研究者データベース

森松 正美(モリマツ マサミ)
獣医学研究院 獣医学部門 応用獣医科学分野
准教授

基本情報

所属

  • 獣医学研究院 獣医学部門 応用獣医科学分野

職名

  • 准教授

科研費研究者番号

  • 70241370

J-Global ID

研究キーワード

  • 獣医生化学   分子遺伝学   実験動物学   Veterinary Biochemistry   Molecular Genetics   Laboratory Animal Science   

研究分野

  • ライフサイエンス / 実験病理学
  • ライフサイエンス / 獣医学
  • ライフサイエンス / 実験動物学

職歴

  • 2013年11月 - 現在 北海道大学大学院獣医学研究科 動物疾病制御学講座実験動物学教室 准教授
  • 2013年 - 現在 Faculty of Veterinary Medicine, Hokkaido University Attending Veterinarian of Animal Facility
  • 2013年 - 現在 Faculty of Veterinary Medicine, Hokkaido University Associate Professor
  • 2012年04月 - 2013年10月 北海道大学遺伝子病制御研究所 附属動物実験施設 施設長
  • 2007年04月 - 2013年10月 北海道大学遺伝子病制御研究所准教授
  • 2012年 - 2013年 北海道大学 遺伝子病制御研究所
  • 2005年 - 2013年 北海道大学 遺伝子病制御研究所
  • 2005年10月 - 2007年03月 北海道大学遺伝子病制御研究所助教授
  • 2004年 - 2005年 北海道大学大学院医学研究科 助手
  • 2004年 - 2005年 Assistant Professor, Hokkaido University Graduate School of Medicine
  • 1997年 - 2004年 岩手大学農学部 助教授
  • 1997年 - 2004年 Associate Professor, Faculty of Agriculture, Iwate University
  • 1996年 - 1997年 University of Texas MD Anderson Cancer Center Visiting Fellow
  • 1992年 - 1997年 北海道大学大学院獣医学研究科 助手
  • 1992年 - 1997年 Assistant Professor, Graduate School of Veterinary Medicine, Hokkaido University
  • 1995年 - 1995年 University of São Paulo School of Medicine Visiting Fellow

学歴

  •         - 1992年03月   北海道大学   大学院獣医学研究科   形態機能学
  •         - 1992年   北海道大学
  •         - 1989年   北海道大学
  •         - 1987年03月   北海道大学   獣医学部   獣医学科

所属学協会

  • JAPANESE ASSOCIATION FOR LABORATORY ANIMAL SCIENCE   THE JAPANESE BIOCHEMICAL SOCIETY   THE MOLECULAR BIOLOGY SOCIETY OF JAPAN   THE JAPANESE SOCIETY OF VETERINARY SCIENCE   The Japanese College of Laboratory Animal Medicine (JCLAM)   The Japanese Association for Laboratory Animal Medicine (JALAM)   日本獣医師会   日本実験動物医学専門医協会   日本実験動物医学会   日本実験動物学会   日本分子生物学会   日本生化学会   日本獣医学会   

研究活動情報

論文

  • Mitsuki Uemura, Kazuhiko Ochiai, Masami Morimatsu, Masaki Michishita, Eri Onozawa, Daigo Azakami, Yumiko Uno, Yasunaga Yoshikawa, Takanori Sasaki, Masami Watanabe, Toshinori Omi
    Veterinary and comparative oncology 2019年09月13日 [査読有り][通常論文]
     
    RAD51 forms a complex with BRCA2 and plays a central role in the DNA damage response pathway that is associated with homologous recombination. The structures of RAD51 and its homologues are highly conserved from prokaryotes to higher eukaryotes. Although a large number of BRCA2 mutations have been reported, there are only a few reports on the mutations of RAD51, which have been shown in humans and dogs. However, several mutations of canine RAD51 were identified from mammary gland tumour tissues in a recent study. Some of these mutations seem to have an influence on the homo-oligomerization or interaction with "Partner and localizer of BRCA2" (PALB2). In this study, we cloned the canine PALB2 homologue and investigated the effect on its interaction with the RAD51 mutants to evaluate the alteration in the function of RAD51 mutants. The A209S and T225S mutants of RAD51 show an attenuation of the interaction between RAD51 and PALB2. These results indicate that the canine RAD51 mutations can potentially alter the homologous recombination pathways in response to DNA damage in dogs.
  • Boonyarattanasoonthorn Tussapon, Elewa Yaser, Hosny Ali, Tag-El-Din-Hassan, Hassan T, Morimatsu Masami, Agui Takashi
    INFECTION GENETICS AND EVOLUTION 73 55 - 65 2019年09月 [査読有り][通常論文]
  • Jinxi Wang, Ruihua Dang, Yoshiki Miyasaka, Kousuke Hattori, Daisuke Torigoe, Tadashi Okamura, Hassan T Tag-Ei-Din-Hassan, Masami Morimatsu, Tomoji Mashimo, Takashi Agui
    PloS one 14 6 e0217132  2019年 [査読有り][通常論文]
     
    The Hirschsprung disease (HSCR) is an inherited disease that is controlled by multiple genes and has a complicated genetic mechanism. HSCR patients suffer from various extents of constipation due to dysplasia of the enteric nervous system (ENS), which can be so severe as to cause complete intestinal obstruction. Many genes have been identified as playing causative roles in ENS dysplasia and HSCR, among them the endothelin receptor type B gene (Ednrb) has been identified to play an important role. Mutation of Ednrb causes a series of symptoms that include deafness, pigmentary abnormalities, and aganglionosis. In our previous studies of three rat models carrying the same spotting lethal (sl) mutation on Ednrb, the haplotype of a region on chromosome (Chr) 2 was found to be responsible for the differing severities of the HSCR-like symptoms. To confirm that the haplotype of the responsible region on Chr 2 modifies the severity of aganglionosis caused by Ednrb mutation and to recreate a rat model with severe symptoms, we selected the GK inbred strain, whose haplotype in the responsible region on Chr 2 resembles that of the rat strain in which severe symptoms accompany the Ednrbsl mutation. An Ednrb mutation was introduced into the GK rat by crossing with F344-Ednrbsl and by genome editing. The null mutation of Ednrb was found to cause embryonic death in F2 progeny possessing the GK haplotype in the responsible region on Chr 2. The results of this study are unexpected, and they provide new clues and animal models that promise to contribute to studies on the genetic regulatory network in the development of ENS and on embryogenesis.
  • Md Atiqul Islam, Daisuke Torigoe, Yayoi Kameda, Takao Irie, Hirokazu Kouguchi, Ryo Nakao, Md Abdul Masum, Osamu Ichii, Yasuhiro Kon, Hassan T Tag-El-Din-Hassan, Masami Morimatsu, Kinpei Yagi, Takashi Agui
    Infection, genetics and evolution : journal of molecular epidemiology and evolutionary genetics in infectious diseases 65 65 - 71 2018年11月 [査読有り][通常論文]
     
    The resistance/susceptibility to Echinococcus multilocularis infection in mice is genetically controlled. However, genetic factors responsible for these differences remain unknown. Our previous study in genetic linkage analysis has revealed that there is a significant quantitative trait locus (QTL) for the establishment of cyst (Emcys1), and a highly significant QTL for the development of protoscolex of E. multilocularis larvae (Empsc1), on mouse chromosomes 6 and 1, respectively. The current study aimed to confirm these QTLs and narrow down the critical genetic region that controls resistance/susceptibility to E. multilocularis infection by establishing congenic and subcongenic lines from C57BL/6 (B6) and DBA/2 (D2) mice. For protoscolex development phenotype, two congenic lines, B6.D2-Empsc1 and D2.B6-Empsc1 were developed, where responsible QTL, Empsc1 was introgressed from D2 into B6 background and vice versa. For cyst establishment phenotype, two congenic lines, B6.D2-Emcys1 and D2.B6-Emcys1 were developed, where responsible QTL, Emcys1 was introgressed from D2 into B6 background and vice versa. Because there was no significant difference in cyst establishment between B6.D2-Emcys1 and D2.B6-Emcys1 mice after challenge with E. multilocularis, it is suggested that the Emcys1 does not solely control the cyst establishment in mouse liver. However, infection experiments with B6.D2-Empsc1 and D2.B6-Empsc1 mice showed a significant difference in protoscolex development in the cyst. It confirms that the Empsc1 controls phenotype of the protoscolex development in the cyst. Subsequently, two subcongenic lines, B6.D2-Empsc1.1 and B6.D2-Empsc1.2 from B6.D2-Emcys1 and one subcongenic line, D2.B6-Empsc1.1 from D2.B6-Empsc1 were developed to narrow down the critical region responsible for protoscolex development. From the results of infection experiments with E. multilocularis in these subcongenic mice, it is concluded that a gene responsible for protoscolex development is located between D1Mit290 (68.1 cM) and D1Mit511 (97.3 cM).
  • Hassan T. Tag-EL-Din-Hassan, Masami Morimatsu, Takashi Agui
    Infection, Genetics and Evolution 62 220 - 232 2018年08月01日 [査読有り][通常論文]
     
    Up-to-date the flavivirus infection in avian taxa is not clearly defined. Several reports have demonstrated that many viruses belonging to Flaviviridae may cause diseases in poultry species however, the susceptibility of other avian species is variable and still unclear. In human and mice, the 2′-5′-oligoadenylate synthetase (OAS) proteins are associated with resistance to the flavivirus infection as well as other virus infections. However, the avian OAS proteins are rarely studied. In our previous studies, we confirmed that the chicken OAS-like protein (chOASL) expressed OAS-enzymatic activity (the classical OAS/RNase L-dependent pathway) as well as the anti-flavivirus activity (the putative OAS/RNase L-independent pathway). Therefore, the current study aimed at functional analysis of avian OAS proteins from duck, goose, and ostrich. The duOASL, goOASL, and osOAS1 proteins expressed enzymatic activity as well as chOASL, whereas osOASL expressed little enzymatic activity. On the other hand, duOASL, goOASL, and osOASL possessed significant antiviral activity against West Nile virus (WNV)-replicon replication as well as chOASL, whereas osOAS1 did not. In addition, similar to chOASL, their antiviral activity was independent of RNase L activation. These results suggest that OASL is the only OAS protein in the duck and goose as well as chicken and possesses both OAS-enzymatic and anti-flavivirus activities, whereas the ostrich possesses both OAS1 and OASL proteins with sharing the functional activities, OAS-enzymatic and anti-flavivirus activities, respectively. It is of interest that the ostrich undergoes differential process in OAS gene evolution from other poultries and thus possesses different molecular mechanism in antiviral activity.
  • Kazuhiko Ochiai, Daigo Azakami, Masami Morimatsu, Hinako Hirama, Shota Kawakami, Takayuki Nakagawa, Masaki Michishita, Ai S Egusa, Takanori Sasaki, Masami Watanabe, Toshinori Omi
    Oncology reports 40 1 488 - 494 2018年07月 [査読有り][通常論文]
     
    Mutations in the p53 gene are associated with more than half of all human cancers. These mutations often cause a disruption of the tumor-suppressor function of p53 and induce genomic instabilities. Wild‑type p53 requires tetramerization to function as an initiator of cell cycle arrest and apoptosis. Although alterations in p53 tetramerization caused by mutation have been well studied, there are few cell lines containing an endogenous mutation in the tetramerization domain of p53. Here, we report the discovery of a canine mammary gland tumor cell line CTB‑m2, which contains the Leu332Gln (L332Q) mutation corresponding to Leu344 in the tetramerization domain of human p53. Although CTB‑m2 cells are genetically heterozygous for the Leu332Gln mutation, the mutant mRNA was almost exclusively expressed. CTB‑m2 cells showed enhanced cell proliferation compared to wild‑type p53-expressing CTB‑m cells of the same lineage. A p53 tetramerization reporter assay showed that the ability of the p53 mutant to form tetramers was significantly lower than that of wild‑type p53. An immunoblot analysis of cross-linked p53 oligomerized forms demonstrated that the L332Q mutant lacked the ability to form tetramers but retained the ability to form dimers. These data suggest that the p53 mutant cell line CTB‑m2 could be a useful tool for analyzing the precise tetramerization mechanisms of p53 and verifying the effects of therapeutic agents against tumors expressing p53 mutants that lack the ability to tetramerize.
  • Raghda M. F. Abbas, Hassan T. Tag-El-Din-Hassan, Tussapon Boonyarattanasoonthorn, Keisuke Aoshima, Masami Morimatsu, Takashi Agui
    Japanese Journal of Veterinary Research 66 2 93 - 103 2018年05月01日 [査読有り][通常論文]
     
    It has been reported that C57BL/6 (B6) mice are resistant to the Sendai virus (SeV) infection, whereas DBA/2 (D2) mice are susceptible, the cause of susceptibility in D2 mice is hyper-inflammatory cytokine production, and three quantitative trait loci (QTLs), Sen1, Sen2, and Sen3 are identified to be responsible for this difference. We previously have verified that the introgression of B6-derived these three QTLs into D2-genetic background increases survival rate and suppresses cytokine production by generating D2.B6-Sen1Sen2Sen3 congenic mice. In this study, we investigated immune cellular responses of D2.B6-Sen1Sen2Sen3 mice after SeV infection. Body weight loss, viral load, immune cells in broncho-alveolar lavage fluid (BALF), and histopathological index of SeV-infected male D2. B6-Sen1Sen2Sen3 mice were comparable to those of B6 mice except for the number of neutrophils in BALF. In contrast, female D2.B6-Sen1Sen2Sen3 mice were divided into survived and non-survived mice after SeV infection. Viral load and macrophage number in BALF in SeV-infected female D2. B6-Sen1Sen2Sen3 mice were comparable to those of B6 mice, whereas the number of total cells, neutrophils, and lymphocytes in BALF were remained in the level of D2 mouse. There was a correlation between body weight loss and these immune cellular responses in SeV-infected female D2.B6-Sen1Sen2Sen3 mice. These results indicate that the introgression of B6 alleles of these three QTLs into D2-genetic background resulted in resistance to SeV infection by optimizing the aggressive immune cellular responses that seen in D2 mice, although there may be other loci responsible for difference between B6 and D2 mice.
  • Shota Kawakami, Kazuhiko Ochiai, Daigo Azakami, Yuiko Kato, Masaki Michishita, Masami Morimatsu, Toshina Ishiguro-Oonuma, Eri Onozawa, Masami Watanabe, Toshinori Omi
    Veterinary research communications 42 1 49 - 56 2018年03月 [査読有り][通常論文]
     
    Glioma is the second most common intracranial neoplasia in dogs, but the pathogenic mechanisms remain unclear. In humans, isocitrate dehydrogenase 1 (IDH1) is frequently mutated in gliomas. Although almost all human IDH1 mutations have been identified as involving the Arg132 codon, few studies have reported structural, functional, and mutational information for canine IDH1. Therefore, in this study, we cloned the canine IDH1 homologue and used PCR mutagenesis to substitute the wildtype (WT) Arg132 with His (R132H) or Ser (R132S). WT and mutated IDH1 were overexpressed in HeLa cells, and their presence was confirmed by immunoblotting and immunocytochemistry using mutation-specific antibodies. The IDH1 activity between WT, R132H, and R132S transfectants was compared by measuring the production of NADH and NADPH. NADPH production in R132H and R132S transfectants was lower than that in WT, but NADH levels were not significantly different. Finally, we detected increased expression of hypoxia inducible factor 1 alpha (HIF-1α) in the R132H and R132S transfectants. These results indicated that the canine IDH1 Arg132 mutation has the potential to induce carcinogenesis in canine somatic cells.
  • Raghda Mohamed Fathi Abbas, Daisuke Torigoe, Yayoi Kameda, Hassan T. Tag-EL-Din-Hassan, Nobuya Sasaki, Masami Morimatsu, Takashi Agui
    INFECTION GENETICS AND EVOLUTION 57 75 - 81 2018年01月 [査読有り][通常論文]
     
    Sendai virus (SeV) is one of the most important pathogens in the specific-pathogen free rodents. It is known that there are some inbred mouse strains susceptible or resistant to SeV infection. The C57BL/6 (B6) and DBA/2 (D2) mice are representative of the resistant and susceptible strains, respectively. Previous study with the quantitative trait locus (QTL) analysis identified three QTLs responsible for resistance or susceptibility to SeV infection on different chromosomes and indicated that resistance or susceptibility to SeV infection was almost predicted by genotypes of these three QTLs. In this paper, to verify the above hypothesis, congenic lines were generated as follows; B6-congenic lines carrying one of the D2 alleles of three QTLs and combination of these three QTLs, and D2-congenic lines carrying single or combination of B6 alleles of three QTLs. All these congenic lines were then challenged with SeV infection. D2 congenic lines introgressed single or combination of B6 alleles of QTLs changed to resistance to SeV infection. Especially, a D2 triple-congenic line became resistant as similar level to B6-parental strain. However, B6-congenic lines introgressed single or combination of D2 alleles of QTLs all remained to be resistant to SeV infection. Both IL-6 and TNF-alpha in broncho-alveolar lavage fluid of D2 triplecongenic line were decreased to the similar level of B6 mice, suggesting that this is a part of factors that D2 triple-congenic line became resistant to the similar level of B6 mice. Data obtained from these congenic mice verified that three QTLs identified previously were indeed responsible for the resistance/susceptibility to SeV infection in B6 and D2 mice.
  • Yoshikazu Fujimoto, Yukiko Tomioka, Kinuyo Ozaki, Keiko Takeda, Haruka Suyama, Sayo Yamamoto, Hiroki Takakuwa, Masami Morimatsu, Toshimitsu Uede, Etsuro Ono
    JOURNAL OF GENERAL VIROLOGY 98 7 1815 - 1822 2017年07月 [査読有り][通常論文]
     
    Herpesvirus entry mediator A (HVEM), nectin-1 and nectin-2 are cellular receptors of glycoprotein D (gD) of herpes simplex virus type-2 (HSV-2). It has been shown that soluble forms of HSV gD receptors have the antiviral potential in cultured cells and transgenic mice. Here, to compare antiviral potential of soluble forms of HVEM, nectin-1 and nectin-2 against HSV-2 infections in vivo, transgenic mice expressing fusion proteins consisting of the entire ectodomain of HVEM, nectin-1 or nectin-2 and the Fc portion of human IgG (HVEMIg, nectin-1Ig and nectin-2Ig, respectively) were intraperitoneally infected with HSV-2. In the infection with 3 MLD50 (50 % mouse lethal dose), effective resistance was not observed in transgenic mice expressing nectin-21g. In a transgenic mouse line with high expression of nectin-11g, significant protection from the infection with 30 and 300 MLD50 was observed (survival rate of 100 and 71 %, respectively). On the other hand, transgenic mice expressing HVEMIg showed a complete resistance to the lethal infection even with 300 MLD50 (survival rate of 100 %). These results demonstrated that HVEMIg could exert effective antiviral activities against HSV-2 infections in vivo as compared with other soluble forms of HSV gD receptors.
  • Yuko Okamatsu-Ogura, Keigo Fukano, Ayumi Tsubota, Junko Nio-Kobayashi, Kyoko Nakamura, Masami Morimatsu, Hiroshi Sakaue, Masayuki Saito, Kazuhiro Kimura
    SCIENTIFIC REPORTS 7 1 6648  2017年07月 [査読有り][通常論文]
     
    We previously reported brown adipocytes can proliferate even after differentiation. To test the involvement of mature adipocyte proliferation in cell number control in fat tissue, we generated transgenic (Tg) mice over-expressing cell-cycle inhibitory protein p27 specifically in adipocytes, using the aP2 promoter. While there was no apparent difference in white adipose tissue (WAT) between wild-type (WT) and Tg mice, the amount of brown adipose tissue (BAT) was much smaller in Tg mice. Although BAT showed a normal cellular morphology, Tg mice had lower content of uncoupling protein 1 (UCP1) as a whole, and attenuated cold exposure-or beta 3-adrenergic receptor (AR) agonist-induced thermogenesis, with a decrease in the number of mature brown adipocytes expressing proliferation markers. An agonist for the beta 3-AR failed to increase the number of proliferating brown adipocytes, UCP1 content in BAT, and oxygen consumption in Tg mice, although the induction and the function of beige adipocytes in inguinal WAT from Tg mice were similar to WT mice. These results show that brown adipocyte proliferation significantly contributes to BAT development and adaptive thermogenesis in mice, but not to induction of beige adipocytes.
  • Yuiko Kato, Kazuhiko Ochiai, Shota Kawakami, Nobuhiro Nakao, Daigo Azakami, Makoto Bonkobara, Masaki Michishita, Masami Morimatsu, Masami Watanabe, Toshinori Omi
    BMC VETERINARY RESEARCH 13 1 170  2017年06月 [査読有り][通常論文]
     
    Background: The pathological condition of canine prostate cancer resembles that of human androgen-independent prostate cancer. Both canine and human androgen receptor (AR) signalling are inhibited by overexpression of the dimerized co-chaperone small glutamine-rich tetratricopeptide repeat-containing protein a (SGTA), which is considered to cause the development of androgen-independency. Reduced expression in immortalised cells (REIC/Dkk-3) interferes with SGTA dimerization and rescues AR signalling. This study aimed to assess the effects of REIC/Dkk-3 and SGTA interactions on AR signalling in the canine androgen-independent prostate cancer cell line CHP-1. Results: Mammalian two-hybrid and Halo-tagged pull-down assays showed that canine REIC/Dkk-3 interacted with SGTA and interfered with SGTA dimerization. Additionally, reporter assays revealed that canine REIC/Dkk-3 restored AR signalling in both human and canine androgen-independent prostate cancer cells. Therefore, we confirmed the interaction between canine SGTA and REIC/Dkk-3, as well as their role in AR signalling. Conclusions: Our results suggest that this interaction might contribute to the development of a novel strategy for androgen-independent prostate cancer treatment. Moreover, we established the canine androgen-independent prostate cancer model as a suitable animal model for the study of this type of treatment-refractory human cancer.
  • Hassan T. Tag-El-Din-Hassan, Nobuya Sasaki, Daisuke Torigoe, Masami Morimatsu, Takashi Agui
    JOURNAL OF INTERFERON AND CYTOKINE RESEARCH 37 2 71 - 80 2017年02月 [査読有り][通常論文]
     
    The oligoadenylate synthetase (OAS) is well known as an antiviral factor against the flavivirus infection in mammals. It is known that the oligoadenylate synthetase-like (ChOAS-L) gene is only present in the chicken genome. It has been shown in the previous report that the ChOAS-L possesses enzymatic activity to convert ATP into 2'-5'-linked oligoadenylates and antiviral activity against West Nile virus (WNV) replicon. Therefore, this study aimed to investigate the relationship between enzymatic and antiviral activities of ChOAS-L. Eight mutated ChOAS-L proteins were generated using either the site-directed mutagenesis or standard polymerase chain reaction protocol. The wild-type and mutated proteins were ectopically expressed in 293FT cells to analyze the enzymatic activity and in BHK-21 and BALB/3T3 cells to analyze the antiviral activity using WNV replicon. The results revealed that all mutated proteins showed no enzymatic activity except for ChOAS-L-A Delta UbL2. However, all mutated proteins showed antiviral activity to inhibit the replication of the WNV replicon except for ChOAS-L-A Delta UbL1/UbL2, which showed a partial inhibition compared to the wild-type ChOAS-L-A or other mutated proteins. These results suggest that the ChOAS-L expresses the antiflavivirus activity in a manner independent of enzymatic activity. Our results propose reconsideration of the mechanism of antiviral activity against the flavivirus replication of ChOAS-L.
  • Enas Elkhateeb, Hassan T. Tag-El-Din-Hassan, Nobuya Sasaki, Daisuke Torigoe, Masami Morimatsu, Takashi Agui
    INFECTION GENETICS AND EVOLUTION 45 393 - 401 2016年11月 [査読有り][通常論文]
     
    The interferon-induced oligoadenylate synthetase (OAS) family is one of the most important immune response proteins to the viral infection. The OAS protein binds dsRNA and is activated to produce 2',5'-oligoadenylates, which lead to the activation of latent form of RNase L, resulting in degradation of cellular and viral RNA and inhibition of viral replication. In mice, the Oas gene family locates on chromosome 5. The mouse Oas gene locus undergoes a recent series of duplication event, leading to the presence of eight paralogs of Oas1 genes (Oas1a through Oas1h) that forms Oas gene cluster with the Oas2, Oas3 and two OasL (OasL1 and OasL2) genes. Previous studies demonstrated that the mouse Oas1b gene conferred resistance to the flavivirus infection in mice; however, the antiviral activity of other mouse Oas1 gene family is still unknown. Therefore, in the present study, we have evaluated the mouse Oas1 paralogs regarding the enzymatic activity and antiviral activity against the two neurotropic flaviviruses, West Nile virus and tick-borne encephalitis virus. The mouse Oas1 genes were cloned from C57BL/6J (B6) as well as the Oas1b derived from feral mouse strain, MSM. The obtained results demonstrated that only Oas1a and Oas1g showed enzymatic activity. Although MSM-derived Oas1b showed antiviral activity to both viruses, all B6-derived OAS paralogs did not show antiviral activity. These results suggest that Oas1a and Oas1g play a role in potentiating viral RNA-induced interferon response in the cell, whereas the Oas1b works as a specific anti-flavivirus factor unless it is mutated. However, the role of other paralogs is unknown and should wait for further investigation. (C) 2016 Elsevier B.V. All rights reserved.
  • Mitsumasa Saito, Sayo Yamamoto, Kinuyo Ozaki, Yukiko Tomioka, Haruka Suyama, Masami Morimatsu, Ken-ichi Nishijima, Shin-ichi Yoshida, Etsuro Ono
    MICROBIAL PATHOGENESIS 99 106 - 110 2016年10月 [査読有り][通常論文]
     
