研究者データベース

木村 享史(キムラ タカシ)
獣医学研究院 獣医学部門 臨床獣医科学分野
教授

基本情報

所属

  • 獣医学研究院 獣医学部門 臨床獣医科学分野

職名

  • 教授

学位

  • 医学博士(北海道大学)

J-Global ID

研究キーワード

  • 獣医病理学   ウイルス学   Pathology   Virology   

研究分野

  • ライフサイエンス / 獣医学
  • ライフサイエンス / 獣医学
  • ライフサイエンス / 獣医学

所属学協会

  • 日本獣医病理学専門家協会   日本獣医病理学会   日本ウイルス学会   American Society for Virology   日本獣医学会   American Society for Virology   Japanese Society of Neuropathology   Japanese Society for Virology   Japanese Society of Veterinary Science   

研究活動情報

論文

  • Hiroki Yamaguchi, Shintaro Kobayashi, Akihiro Ishii, Hirohito Ogawa, Ichiro Nakamura, Ladslav Moonga, Bernard M. Hang'ombe, Aaron S. Mweene, Yuka Thomas, Takashi Kimura, Hirofumi Sawa, Yasuko Orba
    Journal of General Virology 94 6 1357 - 1364 2013年06月01日 [査読有り][通常論文]
     
    To examine polyomavirus (PyV) infection in wildlife, we investigated the presence of PyVs in Zambia with permission from the Zambia Wildlife Authority. We analysed 200 DNA samples from the spleens and kidneys (n5100 each) of yellow baboons and vervet monkeys (VMs) (n550 each). We detected seven PyV genome fragments in 200 DNA samples using a nested broadspectrum PCR method, and identified five full-length viral genomes using an inverse PCR method. Phylogenetic analysis of virally encoded proteins revealed that four PyVs were closely related to either African green monkey PyV or simian agent 12. Only one virus detected from a VM spleen was found to be related, with relatively low nucleotide sequence identity (74 %), to the chimpanzee PyV, which shares 48% nucleotide sequence identity with the human Merkel cell PyV identified from Merkel cell carcinoma. The obtained entire genome of this virus was 5157 bp and had large T- and small t-antigens, and VP1 and VP2 ORFs. This virus was tentatively named vervet monkey PyV 1 (VmPyV1) as a novel PyV. Comparison with other PyVs revealed that VmPyV1, like chimpanzee PyV, had a longer VP1 ORF. To examine whether the VmPyV1 genome could produce viral proteins in cultured cells, the whole genome was transfected into HEK293T cells. We detected VP1 protein expression in the transfected HEK293T cells by immunocytochemical and immunoblot analyses. Thus, we identified a novel PyV genome from VM spleen. © 2013 SGM.
  • Walter Muleya, Boniface Namangala, Martin Simuunza, Ryo Nakao, Noboru Inoue, Takashi Kimura, Kimihito Ito, Chihiro Sugimoto, Hirofumi Sawa
    PARASITES & VECTORS 5 255  2012年11月 [査読有り][通常論文]
     
    Background: Theileriosis, caused by Theileria parva, is an economically important disease in Africa. It is a major constraint to the development of the livestock industry in some parts of eastern, central and southern Africa. In Zambia, theileriosis causes losses of up to 10,000 cattle annually. Methods: Cattle blood samples were collected for genetic analysis of Theileria parva from Isoka and Petauke districts in Zambia. Microsatellite analysis was then performed on all Theileria parva positive samples for PCR using a panel of 9 microsatellite markers. Microsatellite data was analyzed using microsatellite toolkit, GenAlEx ver. 6, Fstat ver. 2.9.3.2, and LIAN computer softwares. Results: The combined percentage of positive samples in both districts determined by PCR using the p104 gene primers was 54.9% (95% CI: 46.7 - 63.1%, 78/142), while in each district, it was 44.8% (95% CI: 34.8 - 54.8%) and 76.1% (95% CI = 63.9 - 88.4%) for Isoka and Petauke districts, respectively. We analyzed the population genetic structure of Theileria parva from a total of 61 samples (33 from Isoka and 28 from Petauke) using a panel of 9 microsatellite markers encompassing the 4 chromosomes of Theileria parva. Wright's F index (F-ST = 0.178) showed significant differentiation between the Isoka and Petauke populations. Linkage disequilibrium was observed when populations from both districts were treated as a single population. When analyzed separately, linkage disequilibrium was observed in Kanyelele and Kalembe areas in Isoka district, Isoka district overall and in Petauke district. Petauke district had a higher multiplicity of infection than Isoka district. Conclusion: Population genetic analyses of Theileria parva from Isoka and Petauke districts showed a low level of genotype exchange between the districts, but a high level of genetic diversity within each district population, implying genetic and geographic sub-structuring between the districts. The sub-structuring observed, along with the lack of panmixia in the populations, could have been due to low transmission levels at the time of sampling. However, the Isoka population was less diverse than the Petauke population.
  • Michihito Sasaki, Agus Setiyono, Ekowati Handharyani, Ibenu Rahmadani, Siswatiana Taha, Sri Adiani, Mawar Subangkit, Hirofumi Sawa, Ichiro Nakamura, Takashi Kimura
    VIROLOGY JOURNAL 9 240  2012年10月 [査読有り][通常論文]
     
    Background: Fruit bats are known to harbor zoonotic paramyxoviruses including Nipah, Hendra, and Menangle viruses. The aim of this study was to detect the presence of paramyxovirus RNA in fruit bats from Indonesia. Methods: RNA samples were obtained from the spleens of 110 fruit bats collected from four locations in Indonesia. All samples were screened by semi-nested broad spectrum reverse transcription PCR targeting the paramyxovirus polymerase (L) genes. Results: Semi-nested reverse transcription PCR detected five previously unidentified paramyxoviruses from six fruit bats. Phylogenetic analysis showed that these virus sequences were related to henipavirus or rubulavirus. Conclusions: This study indicates the presence of novel paramyxoviruses among fruit bat populations in Indonesia.
  • Tadaki Suzuki, Shingo Semba, Yuji Sunden, Yasuko Orba, Shintaro Kobayashi, Kazuo Nagashima, Takashi Kimura, Hideki Hasegawa, Hirofumi Sawa
    MICROBIOLOGY AND IMMUNOLOGY 56 9 639 - 646 2012年09月 [査読有り][通常論文]
     
    JC virus (JCV) belongs to the polyomavirus family of double-stranded DNA viruses and causes progressive multifocal leukoencephalopathy in humans. JCV encodes early proteins (large T antigen, small T antigen, and T' antigen) and four late proteins (agnoprotein, and three viral capsid proteins, VP1, VP2, and VP3). In the current study, a novel function for JCV agnoprotein in the morphogenesis of JC virion particles was identified. It was found that mature virions of agnoprotein-negative JCV are irregularly shaped. Sucrose gradient sedimentation and cesium chloride gradient ultracentrifugation analyses revealed that the particles of virus lacking agnoprotein assemble into irregularly sized virions, and that agnoprotein alters the efficiency of formation of VP1 virus-like particles. An in vitro binding assay and immunocytochemistry revealed that agnoprotein binds to glutathione S-transferase fusion proteins of VP1 and that some fractions of agnoprotein colocalize with VP1 in the nucleus. In addition, gel filtration analysis of formation of VP1-pentamers revealed that agnoprotein enhances formation of these pentamers by interacting with VP1. The present findings suggest that JCV agnoprotein plays a role, similar to that of SV40 agnoprotein, in facilitating virion assembly.
  • Kensuke Nakamura, Masahiro Yamasaki, Tomohiro Osaki, Hiroshi Ohta, Noboru Sasaki, Keisuke Aoshima, Takashi Kimura, Mitsuyoshi Takiguchi
    JOURNAL OF THE AMERICAN ANIMAL HOSPITAL ASSOCIATION 48 5 327 - 330 2012年09月 [査読有り][通常論文]
     
    Bilateral segmental aplasia of the uterine horns with unilateral pyometra and uterine horn torsion were diagnosed in a Pomeranian bitch that presented with chronic abdominal distension and an acute onset of anorexia and lethargy. Because radiographic and ultrasonographic findings revealed the presence of markedly enlarged bilateral uterine horns filled with fluid in the caudal abdomen, a tentative diagnosis of either pyometra or hydrometra with uterine horn torsion was made. Exploratory laparotomy showed bilateral, segmentally distended uterine horns with unilateral uterine horn torsion. Ovariohysterectomy was performed, and bilateral segmental aplasia of the uterine horns with the development of unilateral uterine horn torsion was diagnosed histopathologically. To the authors' knowledge, this is the first report of uterine horn torsion in conjunction with segmental aplasia of the uterine horn in a bitch. (J Am Anim Hosp Assoc 2012; 48:327-330. DOI 10.5326/JAAHA-MS-5771)
  • Nilton Akio Muto, Yuji Sunden, Tomoe Hattori, Daisuke Fujikura, Yosuke Nakayama, Tadaaki Miyazaki, Mitsuo Maruyama, Takashi Kimura, Hirofumi Sawa
    JAPANESE JOURNAL OF INFECTIOUS DISEASES 65 5 383 - 391 2012年09月 [査読有り][通常論文]
     
    This study examined pathological changes in the lung tissues of young and aged mice infected with influenza virus. Young mice inoculated with influenza virus showed body weight loss at 4 days post-infection (dpi), meanwhile body weight decrease started from 9 dpi in the aged mice. We histopathologically examined the lungs of these mice. Immunohistochemical examination revealed that viral antigen-positive bronchiolar and alveolar epithelial cell numbers at 3 dpi were significantly higher in young mice than in the aged ones. Further, viral antigen-positive cells were observed at 9 dpi in the aged mice, but not in the young ones. Diffuse and severe bronchointerstitial pneumonia characterized by the accumulation of polymorphonuclear leukocytes (PMNs) was observed in young mice at 6 dpi. Histopathological changes in the aged mice were milder than those in the young mice. Moreover, T cell and macrophage accumulation in the lungs was significantly higher in the young mice than in the aged mice at 9 dpi. These results suggest that there may be a correlation between the relatively low level of infiltration of PMNs, macrophages, and T lymphocytes and the delayed body weight loss and longer lasting infections observed in the lungs of the aged mice. These findings provide detailed insights into the age-specific course of infection in young and aged populations with associated differences in lung pathology.
  • Shintaro Kobayashi, Yasuko Orba, Hiroki Yamaguchi, Takashi Kimura, Hirofumi Sawa
    NEUROPATHOLOGY 32 4 398 - 405 2012年08月 [査読有り][通常論文]
     
    West Nile virus (WNV) belongs to the Flaviviridae family of viruses and has emerged as a significant cause of viral encephalitis in humans, animals and birds. It has been reported that WNV replication directly induces neuronal injury, followed by neuronal cell death proven as apoptosis. Therefore, it is important to understand the mechanism of neuronal apoptosis caused by this virus to develop strategies to control its pathogenicity. Accumulation of ubiquitinated abnormal proteins has been reported to be associated with neuronal apoptosis in some pathological conditions. A lot of cellular stresses prevent cellular protein quality control mechanisms, resulting in the accumulation of ubiquitinated abnormal proteins. To obtain a better understanding of the mechanisms of WNV-induced neuronal apoptosis, we evaluated the accumulation of ubiquitinated proteins in the WNV-infected neuronal cells. We have observed that WNV infection caused massive neuronal injury in the brains of mice. Viral antigen was detected in the neuronal cytoplasm of the cells exhibiting neuronal apoptosis. Notably, ubiquitinated proteins were detected in WNV-infected neuronal cells. In addition, accumulation of ubiquitinated proteins was markedly enhanced in mouse neuroblastoma, Neuro-2a cells after WNV infection. Our histopathological and in vitro studies suggest that accumulation of ubiquitinated proteins in neuronal cells might be associated with neuronal apoptosis caused by WNV infection.
  • Nilton Akio Muto, Reiko Yoshida, Tadaki Suzuki, Shintaro Kobayashi, Hiroichi Ozaki, Daisuke Fujikura, Rashid Manzoor, Mieko Muramatsu, Ayato Takada, Takashi Kimura, Hirofumi Sawa
    JAPANESE JOURNAL OF VETERINARY RESEARCH 60 2-3 71 - 83 2012年08月 [査読有り][通常論文]
     
    New approaches to the treatment of influenza have been designed based on the highly conserved antigenicity of the M2 envelope protein among influenza A virus strains. The present study examined the anti-viral activities of an anti-M2 ectodomain (M2e) monoclonal antibody (clone rM2ss23), which binds to the M2 proteins of the influenza A virus strains A/Aichi/2/68 (H3N2) (Aichi) and A/PR/8/34 (H1N1) (PR8). The results showed that rM2ss23 bound to both Aichi and PR8 M2 proteins expressed on the cell surface. While the antibody did not prevent virus entry into cells, it significantly inhibited plaque formation by the Aichi strain in a dose-dependent manner when infected cells were cultured in the presence of the antibody. By contrast, the growth of PR8 (H1N1) was not affected by the antibody. A reverse genetics approach revealed that the inhibitory effect of rM2ss23 on the Aichi virus was abolished by replacing the genes encoding the HA and/or M proteins with those of the PR8 strain. These results suggest that rM2ss23 prevents virus release from infected cells and further suggest that the mechanisms underlying the virus budding mediates by HA and M2 proteins might differ between the Aichi and PR8 strains.
  • Walter Muleya, Boniface Namangala, Aaron Mweene, Luke Zulu, Paul Fandamu, Douglas Banda, Takashi Kimura, Hirofumi Sawa, Akihiro Ishii
    VIRUS RESEARCH 163 1 160 - 168 2012年01月 [査読有り][通常論文]
     
