 in macrophages. In this study, we evaluate this role at both cellular and individual levels. Isolated macrophages forcibly expressing Usp2a, a longer splicing variant of USP2, failed to modulate the insulin sensitivity of 3T3-L1 adipocytes. Similarly, macrophage-selective overexpression of Usp2a in mice (Usp2a transgenic mice) had a negligible effect on insulin sensitivity relative to wild type littermates following a three-month high-fat diet. However, Usp2a transgenic mice exhibited fewer M1 macrophages in their mesenteric adipose tissue. Following a six-month high-fat diet, Usp2a transgenic mice exhibited a retarded progression of insulin resistance in their skeletal muscle and liver, and an improvement in insulin sensitivity at an individual level. Although conditioned media from Usp2a-overexpressing macrophages did not directly affect the insulin sensitivity of C2C12 myotubes compared to media from control macrophages, they did increase the insulin sensitivity of C2C12 cells after subsequent conditioning with 3T3-L1 cells. These results indicate that macrophage USP2A hampers obesity-elicited insulin resistance via an adipocyte-dependent mechanism.

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Tunneling Nanotubes and Gap Junctions-Their Role in Long-Range Intercellular Communication during Development, Health, and Disease Conditions.

Ariazi J, Benowitz A, De Biasi V, Den Boer ML, Cherqui S, Cui H, Douillet N, Eugenin EA, Favre D, Goodman S, Gousset K, Hanein D, Israel DI, Kimura S, Kirkpatrick RB, Kuhn N, Jeong C, Lou E, Mailliard R, Maio S, Okafo G, Osswald M, Pasquier J, Polak R, Pradel G, de Rooij B, Schaeffer P, Skeberdis VA, Smith IF, Tanveer A, Volkmann N, Wu Z, Zurzolo C

Frontiers in molecular neuroscience （FRONTIERS MEDIA SA）  10   333 2017年

共著, 査読有り,  ISSN  1662-5099

　概要を見る

Cell-to-cell communication is essential for the organization, coordination, and development of cellular networks and multi-cellular systems. Intercellular communication is mediated by soluble factors (including growth factors, neurotransmitters, and cytokines/chemokines), gap junctions, exosomes and recently described tunneling nanotubes (TNTs). It is unknown whether a combination of these communication mechanisms such as TNTs and gap junctions may be important, but further research is required. TNTs are long cytoplasmic bridges that enable long-range, directed communication between connected cells. The proposed functions of TNTs are diverse and not well understood but have been shown to include the cell-to-cell transfer of vesicles, organelles, electrical stimuli and small molecules. However, the exact role of TNTs and gap junctions for intercellular communication and their impact on disease is still uncertain and thus, the subject of much debate. The combined data from numerous laboratories indicate that some TNT mediate a long-range gap junctional communication to coordinate metabolism and signaling, in relation to infectious, genetic, metabolic, cancer, and age-related diseases. This review aims to describe the current knowledge, challenges and future perspectives to characterize and explore this new intercellular communication system and to design TNT-based therapeutic strategies.

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Tunneling nanotubes and gap junctions–their role in long-range intercellular communication during development, health, and disease conditions

J Ariazi, A Benowitz, V De Biasi, ML Den Boer, S Cherqui, H Cui, ...

Frontiers in molecular neuroscience 10, 333 2017年

Zinc Transporter SLC39A7/ZIP7 Promotes Intestinal Epithelial Self-Renewal by Resolving ER Stress.

Ohashi W, Kimura S, Iwanaga T, Furusawa Y, Irié T, Izumi H, Watanabe T, Hijikata A, Hara T, Ohara O, Koseki H, Sato T, Robine S, Mori H, Hattori Y, Watarai H, Mishima K, Ohno H, Hase K, Fukada T

PLoS genetics 12 （ 10 ） e1006349 2016年10月

研究論文（学術雑誌）, 共著, 査読有り,  ISSN  1553-7390

　概要を見る

Zinc transporters play a critical role in spatiotemporal regulation of zinc homeostasis. Although disruption of zinc homeostasis has been implicated in disorders such as intestinal inflammation and aberrant epithelial morphology, it is largely unknown which zinc transporters are responsible for the intestinal epithelial homeostasis. Here, we show that Zrt-Irt-like protein (ZIP) transporter ZIP7, which is highly expressed in the intestinal crypt, is essential for intestinal epithelial proliferation. Mice lacking Zip7 in intestinal epithelium triggered endoplasmic reticulum (ER) stress in proliferative progenitor cells, leading to significant cell death of progenitor cells. Zip7 deficiency led to the loss of Olfm4+ intestinal stem cells and the degeneration of post-mitotic Paneth cells, indicating a fundamental requirement for Zip7 in homeostatic intestinal regeneration. Taken together, these findings provide evidence for the importance of ZIP7 in maintenance of intestinal epithelial homeostasis through the regulation of ER function in proliferative progenitor cells and maintenance of intestinal stem cells. Therapeutic targeting of ZIP7 could lead to effective treatment of gastrointestinal disorders.