    Group B Streptococcus (GBS) is a leading cause of invasive bacterial infections in human newborns. A key GBS virulence factor is its capsular polysaccharide (CPS), possessing terminal sialic acid residues that suppress host immune response and provide a survival advantage to the pathogen. CPS binds to Siglec-9 expressed on neutrophils, which is expected to down-regulate the immune responsiveness of neutrophils. We hypothesized that a soluble form of Siglec-9 (sSiglec-9) competitively inhibits a binding of CPS to Siglec-9 on immune cells, leading to provide antibacterial benefit against GBS infection in the transgenic mouse line expressing sSiglec-9 (sSiglec-9 Tg). The sSiglec-9 in the sera of sSiglec-9 Tg bound to the sialylated-GBS strains belonging to serotypes Ia, Ib, II, III, IV and V in whole GBS cell ELISA. When GBS cells of serotype III that is a common serotype in late-onset GBS disease (LOD) were intraperitoneally inoculated into sSiglec-9 Tg, sSiglec-9 Tg showed a significant resistance as compared with non-transgenic littermates. Furthermore, GBS serotype III organisms were not detected in cultures of the blood from surviving mice (<1 x 10(3) CFU/ml). These results indicated that sSiglec-9 Tg mice were more efficient in eliminating GBS and survived better after the intraperitoneal challenge with GBS serotype III bacteria. (C) 2016 Elsevier Ltd. All rights reserved.
  • Yoshikazu Fujimoto, Yukiko Tomioka, Hiroki Takakuwa, Gen-Ichiro Uechi, Toshiyo Yabuta, Kinuyo Ozaki, Haruka Suyama, Sayo Yamamoto, Masami Morimatsu, Le Quynh Mai, Tetsu Yamashiro, Toshihiro Ito, Koichi Otsuki, Etsuro Ono
    JOURNAL OF GENERAL VIROLOGY 97 9 2104 - 2116 2016年09月 [査読有り][通常論文]
     
    The nucleoprotein (NP) possesses regions that are highly conserved among influenza A viruses, and has therefore been one of the target viral proteins for development of a universal influenza vaccine. It has been expected that human or humanized antibodies will be made available for the prophylaxis, pre-emptive and acute treatment of viral infection. However, it is still unclear whether anti-NP human antibody can confer protection against influenza virus infection. In this study, we generated transgenic mice expressing anti-NP human mAbs derived from lymphocytes of a patient infected with H5N1 highly pathogenic avian influenza (HPAI) virus, and experimental infections were conducted to examine antiviral effects of the anti-NP antibodies against H5N1 HPAI viral infections with a high fatality rate in mammals. Transgenic mouse lines expressing the anti-NP human mAbs at more than 1 mg ml(-1) showed marked resistance to H5N1 virus infections. In addition, resistance to infection with an H1N1 subtype that shows strong pathogenicity to mice was also confirmed. Although the anti-NP mAbs expressed in the transgenic mice did not neutralize the virus, the mAbs could bind to NP located on the surface of infected cells. These results suggested a possibility that the non-neutralizing anti-NP human mAbs could induce indirect antiviral effects, such as antibody-dependent cellular cytotoxicity or complement-dependent cytotoxicity. Taken together, these results demonstrated that anti-NP human mAbs play an important role in heterosubtypic protection against lethal influenza virus infections in vivo.
  • Yukiko Tomioka, Yoshikazu Fujimoto, Kanji Nakai, Kinuyo Ozaki, Sayo Yamamoto, Haruka Suyama, Masami Morimatsu, Toshihiro Ito, Etsuro Ono
    Biochemistry and Biophysics Reports 5 196 - 202 2016年03月01日 [査読有り][通常論文]
     
    Transgenic mouse lines expressing a soluble form of human nectin-2 (hNectin-2Ig Tg) exhibited distinctive elevation of amylase and lipase levels in the sera. In this study, we aimed to clarify the histopathology and to propose the transgenic mouse lines as new animal model for characteristic pancreatic exocrine defects. The significant increase of amylase and lipase levels in sera of the transgenic lines approximately peaked at 8 weeks old and thereafter, plateaued or gradually decreased. The histopathology in transgenic acinar cells was characterized by intracytoplasmic accumulation of abnormal proteins with decrease of normal zymogen granules. The hNectin-2Ig expression was observed in the cytoplasm of pancreatic acinar cells, which was consistent with zymogen granules. However, signals of hNectin-2Ig were very weak in the transgenic acinar cells with the abnormal cytoplasmic accumulaion. The PCNA-positive cells increased in the transgenic pancreas, which suggested the affected acinar cells were regenerated. Acinar cells of hNectin-2Ig Tg had markedly small number of zymogen granules with remarkable dilation of the endoplasmic reticulum (ER) lumen containing abundant abnormal proteins. In conclusion, hNectin-2Ig Tg is proposed as a new animal model for characteristic pancreatic exocrine defects, which are due to the ER stress induced by expression of mutated cell adhesion molecule that is a soluble form of human nectin-2.
  • Kazuhiko Ochiai, Masami Morimatsu, Yuiko Kato, Toshina Ishiguro-Oonuma, Chihiro Udagawa, Oumaporn Rungsuriyawiboon, Daigo Azakami, Masaki Michishita, Yuichi Ariyoshi, Hideo Ueki, Yasutomo Nasu, Hiromi Kumon, Masami Watanabe, Toshinori Omi
    ONCOTARGET 7 3 3273 - 3286 2016年01月 [査読有り][通常論文]
     
    REIC/DKK-3 is a tumor suppressor, however, its intracellular physiological functions and interacting molecules have not been fully clarified. Using yeast two-hybrid screening, we found that small glutamine-rich tetratricopeptide repeat-containing protein a (SGTA), known as a negative modulator of cytoplasmic androgen receptor (AR) signaling, is a novel interacting partner of REIC/DKK-3. Mammalian two-hybrid and pull-down assay results indicated that the SGTA-REIC/DKK-3 interaction involved the N-terminal regions of both REIC/DKK-3 and SGTA and that REIC/DKK-3 interfered with the dimerization of SGTA, which is a component of the AR complex and a suppressor of dynein motor-dependent AR transport and signaling. A reporter assay in human prostate cancer cells that displayed suppressed AR signaling by SGTA showed recovery of AR signaling by REIC/DKK-3 expression. Considering these results and our previous data that REIC/DKK-3 interacts with the dynein light chain TCTEX-1, we propose that the REIC/DKK-3 protein interferes with SGTA dimerization, promotes dynein-dependent AR transport and then upregulates AR signaling.
  • Kato Y, Ochiai K, Michishita M, Azakami D, Nakahira R, Morimatsu M, Ishiguro-Oonuma T, Yoshikawa Y, Kobayashi M, Bonkobara M, Kobayashi M, Takahashi K, Watanabe M, Omi T
    Veterinary journal (London, England : 1997) 206 2 143 - 148 2015年11月 [査読有り][通常論文]
     
    Although the morbidity of canine prostate cancer is low, the majority of cases present with resistance to androgen therapy and poor clinical outcomes. These pathological conditions are similar to the signs of the terminal stage of human androgen-independent prostate cancer. The co-chaperone small glutamine-rich tetratricopeptide repeat-containing protein alpha (SGTA) is known to be overexpressed in human androgenindependent prostate cancer. However, there is little information about the structure and function of canine SGTA. In this study, canine SGTA was cloned and analysed for its ability to suppress androgen receptor signalling. The full-length open reading frame (ORF) of the canine SGTA gene was amplified by RT-PCR using primers designed from canine-expressed sequence tags that were homologous to human SGTA. The canine SGTA ORF has high homology with the corresponding human (89%) and mouse (81%) sequences. SGTA dimerisation region and tetratricopeptide repeat (TPR) domains are conserved across the three species. The ability of canine SGTA to undergo homodimerisation was demonstrated by a mammalian two-hybrid system and a pull-down assay. The negative impact of canine SGTA on androgen receptor (AR) signalling was demonstrated using a reporter assay in androgen-independent human prostate cancer cell lines. Pathological analysis showed overexpression of SGTA in canine prostate cancer, but not in hyperplasia. A reporter assay in prostate cells demonstrated suppression of AR signalling by canine SGTA. Altogether, these results suggest that canine SGTA may play an important role in the acquisition of androgen independence by canine prostate cancer cells. (C) 2015 Elsevier Ltd. All rights reserved.
  • Toshina Ishiguro-Oonuma, Kazuhiko Ochiai, Kazuyoshi Hashizume, Toshihiko Iwanage, Masami Morimatsu
    JAPANESE JOURNAL OF VETERINARY RESEARCH 63 3 107 - 114 2015年08月 [査読有り][通常論文]
     
    Nuclear factor of kappa light polypeptide gene enhancer in B cells (NF-kappa B) inhibitor zeta (Nfkbiz) is a nuclear inhibitor of NF-kappa B (I kappa B) protein that is also termed as molecule possessing ankyrin repeats induced by lipopolysaccharide, interleukin-1-inducible nuclear ankyrin repeat protein, or I kappa B zeta; We found previously that disrupting the Nfkbiz gene resulted in atopic dermatitis-like lesions in mice, suggesting an important role for Nfkbiz in the skin. In this study, we examined the cellular function of Nfkbiz in keratinocytes. Immunohistochemical analyses for Ki-67 revealed that Nfkbiz(-/-) keratinocytes were hypoproliferative. In skin from Nfkbiz(-/-) mice, the expression of the keratinocyte differentiation markers K10 and filaggrin were reduced, although that of K14 was unchanged. The terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling assay revealed that the frequency of apoptosis was comparable between control and Nfkbiz(-/-) keratinocytes. Interestingly, the subcellular localization of the NF-kappa B subunits and the transcriptional activity of NF-kappa B were not changed in Nfkbiz(-/-) keratinocytes. These findings indicate a novel possible role of Nfkbiz in controlling the proliferation and differentiation of epidermal keratinocytes through NF-kappa B-independent mechanisms.
  • Yasunaga Yoshikawa, Masami Morimatsu, Kazuhiko Ochiai, Toshina Ishiguro-Oonuma, Seiichi Wada, Koichi Orino, Kiyotaka Watanabe
    BMC VETERINARY RESEARCH 11 159  2015年07月 [査読有り][通常論文]
     
    Background: Mammary tumors are the most common tumor type in intact female dogs. Recently, the breast cancer 2 early onset (BRCA2) gene was proposed to be associated with tumorigenesis in dogs. The expression level of BRCA2 is important for its DNA repair function in mammalian cells, and its expression level is linked to tumorigenesis in mammary tissue. However, the expression of canine BRCA2 in mammary tumors is unclear. Results: BRCA2 mRNA levels were compared between seven mammary gland samples and seventeen mammary tumor samples isolated from dogs. The expression level of canine BRCA2 in mammary tumor samples was lower than levels in mammary gland samples. We attempted to identify why the BRCA2 expression level was decreased in mammary tumor samples by promoter sequencing analysis; however, we did not find any mutations in the canine BRCA2 promoter that altered BRCA2 transcription levels. We did detect two types of BRCA2 splice variants in 8 mammary tumor samples. One of the variants induced a frame-shift mutation that could lead to nonsense-mediated mRNA decay, a ubiquitous cellular mechanism that eliminates mRNA containing a premature termination codon. Conclusions: Reduced expression of canine BRCA2 mRNA in mammary tumor samples is a possible mechanism to explain mammary tumor development in dogs. One possible reason for reduced BRCA2 mRNA levels in these tumor samples was nonsense-mediated mRNA decay, not mutations in the BRCA2 promoter region. While it remains unclear why canine BRCA2 expression levels are reduced in mammary tumor samples, this study found that the expression level of BRCA2 was associated with canine mammary tumorigenesis.
  • Ahmed F. Abou-Elnaga, Daisuke Torigoe, Mohamed M. Fouda, Ragab A. Darwish, Usama A. Abou-Ismail, Masami Morimatsu, Takashi Agui
    JAPANESE JOURNAL OF VETERINARY RESEARCH 63 2 53 - 62 2015年05月 [査読有り][通常論文]
     
    Depression is one of the most famous psychiatric disorders in humans in all over the countries and considered a complex neurobehavioral trait and difficult to identify causal genes. Tail suspension test (TST) and forced swimming test (FST) are widely used for assessing depression-like behavior and antidepressant activity in mice. A variety of antidepressant agents are known to reduce immobility time in both TST and FST. To identify genetic determinants of immobility duration in both tests, we analyzed 101 F-2 mice from an intercross between C57BL/6 and DBA/2 strains. Quantitative trait locus (QTL) mapping using 106 microsatellite markers revealed three loci (two significant and one suggestive) and five suggestive loci controlling immobility time in the TST and FST, respectively. Results of QTL analysis suggest a broad description of the genetic architecture underlying depression, providing underpinnings for identifying novel molecular targets for antidepressants to clear the complex genetic mechanisms of depressive disorders.
  • Ishiguro-Oonuma T, Ochiai K, Hashizume K, Morimatsu M
    Biomedical research (Tokyo, Japan) 36 2 103 - 107 2015年04月 [査読有り][通常論文]
     
    Nfkbiz is an inhibitor of nuclear factor KB (IkB) protein localized to the nucleus. We previously found that Nfkbiz gene-disrupted mice showed atopic dermatitis-like lesion, implying the important role of Nfkbiz in skin homeostasis. The purpose of this study was to examine the effect of interferon (IFN)-gamma on Nfkbiz expression in keratinocytes. IFN-gamma induced Nfkbiz expression at a comparable level to IL-1. Promoter analysis revealed that interferon-stimulated response element (ISRE) located in the Nfkbiz promoter region is important for responding to the stimulation. Interestingly, IFN-gamma and IL-1 displayed synergism in terms of inducing Nfkbiz expression. By using selective inhibitors, we found that Janus activated kinase (JAK) 1 and nuclear factor (NF)-kB are important for Nfkbiz expression after IFN-gamma stimulation and for synergism between IFN-gamma and ILL These findings indicate a possible important role of Nfkbiz in modulating the progression of inflammatory diseases in which IFN-gamma and IL-1 are abundant.
  • Kazuhiko Ochiai, Toshina Ishiguro-Oonuma, Yasunaga Yoshikawa, Chihiro Udagawa, Yuiko Kato, Masami Watanabe, Makoto Bonkobara, Masami Morimatsu, Toshinori Omi
    BIOMEDICAL RESEARCH-TOKYO 36 2 155 - 158 2015年04月 [査読有り][通常論文]
     
    Mutations in the breast cancer susceptibility gene BRCA2 leading to the failure of interactions with the recombinase RAD51 are associated with an increased risk of cancer in humans. This interaction depends on the eight BRC repeat (BRC1-8) sequences in BRCA2. We previously reported that canine BRC3 has two polymorphisms (T1425P and K1435R) influencing the interaction with RAD51, and 1435R was identified in mammary tumor dog samples. In this study, we investigated the sequence variations of BRC3 and 4 in 236 dogs of five breeds. Allele frequencies of 1425P and 1435R were 0.063 and 0.314, respectively, and there was no other polymorphism in the sequenced region. A mammalian two-hybrid assay using BRC3-4 sequences demonstrated that 1425P allele reduced the binding strength with RAD51 but 1435R had no effect. These results may provide an insight into the functions of not only individual but also multiple BRC repeats of BRCA2 in dogs.
  • Tomohisa Tanaka, Hirotake Kasai, Atsuya Yamashita, Kaori Okuyama-Dobashi, Jun Yasumoto, Shinya Maekawa, Nobuyuki Enomoto, Toru Okamoto, Yoshiharu Matsuura, Masami Morimatsu, Noboru Manabe, Kazuhiko Ochiai, Kazuto Yamashita, Kohji Moriishi
    JOURNAL OF VIROLOGY 88 22 13352 - 13366 2014年11月 [査読有り][通常論文]
     
    Equine hepacivirus (EHcV) has been identified as a closely related homologue of hepatitis C virus (HCV) in the United States, the United Kingdom, and Germany, but not in Asian countries. In this study, we genetically and serologically screened 31 serum samples obtained from Japanese-born domestic horses for EHcV infection and subsequently identified 11 PCR-positive and 7 seropositive serum samples. We determined the full sequence of the EHcV genome, including the 3' untranslated region (UTR), which had previously not been completely revealed. The polyprotein of a Japanese EHcV strain showed approximately 95% homology to those of the reported strains. HCV-like cis-acting RNA elements, including the stem-loop structures of the 3' UTR and kissing-loop interaction were deduced from regions around both UTRs of the EHcV genome. A comparison of the EHcV and HCV core proteins revealed that Ile(190) and Phe(191) of the EHcV core protein could be important for cleavage of the core protein by signal peptide peptidase (SPP) and were replaced with Ala and Leu, respectively, which inhibited intramembrane cleavage of the EHcV core protein. The loss-of-function mutant of SPP abrogated intramembrane cleavage of the EHcV core protein and bound EHcV core protein, suggesting that the EHcV core protein may be cleaved by SPP to become a mature form. The wild-type EHcV core protein, but not the SPP-resistant mutant, was localized on lipid droplets and partially on the lipid raft-like membrane in a manner similar to that of the HCV core protein. These results suggest that EHcV may conserve the genetic and biological properties of HCV. IMPORTANCE EHcV, which shows the highest amino acid or nucleotide homology to HCV among hepaciviruses, was previously reported to infect horses from Western, but not Asian, countries. We herein report EHcV infection in Japanese-born horses. In this study, HCV-like RNA secondary structures around both UTRs were predicted by determining the whole-genome sequence of EHcV. Our results also suggest that the EHcV core protein is cleaved by SPP to become a mature form and then is localized on lipid droplets and partially on lipid raft-like membranes in a manner similar to that of the HCV core protein. Hence, EHcV was identified as a closely related homologue of HCV based on its genetic structure as well as its biological properties. A clearer understanding of the epidemiology, genetic structure, and infection mechanism of EHcV will assist in elucidating the evolution of hepaciviruses as well as the development of surrogate models for the study of HCV.
  • Yukiko Tomioka, Masami Morimatsu, Ken-ichi Nishijima, Tatsufumi Usui, Sayo Yamamoto, Haruka Suyama, Kinuyo Ozaki, Toshihiro Ito, Etsuro Ono
    BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS 450 1 532 - 537 2014年07月 [査読有り][通常論文]
     
    Tumor-associated MUC1 binds to Siglec-9, which is expected to mediate tumor cell growth and negative immunomodulation. We hypothesized that a soluble form of Siglec-9 (sSiglec-9) competitively inhibits a binding of MUC1 to its receptor molecules like human Siglec-9, leading to provide antitumor benefit against MUC1-expressing tumor, and generated transgenic mouse lines expressing sSiglec-9 (sSiglec-9 Tg). When mammary tumor cells expressing MUC1 were intraperitoneally transplanted into sSiglec-9 Tg, tumor proliferation was slower with the lower histological malignancy as compared with non-transgenic mice. The sSiglec-9 was detected in the ascites caused by the tumor in the sSiglec-9 Tg, and sSiglec-9 and MUC1 were often colocalized on surfaces of the tumor cells. PCNA immunohistochemistry also revealed the reduced proliferation of the tumor cells in sSiglec-9 Tg. In sSiglec-9 Tg with remarkable suppression of tumor proliferation, MUC1 expressions were tend to be reduced. In the ascites of sSiglec-9 Tg bearing the tumor, T cells were uniformly infiltrated, whereas aggregations of degenerative T cells were often observed in the non-transgenic mice. These results suggest that sSiglec-9 has an antitumor benefit against MUC1-expressing tumor in the transgenic mice, which may avoid the negative immunomodulation and/or suppress tumor-associated MUC1 downstream signal transduction, and subsequent tumor proliferation. (C) 2014 Elsevier Inc. All rights reserved.
  • Aya Yoshimura, Manabu Musashi, Takeaki Kaneko, Shunsuke Ohnishi, Chieko Orito, Yukako Kawahara, Satoshi Hashino, Masami Morimatsu, Satoshi Konno, Jiro Arikawa, Tetsuya Ishii, Masaya Sawamura, Ichiro Ueda
    Japanese Journal of Allergology 63 8 1132 - 1139 2014年 [査読有り][通常論文]
     
    Background: Based on a case who developed anaphylaxis after mouse bite which occurred at Hokkaido University, we studied on allergic sensitization prevalence for laboratory animals among students and researchers who are exposed to laboratory rodents and rabbit, for the purpose of allergy prevention, particularly anaphylaxis. Methods: We carried out the health check-up on laboratory animal allergy (LAA) by questionnaires and specific-IgE antibody test for 555 rodents and/or rabbit handlers from whom informed consent was obtained. Result: Prevalence of positive IgE antibody higher than class 1 to mice, rats, hamsters, guinea pigs, and/or rabbits in the examinees was 14.1% (62/441), 17.9% (50/279), 18.8% (6/32), 17.4% (4/23), and 11.3% (12/106), respectively. Moreover, among users of mouse, those who had allergic symptoms during contact with animals resulted in significantly higher positive rate for anti-mouse IgE antibody test than the other (38.1% vs 8.8%, p< 0.01). Conclusion: Health check-up including measurement of specific-IgE antibody against laboratory animals is useful for understanding allergic sensitization.
  • Yukiko Tomioka, Masami Morimatsu, Satoshi Taharaguchi, Sayo Yamamoto, Haruka Suyama, Kinuyo Ozaki, Naoki Iwamori, Etsuro Ono
    BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS 440 4 683 - 688 2013年11月 [査読有り][通常論文]
     
    Transcription factors of alphaherpesviruses not only control the expression of their own viral genes, but also influence the gene expression of mammalian cells. In the course of breeding of the transgenic mouse line (TgIE96) expressing the immediate-early protein IE180 of pseudorabies virus belonging to the subfamily Alphaherpesvirinae, we found that TgIE96 male mice suffered from severe breeding difficulties. Testes of TgIE96 were smaller than that of non-transgenic littermates and abnormal spermatogenesis such as morphological, numerical and functional anomalies of spermatozoa were found in the transgenic mouse line. Expression of IE180 was detected in the germ cells at all stages, especially spermatocytes, and fewer Sertoli cells. In addition, expression of IE180 was also detected in the germinal cells of C57BL/6 mice inoculated with PRV into their testes. These results suggest that IE180 of PRV induces male infertility by abnormal spermatogenesis, which effect morphological, numerical, and functional anomalies of spermatozoa, in transgenic mice. (C) 2013 Elsevier Inc. All rights reserved.
  • Kazuhiko Ochiai, Masami Watanabe, Daigo Azakami, Masaki Michishita, Yasunaga Yoshikawa, Chihiro Udagawa, Pornphimon Metheenukul, Thippayarat Chahomchuen, Hiroshi Aoki, Hiromi Kumon, Masami Morimatsu, Toshinori Omi
    VETERINARY JOURNAL 197 3 769 - 775 2013年09月 [査読有り][通常論文]
     
    REIC/Dkk-3, a member of the human Dickkopf (Dkk) family, plays a role as a suppressor of growth in several human cancers. In this study, the tumour suppression function of canine REIC/Dkk-3 was investigated. The full-length open reading frame of the canine REIC/Dkk-3 homologue was cloned and the tissue distribution of REIC/Dkk-3 mRNA was determined, along with the subcellular localisation of the REIC/Dkk-3 protein in canine cancer cell lines. Expression of REIC/Dkk-3 was lower in mammary gland tumours and in canine mammary carcinoma cell lines than in normal mammary gland tissue. Overexpression of REIC/Dkk-3 induced apoptosis in canine mammary carcinoma cell lines. These results show that expression of REIC/Dkk-3 is downregulated in canine mammary tumours and that one of the functions of this gene is induction of apoptosis. (C) 2013 Elsevier Ltd. All rights reserved.
  • Dominic G. Hildebrand, Eva Alexander, Sebastian Hoerber, Simon Lehle, Kerstin Obermayer, Niels-Arne Muenck, Oliver Rothfuss, Julia-Stefanie Frick, Masami Morimatsu, Ingo Schmitz, Johannes Roth, Jan M. Ehrchen, Frank Essmann, Klaus Schulze-Osthoff
    JOURNAL OF IMMUNOLOGY 190 9 4812 - 4820 2013年05月 [査読有り][通常論文]
     