    The National Livestock Epidemiology and Information Center (NALEIC) in Zambia reported over 132 cases of canine rabies diagnosed by the direct fluorescent antibody test (DFAT) from 2004 to 2009. In this study, the lineage of rabies virus (RABV) in Zambia was determined by phylogenetic analysis of the nucleoprotein (N) and glycoprotein (G) gene sequences. Total RNA was extracted from 87-DFAT brain specimens out of which only 35 (40%) were positive on nested reverse transcription polymerase chain reaction (RT-PCR) for each gene, and 26 being positive for both genes. Positive specimens for the N (n =33) and G (n = 35) genes were used for phylogenetic analysis. Phylogenetic analysis of the N gene showed two phylogenetic clusters in Zambia belonging to the Africa 1b lineage present in eastern and southern Africa. While one cluster exclusively comprised Zambian strains, the other was more heterogeneous regarding the RABV origins and included strains from Tanzania, Mozambique and Zambia. Phylogenetic analysis of the G gene revealed similar RABV strains in different hosts and regions of Zambia. We designed primers for reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay from the consensus sequence of the N gene in an attempt to improve the molecular diagnosis of RABV in Zambia. The specificity and reproducibility of the RI-LAMP assay was confirmed with actual clinical specimens. Therefore, the RI-LAMP assay presented in this study may prove to be useful for routine diagnosis of rabies in Zambia. (C) 2011 Elsevier B.V. All rights reserved.
  • Michihito Sasaki, Eunmi Kim, Manabu Igarashi, Kimihito Ito, Rie Hasebe, Hideto Fukushi, Hirofumi Sawa, Takashi Kimura
    JOURNAL OF BIOLOGICAL CHEMISTRY 286 45 39370 - 39378 2011年11月 [査読有り][通常論文]
     
    Equine herpesvirus-1 (EHV-1), an alpha-herpesvirus of the family Herpesviridae, causes respiratory disease, abortion, and encephalomyelitis in horses. EHV-1 utilizes equine MHC class I molecules as entry receptors. However, hamster MHC class I molecules on EHV-1-susceptible CHO-K1 cells play no role in EHV-1 entry. To identify the MHC class I molecule region that is responsible for EHV-1 entry, domain exchange and site-directed mutagenesis experiments were performed, in which parts of the extracellular region of hamster MHC class I (clone C5) were replaced with corresponding sequences from equine MHC class I (clone A68). Substitution of alanine for glutamine at position 173 (Q173A) within the alpha 2 domain of the MHC class I molecule enabled hamster MHC class I C5 to mediate EHV-1 entry into cells. Conversely, substitution of glutamine for alanine at position 173 (A173Q) in equine MHC class I A68 resulted in loss of EHV-1 receptor function. Equine MHC class I clone 3.4, which possesses threonine at position 173, was unable to act as an EHV-1 receptor. Substitution of alanine for threonine at position 173 (T173A) enabled MHC class I 3.4 to mediate EHV-1 entry into cells. These results suggest that the amino acid residue at position 173 of the MHC class I molecule is involved in the efficiency of EHV-1 entry.
  • Jiancheng Chen, Shuhei Yamada, Yoshiki Hama, Ajaya Kumar Shetty, Takanari Kobayashi, Hiroshi Oda, Kosuke Seiki, Eunmi Kim, Takashi Kimura, Naonori Takahashi, Kazuya I. P. J. Hidari, Takashi Suzuki, Yasuo Suzuki, Kazuyuki Sugahara
    BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS 412 1 136 - 142 2011年08月 [査読有り][通常論文]
     
    The structure and biological activities of a highly sulfated heparan sulfate (HS) extracted from shrimp (Penaeus brasiliensis) heads were characterized. Structurally the shrimp HS was more heterogenous than heparin, although it is still highly sulfated. The molecular mass of the shrimp HS preparation was determined to be 32.3 kDa by gel filtration HPLC. Analysis by surface plasmon resonance demonstrated that various growth/differentiation factors specifically bound to the shrimp HS with comparable affinity. Notably, the shrimp HS had a greater inhibitory effect against infections by dengue virus type 2 as well as Japanese encephalitis virus than heparin. Experiments on anticoagulant activity indicated that the shrimp HS exhibited significant anti-thrombin activity, but less than the commercial heparin. Hence, the HS preparation from shrimp heads, an industrial waste, is a prospective agent for a variety of clinical applications. (C) 2011 Elsevier Inc. All rights reserved.
  • Eunmi Kim, Megumi Okumura, Hirofumi Sawa, Tadaaki Miyazaki, Daisuke Fujikura, Shuhei Yamada, Kazuyuki Sugahara, Michihito Sasaki, Takashi Kimura
    BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS 409 4 717 - 722 2011年06月 [査読有り][通常論文]
     
    Glycosaminoglycans (GAGs) have diverse functions in the body and are involved in viral infection. The purpose of this study was to evaluate the possible roles of the E-disaccharide units GlcA beta 1-3Gal-NAc(4,6-O-disulfate) of chondroitin sulfate (CS), a GAG involved in neuritogenesis and neuronal migration, in Japanese encephalitis virus (JEV) infection. Soluble CS-E (sCS-E) derived from squid cartilage inhibited JEV infection in African green monkey kidney-derived Vero cells and baby hamster kidney-derived BHK cells by interfering with viral attachment. In contrast, sCS-E enhanced viral infection in the mouse neuroblastoma cell line Neuro-2a, despite the fact that viral attachment to Neuro-2a cells was inhibited by sCS-E. This enhancement effect in Neuro-2a cells seemed to be related to increased viral RNA replication and was also observed in a rat infection model in which intracerebral coadministration of sCS-E with JEV in 17-day-old rats resulted in higher brain viral loads than in rats infected without sCS-E administration. These results show the paradoxical effects of sCS-E on JEV infection in different cell types and indicate that potential use of sCS-E as an antiviral agent against JEV infection should be approached with caution considering its effects in the neuron, the major target of JEV. (C) 2011 Elsevier Inc. All rights reserved.
  • Yasuko Orba, Shintaro Kobayashi, Ichiro Nakamura, Akihiro Ishii, Bernard M. Hang'ombe, Aaron S. Mweene, Yuka Thomas, Takashi Kimura, Hirofumi Sawa
    JOURNAL OF GENERAL VIROLOGY 92 4 789 - 795 2011年04月 [査読有り][通常論文]
     
    To investigate polyomavirus infection in wild rodents, we analysed DNA samples from the spleens of 100 wild rodents from Zambia using a broad-spectrum PCR-based assay. A previously unknown polyomavirus genome was identified in a sample from a multimammate mouse (Mastomys species) and the entire viral genome of 4899 bp was :subsequently sequenced. This viral genome contained potential ORFs for the capsid proteins, VP1, VP2 and VP3, and early proteins, small t antigen and large T antigen. Phylogenetic analysis showed that it was a novel member of the family Polyomaviridae, and thus the virus was tentatively named mastomys polyomavirus. After transfection of the viral genome into several mammalian cell lines, transient expression of the VP1 and large T antigen proteins was confirmed by immunoblotting and immunocytochemical analyses. Comparison of large T antigen function in mastomys polyomavirus with that in rhesus monkey polyomavirus SV40 and human polyomavirus JC virus revealed that the large T antigen from mastomys polyomavirus interacted with the tumour suppressor protein pRb, but not with p53.
  • Michihito Sasaki, Rie Hasebe, Yoshinori Makino, Tadaki Suzuki, Hideto Fukushi, Minoru Okamoto, Kazuya Matsuda, Hiroyuki Taniyama, Hirofumi Sawa, Takashi Kimura
    GENES TO CELLS 16 4 343 - 357 2011年04月 [査読有り][通常論文]
     
    The endotheliotropism of equine herpesvirus-1 (EHV-1) leads to encephalomyelitis secondary to vasculitis and thrombosis in the infected horse central nervous system (CNS). To identify the host factors involved in EHV-1 infection of CNS endothelial cells, we performed functional cloning using an equine brain microvascular endothelial cell cDNA library. Exogenous expression of equine major histocompatibility complex (MHC) class I heavy chain genes conferred susceptibility to EHV-1 infection in mouse NIH3T3 cells, which are not naturally susceptible to EHV-1 infection. Equine MHC class I molecules bound to EHV-1 glycoprotein D (gD), and both anti-gD antibodies and a soluble form of gD blocked viral entry into NIH3T3 cells stably expressing the equine MHC class I heavy chain gene (3T3-A68 cells). Treatment with an anti-equine MHC class I monoclonal antibody blocked EHV-1 entry into 3T3-A68 cells, equine dermis (E. Derm) cells and equine brain microvascular endothelial cells. In addition, inhibition of cell surface expression of MHC class I molecules in E. Derm cells drastically reduced their susceptibility to EHV-1 infection. These results suggest that equine MHC class I is a functional gD receptor that plays a pivotal role in EHV-1 entry into equine cells.
  • T. Kimura, M. Sasaki, M. Okumura, E. Kim, H. Sawa
    VETERINARY PATHOLOGY 47 5 806 - 818 2010年09月 [査読有り][通常論文]
     
    Encephalitic flaviviruses are important arthropod-borne pathogens of humans and other animals. In particular, the recent emergence of the West Nile virus (WNV) and Japanese encephalitis virus (JEV) in new geographic areas has caused a considerable public health alert and international concern. Among the experimental in vivo models of WNV and JEV infection, mice and other laboratory rodents are the most thoroughly studied and well-characterized systems, having provided data that are important for understanding the infectious process in humans. Macaca monkeys have also been used as a model for WNV and JEV infection, mainly for the evaluation of vaccine efficacy, although a limited number of published studies have addressed pathomorphology. These animal models demonstrate the development of encephalitis with many similarities to the human disease; however, the histological events that occur during infection, especially in peripheral tissues, have not been fully characterized.
  • Akira Kawaguchi, Tadaki Suzuki, Takashi Kimura, Naoki Sakai, Tokiyoshi Ayabe, Hirofumi Sawa, Hideki Hasegawa
    BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS 398 4 778 - 784 2010年08月 [査読有り][通常論文]
     
    Gene-encoded antimicrobial peptides (AMPS) are an essential component of the innate immune system in many species. Analysis of beta-defensin gene expression in mouse tissue using primers that were specific for conserved sequences located outside of the beta-defensin translated region identified a novel small gene. The novel gene had an open reading frame of 114 bp and encoded a predicted protein of 37 amino acid residues. A search of the genome database revealed that the gene locus and the sequence of exon 1 of this novel gene were similar to subgroup 1 mouse beta-defensins. A small peptide, K17 (FSPQMLQDIIEKKTKIL), derived from the amino acid sequence of this novel gene was synthesized. Circular dichroism (CD) spectroscopic analysis of chemically synthesized peptide demonstrated that the peptide exhibited random coil conformation in aqueous solution, but the peptide adopted helical conformation in the presence of trifluoroethanol or sodium dodecyl sulfate, a membrane-mimicking environment. The peptide exhibited bactericidal activity against Salmonella enterica serovar Typhimurium (Gram negative) and Staphylococcus aureus (Gram positive); it was not cytotoxic in cultures of mammalian cells or hemolytic in cultures of erythrocytes. These results suggested that K17 may be a candidate therapeutic for the treatment of bacterial infection. (C) 2010 Elsevier Inc. All rights reserved.
  • Tadaki Suzuki, Satoko Yamanouchi, Yuji Sunden, Yasuko Orba, Takashi Kimura, Hirofumi Sawa
    JOURNAL OF MEDICAL VIROLOGY 82 7 1229 - 1235 2010年07月 [査読有り][通常論文]
     
    The human polyomavirus JC virus (JCV) infects 70-80% of humans and establishes latent infection in the kidney. In immunosuppressed patients, JCV reactivates and causes a fatal and progressive neurological disease known as progressive multifocal leukoencephalopathy (PML). Over the past three decades, PML has become an important neurological complication in acquired immunodeficiency syndrome (AIDS) patients. Recently, it was reported that patients treated with therapeutics that target the integrin receptor very late antigen (VLA)-4 are at increased risk of developing PML. However, the relationship between Natalizumab and this unexpected onset of PML has yet to be elucidated. Here, we investigated the effect of Natalizumab on the growth of JCV in the permissive human neural cell line IMR-32 following viral inoculation. Natalizumab had no effect either on the expression levels of viral proteins as determined by immunoblot analysis using specific antibodies or on the hemagglutination activity of cellular lysates from infected cells. These results suggest that there is no direct effect of Natalizumab on JCV infectivity in IMR-32 cells in vitro. J. Med. Virol. 82:1229-1235, 2010. (C) 2010 Wiley-Liss, Inc.
  • Rie Hasebe, Tadaki Suzuki, Yoshinori Makino, Manabu Igarashi, Satoko Yamanouchi, Akihiko Maeda, Motohiro Horiuchi, Hirofumi Sawa, Takashi Kimura
    BMC MICROBIOLOGY 10 165  2010年06月 [査読有り][通常論文]
     