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Distinct Roles for the N- and C-terminal Regions of M-Sec in Plasma Membrane Deformation during Tunneling Nanotube Formation

Kimura S, Yamashita M, Yamakami-Kimura M, Sato Y, Yamagata A, Kobashigawa Y, Inagaki F, Amada T, Hase K, Iwanaga T, Ohno H and Fukai S

Scientific Reports （NATURE PUBLISHING GROUP）  6   33548 2016年09月

研究論文（学術雑誌）, 共著, 査読有り,  ISSN  2045-2322

　概要を見る

The tunneling nanotube (TNT) is a structure used for intercellular communication, and is a thin membrane protrusion mediating transport of various signaling molecules and cellular components. M-Sec has potent membrane deformation ability and induces TNT formation in cooperation with the Ral/exocyst complex. Here, we show that the N-terminal polybasic region of M-Sec directly binds phosphatidylinositol (4,5)-bisphosphate for its localization to the plasma membrane during the initial stage of TNT formation. We further report a crystal structure of M-Sec, which consists of helix bundles arranged in a straight rod-like shape, similar to the membrane tethering complex subunits. A positively charged surface in the C-terminal domains is required for M-Sec interaction with active RalA to extend the plasma membrane protrusions. Our results suggest that the membrane-associated M-Sec recruits active RalA, which directs the exocyst complex to form TNTs.

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Osteoprotegerinによる腸管上皮M細胞の分化抑制機構とその意義

木村 俊介, 武藤 麻未, 岩永 敏彦

Journal of Oral Biosciences Supplement （(一社)歯科基礎医学会）  2016   168 - 168 2016年09月

共著,  ISSN  2187-2333

Receptor activator of NFκB ligand(RANKL)によるマウス気管・気管支上皮からのM細胞分化誘導

武藤 麻未, 木村 俊介, 吉沢 早織, 岩永 敏彦, 飯田 順一郎

Journal of Oral Biosciences Supplement （(一社)歯科基礎医学会）  2016   277 - 277 2016年09月

共著,  ISSN  2187-2333

Neural FFA3 activation inversely regulates anion secretion evoked by nicotinic ACh receptor activation in rat proximal colon.

Kaji I, Akiba Y, Konno K, Watanabe M, Kimura S, Iwanaga T, Kuri A, Iwamoto K, Kuwahara A, Kaunitz JD

The Journal of physiology （WILEY-BLACKWELL）  594 （ 12 ） 3339 - 3352 2016年06月

研究論文（学術雑誌）, 共著, 査読有り,  ISSN  0022-3751

　概要を見る

The proximal colonic mucosa is constantly exposed to high concentrations of microbially-produced short-chain fatty acids (SCFAs). Although luminal SCFAs evoke electrogenic anion secretion and smooth muscle contractility via neural and non-neural cholinergic pathways in the colon, the involvement of the SCFA receptor free fatty acid receptor (FFA)3, one of the free fatty acid receptor family members, has not been clarified. We investigated the contribution of FFA3 to cholinergic-mediated secretory responses in rat proximal colon. FFA3 was immunolocalized to enteroendocrine cells and to the enteric neural plexuses. Most FFA3-immunoreactive nerve fibres and nerve endings were cholinergic, colocalized with protein gene product (PGP)9.5, the vesicular ACh transporter, and the high-affinity choline transporter CHT1. In Ussing chambered mucosa-submucosa preparations (including the submucosal plexus) of rat proximal colon, carbachol (CCh)-induced Cl- secretion was decreased by TTX, hexamethonium, and the serosal FFA3 agonists acetate or propionate, although not by an inactive analogue 3-chloropropionate. Serosal application of a selective FFA3 agonist (N-[2-methylphenyl]-[4-furan-3-yl]-2-methyl-5-oxo-1,4,5,6,7,8-hexahydro-quinoline-3-carboxamide; MQC) dose-dependently suppressed the response to CCh but not to forskolin, with an IC50 of 13m. Pretreatment with MQC inhibited nicotine-evoked but not bethanechol-evoked secretion. The inhibitory effect of MQC was reversed by pretreatment with pertussis toxin, indicating that FFA3 acts via the G(i/o) pathway. Luminal propionate induced Cl- secretion via the cholinergic pathway, which was reduced by MQC, as well as by TTX, hexamethonium or removal of the submucosal plexus. These results suggest that the SCFA-FFA3 pathway has a novel anti-secretory function in that it inhibits cholinergic neural reflexes in the enteric nervous system.