    CCL2, also referred to as MCP-1, is critically involved in directing the migration of blood monocytes to sites of inflammation. Consequently, excessive CCL2 secretion has been linked to many inflammatory diseases, whereas a lack of expression severely impairs immune responsiveness. We demonstrate that I kappa B zeta, an atypical I kappa B family member and transcriptional coactivator required for the selective expression of a subset of NF-kappa B target genes, is a key activator of the Ccl2 gene. I kappa B zeta-deficient macrophages exhibited impaired secretion of CCL2 when challenged with diverse inflammatory stimuli, such as LPS or peptidoglycan. These findings were reflected at the level of Ccl2 gene expression, which was tightly coupled to the presence of I kappa B zeta. Moreover, mechanistic insights acquired by chromatin immunoprecipitation demonstrate that I kappa B zeta is directly recruited to the proximal promoter region of the Ccl2 gene and is required for transcription-enhancing histone H3 at lysine-4 trimethylation. Finally, I kappa B zeta-deficient mice showed significantly impaired CCL2 secretion and monocyte infiltration in an experimental model of peritonitis. Together, these findings suggest a distinguished role of I kappa B zeta in mediating the targeted recruitment of monocytes in response to local inflammatory events. The Journal of Immunology, 2013, 190: 4812-4820.
  • 吉村彩, 武藏学, 金子壮朗, 大西俊介, 折戸智恵子, 川原由佳子, 森松正美, 今野哲, 有川二郎
    Campus Health 50 1 316  2013年03月 [査読無し][通常論文]
  • Yasunaga Yoshikawa, Kazuhiko Ochiai, Masami Morimatsu, Yu Suzuki, Seiichi Wada, Takahiro Taoda, Satomi Iwai, Seishiro Chikazawa, Koichi Orino, Kiyotaka Watanabe
    PLOS ONE 7 10 e45833  2012年10月 [査読有り][通常論文]
     
    Mammary tumors are the most common tumor type in both human and canine females. Mutations in the breast cancer susceptibility gene, BRCA2, have been found in most cases of inherited human breast cancer. Similarly, the canine BRCA2 gene locus has been associated with mammary tumors in female dogs. However, deleterious mutations in canine BRCA2 have not been reported, thus far. The BRCA2 protein is involved in homologous recombination repair via its interaction with RAD51 recombinase, an interaction mediated by 8 BRC repeats. These repeats are 26-amino acid, conserved motifs in mammalian BRCA2. Previous structural analyses of cancer-associated mutations affecting the BRC repeats have shown that the weakening of RAD51's affinity for even 1 repeat is sufficient to increase breast cancer susceptibility. In this study, we focused on 2 previously reported canine BRCA2 mutations (T1425P and K1435R) in BRC repeat 3 (BRC3), derived from mammary tumor samples. These mutations affected the interaction of canine BRC3 with RAD51, and were considered deleterious. Two BRC3 mutations (K1440R and K1440E), reported in human breast cancer patients, occur at amino acids corresponding to those of the K1435R mutation in dogs. These mutations affected the interaction of canine BRC3 with RAD51, and may also be considered deleterious. The two BRC3 mutations and a substitution (T1430P), corresponding to T1425P in canine BRCA2, were examined for their effects on human BRC3 function and the results were compared between species. The corresponding mutations and the substitution showed similar results in both human and canine BRC3. Therefore, canine BRCA2 may be a good model for studying human breast cancer caused by BRCA2 mutations.
  • Hiroki Takakuwa, Tetsu Yamashiro, Mai Q. Le, Lien S. Phuong, Hiroichi Ozaki, Ryota Tsunekuni, Tatsufumi Usui, Hiroshi Ito, Masami Morimatsu, Yukiko Tomioka, Tsuyoshi Yamaguchi, Toshihiro Ito, Toshiyuki Murase, Etsuro Ono, Koichi Otsuki
    PREVENTIVE VETERINARY MEDICINE 103 2-3 192 - 200 2012年02月 [査読有り][通常論文]
     
    Repeated epizootics of highly pathogenic avian influenza (HPAI) virus subtype H5N1 were reported from 2003 to 2005 among poultry in Vietnam. More than 200 million birds were killed to control the spread of the disease. Human cases of H5N1 infection have been sporadically reported in an area where repeated H5N1 outbreaks among birds had occurred. Subtype H5N1 strains are established as endemic among poultry in Vietnam, however, insights into how avian influenza viruses including the H5N1 subtype are maintained in endemic areas is not clear. In order to determine the prevalence of different avian influenza viruses (AIVs), including H5N1 circulating among poultry in northern Vietnam, surveillance was conducted during the years 2006-2009. A subtype H5N1 strain was isolated from an apparently healthy duck reared on a farm in northern Vietnam in 2008 and was identified as an HPAI. Although only one H5N1 virus was isolated, it supports the view that healthy domestic ducks play a pivotal role in maintaining and transmitting H5N1 viruses which cause disease outbreaks in northern Vietnam. In addition, a total of 26 AlVs with low pathogenicity were isolated from poultry and phylogenetic analysis of all the eight gene segments revealed their diverse genetical backgrounds, implying that reassortments have occurred frequently among strains in northern Vietnam. It is, therefore, important to monitor the prevalence of influenza viruses among healthy poultry between epidemics in an area where AlVs are endemic. (C) 2011 Elsevier B.V. All rights reserved.
  • Yasunaga Yoshikawa, Masami Morimatsu, Kazuhiko Ochiai, Kento Okuda, Takahiro Taoda, Seishiro Chikazawa, Asako Shimamura, Toshinori Omi, Makoto Bonkobara, Koichi Orino, Kiyotaka Watanabe
    BMC Research Notes 5 173 - 182 2012年 [査読有り][通常論文]
     
    Background: Mammary tumors are the most common tumor type in both human and canine females. In women, carriers of mutations in BRCA2, a tumor suppressor gene product, have a higher risk of breast cancer. Canine BRCA2 has also been suggested to have a relationship with mammary tumors. However, clearly deleterious BRCA2 mutations have not been identified in any canine mammary tumors, as appropriate methods to detect mutations or a consensus BRCA2 sequence have not been reported. Findings. For amplification and sequencing of BRCA2, we designed 14 and 20 PCR primer sets corresponding to the BRCA2 open reading frame (ORF) and all 27 exons, respectively, including exon-intron boundaries of the canine BRCA2 regions, respectively. To define the consensus canine BRCA2 ORF sequence, we used established methods to sequence the full-length canine BRCA2 ORF sequence from two ovaries and a testis obtained from individual healthy mongrel dogs and partially sequence BRCA2 genomic sequences in 20-56 tumor-free dogs, each aged over 6 years. Subsequently, we compared these sequences and seven previously reported sequences, and defined the most common base sequences as the consensus canine BRCA2 ORF sequence. Moreover, we established a detection method for identifying splicing variants. Unexpectedly, we also identified novel splicing variants in normal testes during establishment of these methods. Conclusions: The present analysis methods for determining the BRCA2 base sequence and for detecting BRCA2 splicing variants and the BRCA2 ORF consensus sequence are useful for better understanding the relationship between canine BRCA2 mutation status and cancer risk. © 2010 Yoshikawa et al licensee BioMed Central Ltd.
  • K. Ochiai, Y. Yoshikawa, T. Oonuma, Y. Tomioka, K. Hashizume, M. Morimatsu
    VETERINARY JOURNAL 190 2 293 - 295 2011年11月 [査読有り][通常論文]
     
    In humans, mutations in the gene for the breast cancer susceptibility protein BRCA2 affect its interactions with the recombinase RAD51 and are associated with an increased risk of cancer. This interaction occurs through a series of eight BRC repeat sequences in BRCA2. A mammalian two-hybrid assay using individual BRC repeats demonstrated that BRC6 did not bind to RAD51, whereas there was strong (BRC1, 2 and 4), intermediate (BRC8), or weak (BRC3, 5 and 7) binding of other BRC repeats to RAD51. In serial deletion mutation experiments, binding strengths were increased when the C-terminal BRC repeat was removed from BRC1-8, BRC1-5 and BRC1-3. These results may provide an insight into the effects of missense or truncation mutations in BRCA2 in canine tumours. (C) 2010 Elsevier Ltd. All rights reserved.
  • Shunsuke Meike, Tohru Yamamori, Hironobu Yasui, Masato Eitaki, Akira Matsuda, Masami Morimatsu, Masakazu Fukushima, Yasundo Yamasaki, Osamu Inanami
    MOLECULAR CANCER 10 92 - 100 2011年07月 [査読有り][通常論文]
     
    Background: A novel anticancer drug 1-(3-C-ethynyl-beta-D-ribo-pentofuranosyl)cytosine (ECyd, TAS106) has been shown to radiosensitize tumor cells and to improve the therapeutic efficiency of X-irradiation. However, the effect of TAS106 on cellular DNA repair capacity has not been elucidated. Our aim in this study was to examine whether TAS106 modified the repair capacity of DNA double-strand breaks (DSBs) in tumor cells. Methods: Various cultured cell lines treated with TAS106 were irradiated and then survival fraction was examined by the clonogenic survival assays. Repair of sublethal damage (SLD), which indicates DSBs repair capacity, was measured as an increase of surviving cells after split dose irradiation with an interval of incubation. To assess the effect of TAS106 on the DSBs repair activity, the time courses of gamma-H2AX and 53BP1 foci formation were examined by using immunocytochemistry. The expression of DNA-repair-related proteins was also examined by Western blot analysis and semi-quantitative RT-PCR analysis. Results: In clonogenic survival assays, pretreatment of TAS106 showed radiosensitizing effects in various cell lines. TAS106 inhibited SLD repair and delayed the disappearance of gamma-H2AX and 53BP1 foci, suggesting that DSB repair occurred in A549 cells. Western blot analysis demonstrated that TAS106 down-regulated the expression of BRCA2 and Rad51, which are known as keys among DNA repair proteins in the homologous recombination (HR) pathway. Although a significant radiosensitizing effect of TAS106 was observed in the parental V79 cells, pretreatment with TAS106 did not induce any radiosensitizing effects in BRCA2-deficient V-C8 cells. Conclusions: Our results indicate that TAS106 induces the down-regulation of BRCA2 and the subsequent abrogation of the HR pathway, leading to a radiosensitizing effect. Therefore, this study suggests that inhibition of the HR pathway may be useful to improve the therapeutic efficiency of radiotherapy for solid tumors.
  • Kazuhiko Ochiai, Yasunaga Yoshikawa, Kumiko Yoshimatsu, Toshina Oonuma, Yukiko Tomioka, Eichi Takeda, Jiro Arikawa, Katsumi Mominoki, Toshinori Omi, Kazuyoshi Hashizume, Masami Morimatsu
    FEBS LETTERS 585 12 1771 - 1777 2011年06月 [査読有り][通常論文]
     
    The breast cancer susceptibility protein BRCA2 is essential for recombinational DNA repair. BRCA2 specifically binds to RAD51 via eight BRC repeat motifs and delivers RAD51 to double-stranded DNA breaks. In this study, a mammalian two-hybrid assay and competitive ELISA showed that the interaction between BRC repeat 4 (BRC4) and RAD51 was strengthened by the substitution of a single BRC4 amino acid from valine to isoleucine (V1532I). However, the cancer-associated V1532F mutant exhibited very weak interaction with RAD51. This study used a comparative analysis of BRC4 between animal species to identify V1532 as an important residue that interacts with RAD51. Structured summary of protein interactions: cRAD51 physically interacts with cRAD51 by two hybrid (View interaction) fBRC4 physically interacts with cRAD51 by two hybrid (View interaction) cBRC4 physically interacts with cRAD51 by two hybrid (View interaction) hBRC4 physically interacts with hBRC4 and hRAD51 by competition binding (View Interaction 1, 2) hBRC4 physically interacts with cRAD51 by two hybrid (View interaction) hBRC4 binds to hRAD51 by enzyme linked immunosorbent assay (View interaction) (C) 2011 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.
  • Juan C. Lopez-Ramos, Yukiko Tomioka, Masami Morimatsu, Sayo Yamamoto, Kinuyo Ozaki, Etsuro Ono, Jose M. Delgado-Garcia
    PLOS ONE 5 8 e12123  2010年08月 [査読有り][通常論文]
     
    The cerebellum in transgenic mice expressing pseudorabies virus immediate-early protein IE180 (TgIE96) was substantially diminished in size, and its histoarchitecture was severely disorganized, resulting in severe ataxia. TgIE96 mice can therefore be used as an experimental model to study the involvement of cerebellar circuits in different learning tasks. The performance of three-month-old TgIE96 mice was studied in various behavioral tests, including associative learning (classical eyeblink conditioning), object recognition, spatial orientation (water maze), startle response and prepulse inhibition, and passive avoidance, and compared with that of wild-type mice. Wild-type and TgIE96 mice presented similar reflexively evoked eyeblinks, and acquired classical conditioned eyelid responses with similar learning curves for both trace and delay conditioning paradigms. The two groups of mice also had similar performances during the object recognition test. However, they showed significant differences for the other three tests included in this study. Although both groups of animals were capable of swimming, TgIE96 mice failed to learn the water maze task during the allowed time. The startle response to a severe tone was similar in both control and TgIE96 mice, but the latter were unable to produce a significant prepulse inhibition. TgIE96 mice also presented evident deficits for the proper accomplishment of a passive avoidance test. These results suggest that the cerebellum is not indispensable for the performance of classical eyeblink conditioning and for object recognition tasks, but seems to be necessary for the proper performance of water maze, prepulse inhibition, and passive avoidance tests.
  • Kiyoshi Kano, Ayami Kitamura, Takashi Matsuwaki, Masami Morimatsu, Kunihiko Naito
    MOLECULAR REPRODUCTION AND DEVELOPMENT 77 1 29 - 37 2010年01月 [査読有り][通常論文]
     
    Discoidin domain receptor 2 (DDR2) is a receptor tyrosine kinase (RTK). We recently identified homozygous smallie mutant mice (BKS.HRS. Ddr2(slie/slie)/J, Ddr2(slie/slie) mutants), which lack a functional DDR2. Ddr2(slie/slie) mutant mice are dwarfed and infertile due to peripheral dysregulation of the endocrine system. To understand the role of DDR2 signaling in spermatogenesis, we studied the expression of several receptors, enzymes, and proteins related to spermatogenesis in wild-type and Ddr2(slie/slie) mutant mice at 10 weeks and 5 months of age. DDR2 were expressed in adult wild-type male mice in Leydig cells. The number of differentiated spermatozoa in the seminal fluid was significantly lower in the Ddr2(slie/slie) mutant mice than in the wild-type mice. The number of TUNEL-positive cells was significantly greater in 5-month-old Ddr2(slie/slie) mutants. Testosterone was significantly reduced at 5 months age, but LH was similar in both types of mice at both 10 weeks and 5 months of age. The expression levels of LH receptors (Lhcgr), StAR, P450scc, and Hsd3 beta 6 were significantly different between the two types of mice at 10 weeks of age, but they were significantly reduced in 5-month-old Ddr2(slie/slie) mutants compared to wild-type mice of the same age. DDR2 was expressed in the Leydig cells of adult wild-type male mice. In conclusion, our results indicated that DDR2 signaling plays a critical role in the maintenance of male spermatogenesis.
  • Yukiko Tomioka, Masami Morimatsu, Keiko Amagai, Minako Kuramochi, Yuki Watanabe, Shigeto Kouda, Toshio Wada, Noritaka Kuboki, Etsuro Ono
    MICROBIOLOGY AND IMMUNOLOGY 53 1 8 - 15 2009年01月 [査読有り][通常論文]
     
    Nectin-1 is a Ca(2+)-independent Ig-like cell-cell adhesion molecule and an alphaherpesvirus receptor that binds to virion glycoprotein D by the first Ig-like domain. We have investigated the antiviral potentials of soluble forms of porcine nectin-1 to PRV infection by generating transgenic mice expressing different types of fusion protein. Previously, we reported that mice transgenic for a chimera that carried the entire ectodomain of porcine nectin-1 fused to the Fc portion of porcine IgG1 were more resistant than those transgenic for a chimera that carried the first Ig-like domain fused to the Fc portion. Recently, we generated transgenic mice expressing a fusion protein made of the first Ig-like domain fused to the Fc portion of human IgG1, and reported that they showed a microphthalmia. Here, two transgenic mouse lines expressing the fusion protein were challenged with PRV for comparing their resistances with those of transgenic mice expressing different types of fusion protein. Surprisingly, both transgenic mouse lines showed a high resistance to the viral infection, especially via the i.n. route. Significant resistance of the embryonic fibroblasts was also observed. Altogether, these findings indicated that the fusion protein consisting of the first Ig-like domain fused to the human Fc portion provided a marked resistance against PRV infection to the transgenic mice.
  • Yasunaga Yoshikawa, Masami Morimatsu, Kazuhiko Ochiai, Masashi Nagano, Yukiko Tomioka, Nobuo Sasaki, Kazuyoshi Hashizume, Toshihiko Iwanaga
    AMERICAN JOURNAL OF VETERINARY RESEARCH 69 10 1323 - 1328 2008年10月 [査読有り][通常論文]
     
    Objective-To establish novel polymorphic markers for analysis of loss of heterozygosity (LOH), so as to study the possible involvement of BRCA2 in mammary tumors obtained from dogs. Sample Population-Blood samples, mammary gland specimens, or mammary tumors from 3 tumor-bearing dogs and 10 tumor-free dogs. Procedures-Nucleotide sequence analysis was performed with a DNA autosequencer. Loss of heterozygosity analysis was performed for markers established in the present study. The expression level of canine BRCA2 was quantified by real-time PCR analysis. Results-3 novel microsatellite markers with high heterozygosity rates (> 50%) were established, and the previously reported marker for canine BRCA2 gene locus was improved. These markers were used for the analysis of DNA from formalin-fixed and paraffin-embedded samples. By use of these markers, LOH in canine BRCA2 was identified as a result of recombination. In mammary tumor DNA that corresponded to the LOH-positive dog, the level of canine BRCA2 expression was decreased compared with that of nonneoplastic mammary gland tissue; the open reading frame contained 4 missense variations, 1 insertion variation, and 1 silent variation, some of which were localized to functional domains. Conclusions and Clinical Relevance-3 novel polymorphic markers were developed for LOH analysis of canine BRCA2 and identified a dog with LOH with some variations in the functional domains. These markers could be useful for assessing the relevance of BRCA2 variation in mammary tumors of dogs.
  • Kazuhiko Yoshida, Yukiko Tomioka, Satoru Kase, Masami Morimatsu, Kyoko Shinya, Shigeaki Ohno, Etsuro Ono
    GRAEFES ARCHIVE FOR CLINICAL AND EXPERIMENTAL OPHTHALMOLOGY 246 4 543 - 549 2008年04月 [査読有り][通常論文]
     
    Background Nectins are Ca2+-independent immunoglobulin (Ig)-like cell-cell-adhesion molecules. We have generated transgenic mice expressing a series of soluble forms of nectin-1, and investigated special effects of each soluble form of nectin-1 in vivo. In the course of generating transgenic mice expressing a soluble form of nectin-1 consisting of the first Ig-like domain of nectin-1 and the Fc portion of human IgG1 (PHveC-VhIg), we found that all of the transgenic founder mice showed a microphthalmia. The purpose of this study is to examine functions of the extracellular domains of nectin-1 in eye development using transgenic technology. Methods Eyes of four different transgenic mouse lines expressing each soluble form of nectin-1 were analyzed histologically. Tissue sections were processed with hematoxylin-eosin staining and indirect immunoperoxidase technique. Results All of five transgenic mouse founders expressing PHveC-VhIg, and of three lines expressing PHveC-VpIg made of the first Ig-like domain fused to porcine Fc portions at 5 weeks showed a microphthalmia, but not all of the transgenic mouse lines expressing PHveCIg or PHveCpIg made of the entire ectodomain fused to human or porcine Fc portions. In the abnormal eyes, the vitreous body was almost absent. In PHveC-VhIg-expressing mice at postnatal day 6, each vitreous space was very small. In the neonatal transgenic mice, the vitreous body was almost the same as that of control mice, and PHveC-VhIg was expressed in the optic nerve, conjunctival epithelium, ciliary body, corneal and lens epithelium. At this stage, nectin-1, -3 and -4 were stained in the optic nerve of control mice as well as in that of the transgenic mice. Nectin-1 is faintly stained in the epithelium of the cornea and lens epithelium, but not in the ciliary body. Conclusion Soluble forms of the first Ig-like domain of nectin-1 (PHveC-VhIg and PHveC-VpIg), but not those of the entire ectodomain (PHveCIg and PHveCpIg), lead to microphthalmia and lack of vitreous body in the transgenic mice.
  • Yukiko Tomioka, Taisuke Miyazaki, Satoshi Taharaguchi, Saori Yoshino, Masami Morimatsu, Toshimitsu Uede, Etsuro Ono, Masahiko Watanabe
    EUROPEAN JOURNAL OF NEUROSCIENCE 27 8 2115 - 2132 2008年04月 [査読有り][通常論文]
     
    Pseudorabies virus is an alphaherpesvirus causing fatal neurological diseases in animals. Pseudorabies virus carries a gene encoding immediate-early (IE) protein IE180, which controls the transcription of other viral and host cell genes. Previously, we reported that transgenic expression of IE180 in mice causes severe ataxia and cerebellar deformity. Here we identified profound abnormalities in adult IE180 transgenic mice, including malpositioning of Purkinje cells (PCs), granule cells (GCs) and Bergmann glia (BG), impaired dendritogenesis and synaptogenesis in PCs, disoriented BG fibers, absence of molecular layer interneurons, and increased apoptosis of neurons and glia. In accordance with the cellular defects, we found the expression of IE180 in PCs, GCs and astrocytes during cerebellar development. We next examined transgenic mice expressing truncated IE180 mutants: dlN132 lacking the acidic transcriptional active domain, dlC629 lacking the nuclear localization signal and dlC1081 having all known domains but lacking the carboxyl-terminal sequence. Despite similar expression levels of the transgenes, ataxia and cerebellar defects were only manifested in the dlC1081 transgenic mice but their phenotypes were milder compared with the IE180 transgenic mice. In the dlC1081 transgenic mice, cerebellar neurons and glia were normally positioned but cerebellar size was severely reduced due to GC deficits. Interestingly, dlC1081 was mainly expressed in the GCs with low expression in a few BG. Taken together, the present findings clarified a causal relationship between cerebellar pathology and cellular expression of IE180, and further afforded an experimental insight into different symptomatic severity as a consequence of different cellular defects caused by such cytotoxic viral agents.
  • Etsuro Ono, Yukiko Tomioka, Yuki Watanabe, Keiko Amagai, Masami Morimatsu, Kyoko Shinya, Pierre Cherel
    JOURNAL OF GENERAL VIROLOGY 88 Pt 10 2636 - 2641 2007年10月 [査読有り][通常論文]
     
    Nectin-1 is an alphaherpesvirus receptor that binds to virion glycoprotein D by the first immunoglobulin (lg)-like domain. The possibility of making animals resistant to pseudorabies virus (PRV) infection has been investigated by generating transgenic mice expressing soluble forms of porcine nectin-1. Previously, transgenic mice were generated that expressed a fusion protein made of the entire ectodomain of nectin-1 fused to the Fc portion of human IgG, or the first lg-like domain fused to the Fc portion of porcine IgG. Here, the contribution of the second and third lg-like domains of nectin-1 was analysed by generating transgenic mice expressing the entire ectodomain of nectin-1 fused to the porcine Fc portion. Transgenic mice expressing each of three different fusion proteins were challenged with PRV for comparison of their resistance. Altogether, mice transgenic for a chimera that carried the entire ectodomain were more resistant than those transgenic for a chimera that carried the first lg-like domain.
  • Toshina Oonuma, Masami Morimatsu, Kazuhiko Ochiai, Toshihiko Iwanaga, Kazuyoshi Hashizume
    JOURNAL OF VETERINARY MEDICAL SCIENCE 69 3 279 - 284 2007年03月 [査読有り][通常論文]
     
    Molecule possessing ankyrin-repeats induced by lipopolysaccharide (MAIL) is a nuclear I kappa B protein that is also known as interleukin-1-inducible nuclear ankyrin repeat protein and inhibitor of nuclear factor kappa B zeta (I kappa B zeta). We previously observed that MAIL-deficient mice were affected by atopic dermatitis-like skin lesions and demonstrated the importance of MAIL in the skin. In this study, we investigated MAIL expression in mouse keratinocytes. MAIL mRNA was constitutively expressed in the skin epidermis. MAIL expression was also confirmed in primary keratinocytes and the PAM212 keratinocyte cell line. The inhibitors of nuclear factor kappa B (NF kappa B)-Bay 11-7082 and the I kappa B alpha M supersuppressor-considerably downregulated MAIL expression in the keratinocytes. Immunoreactivity for NF-kappa B components was localized in the cytoplasm and nucleus of normal unstimulated keratinocytes. The expression level of MAIL in the skin did not change following lipopolysaccharide (LPS) administration to mice. Interestingly, in accordance with the in vivo findings, the MAIL expression level did not change following LPS stimulation even in primary keratinocytes; however, MAIL expression was strongly increased by interleukin-1 stimulation. These results collectively suggest that the constitutive expression of MAIL in keratinocytes is controlled, at least in part, by NF-kappa B and that there may be LPS-specific repressive mechanisms that inhibit MAIL induction.
  • Junko Nio, Hiromi Takahashi-Iwanaga, Masami Morimatsu, Yasuhiro Kon, Toshihiko Iwanaga
    HISTOCHEMISTRY AND CELL BIOLOGY 126 1 45 - 56 2006年07月 [査読有り][通常論文]
     