    Background: West Nile virus (WNV) causes viremia after invasion to the hosts by mosquito bite. Endothelial cells could play an important role in WNV spread from the blood stream into the central nervous system and peripheral tissues. Here, we analyzed the capacity of virus-like particles (VLPs) of the highly virulent NY99 6-LP strain (6-LP VLPs) and the low virulence Eg101 strain (Eg VLPs) to cross cultured human endothelial cells. Results: 6-LP VLPs were transported from the apical to basolateral side of endothelial cells, whereas Eg VLPs were hardly transported. The localization of tight junction marker ZO-1 and the integrity of tight junctions were not impaired during the transport of 6-LP VLPs. The transport of 6-LP VLPs was inhibited by treatment with filipin, which prevents the formation of cholesterol-dependent membrane rafts, suggesting the involvement of raft-associated membrane transport. To determine the amino acid residues responsible for the transport of VLPs, we produced mutant VLPs, in which residues of E protein were exchanged between the 6-LP and Eg strains. Double amino acid substitution of the residues 156 and 159 greatly impaired the transport of VLPs. Conclusion: Our results suggest that a transcellular pathway is associated with 6-LP VLPs transport. We also showed that the combination of the residues 156 and 159 plays an important role in the transport of VLPs across endothelial cells.
  • Tadaki Suzuki, Yasuko Orba, Yuki Okada, Yuji Sunden, Takashi Kimura, Shinya Tanaka, Kazuo Nagashima, William W. Hall, Hirofumi Sawa
    PLOS PATHOGENS 6 3 e1000801  2010年03月 [査読有り][通常論文]
     
    Virus infections can result in a range of cellular injuries and commonly this involves both the plasma and intracellular membranes, resulting in enhanced permeability. Viroporins are a group of proteins that interact with plasma membranes modifying permeability and can promote the release of viral particles. While these proteins are not essential for virus replication, their activity certainly promotes virus growth. Progressive multifocal leukoencephalopathy (PML) is a fatal demyelinating disease resulting from lytic infection of oligodendrocytes by the polyomavirus JC virus (JCV). The genome of JCV encodes six major proteins including a small auxiliary protein known as agnoprotein. Studies on other polyomavirus agnoproteins have suggested that the protein may contribute to viral propagation at various stages in the replication cycle, including transcription, translation, processing of late viral proteins, assembly of virions, and viral propagation. Previous studies from our and other laboratories have indicated that JCV agnoprotein plays an important, although as yet incompletely understood role in the propagation of JCV. Here, we demonstrate that agnoprotein possesses properties commonly associated with viroporins. Our findings demonstrate that: (i) A deletion mutant of agnoprotein is defective in virion release and viral propagation; (ii) Agnoprotein localizes to the ER early in infection, but is also found at the plasma membrane late in infection; (iii) Agnoprotein is an integral membrane protein and forms homo-oligomers; (iv) Agnoprotein enhances permeability of cells to the translation inhibitor hygromycin B; (v) Agnoprotein induces the influx of extracellular Ca(2+); (vi) The basic residues at amino acid positions 8 and 9 of agnoprotein key are determinants of the viroporin activity. The viroporin-like properties of agnoprotein result in increased membrane permeability and alterations in intracellular Ca(2+) homeostasis leading to membrane dysfunction and enhancement of virus release.
  • Yasuko Orba, Tadaki Suzuki, Yoshinori Makino, Kanako Kubota, Shinya Tanaka, Takashi Kimura, Hirofumi Sawa
    JOURNAL OF BIOLOGICAL CHEMISTRY 285 2 1544 - 1554 2010年01月 [査読有り][通常論文]
     
    Large T antigen (TAg) of the human polyomavirus JC virus (JCV) possesses DNA binding and helicase activities, which, together with various cellular proteins, are required for replication of the viral genome. We now show that JCV-infected cells expressing TAg accumulate in the G(2) phase of the cell cycle as a result of the activation of ATM- and ATR-mediated G(2) checkpoint pathways. Transient transfection of cells with a TAg expression vector also induced G(2) checkpoint signaling and G(2) arrest. Analysis of TAg mutants with different subnuclear localizations suggested that the association of TAg with cellular DNA contributes to the induction of G(2) arrest. Abrogation of G(2) arrest by inhibition of ATM and ATR, Chk1, and Wee1 suppressed JCV genome replication. In addition, abrogation of the G(2)-M transition by Cdc2 depletion disabled Wee1 depletion induced suppression of JCV genome replication, suggesting that JCV replication is facilitated by G(2) arrest resulting from G(2) checkpoint signaling. Moreover, inhibition of ATM and ATR by caffeine suppressed JCV production. The observation that oligodendrocytes productively infected with JCV in vivo also undergo G(2) arrest suggests that G(2) checkpoint inhibitors such as caffeine are potential therapeutic agents for JCV infection.
  • Akira Kawaguchi, Yasuko Orba, Takashi Kimura, Hidekatsu Iha, Masao Ogata, Takahiro Tsuji, Akira Ainai, Tetsutaro Sata, Takashi Okamoto, William W. Hall, Hirofumi Sawa, Hideki Hasegawa
    BLOOD 114 14 2961 - 2968 2009年10月 [査読有り][通常論文]
     
    Adult T-cell leukemia (ATL) is a T-cell malignancy caused by human T lymphotropic virus type I, and presents as an aggressive leukemia with characteristic widespread leukemic cell infiltration into visceral organs and skin. The molecular mechanisms associated with leukemic cell infiltration are poorly understood. We have used mouse models of ATL to investigate the role of chemokines in this process. Transfer of splenic lymphomatous cells from transgenic to SCID mice reproduces a leukemia and lymphoma that is histologically identical to human disease. It could be shown that lymphomatous cells exhibit specific chemotactic activity in response to stromal cell-derived factor-1 alpha (SDF-1 alpha). Lymphomatous cells exhibited surface expression of CXCR4, the specific receptor of SDF-1 alpha. AMD3100, a CXCR4 antagonist, was found to inhibit both SDF-1 alpha-induced migration and phosphorylation of extracellular signal-related kinase 1/2. Investigation of cultured cells from human ATL patients revealed identical findings. Using the SCID mouse model, it could be demonstrated that AMD3100 inhibited infiltration of lymphomatous cells into liver and lung tissues in vivo. These results demonstrate the involvement of the SDF-1 alpha/CXCR4 interaction as one mechanism of leukemic cell migration and this may provide a novel target as part of combination therapy for ATL. (Blood. 2009; 114: 2961-2968)
  • Rie Hasebe, Michihito Sasaki, Hirofumi Sawa, Ryuichi Wada, Takashi Umemura, Takashi Kimura
    VIROLOGY 393 2 198 - 209 2009年10月 [査読有り][通常論文]
     
    We investigated the mechanism by which equine herpesvirus-1 (EHV-1) enters primary Cultured equine brain microvascular endothelial cells (EBMECs) and equine dermis (E. Derm) cells. EHV-1 colocalized with caveolin in EBMECs and the infection was greatly reduced by the expression of a dominant negative form of equine caveolin-1 (ecavY14F), suggesting that EHV-1 enters EBMECs via caveolar endocytosis. EHV-1 entry into E. Derm cells was significantly reduced by ATP depletion and treatments with lysosomotropic agents. Enveloped virions were detected from E. Derm cells by infectious virus recovery assay after vital internalization, suggesting that EHV-1 enters E. Derm cells via energy- and pH-dependent endocytosis. These results suggest that EHV-1 utilizes multiple endocytic pathways in different cell types to establish productive infection. (C) 2009 Elsevier Inc. All rights reserved.
  • Yasuko Orba, Yuji Sunden, Tadaki Suzuki, Kazuo Nagashima, Takashi Kimura, Shinya Tanaka, Hirofumi Sawa
    VIROLOGY 370 1 173 - 183 2008年01月 [査読有り][通常論文]
     
    The human Polyomavirus JC virus (JCV) utilizes cellular proteins for viral replication and transcription in the host cell nucleus. These cellular proteins represent potential targets for antiviral drugs against the JCV. In this study, we examined the antiviral effects of the pharmacological cyclin-dependent kinase (cdk) inhibitor R-Roscovitine, which has been shown to have antiviral activity against other viruses. We found that Roscovitine significantly inhibited the viral production and cytopathic effects of the JCV in a JCV-infected cell line. Roscovitine attenuated the transcriptional activity of JCV late genes, but not early genes, and also prevented viral replication via inhibiting phosphorylation of the viral early protein, large T antigen. These data suggest that the JCV requires cdks to transcribe late genes and to replicate its own DNA. That Roscovitine exhibited antiviral activity in JCV-infected cells suggests that Roscovitine might have therapeutic utility in the treatment of progressive multifocal leukoencephalopathy (PML). (c) 2007 Elsevier Inc. All rights reserved.
  • T Toyoda, K Ochiai, H Hatai, M Murakami, E Ono, T Kimura, T Umemura
    VETERINARY PATHOLOGY 43 3 294 - 301 2006年05月 [査読有り][通常論文]
     
    Fowl glioma-inducing virus (FGV), which belongs to subgroup A of avian leukosis virus (ALV), shows tumorigenicity and pathogenicity, mainly in the nervous system, and causes astrocytoma and perineurioma. Apart from these neoplasms, cerebellar anomaly was found in chickens infected with FGV in ovo. The study reported here describes the morphologic characteristics of the affected cerebellum. Specific-pathogen-free chickens (C/O) were inoculated with FGV through the yolk sac on the 7th day of incubation. The cerebellar anomaly included diffuse depletion of granular cells of the internal granular layer (IGL), remnants of the external granular layer (EGL), and disorganization of the Purkinje cell layer. These cerebellar changes were observed in all birds except one. In the infected embryos, the EGL was thicker and had an irregular arrangement with a thin molecular layer (ML) and IGL, compared with the control. The granular cells were immunohistochemically positive for ALV common antigen. Immunohistochemical analysis for vimentin revealed disarrangement and decreased number of Bergmann's fibers. Use of the terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling method and electron microscopy indicated that apoptotic granular cells were frequently observed in the EGL and ML. These results suggested that the cerebellar anomaly was hypoplasia, principally resulting from the apoptosis of granular cells in the EGL and ML caused by FGV infection and that the cell loss induced obstruction of granular cell migration and disarrangement of Bergmann's fibers in the ML.
  • R Hasebe, T Kimura, K Nakamura, K Ochiai, K Okazaki, R Wada, T Umemura
    ARCHIVES OF VIROLOGY 151 4 775 - 786 2006年04月 [査読有り][通常論文]
     
    Equine herpesvirus 1 (EHV-1) shows endotheliotropism in the central nervous system (CNS) of infected horses. However, infection of endothelial cells has not been observed in the CNS of infected mice. To explore the basis for this difference in endotheliotropism, we compared the susceptibility of equine brain microvascular endothelial cells (EBMECs) and mouse brain microvascular endothelial cells (MBMECs) to EHV-1 infection. The kinetics of viral growth in EBMECs was typical of a fully productive infection whereas viral infection in MBMECs seemed to be nonproductive. Immunofluorescence microscopy using anti-EHV-1 polyclonal antibody demonstrated viral antigen in infected EBMECs, but not infected MBMECs. EHV-1 immediate early (IE), early (ICP0), and late (gB, gD and gK) transcripts were expressed in infected EBMECs. However, none of these genes was detected in infected MBMECs by reverse transcription-polymerase chain reaction. Electron microscopic examination at the stage of viral entry showed that viral particles were present within uncoated vesicles in the cytoplasm of EBMECs, but absent from those of MBMECs. These results suggest that viral entry is an important determinant of the susceptibility of EBMECs and MBMECs to EHV-1 infection.
  • Efficacy of Intracerebral Immunization against Pseudorabies Virus in Mice
    Shin, J, Sakoda, Y, Kim, J.-H, Tanaka, T, Kida, H, Kimura, T, Ochiai, K, Umemura, T
    Microbiology and Immunology 50 823 - 830 2006年 [査読無し][通常論文]
  • Jae-Ho Shin, Yoshihiro Sakoda, Jae Hoon Kim, Tomohisa Tanaka, Hiroshi Kida, Takashi Kimura, Kenji Ochiai, Takashi Umemura
    MICROBIOLOGY AND IMMUNOLOGY 50 10 823 - 830 2006年 [査読有り][通常論文]
     
    To evaluate the efficacy of intracerebral (IQ immunization, mice were immunized with formalin-inactivated pseudorabies virus (PRV) by either subcutaneous (SQ or IC injection, and then 10(6) plaque-forming units of PRV were introduced into the hindleg of the immunized or non-immunized mice by intramuscular injection. The antibody titer in serum was elevated and boosted by additional immunization via both the SC and IC routes, but was higher after IC immunization. Intracerebrally immunized mice were completely protected from mortality and neurological signs, whereas all the non-immunized and 80% of the subcutaneously immunized mice died after developing neurological signs. In mouse models, IC immunization is more effective at inducing a protective immune response against the transneural spread of PRV than SC immunization.
  • H Hatai, K Ochiai, Y Tomioka, T Toyoda, K Hayashi, M Anada, M Kato, A Toda, K Ohashi, E Ono, T Kimura, T Umemura
    AVIAN PATHOLOGY 34 6 473 - 479 2005年12月 [査読有り][通常論文]
     