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Non-neuronal, but atropine-sensitive ileal contractile responses to short-chain fatty acids: age-dependent desensitization and restoration under inflammatory conditions in mice.

Yajima M, Kimura S, Karaki S, Nio-Kobayashi J, Tsuruta T, Kuwahara A, Yajima T, Iwanaga T

Physiological reports （Wiley-Blackwell Publishing Ltd）  4 （ 7 ） 1 - 11 2016年04月

研究論文（学術雑誌）, 共著, 査読有り,  ISSN  2051-817X

　概要を見る

Intestinal epithelial cells sense short-chain fatty acids (SCFAs) to secrete non-neuronal acetylcholine (ACh). However, the roles of luminal SCFAs and epithelial ACh under normal and pathological conditions remain unknown. We examined ileal contractile responses to SCFAs at different ages and their mucosal cholinergic alterations under inflammatory conditions. Ileal contractile responses to SCFAs in 1-day-old pups to 7-week-old mice were compared using an isotonic transducer, and responses to an intraperitoneal injection of lipopolysaccharide (LPS) were analyzed in 7-week-old mice. The mRNA expression levels of a SCFA activate free fatty acid receptor, acetylcholinesterase (AChE), choline acetyltransferase (Chat), and choline transporter-like protein 4 (CTL4) were measured using real-time quantitative RT-PCR. AChE was analyzed by histochemical and optical enzymatic assays. Atropine-sensitive ileal contractile responses to SCFAs occurred in all 1-day-old pups, but were frequently desensitized after the weaning period. These contractile responses were not inhibited by tetrodotoxin and did not appear when the mucosal layer had been scraped off. Contractile desensitization in 7-week-old mice was abolished in the presence of the AChE inhibitor, eserine, which was consistent with increased AChE activity after weaning. Ileal contractions to SCFAs in adult mice were restored by LPS, which significantly increased the epithelial mRNA expression of Chat and CTL4. Atropine-sensitive ileal contractile responses to SCFAs constitutively occur in the newborn period, and are desensitized during developmental stages following the up-regulated expression of AChE in the villous mucosa, but are restored under inflammatory conditions possibly via the release of epithelial ACh.

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Potential Role of the Formation of Tunneling Nanotubes in HIV-1 Spread in Macrophages.

Hashimoto M, Bhuyan F, Hiyoshi M, Noyori O, Nasser H, Miyazaki M, Saito T, Kondoh Y, Osada H, Kimura S, Hase K, Ohno H, Suzu S

Journal of immunology (Baltimore, Md. : 1950) （AMER ASSOC IMMUNOLOGISTS）  196 （ 4 ） 1832 - 1841 2016年02月

研究論文（学術雑誌）, 共著, 査読有り,  ISSN  0022-1767

　概要を見る

Tunneling nanotubes (TNTs), the long membrane extensions connecting distant cells, have emerged as a novel form of cell-to-cell communication. However, it is not fully understood how and to what extent TNTs contribute to intercellular spread of pathogens including HIV-1. In this study, we show that HIV-1 promotes TNT formation per se via its protein Nef and a cellular protein M-Sec, which appears to mediate approximately half of viral spread among monocyte-derived macrophages (MDMs). A small compound that inhibits M-Sec-induced TNT formation reduced HIV-1 production by almost half in MDMs. Such inhibition was not observed with Nef-deficient mutant HIV-1 that fails to promote TNT formation and replicates less efficiently than the wild-type HIV-1 in MDMs. The TNT inhibitor-sensitive/Nef-promoting viral production was also observed in a T cell line ectopically expressing M-Sec, but not in another M-Sec 2 T cell line. Our results suggest the importance of TNTs in HIV-1 spread among MDMs and might answer the long-standing question how Nef promotes HIV-1 production in a cell type-specific manner.

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