    Galectin is an animal lectin that has high affinity to beta-galactoside of glycoconjugates. In the present study, cellular expression of galectin subtypes in the urinary system of adult mice was examined by in situ hybridization and immunohistochemistry. The major subtype expressed in the murine urinary system was galectin-3, which was expressed continuously from the kidney to the distal end of the urethra. The renal cortex expressed galectin-3 more intensely than the medulla. Renal galectin-3 immunoreactivity was strongest in the cortical collecting ducts, where principal cells were the sole cellular source. All cell layers of the transitional epithelium from the renal pelvis to the urethra strongly expressed galectin-3 at the mRNA and protein levels. An electron microscopic study demonstrated diffuse cytoplasmic localization of galectin-3 in principal cells of the collecting ducts and in the bladder epithelial cells. Urethral galectin-3 expression at the pars spongiosa decreased in intensity near the external urethral orifice, where the predominant subtype of galectin was substituted by galectin-7. The muscular layer of the ureter and urinary bladder contained significant signals for galectin-1. Taken together, the observations indicate that the adult urinary system shows intense and selective expression of galectin-3 in epithelia of the uretic bud- and cloaca-derivatives.
  • RS Fujino, K Tanaka, M Morimatsu, K Tamura, H Kogo, T Hara
    MOLECULAR ENDOCRINOLOGY 20 4 904 - 915 2006年04月 [査読有り][通常論文]
     
    In spermatogenesis, Sertoli cells serve as supporting cells for the proliferation and differentiation of germ cells. However, it appears that Sertoli cell function is regulated by adjacent spermatogonial cells in the testis because expression of lipocalin-2 mRNA, which encodes an iron-siderophore-binding protein, is barely detectable in Sertoli cells of germ cell-deficient W/W-v mice, and more abundantly expressed in jsd/jsd mice. By employing a coculture system comprising immortalized Sertoli cells ( designated as Sertoli-B) and c-Kit(+) spermatogonial cells from 7-d-old mouse testis, we found that lipocalin-2 gene transcription in Sertoli cells is induced by a factor secreted from spermatogonial cells. Transfection of Sertoli-B cells with a series of reporter constructs encompassing an upstream region of the mouse lipocalin-2 gene revealed that a nuclear factor (NF)-kappa B binding consensus sequence in the proximal region of lipocalin-2 gene is responsible for transcriptional activation. A major NF-kappa B component, p65, bound to this region and translocated from the cytoplasm to the nucleus upon stimulation with spermatogonial cell-conditioned medium. Moreover, short interference RNA directed to p65 or a dominant-negative form of I kappa B alpha suppressed the spermatogonial cell factor-mediated transcription of lipocalin-2. However, NF-kappa B-activating inflammatory molecules, such as IL-1 beta and lipopolysaccharide, did not induce lipocalin-2 mRNA in Sertoli-B cells and the expression of lipocalin-2 was unaffected in the testis of I kappa B zeta-deficient mice. These results demonstrate that spermatogonial cells regulate lipocalin-2 gene expression in Sertoli cells in a manner distinct from that employed by immune cells.
  • K Mominoki, M Morimatsu, M Kaijalainen, E Hohtola, R Hissa, M Saito
    COMPARATIVE BIOCHEMISTRY AND PHYSIOLOGY A-MOLECULAR & INTEGRATIVE PHYSIOLOGY 142 4 472 - 477 2005年12月 [査読有り][通常論文]
     
    Haptoglobin (Hp), a hemoglobin-binding protein, is known as an acute phase protein and increases during the acute phase of inflammation in most mammals. We reported previously in brown bears that the mean Hp concentrations were higher in blood samples obtained in winter than those in spring. To examine a possible relation of the seasonal variations of Hp to hibernation, in the present study, we measured the plasma concentrations of Hp as well as some other acute phase proteins (alpha(2)-macroglobulin, alpha(1)-antitrypsin, C-reactive protein) in 6 European brown bears (Ursus arctos), from which blood samples were obtained at 5-6 different months of year including February, the time of hibernation. The Hp concentrations showed clear seasonal variations, being highest in February. The alpha(2)-macroglobulin concentrations also showed a similar but much smaller rise in February, but those of alpha(1)-antitrypsin and C-reactive protein did not show any seasonal variations. Our results suggest that the seasonal variation of plasma Hp concentration in brown bears is associated with a hibernation-specific mechanism more than that of acute phase response. (C) 2005 Elsevier Inc. All rights reserved.
  • Y Yoshikawa, M Morimatsu, K Ochiai, M Nagano, Y Yamane, N Tomizawa, N Sasaki, K Hashizume
    JOURNAL OF VETERINARY MEDICAL SCIENCE 67 10 1013 - 1017 2005年10月 [査読有り][通常論文]
     
    Mammary tumors are the most common tumor type in women as well as in female dogs. The BRCA2 gene encodes a large nuclear protein that is involved in DNA repair, and mutations in the human BRCA2 confer an increased risk of female mammary tumors. The BRCA2 protein acts as a tumor suppressor, and inactivation of BRCA2 by loss of heterozygosity is implicated in mammary carcinogenesis. In this study, to establish an appropriate polymorphic marker for loss of heterozygosity analysis of the canine BRCA2, we analyzed the genomic sequences of the exon 27 regions of 30 mammary-tumor-bearing and 21 tumor-free dogs. In addition to 10204ins/delAAA, which is the only polymorphism previously identified for the canine BRCA2 locus, we discovered four novel single nucleotide polymorphisms. The analysis of these five polymorphisms revealed the presence of four allele types. Since 10204ins/delAAA was the most common of the five polymorphisms identified, we developed a PCR-based assay method to assay for this polymorphism. We believe that this method is valuable for loss of heterozygosity analysis of the canine BRCA2 gene in tumor pathogenesis.
  • Kumiko Takebe, Junko Nio, Masami Morimatsu, Shin-Ichiro Karaki, Atsukazu Kuwahara, Ikuo Kato, Toshihiko Iwanaga
    BIOMEDICAL RESEARCH-TOKYO 26 5 213 - 221 2005年10月 [査読有り][通常論文]
     
    Short-chain fatty acids in the intestinal lumen affect colonic cell proliferation as well as function as an energy source for intestinal epithelial cells. A novel transporter of monocarboxylates, Slc5a8, is expressed abundantly in the colon, where it may participate in the Na+-coupled absorption of short-chain fatty acids produced by bacterial fermentation of dietary fiber. The present study examined the cellular localization of Slc5a8 in the murine gastrointestinal tract and kidney by in situ hybridization and immunohistochemistry. The hybridization signals were recognized in the terminal ileum and whole length of the large intestine, and were especially intense in the distal colon and rectum. The immunoreactivity of Slc5a8 was restricted to the striated border (the brush border) of enterocytes, and was not present in goblet cells, Paneth cells, or lamina propria cells. In the kidney, proximal tubules of both the cortex and the outer stripe of the outer medulla intensely expressed Slc5a8 mRNA, while the distal portions, including the loop of Henle, lacked the signals. The renal Slc5a8 immunoreactivity was localized only in the brush border of proximal tubules, not along the basolateral membrane. Thyroid follicular cells were immunoreactive for Slc5a8, with predominant labeling on the apical membrane. No other organs, including the esophagus, stomach, liver, pancreas, and salivary glands contained any notable signals of Slc5a8. These findings on the cellular and subcellular localization of Slc5a8 under normal conditions are helpful for understanding the physiological and pathological roles of Slc5a8.
  • Y Yoshikawa, M Morimatsu, K Ochiai, M Nagano, Y Yamane, N Tomizawa, N Sasaki, K Hashizume
    BIOMEDICAL RESEARCH-TOKYO 26 3 109 - 116 2005年06月 [査読有り][通常論文]
     
    Mutations in human BRCA2 confer an increased risk of female breast cancer. In this study, we found a novel insertion/deletion polymorphism (10204insAAA causing amino acid change M33321K) in canine BRCA2, which is located in the putative second nuclear localization signal (NLS2) and C-ten-ninal Rad51-binding region. The nuclear localization of the insAAA C-terminus was more efficient than localization of the delAAA sequence when NLS1 was mutated. Strong, comparable Rad51 binding was observed for both the insAAA and delAAA C-termini. Dogs with the insertion/deletion polymorphism will provide a new model for studying the function of BRCA2.
  • T Shiina, A Konno, T Oonuma, H Kitamura, K Imaoka, N Takeda, K Todokoro, M Morimatsu
    JOURNAL OF BIOLOGICAL CHEMISTRY 279 53 55493 - 55498 2004年12月 [査読有り][通常論文]
     
    MAIL (molecule-possessing ankyrin repeats induced by lipopolysaccharide) is a nuclear IkappaB protein that is also termed interleukin-1-inducible nuclear ankyrin repeat protein or inhibitor of nuclear factor kappaB (IkappaB) zeta. In this study, we generated Mail(-/-) mice to investigate the roles of MAIL in whole organisms. Mail(-/-) mice grew normally until 4-8 weeks after birth, when they began to develop lesions in the skin of the periocular region, face, and neck. MAIL mRNA and protein were constitutively expressed in the skin of wild type controls, especially in the keratinocytes. Serum IgE was higher in Mail(-/-) mice than in normal. Histopathological analysis indicated that the Mail(-/-) skin lesions appeared to be atopic dermatitis ( AD) eczema with inflammatory cell infiltration. In addition, markedly elevated expression of some chemokines such as thymus and activation-regulated chemokine was detected in the Mail(-/-) skin lesions, similar to that observed in the skin of patients with AD. In Mail(-/-) mice, MAIL-deficient keratinocytes might be activated to produce chemokines and induce intraepidermal filtration of inflammatory cells, resulting in the onset of the AD-like disease. These findings suggest that MAIL is an essential molecule for homeostatic regulation of skin immunity. The Mail(-/-) mouse is a valuable new animal model for research on AD.
  • K Ochiai, M Morimatsu, Y Yoshikawa, B Syuto, K Hashizume
    BIOMEDICAL RESEARCH-TOKYO 25 6 269 - 275 2004年12月 [査読有り][通常論文]
     
    In humans and mice, the interaction between the breast cancer susceptibility protein, BRCA2, and RAD51 recombinase is essential for DNA repair by homologous recombination, the failure of this process can predispose to cancer. Cells with mutated BRCA2 are hypersensitive to ionizing radiation (IR) and exhibit defective DNA repair. Using yeast and mammalian two-hybrid assays, we demonstrate that canine Rad51 protein interacts specifically with the C-terminus of canine Brca2. In support of the biological significance of this interaction, we found that radiation-induced focus formation of Rad51 in COS-7 cells was compromised by forced expression of the C-terminus of canine Brca2. A similar result was obtained for the murine C-terminus. These data suggest that the C-terminal domain of canine Brca2 functions to bind Rad51 and that this domain contributes to the IR-induced assembly of the Rad51 complex in vivo.
  • T Ito, M Morimatsu, T Oonuma, T Shiina, H Kitamura, B Syuto
    GENE 342 1 137 - 143 2004年11月 [査読有り][通常論文]
     
    IkappaB inhibits nuclear factor kappa B (NF-kappaB), which is known to regulate the expression of various genes, including genes involved in inflammation. Recently, a novel IkappaB family protein, 'molecule possessing ankyrin repeats induced by lipopolysaccharide' (MAIL), was identified. MAIL is a nuclear-acting, inducible protein, unlike typical IkappaB proteins. However, the mechanism of its induction by lipopolysaccharide (LPS) is unclear. Using the LPS-reactive region located upstream from the MAIL gene, we investigated the mechanism of MAIL induction. MAIL expression was strongly regulated by NF-kappaB and partly regulated by CREB. Furthermore, deletion, point mutation and binding analyses revealed that the NF-kappaB binding site located at -229 to -220 bp is a n essential target of MAIL expression. Overexpression of MAIL protein suppressed the LPS-induced promoter activity of the MAIL gene. These data indicate that MAIL expression is strongly upregulated by NF-kappaB, and it is controlled, at least in part, by an autoregulation mechanism. (C) 2004 Elsevier B.V. All rights reserved.
  • D Yamaji, H Kitamura, K Kimura, Y Matsushita, H Okada, T Shiina, M Morimatsu, M Saito
    VETERINARY IMMUNOLOGY AND IMMUNOPATHOLOGY 98 3-4 175 - 184 2004年04月 [査読有り][通常論文]
     
    Molecule possessing ankyrin-repeats induced by lipopolysaccharide (MAIL) is known as an IkappaB protein induced after administration of bacterial lipopolysaccharide, (LPS) to mice. In the present study, we cloned bovine MAIL cDNA and examined its mRNA expression in white blood cells isolated from Holstein cows. Bovine MAIL had more than 80% amino acid identities with murine and human MAILs, highly conserved ankyrin-repeat motifs and PEST-like sequences. Bovine MAIL mRNA was undetectable in isolated peripheral white blood cells, but rapidly induced (< 1h) after stimulation by LPS and lipid A in vitro in a dose-dependent manner. The lipid A-induced MAIL mRNA expression was found in polymorphonuclear cells, monocytes/macrophages and total lymphocytes, but not in T-lymphocytes. MAIL mRNA was also induced in vivo in peripheral blood leukocytes of cows after intramammary injection of Escherichia coli derived from coliform, mastitis. Thus, bovine MAIL, as rodent MAILs, is induced by inflammatory stimuli in specific immune cells in vitro and in vivo, suggesting a role in inflammatory responses to bacterial infection in cattle. (C) 2003 Elsevier B.V. All rights reserved.
  • BH Lee, K Yoshimatsu, A Maeda, K Ochiai, M Morimatsu, K Araki, M Ogino, S Morikawa, J Arikawa
    VIRUS RESEARCH 98 1 83 - 91 2003年12月 [査読有り][通常論文]
     
    We performed yeast two-hybrid screening of a human kidney cell cDNA library to study the biological role of the hantavirus nucleocapsid protein (NP). We found that Seoul virus (SEOV) and Hantaan virus (HTNV) NPs were associated with small ubiquitin-like modifier (SUMO)-1-interacting proteins PIAS 1, PIASxbeta, HIPK2, CHD3, and TTRAP, which interacted with the SUMO-1 conjugating enzyme (Ubc-9) and SUMO-1 in the yeast two-hybrid assay. Interactions between the HIPK2, CHD3, and TTRAP proteins and SEOV NP were also shown in a mammalian two-hybrid assay. However, there was no interaction between PIAS proteins and NP, which was probably due to the inhibitory effect of PIAS on transcription in the mammalian two-hybrid assay. Nevertheless, a co-expression experiment suggested the existence of a PIAS-NP interaction in the cytoplasm. The region spanning amino acids 100-125 of SEOV NP, which represents a critical region for NP-NP polymerization. was found to be responsible for the interaction with SUMO-1-related molecules in both the yeast and mammalian two-hybrid assays. These results add to the information on interactions of hantavirus NP and host cellular proteins. (C) 2003 Elsevier B.V. All rights reserved.
  • T Oonuma, M Morimatsu, K Ochiai, B Syuto
    JOURNAL OF VETERINARY MEDICAL SCIENCE 65 10 1123 - 1126 2003年10月 [査読有り][通常論文]
     
    Mammary tumors are common in cats. As mutations in human Brca2 confer an increased risk of breast cancer, the full-length cDNA of the feline homologue of Brca2 was sequenced to obtain a basis for studying the relationship between its function and susceptibility to mammary tumors. The feline Brca2 cDNA is 10 kb long, and encodes 3,371 amino acids. The amino acid sequence of feline Brca2 shares low homology with the Brca2 of other mammals, e.g., 53% homology with the murine protein. Analysis of the expression pattern of the feline Brca2 gene revealed that, as previously reported for other mammals, it is transcribed in various tissues, including the mammary gland.
  • T Oonuma, M Morimatsu, T Nakagawa, R Uyama, N Sasaki, M Nakaichi, H Tamamura, N Fuji, S Hashimoto, H Yamamura, B Syuto
    JOURNAL OF VETERINARY MEDICAL SCIENCE 65 10 1069 - 1073 2003年10月 [査読有り][通常論文]
     
    It has recently been suggested that the chemokine receptor CXCR4 and its ligand SDF-1 (CXCL12) promote metastasis of various cancers in humans. Since feline mammary tumors also metastasize to distant organs frequently, we used real-time quantitative PCR to examine the expression of feline CXCR4 (fCXCR4) in ten feline mammary tumor cell lines and seven feline mammary tumor tissues, and also the expression of feline SDF-1 (fSDF-1) in various organs. Cell lines derived from metastatic regions expressed more fCXCR4 than those derived from primary tumors. Mammary tumor tissues overexpressed more fCXCR4 than normal mammary tissues. Organs with high levels of fSDF-1 expression represent common sites of metastasis. Migration assays using the feline mammary tumor cell line NAC were also performed to test the activity of TN14003 and TC14012, antagonists of human CXCR4, to antagonize fCXCR4 expressed on NAC cells. TN14003 and TC14012 inhibited migration of NAC cells. We conclude that fCXCR4 may be a therapeutic target for feline mammary tumors.
  • H Kitamura, Y Matsushita, T Iwanaga, K Mori, K Kanehira, D Fujikura, M Morimatsu, M Saito
    ARCHIVES OF HISTOLOGY AND CYTOLOGY 66 1 53 - 62 2003年03月 [査読有り][通常論文]
     
    Molecule possessing ankyrin-repeats induced by lipopolysaccharide (MAIL), a recently cloned nuclear IkappaB protein induced by lipopolysaccharide (LPS) stimulation in lymphoid organs, is involved in the regulation of inflammatory responses. The present in situ hybridization and immunohistochemical analyses revealed the distinct expression of the MAIL mRNA and protein in B-lymphocytes of the white pulp of the spleen and cortical lymphoid follicles of lymph nodes in LPS-injected mice. MAIL signals were also localized in F4/80-positive macrophages in these organs. LPS clearly induced MAIL expression in cultured B-lymphocytes and monocytes/macrophages, but only faintly so in T-lymphocytes, fibroblasts, and endothelial cells. MAIL was also induced by inflammatory cytokines such as interleukin-1 and -6, and tumor necrosis factor in cultured cells. Northern blot, Western blot, and in situ hybridization analyses showed that the major expression product of the Mail gene was a long splicing variant (MAIL-L) rather than a short one, both in lymphoid organs and cultured cells. These results collectively indicate that LPS induces MAIL-L predominantly in B-lymphocytes and macrophages.
  • K Yoshimatsu, BH Lee, K Araki, M Morimatsu, M Ogino, H Ebihara, J Arikawa
    JOURNAL OF VIROLOGY 77 2 943 - 952 2003年01月 [査読有り][通常論文]
     
    Multimerization of the Hantaan virus nucleocapsid protein (NP) in Hantaan virus-infected Vero E6 cells was observed in a competitive enzyme-linked immunosorbent assay (ELISA). Recombinant and truncated NPs of Hantaan, Seoul, and Dobrava viruses lacking the N-terminal 49 amino acids were also detected as multimers. Although truncated NPs of Hantaan virus lacking the N-terminal 154 amino acids existed as a monomer, those of Seoul and Dobrava formed multimers. The multimerized truncated NP antigens of Seoul and Dobrava viruses could detect serotype-specific antibodies, whereas the monomeric truncated NP antigen of Hantaan virus lacking the N-terminal 154 amino acids could not, suggesting that a hantavirus serotype-specific epitope on the NP results in multimerization. The NP-NP interaction was also detected by using a yeast two-hybrid assay. Two regions, amino acids 100 to 125 (region 1) and amino acids 404 to 429 (region 2), were essential for the NP-NP interaction in yeast. The NP of Seoul virus in which the tryptophan at amino acid number 119 was replaced by alanine (W119A mutation) did not multimerize in the yeast two-hybrid assay, indicating that tryptophan 119 in region 1 is important for the NP-NP interaction in yeast. However, W119A mutants expressed in mammalian cells were detected as the multimer by using competitive ELISA. Similarly, the truncated NP of Seoul virus expressing amino acids 155 to 429 showed a homologous interaction in a competitive ELISA but not in the yeast two-hybrid assay, indicating that the C-terminal region is important for the multimerization detected by competitive ELISA. Combined, the results indicate that several steps and regions are involved in multimerization of hantavirus NP.
  • M Suzuki, W Fujimoto, M Goto, M Morimatsu, B Syuto, T Iwanaga
    JOURNAL OF HISTOCHEMISTRY & CYTOCHEMISTRY 50 8 1081 - 1089 2002年08月 [査読有り][通常論文]
     
    Recently, the second mammalian chitinase, designated acidic mammalian chitinase (AMCase), has been identified in human, mouse, and cow. In contrast to the earlier identified macrophage-derived chitinase (chitotriosidase), this chitinase is richly expressed in the gastrointestinal (GI) tract, suggesting its role in digestion of chitin-containing foods as well as defense against chitin-coated microorganisms and parasites. This in situ hybridization study first revealed cellular localization of the gut-type chitinase in the mouse and chicken. In adult mice, the parotid gland, von Ebner's gland, and gastric chief cells, all of which are exocrine cells of the serous type, expressed the gut chitinase mRNA. In the chicken, oxyntico-peptic cells in glandular stomach (proventriculus) and hepatocytes expressed the chitinase mRNA. Because cattle produce the gut chitinase (chitin-binding protein b04) only in the liver, the gut chitinases in mammals and birds have three major sources of production, i.e., the salivary gland, stomach, and liver. During ontogenetic development, the expression level in the parotid gland and stomach of mice increased to the adult level before weaning, whereas in the stomach of chickens intense signals were detectable in embryos from incubation day 7.
  • R Kamata, M Morimatsu, T Suzuki, T Takewaki, H Kobayashi
    ENVIRONMENTAL TOXICOLOGY AND PHARMACOLOGY 12 1 55 - 58 2002年08月 [査読有り][通常論文]
     
    Gel filtration chromatography was performed on cytosol preparation of hen spinal cord to find molecular target(s) for organophosphorus-induced delayed neurotoxicity (OPIDN). Three binding peaks of [H-3]diisopropyl phosphorofluoridate (DFP), an organophosphate that induces OPIDN, were separated from the cytosol preparation. The activities of acetylcholinesterase (AChE) and neuropathy target esterase (NTE) that has been proposed as a screening method for OPIDN eluted in the fractions within these two DFP binding peaks. However, the other peak had none of the activities of AChE and NTE. Therefore, this DFP binding proteins in cytosol may be peculiar to the pathogenesis of OPIDN. (C) 2002 Elsevier Science B.V. All rights reserved.
  • M Suzuki, M Morimatsu, B Syuto
    JOURNAL OF VETERINARY MEDICAL SCIENCE 64 6 477 - 481 2002年06月 [査読有り][通常論文]
     
    Bovine serum contains N-acetyl-D-glucosamine (GlcNAc)-sensitive opsonin inhibitory factors. In the present study, a major component of chitin-binding protein (chitin-binding protein b01, CBPb01) was purified from bovine serum, and identified CBPb01 as bovine IgM by its subunit structure, antigenic properties, and partial sequences. The results of a lectin-binding assay showed that the heavy chain of CBPb01 had a GlcNAc structure, but the commercial IgM did not. It is possible that CBPb01 interconnects through its GlcNAc structure, subsequently forming complexes. We also demonstrated that CBPb01 had opsonin-inhibitory activity, and that this activity was dependent on the binding of CBPb01 to GlcNAc on the zymosan surface. These findings indicate the presence of a kind of IgM that recognizes GlcNAc structure in the regulation of opsonization.
  • H Kitamura, K Kanehira, T Shiina, M Morimatsu, BD Jung, S Akashi, M Saito
    JOURNAL OF VETERINARY MEDICAL SCIENCE 64 5 419 - 422 2002年05月 [査読有り][通常論文]
     
    Molecule possessing ankyrin-repeats induced by lipopolysaccharide (MAIL) is a nuclear IkappaB protein recently identified as a molecule appearing in immunocompetent organs after administration of bacterial lipopolysaccharide (LPS). Participation of Toll-like receptor (TLR) 4, which is a major form of LPS receptors, in the LPS-induced MAIL expression was investigated. When a human myelomonocytic cell line U937 was treated with phorbol 12-myristate 13-acetate for 3 days, the LPS-induced MAIL expression was much potentiated in parallel with an increase in TLR4 expression. The MAIL induction was attenuated when the cells were treated with a neutralizing antibody against TLR4. The in vivo induction of MAIL in the spleen was smaller in mice having a missense mutation of the Tlr4 gene than in normal control mice. These results collectively indicate that TLR4 contributes, at least in part, MAIL induction after LPS stimulation.
  • M Suzuki, M Morimatsu, T Yamashita, T Iwanaga, B Syuto
    FEBS LETTERS 506 2 127 - 130 2001年10月 [査読有り][通常論文]
     