    The complete nucleotide sequence of the avian leukosis virus causing so-called fowl glioma has been previously determined. Primers were designed for detection of the fowl glioma-causal virus (FGV) based on the 3' untranslated region of the viral genome. The provirus and viral RNA of FGV were specifically detected in various organs and tissues, including feather pulp, from experimentally infected birds using nested polymerase chain reaction (PCR) and reverse transcription nested PCR. The prevalence of FGV was evaluated in 131 Japanese fowls of a zoological garden in Japan based on the detection of the FGV genome in feather pulp using PCR and the detection of viral antigen in faeces by enzyme-linked immunosorbent assay. FGV proviral DNA was detected in feather pulp of 52 birds (39.7%) by nested PCR. Later, nine dead birds from among the 52 were histologically diagnosed as having fowl glioma and found to have the proviral DNA in the affected brain. These results demonstrated that the PCR-based detection of FGV in feather pulp is useful for epidemiological studies on fowl glioma.
  • K Matsuda, T Shibata, Y Sakoda, H Kida, T Kimura, K Ochiai, T Umemura
    JOURNAL OF GENERAL VIROLOGY 86 4 1131 - 1139 2005年04月 [査読有り][通常論文]
     
    Neural involvement following infections of influenza viruses can be serious. The neural transport of influenza viruses from the periphery to the central nervous system has been indicated by using mouse models. However, no direct evidence for neuronal infection has been obtained in vitro and the mechanisms of neural transmission of influenza viruses have not been reported. In this study, the transneural transmission of a neurotropic influenza A virus was examined using compartmentalized cultures of neurons from mouse dorsal root ganglia, and the results were compared with those obtained using the pseudorabies virus, a virus with well-established neurotransmission. Both viruses reached the cell bodies of the neurons via the axons. This is the first report on axonal transport of influenza A virus in vitro. In addition, the role of the cytoskeleton (microtubules, microfilaments and intermediate filaments) in the neural transmission of influenza virus was investigated by conducting cytoskeletal perturbation experiments. The results indicated that the transport of avian influenza A virus in the neurons was independent of microtubule integrity but was dependent on the integrity of intermediate filaments, whereas pseudorabies virus needed both for neural spread.
  • T Toyoda, K Ochiai, K Ohashi, Y Tomioka, T Kimura, T Umemura
    VETERINARY PATHOLOGY 42 2 176 - 183 2005年03月 [査読有り][通常論文]
     
    Intraneural perineurioma is an extremely rare condition characterized by perineurial cell proliferation within peripheral nerve (PN) sheaths. In the veterinary field, this entity has been reported only in a dog. We examined multiple enlargements of PNs in 11 chickens (Gallus gallus domesticus) (9 Japanese bantams and 2 specific pathogen-free White Leghorn), which were inoculated with an avian leukosis virus (ALV) causing so-called fowl glioma. All chickens clinically exhibited progressive leg paralysis. Lumbosacral plexus, brachial plexus, and/or spinal ganglion were commonly affected, and these nerves contained a diffuse proliferation of spindle cells arranged concentrically in characteristic onion bulb-like structures surrounded by residual axons and myelin sheaths. The spindle cells were immunohistochemically negative for S-100alpha/beta protein. Electron microscopy revealed that these cells were characterized by short bipolar cytoplasmic processes, occasional cytoplasmic pinocytotic vesicles, and discontinuous basal laminae. These features are consistent with those of intraneural perineurioma. Furthermore, the specific sequence of the ALV was detected in the PN lesions of 8/11 (73%) birds by polymerase chain reaction. These results indicate that the multiple intraneural perineuriomas of chicken may be associated with the ALV-A causing fowl glioma.
  • K Shirato, T Kimura, T Mizutani, H Kariwa, Takashima, I
    JOURNAL OF MEDICAL VIROLOGY 74 3 507 - 513 2004年11月 [査読有り][通常論文]
     
    West Nile (WN) virus is a mosquito-borne flavivirus that can cause lethal encephalitis in humans and horses. The WN virus endemic in New York City (NY) in 1999 caused large-scale mortality of wild birds that was not evident in endemic areas in other parts of the world, and the pathogenesis of the WN virus strain isolated in NY (NY strain) appears to differ from that of previously isolated strains. However, the pathogenesis of NY strain infection remains unclear. This study examined CC (RANTES/CCL5, MIP-1alpha/CCL3, MIP-1beta/CCL4) and CXC (IP-10/CXCL10, B lymphocyte chemoattractant (BLC/CXCL13), and B cell- and monocyte-activating chemokine (BMAC/CXCL14)) chemokine expression during lethal NY strain and non-lethal Eg101 strain infection in mice. We found that the mRNA of the CC chemokines, RANTES, MIP-1alpha, MIP-1beta, and IP-10 was highly up-regulated in the brain of NY strain-infected mice. By contrast, BLC mRNA was not detected in either group of mice, and BMAC mRNA was highly up-regulated in late stage of infection with the non-lethal Eg101 strain relative to levels in NY strain-infected mice. (C) 2004 Wiley-Liss, Inc.
  • Watanabe K, Kadosawa T, Ishiguro T, Takagi S, Ochiai K, Kimura T, Okumura M, Fujinaga T
    The Journal of veterinary medical science / the Japanese Society of Veterinary Science 66 9 1167 - 1170 9 2004年09月 [査読有り][通常論文]
  • Y Tomioka, K Ochiai, K Ohashi, E Ono, T Toyoda, T Kimura, T Umemura
    JOURNAL OF GENERAL VIROLOGY 85 3 647 - 652 2004年03月 [査読有り][通常論文]
     
    So-called fowl glioma is a retroviral infectious disease caused by avian leukosis virus subgroup A (ALV-A). We determined the complete nucleotide sequence of the virus genome. The full-length sequence was consistent with a genetic organization typical of a replication-competent type C retrovirus lacking viral oncogenes. The coding sequences were well conserved with those of replication-competent viruses, but the 3' noncoding regions including LTR were most related to those of replication-defective sarcoma viruses. The U3 region of the LTR had a few deletions and several point mutations compared to that of other ALVs. The promoter activities of the LTRs of glioma-inducing ALV and ALV-A standard strain, RAV-1, were equivalent in chick embryo fibroblasts (CEF), while that of glioma-inducing ALV was significantly lower than that of RAV-1 in human astrocytic cells. These subtle differences of the promoter activity of the LTR may be related to the induction of glial neoplasm.
  • K Matsuda, CH Park, Y Sunden, T Kimura, K Ochiai, H Kida, T Umemura
    VETERINARY PATHOLOGY 41 2 101 - 107 2004年03月 [査読有り][通常論文]
     
    Intranasally inoculated neurotropic influenza viruses in mice infect not only the respiratory tract but also the central nervous system (CNS), mainly the brain stem.(26,34) Previous studies suggested that the route of invasion of virus into the CNS was via the peripheral nervous system, especially the vagus nerve. To evaluate the transvagal transmission of the virus, we intranasally inoculated unilaterally vagectomized mice with a virulent influenza virus (strain 24a5b) and examined the distribution of the viral protein and genome by immunohistochemistry and in situ hybridization over time. An asymmetric distribution of viral antigens was observed between vagal (nodose) ganglia: viral antigen was detected in the vagal ganglion of the vagectomized side 2 days later than in the vagal ganglion of the intact side. The virus was apparently transported from the respiratory mucosa to the CNS directly and decussately via the vagus nerve and centrifugally to the vagal ganglion of the vagectomized side. The results of this study, thus, demonstrate that neurotropic influenza virus,:,an, travels to the CNS mainly via the vagus nerve.
  • T Toyoda, K Ochiai, M Komatsu, T Kimura, T Umemura
    AVIAN PATHOLOGY 33 1 9 - 12 2004年02月 [査読有り][通常論文]
     
    A Hodgson's hawk- eagle ( Spizaetus nipalensis) reared by a falconer showed severe weakness with multiple fractures of bone. It had a history of being fed an all- meat diet. Serological examination revealed a hypocalcaemia ( 72.0 mug/ ml), and hypophosphataemia ( 29.0 mug/ ml). Gross and microscopic examinations demonstrated severe osteodystrophia fibrosa ( fibrous osteodystrophy) characterized by osteoclastic bone resorption and intertrabecular fibrosis with unmineralized trabecular bone containing a large amount of unmineralized osteoid. There was also hypertrophy and hyperplasia of the parathyroid glands, which is consistent with nutritional secondary hyperparathyroidism.
  • K Nakamura, K Ochiai, T Kadosawa, T Kimura, T Umemura
    JOURNAL OF COMPARATIVE PATHOLOGY 130 2-3 205 - 208 2004年02月 [査読有り][通常論文]
     
    A ganglioneuroblastoma of the oral cavlity in a dog was examined histologically and immunohistochemically This rare neoplasm was considered to be derived from ectopic neural crest cells. This is the first report of a canine ectopic ganglioneuroblastoma located in the oral mucosa. (C) 2003 Elsevier Ltd. All rights reserved.
  • T Kimura, R Hasebe, R Mukaiya, K Ochiai, R Wada, T Umemura
    JOURNAL OF COMPARATIVE PATHOLOGY 130 1 41 - 47 2004年01月 [査読有り][通常論文]
     
    Intrauterine infection with equine herpesvirus-1 (EHV-1) has been considered to be the consequence of transplacental transmission of the virus following maternal cell-associated viraemia. In this study the state of EHV-1 gene expression in the placenta of seven naturally aborted equine fetuses was examined. Neither lesions nor viral antigens were detected in the placenta of the fetuses. The amount of infectious virus in the placentas was considerably lower than that in the fetal lungs, which showed pneumonia and typical herpesvirus\ inclusions. Quantitative dot blot hybridization with probes specific for immediate-early (IE), early (ICP0), and late (gD and gK) genes revealed that the placentas expressed the IE gene at a level comparable with that in the lungs; however, expression of the ICP0, gD and gK genes was significantly weaker in the placentas than in the lungs. In-situ hybridization demonstrated that both IE and gK RNAs were distributed mainly in the cytoplasm of trophoblasts. These results suggest that the low level of early and late gene transcription may be related to the limited production of viral progeny and the lack of immunoreactivity for viral antigen in trophoblasts infected with EHV-1. (C) 2003 Elsevier Ltd. All rights reserved.
  • CH Park, K Matsuda, Y Sunden, A Ninomiya, A Takada, H Ito, T Kimura, K Ochiai, H Kida, T Umemura
    VETERINARY MICROBIOLOGY 97 3-4 259 - 268 2003年12月 [査読有り][通常論文]
     
    One-hundred thirty-seven BALB/c mice were intranasally inoculated with neurotropic avian influenza A virus (H5N3). Thirty-nine of these mice died within 16 days post-inoculation (PID) and 98 of the mice recovered from the infection. To investigate whether viral antigens and genomes persist in the central nervous system (CNS) of recovered mice, immunohistochemistry and reverse transcription-polymerase chain reaction (RT-PCR) methods were performed. Histopathologically, mild interstitial pneumonia and non-suppurative encephalomyelitis restricted to the basal part of the frontal lobe of the cerebrum, brain stem and thoracic spinal cord were observed in BALB/c mice until 40 PID. Small amounts of viral antigens were detected in the brain and spinal cord and some viral RNA segments (NA, NP, M, PA, HA, NS, PB1) were intermittently detected in the CNS until 48 PID. Immunosuppression of these mice by dexamethazone (DEX) treatment did not increase the frequency of detection of the lesions, viral antigens or genomes. These findings suggest that viral genomes of neurovirulent influenza virus persist with restricted transcriptive activity in the CNS of the mice even after clinical recovery from the infection. (C) 2003 Elsevier B.V. All rights reserved.
  • Y Tomioka, K Ochiai, K Ohashi, T Kimura, T Umemura
    AVIAN PATHOLOGY 32 6 617 - 624 2003年12月 [査読有り][通常論文]
     
    We have previously isolated an avian leukosis virus (ALV) from a chicken affected with so-called fowl glioma. A resistance-inducing factor test indicated that the isolate was classified into a subgroup A. The distribution and pathogenicity were investigated in C/O specific pathogen free chickens infected in ovo with this virus. Histologically, 11 of 12 (92%) infected birds had non-suppurative encephalitis and three birds (25%) showed the characteristic nodules of fowl glioma at 50 or 100 days of age. Non-suppurative myocarditis with matrix inclusions and atypical myocytes were also noted in nine (75%) of the birds and the ALV antigens were immunohistochemically detected in various general organs as well as the central nervous system and heart. The semi-quantitative determination of the proviral DNA and viral RNA supported the immunohistochemical results and indicated that the virus was likely to replicate especially in myocardial fibres. The isolated ALV failed to induce other neoplastic lesions in this line of chickens within the experimental period of 100 days, despite the broad tissue tropism throughout the body. These results confirmed that this virus was able to induce glioma in embryo-inoculated chickens.
  • Sunden Y, Park CH, Matsuda K, Anagawa A, Kimura T, Ochiai K, Kida H, Umemura T
    The Journal of veterinary medical science / the Japanese Society of Veterinary Science 65 11 1185 - 1188 11 2003年11月 [査読有り][通常論文]
  • T Mizutani, M Kobayashi, Y Eshita, K Shirato, T Kimura, Y Ako, H Miyoshi, T Takasaki, Kurane, I, H Kariwa, T Umemura, Takashima, I
    INSECT MOLECULAR BIOLOGY 12 5 491 - 499 2003年10月 [査読有り][通常論文]
     
    We recently cloned a c-Jun amino-terminal kinase (JNK) sequence from the C6/36 cell line, derived from the mosquito Aedes albopictus. We showed that SP600125, an inhibitor of JNK proteins, inhibits phagocytosis by C6/36 cells, suggesting that the JNK-like protein regulates phagocytosis. Here, we show that C6/36 cells constitutively express low levels of mRNA encoding the antibacterial peptides, cecropin and defensin, but that these mRNAs were up-regulated upon stimulation by lipopolysaccharide (LPS). Thus, the C6/36 cells have properties similar to those of mammalian macrophages. To characterize further the functional properties of C6/36 cells, we have assayed the role of the JNK-like protein in phagocytosis, endocytosis, and viral infection. C6/36 cells phagocytosed bacteria and artificial beads, and this was only slightly up-regulated following LPS stimulation, suggesting that newly stimulated JNK-like protein was not necessary for phagocytosis. SP600125 inhibited the acidification of intracellular compartments, including those involved in the endocytic pathway. Pretreatment of C6/36 cells with SP600125 or bafilomycin A1, but not cytochalasin D, inhibited the entry of West Nile virus (WNV), suggesting that WNV is internalized mainly by endocytosis, and that the JNK signalling pathway is important for endocytic entry. These findings indicate that the JNK-like protein regulates basic physiological functions, including phagocytosis and endocytosis and infection of WNV.
  • HO Kim, T Kimura, K Ochiai, H Yazawa, C Itakura, T Umemura
    JAPANESE JOURNAL OF VETERINARY RESEARCH 51 2 113 - 120 2003年08月 [査読有り][通常論文]
     
    Oligodendrocytes and myelin in the corpus callosum of black tremor and normal hamsters aged over 1.5 years were ultrastructurally examined to determine the myelination index (ratio of myelin thickness/diameter of axon), percentage of naked axons, and proportions of oligodendroglial subtypes (light, medium and dark). The mutant hamsters were remarkably hypomyelinated, with a low myelination index and a high proportion of naked axons, and high proportions of the dark subtypes. Serum concentrations of thyroid hormones (T-3 and T-4) in 6-week-old mutant hamsters were 2-fold (T-3) to 3-fold (T-4) higher than those of age-matched normal animals. However, in the aged animals (over 1.5 years old) only T4 levels of the mutant hamsters were higher in the mutant than normal hamsters. The black tremor hamsters were hypomyelinated throughout their life and high serum level of thyroid hormones might have played a role in the hypomyelination.
  • T Kimura, DE Griffin
    VIROLOGY 311 1 28 - 39 2003年06月 [査読有り][通常論文]
     
    Viral infections of the central nervous system and immune responses to these infections cause a variety of neurological diseases. Infection of weanling mice with Sindbis virus causes acute nonfatal encephalomyelitis followed by clearance of infectious virus, but persistence of viral RNA. Infection with a neuroadapted strain of Sindbis virus (NSV) causes fatal encephalomyelitis, but passive transfer of immune serum after infection protects from fatal disease and infectious virus is cleared. To determine whether persistent NSV RNA is associated with neurological damage, we examined the brains of recovered mice and found progressive loss of the hippocampal gyrus, adjacent white matter, and deep cerebral cortex associated with mononuclear cell infiltration. Mice deficient in CD4(+) T cells showed less tissue loss, while mice lacking CD8(+) T cells showed lesions comparable to those in immunocompetent mice. Mice deficient in both CD4(+) and CD8(+) T cells developed severe tissue loss similar to immunocompetent mice and this was associated with extensive infiltration of macrophages. The number of CD4(+) cells and macrophage/microglial cells, but not CD8(+) cells, infiltrating the hippocampal gyrus was correlated with the number of terminal deoxynucleotidyltransferase-mediated dUTP nick end-labeling positive pyramidal neurons. These results suggest that CD4(+) T cells can promote progressive neuronal death and tissue injury, despite clearance of infectious virus. (C) 2003 Elsevier Science (USA). All rights reserved.
  • Hasebe R, Kimura T, Nakamura K, Okazaki K, Ochiai K, Wada R, Umemura T
    The Journal of veterinary medical science / the Japanese Society of Veterinary Science 64 10 907 - 912 10 2002年10月 [査読有り][通常論文]
  • Kobayashi A, Katagiri S, Kimura T, Ochiai K, Umemura T
    The Journal of veterinary medical science / the Japanese Society of Veterinary Science 64 9 773 - 777 9 2002年09月 [査読有り][通常論文]
  • R Hasebe, T Kimura, E Sato, K Okazaki, K Ochiai, R Wada, T Umemura
    JOURNAL OF COMPARATIVE PATHOLOGY 127 2-3 118 - 125 2002年08月 [査読有り][通常論文]
     
    Little is known about the neuropathogenicity of equine herpesvirus-1 (EHV-1) in mice. No neurological signs were observed in 6-day-old mice inoculated intracerebrally with the HH1 strain (HH1) of EHV-1. However, 6-day-old mice inoculated intracerebrally with a variant derived by serial passage of HH1 in mouse brain showed severe neurological symptoms and eventually died. Histological analyses were performed on 6-day-old mice inoculated with the neuroadapted HH1 (NHH1) and the parental HH1 strain by the intracerebral, intranasal or intraperitoneal route. All routes of inoculation with NHH1 caused encephalitis, but myelitis was observed only in mice inoculated intraperitoneally. Prominent histological findings were perivascular cuffing sometimes associated with small fibrin thrombi, neuronal and glial degeneration and necrosis, and intranuclear inclusion bodies in neurons, glial cells and ependymal cells. Intracerebral and intranasal inoculation, but not intraperitoneal inoculation, with HH1 induced central nervous system (CNS) lesions that were milder than those in mice inoculated with NHH1. The distribution of viral antigen was more widespread in mice inoculated with NHH1 than with HH1 No viral antigen was detected in the CNS of mice inoculated intraperitoneally with HH1. These results indicate that increased viral multiplication and spreading in the CNS were responsible for the enhanced neurovirulence of NHH1. Although EHV-1 has been considered to be primarily endotheliotropic in horses, both NHH1 and HH1 showed tropism for the parenchymal cells of the CNS in mice, namely neurons, glial cells and ependymal cells. (C) 2002 Elsevier Science Ltd. All rights reserved.
  • CH Park, M Ishinaka, A Takada, H Kida, T Kimura, K Ochiai, T Umemura
    ARCHIVES OF VIROLOGY 147 7 1425 - 1436 2002年 [査読有り][通常論文]
     
    A/Hong Kong/483/97 (H5N1) influenza virus (HK483) isolated from the third patient during the outbreak of chicken and human influenza in Hong Kong in 1997 was shown to be neurovirulent in mice. HK483 was inoculated intranasally to mice, and the invasion routes of the virus in the central nervous system (CNS) were investigated by immunohistochemical and in situ hybridization. The pathological changes consisted of bronchopneumonia, ganglionitis, and nonpurulent encephalomyelitis of the brain stem and the anterior part of the thoracic cord. Viral antigens and viral nucleic acids (RNA and mRNA) were demonstrated in the pterygopalatine, trigeminal and superior ganglions prior to or simultaneously with their detection in the CNS. The antigens and nucleic acids were also observed in the olfactory bulb from an early stage of the infection. In the spinal cord, virus-infected cells were first demonstrated in the grey matter of the thoracic cord. The virus, which primarily replicated in the lungs, was considered to invade the thoracic cord via cardiopulmonary splanchnic nerves and sympathetic nerves. These findings indicate that the virus reached the CNS through afferent fibers of the olfactory, vagal, trigeminal, and sympathetic nerves following replication in the respiratory mucosa.
  • T Kimura, Mitsui, I, Y Okada, T Furuya, K Ochiai, T Umemura, C Itakura
    JOURNAL OF COMPARATIVE PATHOLOGY 124 2-3 134 - 141 2001年02月 [査読有り][通常論文]
     
    Adult rabbits were inoculated with liver homogenate from a rabbit haemorrhagic disease (RHD). All experimentally infected rabbits died with typical clinical, gross and histological findings of RHD. Distribution of RHD virus in tissues of the infected rabbits was studied by non-isotopic in-situ hybridization. Both viral plus- and minus-strand RNAs were detected within the cytoplasm of hepatocytes. Kupffer cells and splenic and alveolar macrophages. mainly in morphologically intact cells. Strand-specific reverse transcriptase polymerase chain reaction also demonstrated viral minus-strand RNA as well as plus-strand RNA in the liver, lung and spleen of infected rabbits. Those results suggest that viral replication occurs not only in hepatocytes but also in macrophages. The infected macrophages may contribute to viral dissemination in RHD. (C) 2001 Harcourt Publishers Ltd.
  • Griffin DE, Ubol S, Desprès P, Kimura T, Byrnes A
    Current topics in microbiology and immunology 260 191 - 200 2001年 [査読有り][通常論文]
  • R Mukaiya, T Kimura, K Ochiai, R Wada, T Umemura
    JOURNAL OF COMPARATIVE PATHOLOGY 123 2-3 119 - 125 2000年08月 [査読有り][通常論文]
     
    Equine herpesvirus-1 (EHV-1) infection was demonstrated in the lung tissue of seven aborted fetuses by immunohistochemical labelling and polymerase chain reaction. The placentas of the fetuses were also examined by non-isotopic in-situ hybridization for the EHV-1 glycoprotein B (gB) gene. Positive hybridization signals were observed in the cytoplasm of trophoblasts, especially in microcotyledons, of all seven placentas, and in villous epithelium of the allantochorion of six placentas. Despite the presence of EHV-1 RNA, EHV-1 antigens were not detected in placentas by immunohistochemical examination. The present study represents the first in-vivo demonstration of the EHV-1 gene in equine trophoblasts. The findings suggest direct cell-to-cell spread of EHV-1 from endometrial cells to trophoblasts. (C) 2000 Harcourt Publishers Ltd.
  • T Kimura, DE Griffin
    JOURNAL OF VIROLOGY 74 13 6117 - 6125 2000年07月 [査読有り][通常論文]
     
    Little is known about the role of CD8(+) T cells infiltrating the neural parenchyma during encephalitis induced by neurovirulent Sindbis virus (NSV), NSV preferentially infects neurons in the mouse brain and spinal cord; however, it is generally accepted that neurons can express few if any major histocompatibility complex (MHC) class I molecules. We evaluated the possible roles and interactions of CD8(+) T cells during NSV encephalitis and demonstrated that MHC class I antigen (H2K/D) was expressed on endothelial cells, inflammatory cells, and ependymal cells after intracerebral inoculation of NSV, No immunoreactivity was observed in neurons. On the other hand, in situ hybridization with probes for MHC class I heavy chain, beta 2 microglobulin, and TAP1 and TAP2 mRNAs revealed increased expression in a majority of neurons, as well as in inflammatory cells, endothelial cells, and ependymal cells in the central nervous system of infected mice. NSV-infected neurons may fail to express MHC class I molecules due to a posttranscriptional block or may express only nonclassical MHC class I genes. To better understand the role CD8(+) T cells play during fatal encephalitis induced by NSV, mice lacking functional CD8(+) T cells were studied. The presence or absence of CD8 did not alter outcome, but absence of beta 2 microglobulin improved survival, Interestingly, the intracellular levels of viral RNA decreased more rapidly in immunocompetent mice than in mice without functional CD8(+) T cells, These observations suggest that CD8(+) T cells may act indirectly, possibly via cytokines, to contribute to the clearance of viral RNA in neurons.
  • DC Thach, T Kimura, DE Griffin
    JOURNAL OF VIROLOGY 74 13 6156 - 6161 2000年07月 [査読有り][通常論文]
     
    Neuroadapted Sindbis virus (NSV), given intranasally, caused fatal encephalitis in 100% of adult C57BL/6 mice and 0% of BALB/cBy mice. Most C57BL/6 mice developed severe kyphoscoliosis followed by hind-limb paralysis, while BALB/cBy mice did not. In situ hybridization for detecting NSV RNA and immunohistochemistry for detecting NSV antigen indicated that virus delivered by this route infected neurons of the olfactory region and spread caudally without infection of ependymal cells. Virus antigen was more abundant and infectious virus increased more rapidly and reached higher levels in C57BL/6 mice than in BALB/cBy mice. Surprisingly, infectious virus was cleared faster in C57BL/6 mice, and this was associated with more rapid production of neutralizing antibody. However, viral RNA was cleared more slowly in C57BL/6 mice. In both mouse strains, more infectious virus was present in the lumbar spinal cord than in the cervical spinal cord. These data suggest that genetic susceptibility to fatal NSV encephalomyelitis is determined at least Tn part by the efficiency of viral replication and spread in the central nervous system. The differences identified in this study provide possible phenotypes for mapping genetic loci involved in susceptibility.
  • T Watanabe, N Shinohara, A Sazawa, Y Kobayashi, Y Ogiso, T Kimura, M Takiguchi, J Yasuda, A Hashimoto, T Koyanagi, N Kuzumaki
    CANCER LETTERS 149 1-2 195 - 202 2000年02月 [査読有り][通常論文]
     