    Chitinases are ubiquitous chitin-fragmenting hydrolases. They are synthesized by a vast array of organisms, including those not composed of chitin. Here, we describe a novel serum chitinase (chitin-binding protein b04, CBPb04), which is expressed in bovine liver. Although CBPb04 is secreted as an endocrine chitinase, it shows higher homology with human gastrointestinal tract exocrine chitinase (AMCase) than with macrophage endocrine chitinase (human chitotriosidase). This suggests that cows have a specific defense against chitin-containing microorganisms. CBPb04 mRNA is expressed in hepatocytes. This is the first report of a hepatogenic mammalian chitinase. (C) 2001 Federation of European Biochemical Societies. Published by Elsevier Science B.V. All rights reserved.
  • T Shiina, M Morimatsu, H Kitamura, T Ito, S Kidou, K Matsubara, Y Matsuda, M Saito, B Syuto
    IMMUNOGENETICS 53 8 649 - 655 2001年10月 [査読有り][通常論文]
     
    The Mail (molecule possessing ankyrin repeats induced by lipopolysaccharide) protein is a member of the I kappaB family. It has six ankyrin repeats that are conserved in other I kappaB proteins, such as I kappaB-alpha and Bcl-3. Mail mRNA expression is induced rapidly following lipopolysaccharide (LPS) injection, most notably in the spleen, lung, and lymph nodes of mice, where immune cells, such as lymphocytes and macrophages, are abundant. In this study, we cloned and characterized the Mail gene. The isolated genomic clones span approximately 30 kb and encompass the entire gene. Comparisons with Mail cDNA revealed that the Mail gene consists of 14 exons. Several splice junctions encoding ankyrin repeats are conserved among Mail and other I kappaB family genes. Southern hybridization showed that Mail is a single-copy gene. Using fluorescence in situ hybridization analysis, mouse and rat Mail genes were mapped to Chromosome (Chr) 16C 1.2-C1.3 and Chr 11q21.1, respectively. Primer extension determined the transcription start site of Mail. Sequence analysis of the proximal promoter region revealed the presence of a TATA box and putative transcription factor-binding sites, such as those for NF-kappaB and NF-IL6. This region is sufficient to drive high-level reporter gene expression in LPS-stimulated transfected cells.
  • K Ochiai, M Morimatsu, N Tomizawa, B Syuto
    JOURNAL OF VETERINARY MEDICAL SCIENCE 63 10 1103 - 1108 2001年10月 [査読有り][通常論文]
     
    Mammary tumors are the most common neoplasm in female dogs. Canis canis, and in women. Mutations in human Brca2 confer an increased risk of female breast cancer. Previous studies have shown that the Brca2 tumor suppressor protein interacts with the recombinational repair protein Rad51. We cloned the full-length cDNA of the canine homologues of Brca2 and Rad51 to obtain a basis for studying their relationship with susceptibility to mammary tumors. The canine Brca2 and Rad51 cDNAs are 11 and 1.5 kb long, encoding 3,471 and 339 amino acids, respectively. The amino acid sequence of canine Brca2 showed 68% homology with the human protein., and 58% homology with a murine protein. There were highly conserved regions in the C-terminus of all three proteins, where the Rad51 interacting domain and putative nuclear localization signals are located. Comparing with the partial genomic sequence previously reported, we found possible nuclear polymorphisms in exon 11, some of which result in amino acid substitutions. On the other hand, canine Rad51 protein had extremely high homology (99%) to the human and murine proteins. Expression of both Brca2 and Rad51 was detected in the mammary gland, suggesting that these two genes interact in the canine mammary gland.
  • S Vestri, MM Okamoto, HS de Freitas, RA dos Santos, MT Nunes, M Morimatsu, JC Heimann, UF Machado
    JOURNAL OF MEMBRANE BIOLOGY 182 2 105 - 112 2001年07月 [査読有り][通常論文]
     
    Renal glucose reabsorption is mediated by luminal sodium-glucose cotransporters (SGLTs) and basolateral facilitative glucose transporters (GLUTs). The modulators of these transporters are not known, and their substrates glucose and Na+ are potential candidates. In this study we examined the role of glucose and Na+ filtration rate on gene expression of glucose transporters in renal proximal tubule. SGLT1, SGLT2, GLUT1 and GLUT2 mRNAs were assessed by Northern blotting; and GLUT1 and GLUT2 proteins were assessed by Western blotting. Renal cortex and medulla samples from control rats (C), diabetic rats (D) with glycosuria, and insulin-resistant 15-month old rats (I) without glycosuria; and from normal (NS), low (LS), and high (HS) Na+-diet fed rats were studied. Compared to C and I rats, D rats increased (P < 0.05) gene expression of SGLT2 by similar to 36%, SGLT1 by similar to 20%, and GLUT2 by similar to 100%, and reduced (P < 0.05) gene expression of GLUT1 by more than 50%. Compared to NS rats, HS rats increased (P < 0.05) SCLT2, GLUT2, and GLUT1 expression by <similar to>100%. with no change in SGLT1 mRNA expression, and LS rats increased (P < 0.05) GLUT1 gene expression by <similar to>150% with no changes in other transporters. In summary, the results showed that changes in glucose or Na+ filtrated rate modulate the glucose transporters gene expression in epithelial cells of the renal proximal tubule.
  • K Iwasaki, M Morimatsu, O Inanami, E Uchida, B Syuto, M Kuwabara, M Niiyama
    JOURNAL OF BIOLOGICAL CHEMISTRY 276 12 9400 - 9405 2001年03月 [査読有り][通常論文]
     
    Acute-phase serum proteins were induced by administrating a chicken with turpentine oil. One of these proteins was a new protein that appeared in front of albumin in polyacrylamide disc gel electrophoresis using a 4.5-16% gel. To purify this protein, turpentine-administrated chicken serum was fractionated by ammonium sulfate precipitation at 50% saturation, and the supernatant fraction was chromatographed on a DEAE-Toyopearl 650S column. The purified protein is a mannose-glycoprotein, and its N-terminal sequence, determined by the Edoman method, is not homologous from that of other reported acute-phase proteins. An analysis of physiological function with two different test systems, chemiluminescence measurement and electron spin resonance spectroscopy, showed that the purified protein has antioxidant, activity and inhibits superoxide (O-2(radical anion)) mediated by activation of the receptor. In support of these results, the complete amino acid sequence of 18-B is homologous to the scavenger receptor cysteine-rich (SRCR) family of proteins that participate in the regulation of leukocyte function. 18-B is composed of four SRCR domains, which is different from the previously characterized SRCR family of proteins such as Sp alpha, CD6, and CD163. These findings indicate that turpentine-induced 18-B, a new member of scavenger receptor cysteine-rich family, may be implicated in regulation of cell function in a manner of inhibition of the overproduction of the reactive oxygen species.
  • H Kitamura, K Kanehira, K Okita, M Morimatsu, M Saito
    FEBS LETTERS 485 1 53 - 56 2000年11月 [査読有り][通常論文]
     
    We have identified and characterized a novel member of the ankyrin-repeat family named 'molecule possessing ankyrin-repeats induced by lipopolysaccharide' (MAIL). The C-terminal portion of MAIL shared high sequence homology,vith the I kappaB family. Intraperitoneal injection of lipopolysaccharide (LPS) into mice rapidly (< 0.5 h) induced MAIL mRNA in various tissues, particularly in the spleen, lymph node, and lung. Ectopically expressed MAIL was localized in the nucleus, and remarkably potentiated the LPS-induced mRNA expression and secretion of interleukin (IL)-6 in Swiss 3T3 cells, These findings indicate that MAIL is one of the nuclear I<kappa>B proteins and an activator of IL-6 production. (C) 2000 Federation of European Biochemical Societies. Published by Elsevier Science B.V. All rights reserved.
  • Y Watanabe, M Morimatsu, B Syuto
    JOURNAL OF VETERINARY MEDICAL SCIENCE 62 5 473 - 478 2000年05月 [査読有り][通常論文]
     
    Botulinum C3 enzyme produced by Clostridium botulinum type C and D strains modifies Rho proteins. In a previous study, we observed that the LDH isozyme pattern of neurons treated with C3 enzyme was different from that induced with endogenous growth factor of neurons such as NGF [21]. This type of change is considered to have an advantage in the medical use of C3 enzyme for neural disorder. To determine the functional similarity of C3-treated neurons to control and NGF-treated neurons, we examined the responses of C3-treated neurons to various drugs, including some neurotransmitters, by measuring the rise of intracellular Ca ions into the neurons. The time course of the rise of intracellular Ca ions induced by high concentration of potassium in the C3-treated neurons was similar to that in the NGF-treated neurons. The C3-treated neurons responded to glutamic acid, aspartic acid, kainic acid, gamma-aminobutylic acid, muscarine and ACh with similar time courses and magnitudes as the control neurons. These results suggest that the C3 enzyme induces the functional differentiation of neurons, and that C3 enzyme has the potential for the medical use as an exogenous differentiation-inducing factor of neurons.
  • M Iwase, K Kimura, N Sasaki, R Komagome, K Ishioka, M Morimatsu, T Murakami, M Saito
    RESEARCH IN VETERINARY SCIENCE 68 2 109 - 114 2000年04月 [査読有り][通常論文]
     
    Leptin, the product of the ob gene, is one of the key molecules for the regulation of appetite and whole-body energy balance, and thereby for the pathogenesis of obesity. In an attempt to clarify the roles of leptin in obesity and/or related diseases in companion animals, canine leptin CDNA was cloned by amplifying reverse-transcriptase products of RNA extracted from the adipose tissue of the beagle. A CDNA clone of about 3 kbp contained a 501 bp open reading frame coding a 167-amino acid protein with a 21-amino acid signal peptide. The sequence of a 146-amino acid mature leptin was more than 79 per cent identical to those of other mammals. Northern blot analysis revealed abundant expression of leptin mRNA in adipose tissue, but not in other tissues, in adult beagles. When Chinese hamster ovary cells expressing the rat leptin receptor were stimulated with recombinant canine leptin produced by E. coli, some intracellular signal transduction proteins were phosphorylated, indicating that the recombinant leptin was biologically active. The data reported herein will be helpful for further studies of leptin of the dog in health and disease. (C) Harcourt Publishers Ltd.
  • Y Watanabe, M Morimatsu, B Syuto
    JOURNAL OF VETERINARY MEDICAL SCIENCE 62 3 249 - 254 2000年03月 [査読有り][通常論文]
     
    Changes in the lactate dehydrogenase (LDH) isozyme pattern of primary culture of neurons treated with botulinum C3 enzyme were examined in order to elucidate the functional changes accompanying the morphological change that follows ADP-ribosylation of Rho protein. Primary neurons were prepared from the cerebrum of ICR mouse embryos on day 15. Neurons were cultured in MEM with 10% fetal calf serum at 37 degrees C. In the neurons treated with C3 enzyme, a typical morphological change was observed after 24 hr, and the LDH isozyme pattern was changed after 72 hr. The ratio of H-subunit to M-subunit in LDH was decreased by C3 treatment, suggesting the induction of a state of lower intracellular oxygen consumption in neurons in the primary cultures.
  • K Kimura, N Sasaki, A Asano, J Mizukami, S Kayahashi, T Kawada, T Fushiki, M Morimatsu, T Yoshida, M Saito
    HORMONE AND METABOLIC RESEARCH 32 3 91 - 96 2000年03月 [査読有り][通常論文]
     
    Wild-type or mutated human beta 3-adrenergic receptor (Trp64Arg) cDNAs were stably expressed in mouse 3T3-L1 cells. Saturation binding study using a beta-adrenergic ligand revealed that there was no significant difference in the receptor density and the equilibrium dissociation constant between the two cell lines. However, the ability of the mutant beta 3-adrenergic receptor to accumulate cyclic AMP (cAMP) in response to isoproterenol was much reduced and Kact for cAMP accumulation was lowered as compared to the wild type receptor. The amount of alpha subunit of stimulatory GTP-binding protein (GS alpha) and adenylyl cyclase activity in response to forskolin were not different in the two cell lines. The responses of the mutant receptor to epinephrine, norepinephrine and L-755,507, a highly specific agonist for human beta 3-adrenergic receptor, were also reduced, but the reduction of Kact for L-755,507 was more evident than other agonists tested. The cAMP accumulation in response to some conventional beta 3 agonists was less than 10% of that to isoproterenol even in the cells expressing the wild type receptor. These results suggest that the Trp64Arg mutant beta 3-adrenergic receptor has less ability to stimulate adenylyl cyclase, and that lipolytic activity through the beta 3-adrenergic receptor by catecholamines in subjects carrying this mutation might be suppressed.
  • M Morimatsu, G Donoho, P Hasty
    CANCER RESEARCH 58 15 3441 - 3447 1998年08月 [査読有り][通常論文]
     
    The putative Brca2-MmRad51 interaction is analyzed in mouse cells deleted for the COOH terminus of Brca2 (amino acids 3140-3328), which contains a region that associates with MmRad51 by yeast two-hybrid. These cells are hypersensitive to gamma-radiation (suggesting defective recombinational repair) but not UV light (suggesting intact nucleotide excision repair) and maintain the G(1)-S and G(2)-M checkpoints after exposure to gamma-irradiation. Cells deleted for the COOH terminus of Brca2 progress through the cell cycle at a similar rate as wild-type cells but undergo senescence more rapidly. These data support the hypothesis that deletion of Brca2 stimulates cancer by defective MmRad51-mediated DNA repair and not by defective cell cycle regulation.
  • H Abe, Y Kawakita, T Miyashige, M Morimatsu, M Saito
    JOURNAL OF VETERINARY MEDICAL SCIENCE 60 6 769 - 771 1998年06月 [査読有り][通常論文]
     
    The nucleotide sequence of cDNA coding the 38-amino acid of the C-terminal domain of insulin-responsive glucose transporter (GLUT4) was determined by the method of reverse transcription-polymerase chain reaction in the sheep, goat and pig, and compared with that of bovine which have been shown to have a unique amino acid conversion of Asn508 to His. The deduced amino acid sequence was completely identical in the three species, and did not have the amino acid conversion at position 508. Western blot analysis confirmed that an antiserum raised against a rat C-terminal peptide cross-reacted efficiently with GLUT4 of these livestock mammals.
  • H Kitamura, S Okamoto, Y Shimamoto, M Morimatsu, A Terao, M Saito
    CELLULAR AND MOLECULAR LIFE SCIENCES 54 3 282 - 287 1998年03月 [査読有り][通常論文]
     
    Centrally given interleukin (IL)-1 is known to induce a rapid rises in blood IL-6. To extend this and to examine the mechanism by which this occurs, the effects of intracerebroventricular (icv) injection of human recombinant IL-1 beta on mRNA expression of IL-6 and tumour necrosis factor (TNF) in the spleen and liver were examined in rats. Icy injection of IL-1 produced a rapid rise of the tissue mRNA levels of IL-6 and TNF in both organs, prior to and/or in parallel with an increase in their serum levels. Pretreatment with chlorisondamine, a ganglionic blocking agent, inhibited the IL-6 responses, while it had little influence on the TNF responses. The results suggest that brain IL-1 induces peripheral production of IL-6, but not of TNF, through autonomic nervous system activation.
  • Kitamura H, Okamoto S, Shimamoto Y, Terao A, Morimatsu M, Saito M
    Cell Mol Life Sci 54 3 282 - 287 1998年03月 [査読有り][通常論文]
  • A Asano, M Morimatsu, H Nikami, T Yoshida, M Saito
    BIOCHEMICAL JOURNAL 328 179 - 183 1997年11月 [査読有り][通常論文]
     
    Cold exposure produces adaptive hyperplasia and growth of brown adipose tissue (BAT), the major site of non-shivering thermogenesis in rodents, associated with increased angiogenesis in this tissue. Vascular endothelial growth factor (VEGF), one of the most potent angiogenic factors, was found to be expressed abundantly in BAT of the rat. When rats were exposed to cold at 4 degrees C, the VEGF mRNA level in BAT was increased by 2-3-fold in 1-4 h, but returned to the basal level within 24 h. VEGF expression in other tissues such as heart, kidney and lung did not change after cold exposure. The cold-induced increase in VEGF mRNA was abolished by surgical sympathetic denervation, but mimicked by administration of noradrenaline or a beta(3)-adrenoceptor agonist CL316,243, indicating the critical role of the beta-adrenergic pathway in VEGF expression in BAT. Among three isoforms of VEGF, the mRNA of a short form (VEGF120) lacking heparin-binding activity was preferentially increased after cold exposure and treatment with the adrenergic agonists. These results suggest that cold exposure activates the sympathetic nerves and leads to a rapid increase in synthesis of VEGF in BAT, which in turn stimulates the proliferation of surrounding vascular endothelial cells.
  • H Kitamura, A Konno, M Morimatsu, BD Jung, K Kimura, M Saito
    BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS 238 3 707 - 711 1997年09月 [査読有り][通常論文]
     
    When mice were subjected to restriction of movement in a small cylinder (immobilization stress), the serum interleukin (IL)-6 level rose in 1 h, following increased expression of IL-6 mRNA in both the liver and the spleen. The IL-6 mRNA induction was much greater in the liver than in the spleen when compared on a whole-organ basis. Intraperitoneal injection of bacterial lipopolysaccharide (LPS) also increased IL-6 mRNA expression in these organs, but more preferentially in the spleen. Immunohistochemical examinations of liver tissue using an antibody against murine IL-6 revealed that immobilization stress induced IL-6 mainly in hepatic parenchymal cells, whereas LPS injection did so only in sinusoidal mononuclear cells. These results indicate that immobilization stress induces IL-6 production in the liver, especially in hepatic parenchymal cells, probably by a different mechanism from that for IL-6 induction by LPS. (C) 1997 Academic Press.
  • SK Sharan, M Morimatsu, U Albrecht, DS Lim, E Regel, C Dinh, A Sands, G Eichele, P Hasty, A Bradley
    NATURE 386 6627 804 - 810 1997年04月 [査読有り][通常論文]
     
    Inherited mutations in the human BRCA2 gene cause about half of the cases of early-onset breast cancer. The embryonic expression pattern of the mouse Brca2 gene is now defined and an interaction identified of the Brca2 protein with the DNA-repair protein Rad51. Developmental arrest in Brca2-deficient embryos, their radiation sensitivity, and the association of Brca2 with Rad51 indicate that Brca2 may be an essential cofactor in the Rad51-dependent DNA repair of double-strand breaks, thereby explaining the tumour-suppressor function of Brca2.
  • H Abe, M Morimatsu, H Nikami, T Miyashige, M Saito
    JOURNAL OF ANIMAL SCIENCE 75 1 182 - 188 1997年01月 [査読有り][通常論文]
     
    Insulin-responsive glucose transporter GLUT4, is a member of the glucose transporter family (GLUT) and is present exclusively in muscle and adipose tissue. It is a target of insulin action in humans and rodents. To clarify the molecular structure of bovine GLUT4, its GLUT4 cDNA was cloned by the RT-PCR method. Several cDNA clones corresponding to the different regions of GLUT4 were obtained by amplifying reverse-transcriptase products of RNA extracted from Holstein cattle skeletal muscle. Nucleotide sequence analysis of the cDNA clones revealed that bovine GLUT4 cDNA was composed of 2,656 base pairs with a coding region for a 509 amino acid protein. The deduced amino acid sequence was 64% and 92% identical with bovine GLUT1 (GLUT ubiquitously expressed in all tissues) and rat GLUT4, respectively. Although the amino acid sequence of the GLUT4 COOH-terminal region is highly conserved among the species so far reported, one amino acid (Asp) of this region was replaced by His in bovine GLUT4. The tissue distribution of GLUT4, was also examined by Northern blot analysis using a probe prepared from the bovine cDNA. GLUT4 mRNA was detected in skeletal muscle, heart, and adipose tissue, but not in liver, kidney, lung, brain, or spleen. Such a distribution is essentially the same as in humans and rodents, suggesting that GLUT4 is an insulin-responsive glucose transporter in cattle.
  • K Mominoki, H Tsuruga, M Morimatsu, M Saito
    COMPARATIVE BIOCHEMISTRY AND PHYSIOLOGY A-PHYSIOLOGY 114 4 349 - 353 1996年08月 [査読有り][通常論文]
     
    Haptoglobin (Hp), a hemoglobin-binding protein, is known as an acute phase protein increasing in blood during inflammation in most mammals. On the basis of our previous studies on purification and characterization of bear Hp (Comp. Biochem. Physiol. 110B, 785-789, 1995), in this study, we developed an immunoassay method to measure serum Hp level in bear, and measured the concentration of Hp in blood samples collected from 84 reared and 25 wild brown bears in Hokkaido, Japan. The mean serum Hp concentration was 0.94 +/- 0.25 mg/ml in wild bears, which is nearly equal to those reported in other species. In reared bears, the Hp concentration was apparently higher (3.82 +/- 0.29 mg/ml), although total protein and albumin concentrations were nearly equal in the two groups. A significant seasonal variation of serum Hp, low in spring and high in autumn and winter, was found in reared bears. Possible factors participating in the seasonal variation were discussed with special references to hibernation.
  • M Kobayashi, H Nikami, M Morimatsu, M Saito
    NEUROSCIENCE LETTERS 213 2 103 - 106 1996年08月 [査読有り][通常論文]
     
    The localization of glucose transporters (GLUTs) was examined in various regions of the rat brain. The mRNA of GLUT1 and GLUT3 were found ubiquitously in every brain region (cortex, hippocampus, midbrain, striatum, hypothalamus, medulla oblongata and cerebellum). The mRNA and protein of GLUT4, an insulin-regulatable glucose transporter in peripheral tissues, were also identified, particularly abundantly in the cerebellum. In situ hybridization analysis revealed that GLUT4 mRNA was present in some discrete cells, such as Purkinje cells in the cerebellum, the vestibular nucleus in the medulla oblongata and also in ependymal cells along the cerebral ventricles. The GLUT4 mRNA level in the cerebellum changed little in fasted or experimentally induced diabetic rats while those in adipose tissues decreased much. The results suggest that insulin-sensitive glucose uptake may occur in some specific cells of the brain but is regulated in a different manner from those in peripheral cells.
  • H KITAMURA, Y SHIMAMOTO, M MORIMATSU, A TERAO, M SAITO
    BIOMEDICAL RESEARCH-TOKYO 16 5 353 - 356 1995年10月 [査読有り][通常論文]
     
    To examine effects of brain cytokines on hepatic acute phase protein synthesis, the plasma level of haptoglobin (Hp) was measured after intracerebroventricular (icv) administration of human recombinant interleukin (Il)-1 beta, IL-6 and tumor necrosis factor (TNF)-alpha in rats. Icv injection of IL-1 or TNF (100 ng/rat, 3 times at every 3 h) produced a significant increase in plasma Hp level 12-24 h after the first injection. Neither intraperitoneal injection of these cytokines at the same doses nor icv injection of IL-6 affected the plasma Hp level. These results indicate that brain IL-1 and TNF, but not IL-6, can induce the acute phase response of some plasma proteins.
  • N NAKAGAWATOSA, M MORIMATSU, M KAWASAKI, H NAKATSUJI, B SYUTO, M SAITO
    JOURNAL OF VETERINARY MEDICAL SCIENCE 57 2 219 - 223 1995年04月 [査読有り][通常論文]
     
    The hepatic synthesis of acute phase proteins in ruminants has been suggested to be regulated by some mechanisms different from those in other species such as rodents and human. To explore possible regulatory factors unique to ruminants, we examined effects of interleukin (IL)-6, IL-1 and tumor necrosis factor (TNF), on haptoglobin (Hp) synthesis using a primary culture system of bovine hepatocytes. After bovine primary cultured hepatocytes were incubated in the presence of various concentrations of the cytokines, the synthesis and mRNA level of haptoglobin and albumin were measured by labeling with [S-35]-methionine and immunoprecipitation, and by Northern blot analysis, respectively. Hp synthesis was dose-dependently increased by recombinant human (rh) IL-6, and also by rhTNF-alpha, but to a less extent, while it was not affected by rhlL-1 beta. The stimulatory effect is mainly pretranslational, because mRNA level of Hp changed in parallel with protein synthesis. In contrast, albumin synthesis was suppressed by these three cytokines similarly. These results are inconsistent with the previously proposed view that TNF and IL-l overlap in their pathways leading to the transcriptional activation of many acute phase protein genes. In conclusion, there is a species-specific unique signaling system, especially for TNF, in transcriptional activation of bovine Hp gene.
  • K MOMINOKI, N NAKAGAWATOSA, M MORIMATSU, B SYUTO, M SAITO
    COMPARATIVE BIOCHEMISTRY AND PHYSIOLOGY B-BIOCHEMISTRY & MOLECULAR BIOLOGY 110 4 785 - 789 1995年04月 [査読有り][通常論文]
     