    To investigate the suppressive effect of dominant negative H-ras mutant N116Y on transformed phenotypes, we established two N116Y ras mutant stable transfectant clones (C5, C13) of human bladder cancer cell line, UMUC-2. These N116Y ras mutant transfectants, especially the C5 cells, showed a dramatic change of cellular morphology and significantly reduced growth in soft agar compared to their control. Furthermore, phosphorylation of the Jun NH2-terminal kinase (JNK) was significantly decreased in these transfectants compared to the control. These results suggest that the N116Y-induced suppression of transformed phenotypes in UMUC-2 cells is associated with inhibition of JNK phosphorylation, (C) 2000 Elsevier Science Ireland Ltd. All rights reserved.
  • Nishino Y, Funaba M, Fukushima R, Mizutani T, Kimura T, Iizuka R, Hirami H, Hara M
    The Journal of veterinary medical science / the Japanese Society of Veterinary Science 61 10 1167 - 1170 10 1999年10月 [査読有り][通常論文]
  • K Ochiai, T Kimura, K Uematsu, T Umemura, C Itakura
    JOURNAL OF WILDLIFE DISEASES 35 4 766 - 769 1999年10月 [査読有り][通常論文]
     
    We collected 430 harvested ducks (Anas sp. and Aythya sp.) from nine prefectures in Japan between 1994 and 1997. Fifteen (4%) of 363 birds harvested during and after hunting seasons had one lead pellet each in the proventriculus and gizzard. In addition, 32 (34%) of 93 swans (Cygnus sp.) and two of 14 geese (Anser sp.) found dead from various wetlands had lesions consistent with lead poisoning. One to nine swans suspected of having toxicosis from ingestion of lead shot were found dead each year. Twenty-seven (84%) of the 32 lead-exposured swans were found in Hokkaido Prefecture. We concluded that lead poisoning is still a serious threat to waterfowl in Japan and that there is considerable need for environmental improvement concerning this problem.
  • E Handharyani, K Ochiai, T Kadosawa, T Kimura, T Umemura
    JOURNAL OF VETERINARY DIAGNOSTIC INVESTIGATION 11 5 474 - 478 1999年09月 [査読有り][通常論文]
  • K Asano, M Murakami, D Endo, T Kimura, T Fujinaga
    AMERICAN JOURNAL OF VETERINARY RESEARCH 60 7 860 - 864 1999年07月 [査読有り][通常論文]
     
    Objective-To determine complimentary DNA (cDNA) sequence and tissue distribution of canine brain natriuretic peptide (BNP), and to investigate whether synthesis of canine BNP increases in association with cardiovascular dysfunction. Animals-5 healthy adult mixed-breed dogs and 3 healthy adult Beagles. Procedure-Total RNA was extracted from normal canine hearts and was used in a reverse transcription-polymerase chain reaction IRT-PCR) procedure to isolate canine BNP cDNA. Sequence of the isolated cDNA was analyzed. Gene expression of canine BNP in various tissues from 2 mixed-breed dogs was investigated, using RT-PCR and northern blot analyses. Moreover, messenger RNA (mRNA) expression of canine BNP, using northern blot analysis, was compared between grossly normal hearts from 3 Beagles and hearts from 3 mixed-breed dogs with acute myocardial infarction created by surgical ligation. Results-The cDNA sequence and deduced amino acid residues of canine BNP precursor were 420 base pairs and 140 residues, respectively. Messenger RNA expression of canine BNP was delectable in the atria but not in the ventricles and the other tissues. Messenger RNA expression of canine BNP was, however, detectable in the infarcted portion of the ventricles. The amount of canine BNP mRNA in the infarcted ventricles was significantly increased, compared with that of noninfarcted ventricles. Conclusion-The cDNA sequence of canine BNP was determined. Expression of canine BNP mRNA was detected not only in the atria but also in infarcted ventricles. Synthesis of canine BNP increases in association with ischemic myocardial injury. Canine BNP may be used as an indicator of severity of ventricular myocardial injury.
  • E Hayashida, K Ochiai, T Kadosawa, T Kimura, T Umemura
    JOURNAL OF COMPARATIVE PATHOLOGY 121 1 71 - 76 1999年07月 [査読有り][通常論文]
     
    An 8-year-old female German Shepherd dog showed first order Horner's syndrome associated with progressive right-sided hemiplegia and megaoesophagus. Intramedullary and leptomeningeal arteriovenous malformation (AVM) was identified in the cervical spinal cord. The morphological characteristics were arteriovenous shunting, intramedullary multiple thromboses and haemorrhage, non-inflammatory necrosis of white and grey matter around the shunt, and intervening neural gliosis with neovascularization. These findings suggested that the malformation induced a focal circulatory disturbance within the cervical spinal cord and that fatal thrombosis was responsible for the sudden onset of the nervous signs and progressive neurological deterioration. This is the first report of intramedullary spinal AVM in a dog. (C) 1999 W.B. Saunders Company Limited.
  • K Ochiai, K Ohashi, T Mukai, T Kimura, T Umemura, C Itakura
    VETERINARY RECORD 145 3 79 - 81 1999年07月 [査読有り][通常論文]
  • D Mikami, JH Park, T Kimura, K Ochiai, C Itakura
    RESEARCH IN VETERINARY SCIENCE 66 3 237 - 242 1999年06月 [査読有り][通常論文]
     
    Twenty young rabbits (eleven 2-week-old and nine 4-week-old) were experimentally infected with rabbit haemorrhagic disease virus (RHDV) to clarify susceptibility. They were killed chronologically up to 96 hours post-inoculation (PI) and examined for lesions. All inoculated rabbits were clinically normal, but grossly minute white or grey spots were detected throughout the liver. Histologically, the lesions consisted of aggregates of lymphocytes, macrophages and heterophils, with or without acidophilic bodies and necrotic hepatocytes. Immunohistochemically, RHDV antigens were found in the degenerated hepatocytes and in macrophages. The cellular aggregates were considered to be a reaction to necrotic hepatocytes infected with RHDV. It was concluded that some hepatocytes are susceptible to RHDV in young rabbits.
  • T Hirai, M Mizutani, T Kimura, K Ochiai, T Umemura, C Itakura
    TOXICOLOGIC PATHOLOGY 27 3 348 - 353 1999年05月 [査読有り][通常論文]
     
    The neurotoxic effects of 2,5-hexanedione (2,5-HD) were investigated using neurofilament (NF)-deficient (Quv) Japanese quail in comparison with normal Japanese quail. Both Quv and normal Japanese quail were inoculated intraperitoneally with 350 mg/kg/day 2,5-HD for 6 consecutive wk. The results of 2,5-HD exposure differed substantially between the 2 strains of Japanese quail. The 2,5 HD-exposed normal quail showed leg paralysis about 4 wk after initiation of dosing. Some treated normal quail fell into dysstasia and died of nutritional disturbances. Histologically, 2,5-HD-treated normal quail had NF-rich axonal swellings and degeneration in the distal parts of the peripheral nerves, spinal cord, and cerebellar peduncles. In contrast, 2,5-HD-injected Quv quail showed tonic convulsion, ataxia gait, severe quivering, and excitation about 2-3 days after administration. Some treated Quv birds died immediately after systemic tonic convulsion, probably because of asphyxia. Although all treated Quv quail showed neurologic signs, there were no recognizable 2,S-HD-induced lesions in the nervous system. After about 4-6 wk of dosing, 2,5-HD induced distal axonopathy in normal quail and acute neurotoxicity in Quv quail.
  • T Mizutani, M Ogino, Y Nishino, T Kimura, H Inagaki, D Hayasaka, H Kariwa, Takashima, I
    JAPANESE JOURNAL OF VETERINARY RESEARCH 46 4 165 - 169 1999年02月 [査読有り][通常論文]
     
    There are few copies of Borna disease virus (BDV) genome in peripheral blood mononuclear cells and no reliable standard reverse transcriptase-polymerase chain reaction (RT-PCR) method for the detection of BDV RNA, which is both highly sensitive and free of contamination. Single-step RT-PCR, in which both reverse transcription and amplification by Tag DNA polymerase work efficiently in a single buffer, was applied to detect the p24 region of BDV RNA in vitro and in vivo. Using in vitro synthesized RNA, it was demonstrated that at least 100 copies of BDV RNA could be detected and the sensitivity and specificity were nearly equal to those obtained by RT-nested PCR. We could detect BDV RNA from more than 1 pg of cellular RNA obtained from BDV-persistently infected MDCK cells. Furthermore, this method was successfully performed on brain specimens obtained from a BDV-infected rat at II weeks post-inoculation. This single-step RT-PCR method will be convenient for detecting limited amounts of BDV RNA in various cells and tissue samples.
  • T Mizutani, M Ogino, Y Nishino, T Kimura, H Kariwa, K Tsujimura, H Inagaki, Takashima, I
    JAPANESE JOURNAL OF VETERINARY RESEARCH 46 2-3 73 - 81 1998年11月 [査読有り][通常論文]
     
    For detecting Borna disease virus (BDV) genomic stranded RNA, single-tube reverse transcription-polymerase chain reaction (St RT-PCR) was developed to equal the sensitivity of RT-nested PCR but with reduced risk of contamination. BDV-genomic stranded RNA was synthesized in vitro using plasmid cDNA of BDV p24 region as a template and RNA was also extracted from BDV-persistently infected MDCK (MDCK/BDV) cells. Both RNAs were amplified by St RT-PCR in which a single round of RT and a single round of PCR were performed in the same tube. Ten copies of synthesized RNA could be amplified by St RT-PCR, indicating that St RT-PCR method is as sensitive as the ordinary RT-nested PCR method. Furthermore, this method was applied to quantify the exact copy number of genomic RNA in MDCK/BDV cells. Signals were obtained from the samples containing more than 1 pg total cellular RNA. From the results, approximately 100 copies of BDV genomic RNA exist in one MDCK/BDV cell. BDV genomic RNA from the in viva RNA samples using St RT-PCR, indicating this method is applicable for the epidemiological study of BDV without contamination.
  • KO Cho, T Kimura, K Ochiai, C Itakura
    AVIAN PATHOLOGY 27 1 100 - 102 1998年02月 [査読有り][通常論文]
     
    Spontaneous gizzard adenocarcinoma was found in a 15-year-old male Humboldt penguin (Spheniscus humboldti). The lesser curvature of the gizzard wall was extensively and markedly thickened with a hard white tumour mass. Histopathologically, the tumour in the gizzard consisted of two structures: small to large acini lined by cuboidal or flattened cells with poor stroma in the mucosa, and undifferentiated tumour cells arranged singly or in cords with acinar structures separated by dense fibrous tissue in the muscular layer. Small white metastatic tumours of the intestine and the pancreas had a similar histological appearance to that in the muscular layer of the gizzard. From the location of tumour in the gizzard and the stainability with combined alcian blue pH 2.5-periodic acid Schiff, it was considered that this tumour might originate from the lining cells of the tip of the gizzard mucosal folds.
  • Y Okada, K Ochiai, K Osaki, T Kimura, C Itakura
    JOURNAL OF COMPARATIVE PATHOLOGY 118 1 69 - 74 1998年01月 [査読有り][通常論文]
     
    Bronchiolar-alveolar carcinoma was observed in the lung of an 8-year-old Holstein cow. Grossly, the lung contained multiple tumour masses, which were solid, yellowish-white in colour, and firm in consistency. These tumours also occurred in the liver, pancreas, uterus and lymph nodes in the thoracic, abdominal and pelvic cavities. Histologically, the masses were composed of abundant fibrous stroma and proliferating atypical cuboidal epithelial cells, occasionally forming glandular structures. Electron microscopy revealed that the neoplastic cells had microvilli and lamellar bodies in the cytoplasm, suggesting that they originated from immature respiratory epithelial cells differentiating towards either Clara cells or type II pneumocytes. (C) 1998 W.B. Saunders Company Limited.
  • KO Cho, D Endoh, T Kimura, K Ochiai, C Itakura
    AVIAN PATHOLOGY 26 4 707 - 720 1997年12月 [査読有り][通常論文]
     
    Skin tissues were obtained from 12 chickens showing Marek's disease (MD) skin tumour induced by a very virulent MD/5 strain of Marek's disease virus serotype 1 (MDV1) and taken from 24 field broiler chickens (8 to 9-weeks-old) manifesting MD skin tumour. They were examined by the immunohistochemical method in order to examine the topographic expression of MDV1-specific phosphorylated proteins (PP) and by the reverse-transcription polymerase chain reaction following Southern hybridization to analyse their gene transcriptional activity. PP were massively detected and their gene transcripts were abundantly expressed only in the necrotic areas of some experimentally-induced MD cutaneous lymphoma lesions. Strong PP-positive cells were degenerate or necrotic lymphoblasts. The other field or experimentally-induced MD cutaneous lymphoma lesions, even those consisting almost completely of tumorous lymphoblasts, had only a few PP-positive lymphoid cells and few gene transcripts. Necrotic areas of feather pulp MD lymphoma lesions consisting mainly of degenerate or necrotic lymphoblasts and cytolycic feather follicle epithelium (FFE) were also strongly positive for PP. PPs and their gene transcripts were detectable only in the cytolytically infected cells such as FFE and tumorous lymphoblasts converting to the abortic state.
  • T Hirai, M Mubarak, T Kimura, K Ochiai, C Itakura
    VETERINARY PATHOLOGY 34 3 232 - 234 1997年05月 [査読有り][通常論文]
     