    Haptoglobin (Hp), a hemoglobin-binding protein in plasma, consists of alpha and beta subunits and has a tetra-chain arrangement (beta-alpha-alpha-beta) connected by disulfide bridges in most mammals so far examined, Dog Hp has been reported to be unique compared with other Hps in respect that (1) the two alpha beta units are joined by a non-covalent interaction rather than a disulfide bridge and (2) the alpha chain has an oligosaccharide-binding sequence (Asn-X-Ser/Thr) and is glycosylated, To determine whether the unique structures of dog Hp are common in the Carnivora, we purified Hps from sera of bear and cat, and analyzed their subunit structure and partial amino acid sequences, The analyses by gel filtration and sodium dodecyl sulfate-polyacrylamide gel electrophoresis, under both reducing and non-reducing conditions, revealed that bear and cat Hps have similar subunit arrangements to dog Hp, suggesting the absence of a disulfide bridge between two a chains, This was confirmed by amino acid sequence analysis of the alpha chains: that is, Cys(15) participating in the inter-a chain disulfide bridge was replaced by Val in bear or Leu in cat and dog, Thus, the unique subunit arrangement of Hp reported in dog may be common in the Carnivora, In contrast to dog Hp, however, alpha chains of bear and cat Hps were found not to have the typical oligosaccharide binding sequence on their alpha chains and were not glycosylated.
  • NAGASE, I, N SASAKI, K TSUKAZAKI, T YOSHIDA, M MORIMATSU, M SAITO
    JAPANESE JOURNAL OF VETERINARY RESEARCH 42 3-4 137 - 146 1994年12月 [査読有り][通常論文]
     
    When mammals are exposed to a cold environment for a long time, the capacity of nonshivering thermogenesis by brown adipose tissue (BAT) increases in association with the increased expression of some specific proteins and tissue hyperplasia, which are totally dependent on sympathetic innervation to this tissue. To clarify roles of the beta-adrenergic mechanism in BAT hyperplasia, the effects of chronic administration of various beta-adrenergic agonists on BAT were examined in rats, especially focusing on some agonists to the beta 3-adrenoceptor which is present specifically in adipocytes. Chronic administration of noradrenaline or isoproterenol for 7-10 days produced a marked increase in the tissue contents of DNA, total protein, mitochondrial uncoupling protein, and insulin-regulatable glucose transporter protein. The trophic effects of noradrenaline and isoproterenol were mimicked by chronic administration of beta 3-adrenergic agonists, such as CL316,243, BRL 26830A, and ICI D7114. These results suggest that the beta 3-adrenoceptor plays important roles for hyperplasia of BAT, and thereby increasing in the capacity of thermogenesis.
  • N NAKAGAWATOSA, M MORIMATSU, K MOMINOKI, H NAKATSUJI, B SYUTO, M SAITO
    JOURNAL OF VETERINARY MEDICAL SCIENCE 56 1 125 - 129 1994年02月 [査読有り][通常論文]
     
    We describe a technique for isolation and primary culture of bovine hepatocytes, and their metabolic characterization. Hepatocytes were isolated from the caudate lobe of bovine liver by perfusion with 0.25 mM ethyleneglycol tetraacetic acid and 0.05% collagenase. The viability and yield of the cells were 70-92% and 0.1-3.6 x 10(7) cells/g liver, respectively. When the isolated hepatocytes were cultured in Williams' medium E, they began to spread in 3 hr and formed monolayers in 24 hr. These monolayers were retained for at least 6 days. To monitor the metabolic activities specific to liver, synthesis and secretion of albumin were measured by labeling with [S-35]-methionine and immunoprecipitation. This activity was low in isolated hepatocytes, but increased after culturing 1-3 days, and decreased again after 6 days. Glycogenolytic activity was also assessed by measuring glucose release to the medium by stimulation with epinephrine. The glycogenolytic response to epinephrine was also enhanced by culturing the hepatocytes 1-3 days, but was decreased after 6 days. Since the isolated bovine hepatocytes retained the liver-specific activities of albumin synthesis and glycogenolysis for several days in culture, these cells are useful for cellular and molecular studies on the functions of bovine liver.
  • S YAMAMOTO, K TAGATA, Y ISHIKAWA, H SANTSUKA, M YAMADA, M MORIMATSU, M NAIKI
    VETERINARY IMMUNOLOGY AND IMMUNOPATHOLOGY 36 3 257 - 264 1993年04月 [査読有り][通常論文]
     
    This paper describes the avidity of IgG antibody used for preparation of latex sensitized with IgG antibody (IgG-sensitized latex) and the agglutinability of IgG-sensitized latex in slide reversed passive latex agglutination (RPLA). Using immunodiffusion techniques, it was found that anti-canine C-reactive protein (CRP) sera from four rabbits immunized with canine CRP had the same antibody titers. However, the antibodies had different levels of avidity. When lattices were sensitized under the same condition with the IgG antibodies of different avidity levels separated from the above-mentioned antisera using Protein A and canine CRP-Sepharose 4B immunosorbent, these demonstrated different patterns of agglutinability in slide RPLA. The latex sensitized with IgG antibody of higher avidity demonstrated a stronger agglutinability.
  • S YAMAMOTO, N ABE, H SANTSUKA, T SHIDA, K KISHIDA, S KUWAJIMA, M YAMADA, M MORIMATSU, M NAIKI
    VETERINARY IMMUNOLOGY AND IMMUNOPATHOLOGY 36 3 293 - 301 1993年04月 [査読有り][通常論文]
     
    The canine C-reactive protein (CRP) fraction isolated from canine acute-phase serum on a phosphorylcholine-Sepharose 4B column was further subjected to Sephacryl S-300 gel filtration. A canine CRP fraction not containing IgM was then obtained. The antisera, obtained after several immunizations with this canine CRP fraction, contained nonspecific antibodies that reacted with albumin, transferrin and IgG in addition to CRP. This antiserum could be easily changed to monospecific canine CRP serum, when it was subjected to absorption for only 15 min using glutaraldehyde-insolubilized normal canine serum protein containing 3.5 mug ml-1 of CRP. Pure canine CRP was isolated with a recovery rate of 95% from canine acute-phase serum by affinity chromatography using specific anti-canine CRP antibody.
  • N TOSA, M MORIMATSU, M NAKAGAWA, F MIYOSHI, E UCHIDA, M NIIYAMA, B SYUTO, M SAITO
    JOURNAL OF VETERINARY MEDICAL SCIENCE 55 1 27 - 31 1993年02月 [査読有り][通常論文]
     
    Polyacrylamide gel electrophoretic analysis of canine serum protein has revealed that the administration of anthelmintics elicits an increase in a certain serum protein. This protein, named PT60, was partially purified by ammonium sulfate fractionation and preparative electrophoresis. The purified PT60 gave a single band with the molecular size of 53 kDa in sodium dodecyl sulfate-polyacrylamide gel electrophoresis under non-reducing conditions. After reduction with 2-mercaptoethanol, two bands appeared at 35 kDa and 17 kDa, indicating that PT60 consists of two subunits which are linked with each other by disulfide bonds. PT60 had the capacity to bind to hemoglobin. In an immunodiffusion test, an antiserum against PT60 cross-reacted with canine haptoglobin (Hp). N-terminal amino acid sequences of two PT60 subunits were identical to those of alpha and beta subunits of canine Hp, respectively. Thus, PT60 was identified as Hp.
  • Y NAKAJIMA, E MOMOTANI, T MURAKAMI, Y ISHIKAWA, M MORIMATSU, M SAITO, H SUZUKI, K YASUKAWA
    VETERINARY IMMUNOLOGY AND IMMUNOPATHOLOGY 35 3-4 385 - 391 1993年01月 [査読有り][通常論文]
     
    Interleukin-6 (IL-6) is a major inducer of acute phase proteins in human and murine species. However, the effects of IL-6 have not yet been investigated in cattle. Following continuous infusion of recombinant human IL,6, serum concentrations of bovine haptoglobin and fibrinogen increased in a manner similar to those in cattle with acute phase reaction. In contrast, C-reactive protein and alpha1-acid glycoprotein, as well as the other hematologic parameters, did not change significantly. Intravenous administration of recombinant human IL-6 resulted in only a mild and transient increase of bovine haptoglobin. These results suggest that the regulation of acute phase protein production in cattle is similar, but not identical, to that observed in human and murine species.
  • M MORIMATSU, M SARIKAPUTI, B SYUTO, M SAITO, S YAMAMOTO, M NAIKI
    VETERINARY IMMUNOLOGY AND IMMUNOPATHOLOGY 33 4 365 - 372 1992年09月 [査読有り][通常論文]
     
    Using purified bovine haptoglobin (Hp) and specific antisera, a single radial immunodiffusion (SRID) assay method has been developed to measure the serum Hp level in cattle. Bovine Hp is a highly polymerized protein showing heterogeneous molecular forms in serum. After treatment with cysteine or glutathione, Hp was partially reduced to a homogeneous form, suitable for SRID assay. This method gives values comparable to those obtained by hemoglobin-binding capacity assay, and has the advantage of being simple and convenient. Although serum Hp was not detectable in healthy cattle, it was found more than 50-fold after invasive surgery, indicating that Hp is a characteristic acute-phase protein in cattle.
  • S YAMAMOTO, K TAGATA, Y ISHIKAWA, H FUJISE, H NAGAHATA, M YAMADA, T SAKANO, M MORIMATSU, M NAIKI
    VETERINARY RESEARCH COMMUNICATIONS 16 4 265 - 272 1992年08月 [査読有り][通常論文]
     
    A method is described for preparing latex particles sensitized with IgG antibody (IgG-sensitized latex) applicable to the slide reversed passive agglutination (RPLA) test. Soap-free latex (latex) was sensitized with IgG which had been isolated from rabbit anti-bovine lactoferrin serum using protein A Sepharose CL 4B. Unadsorbed protein-binding sites on the surface of latex were blocked with bovine serum albumin (BSA). IgG-sensitized latex that gave better agglutination in RPLA could be selectively obtained by centrifugation at 19 900g for 15 min in 0.01 mol/L glycine buffer (pH 7.3; specific gravity 1.042) containing 3% NaCl, 5% saccharose and 2% choline chloride. By dispersing this IgG-sensitized latex in 0.01 mol/L glycine buffer (pH 7.3) containing 1-2% BSA, a uniformly suspended, highly reactive, readily agglutinable preparation was obtained.
  • M SARIKAPUTI, M MORIMATSU, S YAMAMOTO, B SYUTO, M SAITO, M NAIKI
    JAPANESE JOURNAL OF VETERINARY RESEARCH 40 1 1 - 12 1992年05月 [査読有り][通常論文]
     
    A semi-quantitative latex agglutination test for bovine serum CRP levels has been established by mixing diluted serum (or diluted standard serum) with a 1% latex suspension containing 0.489-mu-m latex particles coated with affinity-purified antibody at a ratio of 20-mu-g/mg latex. The agglutination was performed on a glass slide in a moist chamber at room temperature with 45 min. incubation. This test is reliable, reproducible and the results correlate with those of the single radial immunodiffusion (SRID) test. The effect of low temperature storage on CRP concentration revealed a 30% degradation of CRP during 2 years storage at 4-degrees-C. The possible role of EDTA addition to prevent a decrease in serum CRP concentration by freezing and thawing is also discussed.
  • A WATANABE, M MORIMATSU, K YOSHIMATSU, S YAMAMOTO, A TERAO, K TSUKAZAKI, M SAITO, M NAIKI
    JOURNAL OF SMALL ANIMAL PRACTICE 33 2 71 - 77 1992年02月 [査読有り][通常論文]
     
    C-reactive protein (CRP) was isolated from sera from healthy cats by calcium-dependent affinity chromatography on phosphorylcholine derivatives of bovine albumin-coupled Toyopearl, followed by an anion-exchange chromatography using DEAE cellulose. It was identified as CRP by its immunochemical cross reactivity with human CRP. The molecular weight of cat CRP was approximately 100 kilodaltons (kDa) and composed of two glycosylated subunits (23 kDa) and three non-glycosylated subunits (20 kDa) with non-covalent association. Under electron microscopic examination, cat CRP had a pentameric disc-like configuration which is characteristic of CRP. Immunoelectrophoresis and isoelectric focusing showed that cat CRP is an acidic alpha-1-globulin (pI 4.1 to 4.3). Serum concentrations of CRP in cats and kittens were measured by single radial immunodiffusion. In 12 healthy cats from various sources, values ranged from 38 to 168-mu-g/ml. In kittens, serum CRP levels also showed a wide distribution, 81 per cent of them were less than 40-mu-g/ml.
  • S YAMAMOTO, K TAGATA, H NAGAHATA, Y ISHIKAWA, M MORIMATSU, M NAIKI
    VETERINARY IMMUNOLOGY AND IMMUNOPATHOLOGY 30 4 329 - 339 1992年01月 [査読有り][通常論文]
     
    C-reactive protein (CRP) was isolated from the acute phase serum of dogs subjected to surgical stimulation. Its properties were characterized. Canine CRP was isolated by ion-exchange chromatography using DEAE-Sephaeel and DEAE-Sephadex A-50 and affinity chromatography using protein A-Sepharose CL 4B in combination with agar-block electrophoresis. In immunoelectrophoresis, canine CRP had the same gamma-mobility as human gamma-type CRP. The molecular weight of purifined canine CRP was estimated by gel filtration and sodium dodecyl sulfate polyacrylamide gel electrophoresis to be approximately 157 000 and 155 000 respectively. This CRP was a thermolabile protein which completely lost its antigenicity by heating at 70-degrees-C for 15 min. The serum concentration of CRP in normal beagle dogs ranged from 0.198 to 0.826-mu-g ml-1 (0.486 +/- 0.170-mu-g ml-1). The concentration was acutely increased by surgery as it was in man and was rapidly decreased with convalescence. Dogs can be a useful animal model for investigation of the mechanism of CRP production and the function of CRP.
  • M MORIMATSU, A WATANABE, K YOSHIMATSU, T FUJINAGA, M OKUBO, M NAIKI
    JOURNAL OF DAIRY RESEARCH 58 3 257 - 261 1991年08月 [査読有り][通常論文]
     
    C-reactive protein (CRP) and serum amyloid P component (SAP), which are known to increase in sera from humans and many other animals with acute inflammation caused by infection, toxic drug administration or injury, were previously purified from bovine serum. These serum levels were determined by enzyme-linked immunosorbent assay (ELISA) using specific antiserum to bovine CRP or SAP which was prepared by immunizing rabbits and goats with each purified protein. Among 68 healthy Holstein cows, 45 non-lactating cows had levels of CRP and SAP of 20.6 +/- 1.4 and 27.6 +/- 1.3-mu-g/ml respectively; 23 lactating cows had higher levels of CRP and SAP (76.0 +/- 13.6 and 38.3 +/- 5.5-mu-g/ml respectively). In the latter group, there was a high correlation between milk yield and serum CRP levels (P < 0.001). From these observations, it was assumed that lactation might stimulate CRP synthesis rather than SAP synthesis in bovine liver as an acute phase reaction, and that CRP might be called a lactation-associated protein.
  • M MORIMATSU, B SYUTO, N SHIMADA, T FUJINAGA, S YAMAMOTO, M SAITO, M NAIKI
    JOURNAL OF BIOLOGICAL CHEMISTRY 266 18 11833 - 11837 1991年06月 [査読有り][通常論文]
     
    A macromolecular hemoglobin-binding protein, which was not detectable in normal bovine sera but appeared during acute phase inflammation, was purified, characterized, and designated as bovine haptoglobin (Hp). The purified protein had a molecular mass of 1,000-2,000 kDa, and was composed of two kinds of peptides, a 20-kDa peptide (alpha-chain) and a 35-kDa glycopeptide (beta-chain) linked by disulfide bonds. Amino acid composition and N-terminal sequence analyses revealed that both peptides were homologous to each counterpart of human Hp. Studies using some reducing reagents proved that highly polymerized Hp in serum was composed of 2-20 polymerized forms of alpha-2-beta-2 tetramer. Hp could bind one molecule of hemoglobin/alpha-2-beta-2 unit. Hp with smaller sizes obtained from native Hp by partial reduction with cysteine showed almost the same Hb-binding capacity.
  • M SARIKAPUTI, M MORIMATSU, B SYUTO, M SAITO, M NAIKI
    INTERNATIONAL JOURNAL OF BIOCHEMISTRY 23 10 1137 - 1142 1991年 [査読有り][通常論文]
     
    1. A new purification procedure was started with salting-out fractionation of serum proteins at 45-75% saturated ammonium sulfate concentration, followed by HE agarose affinity chromatography by which calcium-dependently bound CRP and SAP were purely eluted with EDTA-containing buffer. 2. Pure CRP and SAP were finally separated by DEAE-5PW HPLC. 3. This procedure gave recovery of 15 and 26%, and fold purification of 2650 and 2400 for CRP and SAP. respectively. 4. Each subunit of CRP and SAP had one intrasubunit disulfide bond, determined by reduction and carboxymethylation.
  • M MORIMATSU, H SAKAI, K YOSHIMATSU, O MINOWA, S YAMAMOTO, K YATOMI, T FUJINAGA, M NAIKI
    JAPANESE JOURNAL OF VETERINARY SCIENCE 51 4 723 - 732 1989年08月 [査読有り][通常論文]

講演・口頭発表等

その他活動・業績

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    植村 光希, 落合 和彦, 森松 正美, 柏木 伸旭, 宇埜 友美子, 道下 正貴, 呰上 大吾, 小野沢 栄里, 近江 俊徳 日本獣医学会学術集会講演要旨集 162回 479 -479 2019年08月 [査読無し][通常論文]
  • イヌ乳腺腫瘍細胞株で生じたp53遺伝子変異の検出と機能解析
    落合 和彦, 平間 日奈子, 森松 正美, 川上 翔太, 道下 正貴, 中川 貴之, 呰上 大吾, 稲垣 健志, 坂本 敦司, 近江 俊徳 日本獣医学会学術集会講演要旨集 161回 431 -431 2018年08月 [査読無し][通常論文]
  • C57BL/6マウスよりセンダイウイルス感染抵抗性遺伝子座を導入されたDBA/2コンジェニックマウスのセンダイウイルス感染に対する免疫応答
    Abbas Raghda, Tag-EL-Din-Hassan Hassan, Boonyarattanasoonthorn Tussapon, 青島 圭佑, 森松 正美, 安居院 高志 日本獣医学会学術集会講演要旨集 161回 476 -476 2018年08月 [査読無し][通常論文]
  • 森松 正美 実験動物と環境 26 (1) 47 -51 2018年04月 [査読無し][通常論文]
  • 伊達衆, 中尾亮, ISLAM Md, 孝口裕一, 入江隆夫, 森松正美, 安居院高志, 八木欣平, 片倉賢 日本獣医学会学術集会講演要旨集 160th 344 2017年08月30日 [査読無し][通常論文]
  • ISLAM Md, 孝口裕一, 入江隆夫, 亀田弥生, 鳥越大輔, 中尾亮, TAG‐EL‐DIN‐HASSAN Hassan, 森松正美, 八木欣平, 安居院高志 日本獣医学会学術集会講演要旨集 160th 517 2017年08月30日 [査読無し][通常論文]
  • Aya Yoshimura, Manabu Musashi, Takeaki Kaneko, Shunsuke Ohnishi, Chieko Orito, Yukako Kawahara, Satoshi Hashino, Masami Morimatsu, Satoshi Konno, Jiro Arikawa, Tetsuya Ishii, Masaya Sawamura, Ichiro Ueda Japanese Journal of Allergology 63 (8) 1132 -1139 2014年 [査読無し][通常論文]
     
    Background: Based on a case who developed anaphylaxis after mouse bite which occurred at Hokkaido University, we studied on allergic sensitization prevalence for laboratory animals among students and researchers who are exposed to laboratory rodents and rabbit, for the purpose of allergy prevention, particularly anaphylaxis. Methods: We carried out the health check-up on laboratory animal allergy (LAA) by questionnaires and specific-IgE antibody test for 555 rodents and/or rabbit handlers from whom informed consent was obtained. Result: Prevalence of positive IgE antibody higher than class 1 to mice, rats, hamsters, guinea pigs, and/or rabbits in the examinees was 14.1% (62/441), 17.9% (50/279), 18.8% (6/32), 17.4% (4/23), and 11.3% (12/106), respectively. Moreover, among users of mouse, those who had allergic symptoms during contact with animals resulted in significantly higher positive rate for anti-mouse IgE antibody test than the other (38.1% vs 8.8%, p< 0.01). Conclusion: Health check-up including measurement of specific-IgE antibody against laboratory animals is useful for understanding allergic sensitization.
  • 小原徹, 高尾尊身, 佐加良英治, 朱宮正剛, 米川博通, 室田宏之, 森松正美 実験動物と環境 21 (41) 47 -53 2013年04月01日 [査読無し][通常論文]
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  • 落合和彦, 吉川泰永, 近江俊徳, 森松正美 日本病態生理学会雑誌 21 (1) 33 -40 2012年05月20日 [査読無し][通常論文]
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  • 落合和彦, 森松正美, 吉川泰永, 宇賀聡, 首藤文栄, 橋爪一善 獣医生化学 41 (2) 51 -59 2004年10月30日 [査読無し][通常論文]
  • 吉川泰永, 森松正美, 落合和彦, 永野昌志, 冨沢伸行, 山根義久, 橋爪一善 日本獣医学会学術集会講演要旨集 138th 182 2004年08月01日 [査読無し][通常論文]
  • BH Lee, K Yoshimatsu, A Maeda, K Ochiai, M Morimatsu, K Araki, M Ogino, S Morikawa, J Arikawa VIRUS RESEARCH 98 (1) 83 -91 2003年12月 [査読無し][通常論文]
     
    We performed yeast two-hybrid screening of a human kidney cell cDNA library to study the biological role of the hantavirus nucleocapsid protein (NP). We found that Seoul virus (SEOV) and Hantaan virus (HTNV) NPs were associated with small ubiquitin-like modifier (SUMO)-1-interacting proteins PIAS 1, PIASxbeta, HIPK2, CHD3, and TTRAP, which interacted with the SUMO-1 conjugating enzyme (Ubc-9) and SUMO-1 in the yeast two-hybrid assay. Interactions between the HIPK2, CHD3, and TTRAP proteins and SEOV NP were also shown in a mammalian two-hybrid assay. However, there was no interaction between PIAS proteins and NP, which was probably due to the inhibitory effect of PIAS on transcription in the mammalian two-hybrid assay. Nevertheless, a co-expression experiment suggested the existence of a PIAS-NP interaction in the cytoplasm. The region spanning amino acids 100-125 of SEOV NP, which represents a critical region for NP-NP polymerization. was found to be responsible for the interaction with SUMO-1-related molecules in both the yeast and mammalian two-hybrid assays. These results add to the information on interactions of hantavirus NP and host cellular proteins. (C) 2003 Elsevier B.V. All rights reserved.
  • 富山美奈子, 小藤田久義, 森松正美, 首藤文栄 獣医生化学 40 (2) 47 -53 2003年10月30日 [査読無し][通常論文]
  • Toshina Oonuma, Masami Morimatsu, Kazuhiko Ochiai, Bunei Syuto Journal of Veterinary Medical Science 65 (10) 1123 -1126 2003年10月 [査読無し][通常論文]
     
    Mammary tumors are common in cats. As mutations in human Brca2 confer an increased risk of breast cancer, the full-length cDNA of the feline homologue of Brca2 was sequenced to obtain a basis for studying the relationship between its function and susceptibility to mammary tumors. The feline Brca2 cDNA is 10 kb long, and encodes 3,371 amino acids. The amino acid sequence of feline Brca2 shares low homology with the Brca2 of other mammals, e.g., 53% homology with the murine protein. Analysis of the expression pattern of the feline Brca2 gene revealed that, as previously reported for other mammals, it is transcribed in various tissues, including the mammary gland.
  • T Oonuma, M Morimatsu, T Nakagawa, R Uyama, N Sasaki, M Nakaichi, H Tamamura, N Fuji, S Hashimoto, H Yamamura, B Syuto JOURNAL OF VETERINARY MEDICAL SCIENCE 65 (10) 1069 -1073 2003年10月 [査読無し][通常論文]
     
    It has recently been suggested that the chemokine receptor CXCR4 and its ligand SDF-1 (CXCL12) promote metastasis of various cancers in humans. Since feline mammary tumors also metastasize to distant organs frequently, we used real-time quantitative PCR to examine the expression of feline CXCR4 (fCXCR4) in ten feline mammary tumor cell lines and seven feline mammary tumor tissues, and also the expression of feline SDF-1 (fSDF-1) in various organs. Cell lines derived from metastatic regions expressed more fCXCR4 than those derived from primary tumors. Mammary tumor tissues overexpressed more fCXCR4 than normal mammary tissues. Organs with high levels of fSDF-1 expression represent common sites of metastasis. Migration assays using the feline mammary tumor cell line NAC were also performed to test the activity of TN14003 and TC14012, antagonists of human CXCR4, to antagonize fCXCR4 expressed on NAC cells. TN14003 and TC14012 inhibited migration of NAC cells. We conclude that fCXCR4 may be a therapeutic target for feline mammary tumors.
  • H Kitamura, Y Matsushita, T Iwanaga, K Mori, K Kanehira, D Fujikura, M Morimatsu, M Saito ARCHIVES OF HISTOLOGY AND CYTOLOGY 66 (1) 53 -62 2003年03月 [査読無し][通常論文]
     
    Molecule possessing ankyrin-repeats induced by lipopolysaccharide (MAIL), a recently cloned nuclear IkappaB protein induced by lipopolysaccharide (LPS) stimulation in lymphoid organs, is involved in the regulation of inflammatory responses. The present in situ hybridization and immunohistochemical analyses revealed the distinct expression of the MAIL mRNA and protein in B-lymphocytes of the white pulp of the spleen and cortical lymphoid follicles of lymph nodes in LPS-injected mice. MAIL signals were also localized in F4/80-positive macrophages in these organs. LPS clearly induced MAIL expression in cultured B-lymphocytes and monocytes/macrophages, but only faintly so in T-lymphocytes, fibroblasts, and endothelial cells. MAIL was also induced by inflammatory cytokines such as interleukin-1 and -6, and tumor necrosis factor in cultured cells. Northern blot, Western blot, and in situ hybridization analyses showed that the major expression product of the Mail gene was a long splicing variant (MAIL-L) rather than a short one, both in lymphoid organs and cultured cells. These results collectively indicate that LPS induces MAIL-L predominantly in B-lymphocytes and macrophages.
  • 山地大介, 北村浩, 松下由紀子, 椎名貴彦, 森松正美, 斉藤昌之 獣医生化学 39 (2) 13 -18 2002年10月31日 [査読無し][通常論文]
  • 三井隆行, 菊佳男, 福田茂夫, 田中隆志, 伊藤寿浩, 森松正美, 首藤文栄, 高橋淳吉, 吉野知男 日本獣医学会学術集会講演要旨集 134th 149 -149 2002年08月20日 [査読無し][通常論文]
  • R Kamata, M Morimatsu, T Suzuki, T Takewaki, H Kobayashi ENVIRONMENTAL TOXICOLOGY AND PHARMACOLOGY 12 (1) 55 -58 2002年08月 [査読無し][通常論文]
     
    Gel filtration chromatography was performed on cytosol preparation of hen spinal cord to find molecular target(s) for organophosphorus-induced delayed neurotoxicity (OPIDN). Three binding peaks of [H-3]diisopropyl phosphorofluoridate (DFP), an organophosphate that induces OPIDN, were separated from the cytosol preparation. The activities of acetylcholinesterase (AChE) and neuropathy target esterase (NTE) that has been proposed as a screening method for OPIDN eluted in the fractions within these two DFP binding peaks. However, the other peak had none of the activities of AChE and NTE. Therefore, this DFP binding proteins in cytosol may be peculiar to the pathogenesis of OPIDN. (C) 2002 Elsevier Science B.V. All rights reserved.
  • M Suzuki, W Fujimoto, M Goto, M Morimatsu, B Syuto, T Iwanaga JOURNAL OF HISTOCHEMISTRY & CYTOCHEMISTRY 50 (8) 1081 -1089 2002年08月 [査読無し][通常論文]
     
    Recently, the second mammalian chitinase, designated acidic mammalian chitinase (AMCase), has been identified in human, mouse, and cow. In contrast to the earlier identified macrophage-derived chitinase (chitotriosidase), this chitinase is richly expressed in the gastrointestinal (GI) tract, suggesting its role in digestion of chitin-containing foods as well as defense against chitin-coated microorganisms and parasites. This in situ hybridization study first revealed cellular localization of the gut-type chitinase in the mouse and chicken. In adult mice, the parotid gland, von Ebner's gland, and gastric chief cells, all of which are exocrine cells of the serous type, expressed the gut chitinase mRNA. In the chicken, oxyntico-peptic cells in glandular stomach (proventriculus) and hepatocytes expressed the chitinase mRNA. Because cattle produce the gut chitinase (chitin-binding protein b04) only in the liver, the gut chitinases in mammals and birds have three major sources of production, i.e., the salivary gland, stomach, and liver. During ontogenetic development, the expression level in the parotid gland and stomach of mice increased to the adult level before weaning, whereas in the stomach of chickens intense signals were detectable in embryos from incubation day 7.
  • M Suzuki, M Morimatsu, B Syuto JOURNAL OF VETERINARY MEDICAL SCIENCE 64 (6) 477 -481 2002年06月 [査読無し][通常論文]
     
    Bovine serum contains N-acetyl-D-glucosamine (GlcNAc)-sensitive opsonin inhibitory factors. In the present study, a major component of chitin-binding protein (chitin-binding protein b01, CBPb01) was purified from bovine serum, and identified CBPb01 as bovine IgM by its subunit structure, antigenic properties, and partial sequences. The results of a lectin-binding assay showed that the heavy chain of CBPb01 had a GlcNAc structure, but the commercial IgM did not. It is possible that CBPb01 interconnects through its GlcNAc structure, subsequently forming complexes. We also demonstrated that CBPb01 had opsonin-inhibitory activity, and that this activity was dependent on the binding of CBPb01 to GlcNAc on the zymosan surface. These findings indicate the presence of a kind of IgM that recognizes GlcNAc structure in the regulation of opsonization.
  • Hiroshi Kitamura, Katsushi Kanehira, Takahiko Shiina, Masami Morimatsu, Bae Dong Jung, Sachiko Akashi, Masayuki Saito Journal of Veterinary Medical Science 64 (5) 419 -422 2002年05月 [査読無し][通常論文]
     
    Molecule possessing ankyrin-repeats induced by lipopolysaccharide (MAIL) is a nuclear IκB protein recently identified as a molecule appearing in immunocompetent organs after administration of bacterial lipopolysaccharide (LPS). Participation of Toll-like receptor (TLR) 4, which is a major form of LPS receptors, in the LPS-induced MAIL expression was investigated. When a human myelomonocytic cell line U937 was treated with phorbol 12-myristate 13-acetate for 3 days, the LPS-induced MAIL expression was much potentiated in parallel with an increase in TLR4 expression. The MAIL induction was attenuated when the cells were treated with a neutralizing antibody against TLR4. The in vivo induction of MAIL in the spleen was smaller in mice having a missense mutation of the Tlr4 gene than in normal control mice. These results collectively indicate that TLR4 contributes, at least in part, MAIL induction after LPS stimulation.
  • 鈴木 雅子, 森松 正美, 山下 哲郎, 岩永 敏彦, 首藤 文榮 獣医生化学 = Veterinary biochemistry 38 (2) 2001年10月31日 [査読無し][通常論文]
  • 山地 大介, 北村 浩, 森 浩志, 岩永 敏彦, 松下 由紀子, 椎名 貴彦, 森松 正美, 斉藤 昌之 獣医生化学 = Veterinary biochemistry 38 (2) 2001年10月31日 [査読無し][通常論文]
  • 金平克史, 北村浩, 岩永敏彦, 藤倉大輔, 森松正美, 斉藤昌之 獣医生化学 38 (2) 19 -25 2001年10月31日 [査読無し][通常論文]
  • T Shiina, M Morimatsu, H Kitamura, T Ito, S Kidou, K Matsubara, Y Matsuda, M Saito, B Syuto IMMUNOGENETICS 53 (8) 649 -655 2001年10月 [査読無し][通常論文]
     
    The Mail (molecule possessing ankyrin repeats induced by lipopolysaccharide) protein is a member of the I kappaB family. It has six ankyrin repeats that are conserved in other I kappaB proteins, such as I kappaB-alpha and Bcl-3. Mail mRNA expression is induced rapidly following lipopolysaccharide (LPS) injection, most notably in the spleen, lung, and lymph nodes of mice, where immune cells, such as lymphocytes and macrophages, are abundant. In this study, we cloned and characterized the Mail gene. The isolated genomic clones span approximately 30 kb and encompass the entire gene. Comparisons with Mail cDNA revealed that the Mail gene consists of 14 exons. Several splice junctions encoding ankyrin repeats are conserved among Mail and other I kappaB family genes. Southern hybridization showed that Mail is a single-copy gene. Using fluorescence in situ hybridization analysis, mouse and rat Mail genes were mapped to Chromosome (Chr) 16C 1.2-C1.3 and Chr 11q21.1, respectively. Primer extension determined the transcription start site of Mail. Sequence analysis of the proximal promoter region revealed the presence of a TATA box and putative transcription factor-binding sites, such as those for NF-kappaB and NF-IL6. This region is sufficient to drive high-level reporter gene expression in LPS-stimulated transfected cells.
  • K Ochiai, M Morimatsu, N Tomizawa, B Syuto JOURNAL OF VETERINARY MEDICAL SCIENCE 63 (10) 1103 -1108 2001年10月 [査読無し][通常論文]
     
    Mammary tumors are the most common neoplasm in female dogs. Canis canis, and in women. Mutations in human Brca2 confer an increased risk of female breast cancer. Previous studies have shown that the Brca2 tumor suppressor protein interacts with the recombinational repair protein Rad51. We cloned the full-length cDNA of the canine homologues of Brca2 and Rad51 to obtain a basis for studying their relationship with susceptibility to mammary tumors. The canine Brca2 and Rad51 cDNAs are 11 and 1.5 kb long, encoding 3,471 and 339 amino acids, respectively. The amino acid sequence of canine Brca2 showed 68% homology with the human protein., and 58% homology with a murine protein. There were highly conserved regions in the C-terminus of all three proteins, where the Rad51 interacting domain and putative nuclear localization signals are located. Comparing with the partial genomic sequence previously reported, we found possible nuclear polymorphisms in exon 11, some of which result in amino acid substitutions. On the other hand, canine Rad51 protein had extremely high homology (99%) to the human and murine proteins. Expression of both Brca2 and Rad51 was detected in the mammary gland, suggesting that these two genes interact in the canine mammary gland.
  • M Suzuki, M Morimatsu, T Yamashita, T Iwanaga, B Syuto FEBS LETTERS 506 (2) 127 -130 2001年10月 [査読無し][通常論文]
     
    Chitinases are ubiquitous chitin-fragmenting hydrolases. They are synthesized by a vast array of organisms, including those not composed of chitin. Here, we describe a novel serum chitinase (chitin-binding protein b04, CBPb04), which is expressed in bovine liver. Although CBPb04 is secreted as an endocrine chitinase, it shows higher homology with human gastrointestinal tract exocrine chitinase (AMCase) than with macrophage endocrine chitinase (human chitotriosidase). This suggests that cows have a specific defense against chitin-containing microorganisms. CBPb04 mRNA is expressed in hepatocytes. This is the first report of a hepatogenic mammalian chitinase. (C) 2001 Federation of European Biochemical Societies. Published by Elsevier Science B.V. All rights reserved.
  • 北村浩, 金平克史, 岩永敏彦, 藤倉大輔, 山地大介, 松下由紀子, 森松正美, 斉藤昌之 日本獣医学会学術集会講演要旨集 132nd 140 2001年09月07日 [査読無し][通常論文]
  • 北村浩, 金平克史, 岩永敏彦, 椎名貴彦, 松下由紀子, 藤倉大輔, 赤司祥子, 森松正美, 斉藤昌之 生化学 73 (8) 1042 2001年08月25日 [査読無し][通常論文]
  • S Vestri, MM Okamoto, HS de Freitas, RA dos Santos, MT Nunes, M Morimatsu, JC Heimann, UF Machado JOURNAL OF MEMBRANE BIOLOGY 182 (2) 105 -112 2001年07月 [査読無し][通常論文]
     
    Renal glucose reabsorption is mediated by luminal sodium-glucose cotransporters (SGLTs) and basolateral facilitative glucose transporters (GLUTs). The modulators of these transporters are not known, and their substrates glucose and Na+ are potential candidates. In this study we examined the role of glucose and Na+ filtration rate on gene expression of glucose transporters in renal proximal tubule. SGLT1, SGLT2, GLUT1 and GLUT2 mRNAs were assessed by Northern blotting; and GLUT1 and GLUT2 proteins were assessed by Western blotting. Renal cortex and medulla samples from control rats (C), diabetic rats (D) with glycosuria, and insulin-resistant 15-month old rats (I) without glycosuria; and from normal (NS), low (LS), and high (HS) Na+-diet fed rats were studied. Compared to C and I rats, D rats increased (P < 0.05) gene expression of SGLT2 by similar to 36%, SGLT1 by similar to 20%, and GLUT2 by similar to 100%, and reduced (P < 0.05) gene expression of GLUT1 by more than 50%. Compared to NS rats, HS rats increased (P < 0.05) SCLT2, GLUT2, and GLUT1 expression by <similar to>100%. with no change in SGLT1 mRNA expression, and LS rats increased (P < 0.05) GLUT1 gene expression by <similar to>150% with no changes in other transporters. In summary, the results showed that changes in glucose or Na+ filtrated rate modulate the glucose transporters gene expression in epithelial cells of the renal proximal tubule.
  • K Iwasaki, M Morimatsu, O Inanami, E Uchida, B Syuto, M Kuwabara, M Niiyama JOURNAL OF BIOLOGICAL CHEMISTRY 276 (12) 9400 -9405 2001年03月 [査読無し][通常論文]
     
    Acute-phase serum proteins were induced by administrating a chicken with turpentine oil. One of these proteins was a new protein that appeared in front of albumin in polyacrylamide disc gel electrophoresis using a 4.5-16% gel. To purify this protein, turpentine-administrated chicken serum was fractionated by ammonium sulfate precipitation at 50% saturation, and the supernatant fraction was chromatographed on a DEAE-Toyopearl 650S column. The purified protein is a mannose-glycoprotein, and its N-terminal sequence, determined by the Edoman method, is not homologous from that of other reported acute-phase proteins. An analysis of physiological function with two different test systems, chemiluminescence measurement and electron spin resonance spectroscopy, showed that the purified protein has antioxidant, activity and inhibits superoxide (O-2(radical anion)) mediated by activation of the receptor. In support of these results, the complete amino acid sequence of 18-B is homologous to the scavenger receptor cysteine-rich (SRCR) family of proteins that participate in the regulation of leukocyte function. 18-B is composed of four SRCR domains, which is different from the previously characterized SRCR family of proteins such as Sp alpha, CD6, and CD163. These findings indicate that turpentine-induced 18-B, a new member of scavenger receptor cysteine-rich family, may be implicated in regulation of cell function in a manner of inhibition of the overproduction of the reactive oxygen species.
  • 北村浩, 金平克史, 沖田圭介, 岩永敏彦, 椎名貴彦, 藤倉大輔, 森松正美, 斉藤昌之 生化学 72 (8) 1017 2000年08月25日 [査読無し][通常論文]
  • Y Watanabe, M Morimatsu, B Syuto JOURNAL OF VETERINARY MEDICAL SCIENCE 62 (5) 473 -478 2000年05月 [査読無し][通常論文]
     
    Botulinum C3 enzyme produced by Clostridium botulinum type C and D strains modifies Rho proteins. In a previous study, we observed that the LDH isozyme pattern of neurons treated with C3 enzyme was different from that induced with endogenous growth factor of neurons such as NGF [21]. This type of change is considered to have an advantage in the medical use of C3 enzyme for neural disorder. To determine the functional similarity of C3-treated neurons to control and NGF-treated neurons, we examined the responses of C3-treated neurons to various drugs, including some neurotransmitters, by measuring the rise of intracellular Ca ions into the neurons. The time course of the rise of intracellular Ca ions induced by high concentration of potassium in the C3-treated neurons was similar to that in the NGF-treated neurons. The C3-treated neurons responded to glutamic acid, aspartic acid, kainic acid, gamma-aminobutylic acid, muscarine and ACh with similar time courses and magnitudes as the control neurons. These results suggest that the C3 enzyme induces the functional differentiation of neurons, and that C3 enzyme has the potential for the medical use as an exogenous differentiation-inducing factor of neurons.
  • 鈴木雅子, 竹内由香, 海老原恵, 森松正美, 首藤文栄 獣医生化学 37 (1) 11 -15 2000年04月30日 [査読無し][通常論文]
  • M Iwase, K Kimura, N Sasaki, R Komagome, K Ishioka, M Morimatsu, T Murakami, M Saito RESEARCH IN VETERINARY SCIENCE 68 (2) 109 -114 2000年04月 [査読無し][通常論文]
     
    Leptin, the product of the ob gene, is one of the key molecules for the regulation of appetite and whole-body energy balance, and thereby for the pathogenesis of obesity. In an attempt to clarify the roles of leptin in obesity and/or related diseases in companion animals, canine leptin CDNA was cloned by amplifying reverse-transcriptase products of RNA extracted from the adipose tissue of the beagle. A CDNA clone of about 3 kbp contained a 501 bp open reading frame coding a 167-amino acid protein with a 21-amino acid signal peptide. The sequence of a 146-amino acid mature leptin was more than 79 per cent identical to those of other mammals. Northern blot analysis revealed abundant expression of leptin mRNA in adipose tissue, but not in other tissues, in adult beagles. When Chinese hamster ovary cells expressing the rat leptin receptor were stimulated with recombinant canine leptin produced by E. coli, some intracellular signal transduction proteins were phosphorylated, indicating that the recombinant leptin was biologically active. The data reported herein will be helpful for further studies of leptin of the dog in health and disease. (C) Harcourt Publishers Ltd.
  • Y Watanabe, M Morimatsu, B Syuto JOURNAL OF VETERINARY MEDICAL SCIENCE 62 (3) 249 -254 2000年03月 [査読無し][通常論文]
     
    Changes in the lactate dehydrogenase (LDH) isozyme pattern of primary culture of neurons treated with botulinum C3 enzyme were examined in order to elucidate the functional changes accompanying the morphological change that follows ADP-ribosylation of Rho protein. Primary neurons were prepared from the cerebrum of ICR mouse embryos on day 15. Neurons were cultured in MEM with 10% fetal calf serum at 37 degrees C. In the neurons treated with C3 enzyme, a typical morphological change was observed after 24 hr, and the LDH isozyme pattern was changed after 72 hr. The ratio of H-subunit to M-subunit in LDH was decreased by C3 treatment, suggesting the induction of a state of lower intracellular oxygen consumption in neurons in the primary cultures.
  • K Kimura, N Sasaki, A Asano, J Mizukami, S Kayahashi, T Kawada, T Fushiki, M Morimatsu, T Yoshida, M Saito HORMONE AND METABOLIC RESEARCH 32 (3) 91 -96 2000年03月 [査読無し][通常論文]
     
    Wild-type or mutated human beta 3-adrenergic receptor (Trp64Arg) cDNAs were stably expressed in mouse 3T3-L1 cells. Saturation binding study using a beta-adrenergic ligand revealed that there was no significant difference in the receptor density and the equilibrium dissociation constant between the two cell lines. However, the ability of the mutant beta 3-adrenergic receptor to accumulate cyclic AMP (cAMP) in response to isoproterenol was much reduced and Kact for cAMP accumulation was lowered as compared to the wild type receptor. The amount of alpha subunit of stimulatory GTP-binding protein (GS alpha) and adenylyl cyclase activity in response to forskolin were not different in the two cell lines. The responses of the mutant receptor to epinephrine, norepinephrine and L-755,507, a highly specific agonist for human beta 3-adrenergic receptor, were also reduced, but the reduction of Kact for L-755,507 was more evident than other agonists tested. The cAMP accumulation in response to some conventional beta 3 agonists was less than 10% of that to isoproterenol even in the cells expressing the wild type receptor. These results suggest that the Trp64Arg mutant beta 3-adrenergic receptor has less ability to stimulate adenylyl cyclase, and that lipolytic activity through the beta 3-adrenergic receptor by catecholamines in subjects carrying this mutation might be suppressed.
  • 金平克史, 北村浩, 沖田圭介, 岩永敏彦, 藤倉大輔, 森松正美, 斉藤昌之 日本獣医学会学術集会講演要旨集 129th 165 -165 2000年03月01日 [査読無し][通常論文]
  • Y Tabata, K Yotsuya, Y Torii, K Aoyagi, M Morimatsu, B Syuto JAPANESE JOURNAL OF INFECTIOUS DISEASES 53 (1) 32 -45 2000年02月 [査読無し][通常論文]
  • 北村浩, 沖田圭介, 金平克史, 藤倉大輔, 岩永敏彦, 森松正美, 斉藤昌之 日本分子生物学会年会プログラム・講演要旨集 22nd 482 1999年11月22日 [査読無し][通常論文]
  • 鈴木恵子, 山下哲郎, 森松正美, 岡達三, 首藤文栄 家畜生化学 36 (1) 37 -42 1999年04月30日 [査読無し][通常論文]
  • M Morimatsu, G Donoho, P Hasty CANCER RESEARCH 58 (15) 3441 -3447 1998年08月 [査読無し][通常論文]
     
    The putative Brca2-MmRad51 interaction is analyzed in mouse cells deleted for the COOH terminus of Brca2 (amino acids 3140-3328), which contains a region that associates with MmRad51 by yeast two-hybrid. These cells are hypersensitive to gamma-radiation (suggesting defective recombinational repair) but not UV light (suggesting intact nucleotide excision repair) and maintain the G(1)-S and G(2)-M checkpoints after exposure to gamma-irradiation. Cells deleted for the COOH terminus of Brca2 progress through the cell cycle at a similar rate as wild-type cells but undergo senescence more rapidly. These data support the hypothesis that deletion of Brca2 stimulates cancer by defective MmRad51-mediated DNA repair and not by defective cell cycle regulation.
  • H Abe, Y Kawakita, T Miyashige, M Morimatsu, M Saito JOURNAL OF VETERINARY MEDICAL SCIENCE 60 (6) 769 -771 1998年06月 [査読無し][通常論文]
     
    The nucleotide sequence of cDNA coding the 38-amino acid of the C-terminal domain of insulin-responsive glucose transporter (GLUT4) was determined by the method of reverse transcription-polymerase chain reaction in the sheep, goat and pig, and compared with that of bovine which have been shown to have a unique amino acid conversion of Asn508 to His. The deduced amino acid sequence was completely identical in the three species, and did not have the amino acid conversion at position 508. Western blot analysis confirmed that an antiserum raised against a rat C-terminal peptide cross-reacted efficiently with GLUT4 of these livestock mammals.
  • もみ木勝巳, 森松正美, 斉藤昌之 家畜生化学 34 (2) 45 -53 1997年12月 [査読無し][通常論文]
  • H Kitamura, A Konno, M Morimatsu, BD Jung, K Kimura, M Saito BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS 238 (3) 707 -711 1997年09月 [査読無し][通常論文]
     
    When mice were subjected to restriction of movement in a small cylinder (immobilization stress), the serum interleukin (IL)-6 level rose in 1 h, following increased expression of IL-6 mRNA in both the liver and the spleen. The IL-6 mRNA induction was much greater in the liver than in the spleen when compared on a whole-organ basis. Intraperitoneal injection of bacterial lipopolysaccharide (LPS) also increased IL-6 mRNA expression in these organs, but more preferentially in the spleen. Immunohistochemical examinations of liver tissue using an antibody against murine IL-6 revealed that immobilization stress induced IL-6 mainly in hepatic parenchymal cells, whereas LPS injection did so only in sinusoidal mononuclear cells. These results indicate that immobilization stress induces IL-6 production in the liver, especially in hepatic parenchymal cells, probably by a different mechanism from that for IL-6 induction by LPS. (C) 1997 Academic Press.
  • SK Sharan, M Morimatsu, U Albrecht, DS Lim, E Regel, C Dinh, A Sands, G Eichele, P Hasty, A Bradley NATURE 386 (6627) 804 -810 1997年04月 [査読無し][通常論文]
     
    Inherited mutations in the human BRCA2 gene cause about half of the cases of early-onset breast cancer. The embryonic expression pattern of the mouse Brca2 gene is now defined and an interaction identified of the Brca2 protein with the DNA-repair protein Rad51. Developmental arrest in Brca2-deficient embryos, their radiation sensitivity, and the association of Brca2 with Rad51 indicate that Brca2 may be an essential cofactor in the Rad51-dependent DNA repair of double-strand breaks, thereby explaining the tumour-suppressor function of Brca2.
  • Adrenergic activation of vascular endothelial growth factor mRNA expression in ratbrown adipose tissue : implication in cold-induced angiogenesis. (jointly worked)
    Biochemical Journal 328 179 -183 1997年 [査読無し][通常論文]
  • H Abe, M Morimatsu, H Nikami, T Miyashige, M Saito JOURNAL OF ANIMAL SCIENCE 75 (1) 182 -188 1997年01月 [査読無し][通常論文]
     