    A 13-year-old male Shetland Sheepdog with progressive exophthalmos had a neoplastic mass in the ocular adnexa. Histologically, this neoplasm was composed of duct-forming epithelial cells with decapitation secretion. Tumor cells invaded the globe through the tunica conjunctiva and replaced the vitreous body. The cornea, iris, ciliary body, and retina were extensively destroyed. Both the epithelial and spindle-shaped myoepithelial cells showed nuclear atypia and mitotic activity in the globe. The primary tumor was diagnosed as adenocarcinoma, probably originating from apocrine sweat glands of the eyelid, and the infiltrating intraocular neoplasm was diagnosed as a malignant mixed tumor.
  • 伊藤謙一, 川嶋和晴, 大庭芳和, 播谷亮, 木村享史, 板倉智敏
    日本獣医師会雑誌 50 523 - 526 1997年 [査読有り][通常論文]
  • S Tanaka, IS Braga, T Kimura, K Ochiai, C Itakura, M Mizutani
    JOURNAL OF COMPARATIVE PATHOLOGY 115 2 139 - 150 1996年08月 [査読有り][通常論文]
     
    Cryostat sections of myofibres from the Musculus pectoralis thoracicus of a newly established mutant strain (LWC) of Japanese quail with a myotonic dystrophy-like myopathy were labelled with antibody against myosin heavy chain (MHC) isoforms and neural cell adhesion molecule (N-CAM). The characteristic lesions found in sections of muscle of LWC quail stained with haematoxylin and eosin were type 2B fibre atrophy, sarcoplasmic masses, and ring fibres. Immunohistochemical examination failed to distinguish type 2A and 2B fibres in the LWC quail. Antibody to adult fast MHC, which reacted only with type 2A fibres in normal quail, reacted in LWC quail with type 2B fibres, and to a limited degree with type 2A fibres. Sarcoplasmic masses reacted with both fast and slow MHC antibodies. Some masses also reacted with N-CAM antibody, but apparently independently of similar reactions in fibres. These findings suggest that the changes observed in the myofibres of the LWC quail were not neurogenic but represented defects in both the plasma membrane and intermediate filaments. (C) 1996 W.B. Saunders Company Limited
  • Cho KO, Mubarak M, Kimura T, Ochiai K, Itakura C
    Avian pathology : journal of the W.V.P.A 25 325 - 343 2 1996年06月 [査読有り][通常論文]
  • S Tanaka, IS Braga, T Kimura, K Ochiai, C Itakura, M Mizutani
    JOURNAL OF COMPARATIVE PATHOLOGY 114 3 325 - 337 1996年04月 [査読有り][通常論文]
     
    Ultrastructural study of muscles taken from a mutant (LWC) strain of Japanese quail with myotonia showed type 2 fibre atrophy, ring fibre formation, sarcoplasmic masses, and ''moth-eaten'' fibres. In these abnormal fibres, the most characteristic feature was the loss of interconnection among the myofibrils, mitochondria, and T tubules. Apparently normal muscle fibres often showed mild changes, such as proliferation of T tubules and enlarged sarcoplasmic areas with increased glycogen granules and ribosomes at the periphery of the fibres. The study suggested that one possible cause of these ultrastructural changes was a defect in cytoskeleton of muscle cells, especially in intermediate filaments. (C) 1996 W.B. Saunders Company Limited
  • IS BRAGA, S TANAKA, T KIMURA, C ITAKURA, M MIZUTANI
    JOURNAL OF COMPARATIVE PATHOLOGY 113 2 131 - 143 1995年08月 [査読有り][通常論文]
     
    The progression of the pathological changes that occur in the skeletal muscle was examined in 19 Japanese quail of the LWC strain, affected with an autosomal dominant inherited muscular disorder producing electrical myotonia. The muscle samples were obtained every 10 days from 20 to 70 days of age. Muscle samples from 18 age-matched commercial quail were used as normal controls. Characteristic histological lesions found in the skeletal muscles included sarcoplasmic masses, ringed fibres, internal migration of nuclei and fibre size variation. These lesions, which mainly occurred in the proximal muscles, appeared first in the pectoral region and later in the muscles of the thoracic and pelvic limbs. The most predominant lesion observed at all ages consisted of sarcoplasmic masses. The presence of histological changes did not affect muscle fibre typing by two staining methods, for myosin ATPase at pH 4.5, and by NADH-TR stain. The histological changes were observed in type 2A and less commonly in 2B fibres, but not in type 1. The pectoralis thoracicus muscle, in which lesions were particularly common, showed abnormally large type 2B muscle fibres at 20 days of age. These fibres began to decrease in size at 30 days of age, and at 70 days had become strikingly atrophic, their diameter being only about half that observed at 20 days. The atrophic type 2B muscle fibres were eventually replaced by lipocytes. Chronological staging of the histopathological changes in muscle was impossible since no inter-relationship was observed between the age of the quail, the severity of clinical signs and the extent of muscle lesions. This variability in the severity and age of onset may have been due to the variable expression or incomplete penetrance of the defective gene. Because the disorder is hereditary and progressive in nature, it can be classified as a type of progressive muscular dystrophy. (C) 1995 Academic Press Limited
  • T KIMURA, J KIMURAKURODA, K NAGASHIMA, K YASUI
    ARCHIVES OF VIROLOGY 139 3-4 239 - 251 1994年 [査読有り][通常論文]
     
    The susceptibility of fourteen established cell lines to infection with Japanese encephalitis virus (JEV) was assayed using an indirect fluorescent antibody technique. In kinetic studies, the degree of binding and internalization of JEV allowed the identification of high susceptibility and low-susceptibility cells. Scatchard analysis showed that JEV specifically bound to high-susceptibility Vero cells with greater affinity than to low-susceptibility NRK cells. Microinjection of viral genomic RNA into NRK cells induced highly efficient production of viral antigen and infectious virions. A hemagglutinin-inhibiting monoclonal antibody against JEV (MAb 301) inhibited the binding of JEV to the Vero and NRK cells. JEV was found to bind to a 74K molecule present in the membrane fraction of Vero cells and this binding was inhibited by MAb 301. Importantly, the 74K molecule was not detected in the membrane faction of NRK cells. These results suggest that early events in the JEV-cell interaction influence the susceptibility of cells to infection, and in particular suggests that the 74K molecule may be a possible candidate or component of the cellular receptor for JEV.
  • キスカル酸レセプター刺激によるイノシトールリン脂質代謝応答の加齢ラット海馬での増大.
    斎藤巨志, 中川勇三, 石原高文, 木村享史, 長嶋和郎
    神経化学 29 472 - 473 1990年 [査読有り][通常論文]

書籍

  • 動物病理学総論第3版.日本獣医病理学会編
    木村 享史 (担当:分担執筆範囲:第7章第4節 炎症のメディエーター p.129-135,第7章第6節第3項 肉芽腫性炎症に分類される疾患p.148-152)
    文永堂出版 2013年04月
  • 動物病理学各論第2版.日本獣医病理学会編.
    木村 享史 (担当:分担執筆範囲:第1章第2節第4項 リンパ管 p.26-27,第1章第2節第5項 血管およびリンパ管の腫瘍p.28-29,第3章第2節 腹腔,腹膜 p.77-81)
    文永堂出版 2010年03月
  • 新獣医学辞典.新獣医学辞典編集委員会編.
    木村 享史 (担当:分担執筆範囲:壊死後性肝硬変 p.122,間質性肺炎 p.238,奇異性塞栓症 p.259,クロイツフェルト‐ヤコブ病 p.340)
    チクサン出版社 2008年01月
  • 動物病理カラーアトラス.日本獣医病理学会編.
    木村 享史 (担当:分担執筆範囲:第5編 消化器系Ⅱ 5-8. 兎出血病 p.115)
    文永堂出版 2007年02月

講演・口頭発表等

  • ウイルス感染と免疫  [招待講演]
    木村 享史
    日本獣医病理学会スライドセミナー“炎症と免疫” 2013年03月 公開講演,セミナー,チュートリアル,講習,講義等 東京大学
  • Seminar Public Awareness “Surveillance of Potential Pathogens in Fruitbats in Indonesia”  [招待講演]
    Takashi Kimura
    Joint Seminar between National Committee on Zoonosis and Bogor Agricultural University 2013年03月 公開講演,セミナー,チュートリアル,講習,講義等 IPB International Convention Center, Bogor, Indonesia
  • MHC class I を介したウマヘルペスウイルス1型の細胞内侵入機構  [招待講演]
    木村 享史
    第6回獣医学科特別セミナー 2011年08月 公開講演,セミナー,チュートリアル,講習,講義等 山口大学
  • Immune responses to Sindbis virus infection determine the outcome of acute encephalitis in mice.  [通常講演]
    Takashi Kimura
    Joint Symposium between Hokkaido University Graduate School of Veterinary Medicine and Seoul National University College of Veterinary Medicine-Second COE International Symposium for Zoonosis Control- 2003年12月 シンポジウム・ワークショップパネル(指名) Seoul National University, Korea
  • 中枢神経系におけるウイルス感染の免疫学的制御  [招待講演]
    木村 享史
    2003年度第5回生物研セミナー 2003年11月 公開講演,セミナー,チュートリアル,講習,講義等 麻布大学

作品等

  • Studies on the pathogenesis of encephalomyelitis caused by equine herpesvirus-1 infection
    2000年 -2002年

共同研究・競争的資金等の研究課題

  • 文部科学省:科学研究費補助金(基盤研究(B))
    研究期間 : 2012年 -2015年 
    代表者 : 木村 享史
  • 文部科学省:科学研究費補助金(基盤研究(B))
    研究期間 : 2008年 -2011年 
    代表者 : 木村 享史, 澤 洋文
     
    ウマヘルペスウイルス1型(EHV-1)はアルファヘルペスウイルス亜科に属し、ウマに呼吸器症状、流産、脳脊髄炎を惹き起こす。臨床上特に問題となる流産と脳脊髄炎では、妊娠子宮、中枢神経系特異的に血管内皮細胞へのウイルス感染が生じ、それに続発する血管炎、血栓形成、循環障害が病態発生に大きな役割を果たしている。従って、血管内皮細胞へのウイルス感染は流産、脳脊髄炎の発症において最も重要なステップであるにも関わらず、その感受性を規定するウイルスレセプターに関しては不明な点が多い。本研究では、ウマ脳血管内皮細胞からクローニングしたEHV-1レセプターの機能解析を行い、レセプター遺伝子導入によるマウスモデルの作製とウイルスエンベロープ蛋白-レセプター相互作用を利用した感染予防法の検討を行う。
  • 文部科学省:科学研究費補助金(挑戦的萌芽研究)
    研究期間 : 2009年 -2010年 
    代表者 : 木村 享史
     
    日本脳炎ウイルス(JEV)感染ラットの脳内では低レベルのウイルスゲノムRNA複製が持続している。この感染状態に類似したin vitroシステムを確立する目的で、以下の実験を行った。ラット神経細胞由来株化細胞として、温度感受性SV40T抗原で不死化されたCSM14.1を使用した。JEVの構造蛋白遺伝子を組み込んだ発現プラスミドを作製し、ウエストナイルウイルス(WNV)非構造蛋白遺伝子とEGFP発現カセットを有するWNVレプリコン(ペンシルベニア大学より分与)と共に293T細胞へ導入することにより、上清中に産生されるVLPを回収した。CSM14.1細胞にVLPを感染させた場合、31℃にて維持した細胞では13.6%がGFP陽性を示した。一方、37℃下で72時間培養し神経系分化マーカーが発現した細胞では、VLPの感染により51%がGFP陽性を示し、37℃下での培養によってVLP感染率が低下することが明らかとなった。また、37℃にて培養した細胞では、31℃で維持した細胞と比較して、エンドサイトーシスによるJEV particleの取り込み効率に差異があることが示唆された。従って、レプリコンRNAの導入には31℃にて維持した細胞を使用することが適切であることが確認された。JEV感染ラットの脳内ではセロトニン産生酵素であるトリプトファン水解酵素(TPH)の遺伝子発現が上昇している。F344...
  • 文部科学省:科学研究費補助金(基盤研究(C))
    研究期間 : 2006年 -2007年 
    代表者 : 木村 享史, 好井 健太朗, 澤 洋文
     
    1.ウエストナイルウイルス(WNV)が血液脳関門(Blood-brain barrier; BBB)を通過する機序には不明な点が多い。本研究では、ヒト臍静脈内皮細胞(HUVEC)をTranswell filterの上面に接着、単層培養を行い、BBBのin vitroモデルとして使用した。2.pCMVベクターにWNV NY99 6LP株およびEg101株の構造遺伝子(C、PrM、E)を組み込んだ発現プラスミドを作製した。これとWNVレプリコン(WNV非構造蛋白遺伝子全長と緑色蛍光蛋白EGFP発現カセットを有する)をBHK細胞に共導入し、ウイルス様粒子(Virus-like particles; VLPs)を作製した。3.WNV強毒株である 6LP株のVLPs(6LP-VLPs)および弱毒株であるg101株のVLPs(Eg-VLPs)をin vitro BBBモデルに感染させ、BBBを透過したVLPsの量を24時間後に測定した。Transwell内に接種した6LP-VLPsの約1/10量が下方のチャンバーから検出されたが、同様に接種したEg-VLPsは下方のチャンバーからほとんど検出されなかった。このように両者のin vitroBBB透過性には有意な差が認められ、ウイルスの毒力とBBB透過性との関連性が示唆された。次にHUVECに6LP-VLPsを感染させ、タイトジャンクション(...
  • 文部科学省:科学研究費補助金(基盤研究(B))
    研究期間 : 2004年 -2006年 
    代表者 : 鈴木 正嗣, 梶 光一, 山内 貴義, 有川 二郎, 横山 真弓, 堀内 基広, 木村 享史
     
    1.ニホンジカ(976頭)とニホンイノシシ(87頭)から得た血清を用い,ELISA法とウエスタンブロット法によりE型肝炎に対する血清学的診断を行った.その結果,それぞれの種で2.6%および2.3%が抗体陽性と判断された.イノシシにおいては,RT-PCR法により3頭でHEV-RNAが確認されたが,捕獲状況からこれらは同腹子と考えられた2.野生のニホンイノシシ(88頭)から得た血清を用い,ELISA法と顕微鏡下凝集試験(MAT)により,レプトスピフに対する抗体の検出を行った.その結果,12検体(13.6%)が抗体陽性と判断され,さらにMATでは血清型copenhageni, australis, bratislavaの抗体が検出された.そのため,捕獲ならびに解体にともなう人や猟犬に対する感染リスクが示唆された.3.野生のニホンジカ173頭を材料に,糞便培養法,LAMP法,nested PCR法によりヨーネ菌の検出を試みた.その結果,糞便培養法では菌は確認されなかった.また,LAMP法とnested PCR法でもヨーネ菌DNAは検出されなかった.4.北海道産の野生ニホンジカ(エゾシカ)においては,末梢血よりRickettsia helveticaとEhrlichiamurisの遺伝子が確認された.5.岩手県で捕獲された野生ニホンジカからは,槍形吸虫Dicrocoelium chin...
  • 文部科学省:科学研究費補助金(若手研究(B))
    研究期間 : 2002年 -2003年 
    代表者 : 木村 享史
     
    ウマヘルペスウイルス1型(EHV-1)の神経向性は他のヘルペスウイルスに比較して非常に弱いことが知られているが、脳継代によって作出したEHV-1神経馴化株(NHH1)は親株(HH1)に比べて強い神経病原性を示す。脳継代による変異領域を同定する目的で、両ウイルス株のDNAを比較解析した。LA-PCR法によって13Kbに至るウイルスDNA断片を複数増幅し、RFLP解析を行ったが、両ウイルス間に差異は検出されなかった。平成14年度の解析により、NHH1はHH1に比べてより広範囲な組織向性を示すことが明らかとなっているため、細胞への吸着、侵入に関与するエンベロープ蛋白に変異が生じている可能性が考えられた。そこで、NHH1、HH1のgB遺伝子、gD遺伝子をそれぞれシークエンスしたが、両ウイルス株間でコードされるアミノ酸配列に相違は認められなかった。平成14年度に行った解析により、NHH1、HH1の示す神経侵襲性の差異が脳血管内皮細胞への感染性によって一部規定される可能性が考えられた。また、馬のEHV-1脳脊髄炎においても血行性にCNSに到達したウイルスが脳血管内皮細胞に感染することが知られている。そこで、馬、マウスの大脳皮質より脳微小血管内皮細胞(BMEC)を分離、培養し、HH1株に対する感受性を検討した。その結果、馬BMECはEHV-1感受性であり、エンドサイトーシスによってウイルス...
  • 文部科学省:科学研究費補助金(特定領域研究(C))
    研究期間 : 2001年 -2001年 
    代表者 : 木村 享史
     
    神経強毒性シンドビスウイルス(NSV)は成マウスに致死性(死亡率70〜100%)の急性ウイルス性脳炎を引き起こすが、耐過マウスには海馬組織欠損を特徴とする水頭症様病変が頻繁に認められる。本研究はこの海馬組織傷害に関与する細胞性免疫因子の同定を目的として行われた。C57BL/6マウス(B6)および同系のCD4遺伝子欠損マウス(CD4 -/-)、CD8α遺伝子欠損マウス(CD8 -/-)、recombination activating gene 1遺伝子欠損マウス(RAG -/-)にNSVを脳内接種し、接種後24時間目に抗NSV免疫血清を腹腔内投与することによってマウスを人為的に耐過させた。B6マウスにはマクロファージ、リンパ球浸潤を伴った海馬組織欠損が認められ、同病変は脳内から感染性ウイルスが消失した後に形成された。CD4 -/-マウスの海馬組織欠損は軽度であったが、CD8 -/-マウスはB6マウスと同様の水頭症病変を形成した。一部のRAG -/-マウスにおいてもマクロファージ浸潤を伴った重度の水頭症病変が形成された。TUNEL法陽性海馬分子層ニューロン数は海馬組織中に浸潤するCD4陽性細胞数およびマクロファージ数と比例したが、CD8陽性細胞数との相関は認められなかった。なお、抗CD4および抗CD8モノクローナル抗体投与によって作成したCD4陽性細胞欠損マウス、CD8陽性細胞...
  • 文部科学省:科学研究費補助金(奨励研究(A))
    研究期間 : 1996年 -1996年 
    代表者 : 木村 享史
     
    ヒトIL-2 (hIL-2)ゲノムDNAを導入して作製したトランスジェニックマウス(hIL-2マウス)を免疫組織化学的、分子生物学的に解析し、以下の知見を得た。1.hIL-2マウスの小脳病変の免疫組織化学的検索5週齢のhIL-2マウスの小脳凍結切片を抗Thy-1抗体を用いた酵素抗体法によって検索したところ、クモ膜下腔に集簇するリンパ球のうち多くのものはThy-1陽性を呈した。2.hIL-2発現細胞の同定9日齢のhIL-2マウスの小脳パラフィン包埋切片に対し、hIL-2遺伝子を特異的に増幅するプライマー対を用いたRT-in situPCR法を施行した。hIL-2mRNAの5'末端を増幅するプライマー対と、第2エクソンから第3エクソンにわたる領域を増幅するプライマー対の2種類を用いて解析を行った結果、いずれのプライマー対を用いた場合においても、プルキンエ細胞層に限局して弱いシグナルが観察された。クモ膜下腔に浸潤している少数の炎症性細胞にシグナルは認められなかった。3.小脳において発現しているhIL-2mRNAの解析hIL-2マウス小脳mRNAからRT-PCR法によって第1イントロンを含んだhIL-2primary transcriptを増幅した。増幅したcDNA (293bp)をpGEM-4Zベクターにクローニングし、ジデオキシ法による塩基配列の解読を行った。その結果、第1イント...
  • 文部科学省:科学研究費補助金(一般研究(B), 基盤研究(B))
    研究期間 : 1995年 -1996年 
    代表者 : 板倉 智敏, 木村 亨史, 落合 謙爾
     
    1.ニューロフィラメント(NF)欠損ウズラ(Quv)の本態についてQuvの末梢神経線維における軸索と髄鞘の変化,およびその相関関係を調べた.無髄神経線維では,NF欠損を代償してマイクロチューブ(MT)数が増加しているようであった.有髄神経線維では,軸索の大きさが髄鞘層数の規定因子ではないと判断された.次に,NFの神経線維再生への関与を調べた.このため,Quv坐骨神経を鉗子で押して砕き,2週間後にこれより末梢部位を形態学的に調べた.この結果,Quvでは正常対照例より再生神経線維が少なく,NF欠損はその再生を遅延させることが明らかとなった.さらに,NFの欠損が三量体型GTP結合(G)蛋白質に影響を及ぼすか否かを検討した.G蛋白質はQuvでは正常ウズラよりも増量していたが,これがQuvの振戦症状発現に関与しているか否かは明らかではない.2.NF欠損ウズラ(Quv)に対する神経毒の影響Quvに対して,神経毒であるアクリルアミドに続いてtri-ortho-cresyl-phosphate(TOCP)の影響を調べた.TOCPの連続投与は,正常ウズラの脊髄白質の神経路に遠位軸索症を引き起こしたが,Quvにはその変化は生じなかった.よってNFの存在がその病変発現に関与していることは明らかであった.一方,TOCPの正常およびQuvウズラへの投与は,軸索終末変性を両者に引き起こし,この変性病変発...
  • 文部科学省:科学研究費補助金(奨励研究(A))
    研究期間 : 1995年 -1995年 
    代表者 : 木村 享史
     
    ヒトIL-2ゲノムDNAを導入して作製したトランスジェニックマウス(hIL-2マウス)を解析し、以下の知見を得た。1.hIL-2マウスの組織学的検索9日令、5週令および12週令の雄マウスについて病理組織学的検索を行ったところ、小脳、皮膚に限局して病変が認められた。9日令の小脳では、クモ膜下腔にリンパ球、マクロファージ、好中球の集簇が認められた。5週令、12週令ではクモ膜下腔のリンパ球浸潤に加え脂肪顆粒細胞が小脳実質内に浸潤し、これに伴って小脳皮質の菲薄化、小脳小葉の縮小が認められた。9日令の皮膚には変化が認められなかったが、5週令、12週令の皮膚では、真皮における毛包および血管を中心としたリンパ球、マクロファージ浸潤ならびに毛包、皮脂腺の消失が生じていた(第120回日本獣医学会発表、1995年11月)。2.hIL-2発現組織の同定9日令、5週令、12週令のhIL-2マウスの前進諸臓器ならびに末梢血白血球からtotal RNAを抽出し、ヒトhIL-2を特異的に増幅するプライマーペアを用いてRT-PCRを行ったところ、9日令、5週令の小脳、皮膚12週令の皮膚、体表リンパ節に特異的に導入遺伝子の発現が確認された(第120回日本獣医学会発表、1995年11月)。3.抗神経組織抗体の存在の有無の検討C57BL/6マウス小脳ホモジネートをhIL-2マウス血清を抗体として用いたImmuno...
  • Pathogenesis of viral infections

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  • 獣医科学特別研究
    開講年度 : 2018年
    課程区分 : 博士後期課程
    開講学部 : 獣医学研究科
  • 神経・運動器病理学
    開講年度 : 2018年
    課程区分 : 学士課程
    開講学部 : 獣医学部
    キーワード : 神経病理学、内分泌病理学、骨病理学、骨格筋病理学
  • 獣医科学特論演習
    開講年度 : 2018年
    課程区分 : 博士後期課程
    開講学部 : 獣医学研究科
  • 泌尿器病理学
    開講年度 : 2018年
    課程区分 : 学士課程
    開講学部 : 獣医学部
    キーワード : 腎病理学、生殖器病理学、皮膚病理学
  • 先端獣医科学特論B 獣医病理学特論
    開講年度 : 2018年
    課程区分 : 博士後期課程
    開講学部 : 獣医学研究科
  • 一般教育演習(フレッシュマンセミナー)
    開講年度 : 2018年
    課程区分 : 学士課程
    開講学部 : 全学教育
    キーワード : 動物、ヒト、病気、病因、病態、病理、症状、予防、治療
  • 先端獣医科学特論A 国際獣医科学特論:臨床獣医学
    開講年度 : 2018年
    課程区分 : 博士後期課程
    開講学部 : 獣医学研究科
  • 病理学総論
    開講年度 : 2018年
    課程区分 : 学士課程
    開講学部 : 獣医学部
    キーワード : 変性、壊死、アポトーシス、炎症、腫瘍、先天異常
  • 獣医科学特別研究
    開講年度 : 2018年
    課程区分 : 博士後期課程
    開講学部 : 獣医学院
  • 病理学総論実習
    開講年度 : 2018年
    課程区分 : 学士課程
    開講学部 : 獣医学部
    キーワード : 動物病理学,病理解剖,病理組織学,細胞傷害,細胞・組織の適応,循環障害,炎症,腫瘍
  • 獣医科学特論演習
    開講年度 : 2018年
    課程区分 : 博士後期課程
    開講学部 : 獣医学院
  • 病理学各論実習
    開講年度 : 2018年
    課程区分 : 学士課程
    開講学部 : 獣医学部
    キーワード : 疾患,病理学,診断学
  • 獣医科学基礎科目 臨床疾病学特論
    開講年度 : 2018年
    課程区分 : 博士後期課程
    開講学部 : 獣医学院
  • 先端獣医科学科目 獣医病理学特論
    開講年度 : 2018年
    課程区分 : 博士後期課程
    開講学部 : 獣医学院
  • 総合専門臨床特論
    開講年度 : 2018年
    課程区分 : 博士後期課程
    開講学部 : 獣医学院
  • 獣医病理診断学演習
    開講年度 : 2018年
    課程区分 : 学士課程
    開講学部 : 獣医学部
  • 研究・臨床セミナー
    開講年度 : 2018年
    課程区分 : 学士課程
    開講学部 : 獣医学部
  • アドバンスト演習
    開講年度 : 2018年
    課程区分 : 学士課程
    開講学部 : 獣医学部

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  • 2009年 - 現在   日本獣医病理学専門家協会   評議員
  • 2006年 - 現在   日本獣医病理学会   評議員
  • 2006年 - 現在   日本獣医学会   評議員


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