    Insulin-responsive glucose transporter GLUT4, is a member of the glucose transporter family (GLUT) and is present exclusively in muscle and adipose tissue. It is a target of insulin action in humans and rodents. To clarify the molecular structure of bovine GLUT4, its GLUT4 cDNA was cloned by the RT-PCR method. Several cDNA clones corresponding to the different regions of GLUT4 were obtained by amplifying reverse-transcriptase products of RNA extracted from Holstein cattle skeletal muscle. Nucleotide sequence analysis of the cDNA clones revealed that bovine GLUT4 cDNA was composed of 2,656 base pairs with a coding region for a 509 amino acid protein. The deduced amino acid sequence was 64% and 92% identical with bovine GLUT1 (GLUT ubiquitously expressed in all tissues) and rat GLUT4, respectively. Although the amino acid sequence of the GLUT4 COOH-terminal region is highly conserved among the species so far reported, one amino acid (Asp) of this region was replaced by His in bovine GLUT4. The tissue distribution of GLUT4, was also examined by Northern blot analysis using a probe prepared from the bovine cDNA. GLUT4 mRNA was detected in skeletal muscle, heart, and adipose tissue, but not in liver, kidney, lung, brain, or spleen. Such a distribution is essentially the same as in humans and rodents, suggesting that GLUT4 is an insulin-responsive glucose transporter in cattle.
  • 阿部啓之, 森松正美, 斉藤昌之 家畜生化学 33 (2) 1 -10 1996年12月 [査読無し][通常論文]
  • 斉藤昌之, 森松正美, 阿部啓之 食肉に関する助成研究調査成果報告書 14(1995) 185 -189 1996年12月 [査読無し][通常論文]
  • 木村 和弘, 岡田 尚子, 森松 正美, 斉藤 昌之 神経化学 35 (3) 540 -541 1996年09月05日 [査読無し][通常論文]
  • M Kobayashi, H Nikami, M Morimatsu, M Saito NEUROSCIENCE LETTERS 213 (2) 103 -106 1996年08月 [査読無し][通常論文]
     
    The localization of glucose transporters (GLUTs) was examined in various regions of the rat brain. The mRNA of GLUT1 and GLUT3 were found ubiquitously in every brain region (cortex, hippocampus, midbrain, striatum, hypothalamus, medulla oblongata and cerebellum). The mRNA and protein of GLUT4, an insulin-regulatable glucose transporter in peripheral tissues, were also identified, particularly abundantly in the cerebellum. In situ hybridization analysis revealed that GLUT4 mRNA was present in some discrete cells, such as Purkinje cells in the cerebellum, the vestibular nucleus in the medulla oblongata and also in ependymal cells along the cerebral ventricles. The GLUT4 mRNA level in the cerebellum changed little in fasted or experimentally induced diabetic rats while those in adipose tissues decreased much. The results suggest that insulin-sensitive glucose uptake may occur in some specific cells of the brain but is regulated in a different manner from those in peripheral cells.
  • K Mominoki, H Tsuruga, M Morimatsu, M Saito COMPARATIVE BIOCHEMISTRY AND PHYSIOLOGY A-PHYSIOLOGY 114 (4) 349 -353 1996年08月 [査読無し][通常論文]
     
    Haptoglobin (Hp), a hemoglobin-binding protein, is known as an acute phase protein increasing in blood during inflammation in most mammals. On the basis of our previous studies on purification and characterization of bear Hp (Comp. Biochem. Physiol. 110B, 785-789, 1995), in this study, we developed an immunoassay method to measure serum Hp level in bear, and measured the concentration of Hp in blood samples collected from 84 reared and 25 wild brown bears in Hokkaido, Japan. The mean serum Hp concentration was 0.94 +/- 0.25 mg/ml in wild bears, which is nearly equal to those reported in other species. In reared bears, the Hp concentration was apparently higher (3.82 +/- 0.29 mg/ml), although total protein and albumin concentrations were nearly equal in the two groups. A significant seasonal variation of serum Hp, low in spring and high in autumn and winter, was found in reared bears. Possible factors participating in the seasonal variation were discussed with special references to hibernation.
  • 岡田 尚子, 森松 正美, 木村 和弘, 斉藤 昌之 日本分子生物学会年会プログラム・講演要旨集 19 (0) 1996年08月01日 [査読無し][通常論文]
  • Fujino R-S, Tanaka K, Morimatsu M, Tamura K, Kogo H, Hara T Mol Endocrinol. 20 (4) 904 -915 1996年 [査読無し][通常論文]
  • 斉藤昌之, 森松正美, 阿部啓之 食肉に関する助成研究調査成果報告書 13(1994) 160 -166 1995年12月 [査読無し][通常論文]
  • 森松正美, 中川紀子, 川崎昌美, 斉藤昌之 北海道獣医師会雑誌 39 (9) 173 -177 1995年09月 [査読無し][通常論文]
  • K MOMINOKI, N NAKAGAWATOSA, M MORIMATSU, B SYUTO, M SAITO COMPARATIVE BIOCHEMISTRY AND PHYSIOLOGY B-BIOCHEMISTRY & MOLECULAR BIOLOGY 110 (4) 785 -789 1995年04月 [査読無し][通常論文]
     
    Haptoglobin (Hp), a hemoglobin-binding protein in plasma, consists of alpha and beta subunits and has a tetra-chain arrangement (beta-alpha-alpha-beta) connected by disulfide bridges in most mammals so far examined, Dog Hp has been reported to be unique compared with other Hps in respect that (1) the two alpha beta units are joined by a non-covalent interaction rather than a disulfide bridge and (2) the alpha chain has an oligosaccharide-binding sequence (Asn-X-Ser/Thr) and is glycosylated, To determine whether the unique structures of dog Hp are common in the Carnivora, we purified Hps from sera of bear and cat, and analyzed their subunit structure and partial amino acid sequences, The analyses by gel filtration and sodium dodecyl sulfate-polyacrylamide gel electrophoresis, under both reducing and non-reducing conditions, revealed that bear and cat Hps have similar subunit arrangements to dog Hp, suggesting the absence of a disulfide bridge between two a chains, This was confirmed by amino acid sequence analysis of the alpha chains: that is, Cys(15) participating in the inter-a chain disulfide bridge was replaced by Val in bear or Leu in cat and dog, Thus, the unique subunit arrangement of Hp reported in dog may be common in the Carnivora, In contrast to dog Hp, however, alpha chains of bear and cat Hps were found not to have the typical oligosaccharide binding sequence on their alpha chains and were not glycosylated.
  • 中尾敦子, 四谷芳, 森松正美, 岡田啓司, 稲波修, 首藤文栄 家畜生化学 32 (1) 31 -36 1995年03月 [査読無し][通常論文]
  • 塩川嗣章, 森松正美, 稲波修, 岡田啓司, 志賀あき郎, 首藤文栄 家畜生化学 32 (1) 37 -41 1995年03月 [査読無し][通常論文]
  • Biomedical Research 16 (5) 353 1995年 [査読無し][通常論文]
  • NAGASE, I, N SASAKI, K TSUKAZAKI, T YOSHIDA, M MORIMATSU, M SAITO JAPANESE JOURNAL OF VETERINARY RESEARCH 42 (3-4) 137 -146 1994年12月 [査読無し][通常論文]
     
    When mammals are exposed to a cold environment for a long time, the capacity of nonshivering thermogenesis by brown adipose tissue (BAT) increases in association with the increased expression of some specific proteins and tissue hyperplasia, which are totally dependent on sympathetic innervation to this tissue. To clarify roles of the beta-adrenergic mechanism in BAT hyperplasia, the effects of chronic administration of various beta-adrenergic agonists on BAT were examined in rats, especially focusing on some agonists to the beta 3-adrenoceptor which is present specifically in adipocytes. Chronic administration of noradrenaline or isoproterenol for 7-10 days produced a marked increase in the tissue contents of DNA, total protein, mitochondrial uncoupling protein, and insulin-regulatable glucose transporter protein. The trophic effects of noradrenaline and isoproterenol were mimicked by chronic administration of beta 3-adrenergic agonists, such as CL316,243, BRL 26830A, and ICI D7114. These results suggest that the beta 3-adrenoceptor plays important roles for hyperplasia of BAT, and thereby increasing in the capacity of thermogenesis.
  • 中川紀子, 森松正美, もみの木勝巳, 中辻浩喜, 首藤文栄, 斉藤昌之 家畜生化学研究会報 30 (2) 51 -56 1993年10月 [査読無し][通常論文]
  • 中川紀子, 森松正美, 川崎昌美, 中辻浩喜, 首藤文栄, 斉藤昌之 日本獣医学会学術集会講演要旨集 116th 77 1993年08月 [査読無し][通常論文]
  • S YAMAMOTO, K TAGATA, Y ISHIKAWA, H SANTSUKA, M YAMADA, M MORIMATSU, M NAIKI VETERINARY IMMUNOLOGY AND IMMUNOPATHOLOGY 36 (3) 257 -264 1993年04月 [査読無し][通常論文]
     
    This paper describes the avidity of IgG antibody used for preparation of latex sensitized with IgG antibody (IgG-sensitized latex) and the agglutinability of IgG-sensitized latex in slide reversed passive latex agglutination (RPLA). Using immunodiffusion techniques, it was found that anti-canine C-reactive protein (CRP) sera from four rabbits immunized with canine CRP had the same antibody titers. However, the antibodies had different levels of avidity. When lattices were sensitized under the same condition with the IgG antibodies of different avidity levels separated from the above-mentioned antisera using Protein A and canine CRP-Sepharose 4B immunosorbent, these demonstrated different patterns of agglutinability in slide RPLA. The latex sensitized with IgG antibody of higher avidity demonstrated a stronger agglutinability.
  • S YAMAMOTO, N ABE, H SANTSUKA, T SHIDA, K KISHIDA, S KUWAJIMA, M YAMADA, M MORIMATSU, M NAIKI VETERINARY IMMUNOLOGY AND IMMUNOPATHOLOGY 36 (3) 293 -301 1993年04月 [査読無し][通常論文]
     
    The canine C-reactive protein (CRP) fraction isolated from canine acute-phase serum on a phosphorylcholine-Sepharose 4B column was further subjected to Sephacryl S-300 gel filtration. A canine CRP fraction not containing IgM was then obtained. The antisera, obtained after several immunizations with this canine CRP fraction, contained nonspecific antibodies that reacted with albumin, transferrin and IgG in addition to CRP. This antiserum could be easily changed to monospecific canine CRP serum, when it was subjected to absorption for only 15 min using glutaraldehyde-insolubilized normal canine serum protein containing 3.5 mug ml-1 of CRP. Pure canine CRP was isolated with a recovery rate of 95% from canine acute-phase serum by affinity chromatography using specific anti-canine CRP antibody.
  • 中川紀子, 森松正美, 中辻浩喜, 首藤文栄, 斉藤昌之 日本獣医学会学術集会講演要旨集 115th 110 1993年04月 [査読無し][通常論文]
  • 森松正美, 中川紀子, もみの木勝巳, 首藤文栄, 内貴正治, 斉藤昌之 家畜生化学研究会報 30 (1) 23 -30 1993年03月 [査読無し][通常論文]
  • N TOSA, M MORIMATSU, M NAKAGAWA, F MIYOSHI, E UCHIDA, M NIIYAMA, B SYUTO, M SAITO JOURNAL OF VETERINARY MEDICAL SCIENCE 55 (1) 27 -31 1993年02月 [査読無し][通常論文]
     
    Polyacrylamide gel electrophoretic analysis of canine serum protein has revealed that the administration of anthelmintics elicits an increase in a certain serum protein. This protein, named PT60, was partially purified by ammonium sulfate fractionation and preparative electrophoresis. The purified PT60 gave a single band with the molecular size of 53 kDa in sodium dodecyl sulfate-polyacrylamide gel electrophoresis under non-reducing conditions. After reduction with 2-mercaptoethanol, two bands appeared at 35 kDa and 17 kDa, indicating that PT60 consists of two subunits which are linked with each other by disulfide bonds. PT60 had the capacity to bind to hemoglobin. In an immunodiffusion test, an antiserum against PT60 cross-reacted with canine haptoglobin (Hp). N-terminal amino acid sequences of two PT60 subunits were identical to those of alpha and beta subunits of canine Hp, respectively. Thus, PT60 was identified as Hp.
  • Y NAKAJIMA, E MOMOTANI, T MURAKAMI, Y ISHIKAWA, M MORIMATSU, M SAITO, H SUZUKI, K YASUKAWA VETERINARY IMMUNOLOGY AND IMMUNOPATHOLOGY 35 (3-4) 385 -391 1993年01月 [査読無し][通常論文]
     
    Interleukin-6 (IL-6) is a major inducer of acute phase proteins in human and murine species. However, the effects of IL-6 have not yet been investigated in cattle. Following continuous infusion of recombinant human IL,6, serum concentrations of bovine haptoglobin and fibrinogen increased in a manner similar to those in cattle with acute phase reaction. In contrast, C-reactive protein and alpha1-acid glycoprotein, as well as the other hematologic parameters, did not change significantly. Intravenous administration of recombinant human IL-6 resulted in only a mild and transient increase of bovine haptoglobin. These results suggest that the regulation of acute phase protein production in cattle is similar, but not identical, to that observed in human and murine species.
  • M MORIMATSU, M SARIKAPUTI, B SYUTO, M SAITO, S YAMAMOTO, M NAIKI VETERINARY IMMUNOLOGY AND IMMUNOPATHOLOGY 33 (4) 365 -372 1992年09月 [査読無し][通常論文]
     
    Using purified bovine haptoglobin (Hp) and specific antisera, a single radial immunodiffusion (SRID) assay method has been developed to measure the serum Hp level in cattle. Bovine Hp is a highly polymerized protein showing heterogeneous molecular forms in serum. After treatment with cysteine or glutathione, Hp was partially reduced to a homogeneous form, suitable for SRID assay. This method gives values comparable to those obtained by hemoglobin-binding capacity assay, and has the advantage of being simple and convenient. Although serum Hp was not detectable in healthy cattle, it was found more than 50-fold after invasive surgery, indicating that Hp is a characteristic acute-phase protein in cattle.
  • 森松正美, 土佐紀子, 内貴正治, 斉藤昌之 家畜生化学研究会報 (29) 61 -68 1992年09月 [査読無し][通常論文]
  • S YAMAMOTO, K TAGATA, Y ISHIKAWA, H FUJISE, H NAGAHATA, M YAMADA, T SAKANO, M MORIMATSU, M NAIKI VETERINARY RESEARCH COMMUNICATIONS 16 (4) 265 -272 1992年08月 [査読無し][通常論文]
     
    A method is described for preparing latex particles sensitized with IgG antibody (IgG-sensitized latex) applicable to the slide reversed passive agglutination (RPLA) test. Soap-free latex (latex) was sensitized with IgG which had been isolated from rabbit anti-bovine lactoferrin serum using protein A Sepharose CL 4B. Unadsorbed protein-binding sites on the surface of latex were blocked with bovine serum albumin (BSA). IgG-sensitized latex that gave better agglutination in RPLA could be selectively obtained by centrifugation at 19 900g for 15 min in 0.01 mol/L glycine buffer (pH 7.3; specific gravity 1.042) containing 3% NaCl, 5% saccharose and 2% choline chloride. By dispersing this IgG-sensitized latex in 0.01 mol/L glycine buffer (pH 7.3) containing 1-2% BSA, a uniformly suspended, highly reactive, readily agglutinable preparation was obtained.
  • M SARIKAPUTI, M MORIMATSU, S YAMAMOTO, B SYUTO, M SAITO, M NAIKI JAPANESE JOURNAL OF VETERINARY RESEARCH 40 (1) 1 -12 1992年05月 [査読無し][通常論文]
     
    A semi-quantitative latex agglutination test for bovine serum CRP levels has been established by mixing diluted serum (or diluted standard serum) with a 1% latex suspension containing 0.489-mu-m latex particles coated with affinity-purified antibody at a ratio of 20-mu-g/mg latex. The agglutination was performed on a glass slide in a moist chamber at room temperature with 45 min. incubation. This test is reliable, reproducible and the results correlate with those of the single radial immunodiffusion (SRID) test. The effect of low temperature storage on CRP concentration revealed a 30% degradation of CRP during 2 years storage at 4-degrees-C. The possible role of EDTA addition to prevent a decrease in serum CRP concentration by freezing and thawing is also discussed.
  • A WATANABE, M MORIMATSU, K YOSHIMATSU, S YAMAMOTO, A TERAO, K TSUKAZAKI, M SAITO, M NAIKI JOURNAL OF SMALL ANIMAL PRACTICE 33 (2) 71 -77 1992年02月 [査読無し][通常論文]
     
    C-reactive protein (CRP) was isolated from sera from healthy cats by calcium-dependent affinity chromatography on phosphorylcholine derivatives of bovine albumin-coupled Toyopearl, followed by an anion-exchange chromatography using DEAE cellulose. It was identified as CRP by its immunochemical cross reactivity with human CRP. The molecular weight of cat CRP was approximately 100 kilodaltons (kDa) and composed of two glycosylated subunits (23 kDa) and three non-glycosylated subunits (20 kDa) with non-covalent association. Under electron microscopic examination, cat CRP had a pentameric disc-like configuration which is characteristic of CRP. Immunoelectrophoresis and isoelectric focusing showed that cat CRP is an acidic alpha-1-globulin (pI 4.1 to 4.3). Serum concentrations of CRP in cats and kittens were measured by single radial immunodiffusion. In 12 healthy cats from various sources, values ranged from 38 to 168-mu-g/ml. In kittens, serum CRP levels also showed a wide distribution, 81 per cent of them were less than 40-mu-g/ml.
  • S YAMAMOTO, K TAGATA, H NAGAHATA, Y ISHIKAWA, M MORIMATSU, M NAIKI VETERINARY IMMUNOLOGY AND IMMUNOPATHOLOGY 30 (4) 329 -339 1992年01月 [査読無し][通常論文]
     
    C-reactive protein (CRP) was isolated from the acute phase serum of dogs subjected to surgical stimulation. Its properties were characterized. Canine CRP was isolated by ion-exchange chromatography using DEAE-Sephaeel and DEAE-Sephadex A-50 and affinity chromatography using protein A-Sepharose CL 4B in combination with agar-block electrophoresis. In immunoelectrophoresis, canine CRP had the same gamma-mobility as human gamma-type CRP. The molecular weight of purifined canine CRP was estimated by gel filtration and sodium dodecyl sulfate polyacrylamide gel electrophoresis to be approximately 157 000 and 155 000 respectively. This CRP was a thermolabile protein which completely lost its antigenicity by heating at 70-degrees-C for 15 min. The serum concentration of CRP in normal beagle dogs ranged from 0.198 to 0.826-mu-g ml-1 (0.486 +/- 0.170-mu-g ml-1). The concentration was acutely increased by surgery as it was in man and was rapidly decreased with convalescence. Dogs can be a useful animal model for investigation of the mechanism of CRP production and the function of CRP.
  • M MORIMATSU, A WATANABE, K YOSHIMATSU, T FUJINAGA, M OKUBO, M NAIKI JOURNAL OF DAIRY RESEARCH 58 (3) 257 -261 1991年08月 [査読無し][通常論文]
     
    C-reactive protein (CRP) and serum amyloid P component (SAP), which are known to increase in sera from humans and many other animals with acute inflammation caused by infection, toxic drug administration or injury, were previously purified from bovine serum. These serum levels were determined by enzyme-linked immunosorbent assay (ELISA) using specific antiserum to bovine CRP or SAP which was prepared by immunizing rabbits and goats with each purified protein. Among 68 healthy Holstein cows, 45 non-lactating cows had levels of CRP and SAP of 20.6 +/- 1.4 and 27.6 +/- 1.3-mu-g/ml respectively; 23 lactating cows had higher levels of CRP and SAP (76.0 +/- 13.6 and 38.3 +/- 5.5-mu-g/ml respectively). In the latter group, there was a high correlation between milk yield and serum CRP levels (P < 0.001). From these observations, it was assumed that lactation might stimulate CRP synthesis rather than SAP synthesis in bovine liver as an acute phase reaction, and that CRP might be called a lactation-associated protein.
  • M MORIMATSU, B SYUTO, N SHIMADA, T FUJINAGA, S YAMAMOTO, M SAITO, M NAIKI JOURNAL OF BIOLOGICAL CHEMISTRY 266 (18) 11833 -11837 1991年06月 [査読無し][通常論文]
     
    A macromolecular hemoglobin-binding protein, which was not detectable in normal bovine sera but appeared during acute phase inflammation, was purified, characterized, and designated as bovine haptoglobin (Hp). The purified protein had a molecular mass of 1,000-2,000 kDa, and was composed of two kinds of peptides, a 20-kDa peptide (alpha-chain) and a 35-kDa glycopeptide (beta-chain) linked by disulfide bonds. Amino acid composition and N-terminal sequence analyses revealed that both peptides were homologous to each counterpart of human Hp. Studies using some reducing reagents proved that highly polymerized Hp in serum was composed of 2-20 polymerized forms of alpha-2-beta-2 tetramer. Hp could bind one molecule of hemoglobin/alpha-2-beta-2 unit. Hp with smaller sizes obtained from native Hp by partial reduction with cysteine showed almost the same Hb-binding capacity.
  • 森松正美, SARIKAPUTI M, 島田尚美, 首藤文栄, 斎藤昌之, 藤永徹, 中辻浩喜, 大久保正彦, 内貴正治 日本獣医学会学術集会講演要旨集 111th 302 1991年03月 [査読無し][通常論文]
  • M SARIKAPUTI, M MORIMATSU, B SYUTO, M SAITO, M NAIKI INTERNATIONAL JOURNAL OF BIOCHEMISTRY 23 (10) 1137 -1142 1991年 [査読無し][通常論文]
     
    1. A new purification procedure was started with salting-out fractionation of serum proteins at 45-75% saturated ammonium sulfate concentration, followed by HE agarose affinity chromatography by which calcium-dependently bound CRP and SAP were purely eluted with EDTA-containing buffer. 2. Pure CRP and SAP were finally separated by DEAE-5PW HPLC. 3. This procedure gave recovery of 15 and 26%, and fold purification of 2650 and 2400 for CRP and SAP. respectively. 4. Each subunit of CRP and SAP had one intrasubunit disulfide bond, determined by reduction and carboxymethylation.
  • 森松正美, SARIKAPUTI M, 内貴正治 家畜生化学研究会報 (25) 29 -36 1990年11月 [査読無し][通常論文]
  • M MORIMATSU, H SAKAI, K YOSHIMATSU, O MINOWA, S YAMAMOTO, K YATOMI, T FUJINAGA, M NAIKI JAPANESE JOURNAL OF VETERINARY SCIENCE 51 (4) 723 -732 1989年08月 [査読無し][通常論文]

特許

共同研究・競争的資金等の研究課題

  • 腫瘍におけるBRCA2の変異と結合分子群の機能解明を目的とした新規モデルの作出
    日本学術振興会:科学研究費補助金(基盤研究(B))
    研究期間 : 2018年 -2021年 
    代表者 : 森松 正美
  • 組換え酵素Rad51と癌抑制タンパク質BRCA2の結合の種間比較に基づく分子創薬
    日本学術振興会:科学研究費補助金(基盤研究(C))
    研究期間 : 2014年 -2016年 
    代表者 : 森松 正美
  • 文部科学省:科学研究費補助金(基盤研究(C))
    研究期間 : 2011年 -2013年 
    代表者 : 森松 正美, 落合 和彦
     
    本研究の目的は、1.イヌの乳腺腫瘍の発症機構について、遺伝性乳癌原因遺伝子BRCA2の変異を基点とするゲノム不安定化に焦点をあてて、腫瘍細胞において特定の遺伝子にコピー数の変化が認められるかどうかを調べること、2.変異が予想されるc-MYC(MYC)等の癌遺伝子やいくつかの癌抑制遺伝子、ゲノム安定化遺伝子の機能解析を通じてイヌの乳腺腫瘍発症機構の解明をめざすこと、とした。イヌのMYCについて、cDNAクローニングを行って全長配列を獲得し、レトロウイルスベクターに組み換えて遺伝子導入による解析方法を確立した。さらに、MYC等を対象にイヌ特有の機能に関与する構造の予測を行うため、イヌとヒトのMYCでアミノ酸配列を比較するとともに、SWISS-PROTやUCSF-chimera等のアプリケーションを利用した構造モデリングを行って解析した。それにより、イヌMYCの立体構造上の特徴を明らかにし、特に分子表面に存在するアミノ酸の位置情報から、イヌc-MYCで特有の機能を発現する可能性のあるアミノ酸の候補を推定した。液体窒素に凍結保存してあった乳腺腫瘍細胞株について、それらの保存状況を確認するためにすべての株を解凍、培養したところ、いずれも良好に増殖したので、再度、新鮮な凍結ストックを作製した。これらを研究分担者との間でやりとりし、保存した。さらに、いくつかの乳腺腫瘍組織や正常乳腺組織といった材料を分担者との間で共有し、これらのいくつかについては、ゲノムDNAやRNAの分離を行った。ゲノムコピー数の変化を調べるためにリアルタイムPCRの装置を購入するとともに、単一コピー遺伝子をイヌゲノムのデータベースで検索し、ヒポキサンチン-グアニンホスホリボシルトランスフェラーゼ(hprt)を候補としてリアルタイムPCRによる測定条件を検討した。
  • 文部科学省:科学研究費補助金(基盤研究(B))
    研究期間 : 2007年 -2009年 
    代表者 : 森松 正美, 富岡 幸子, 重茂 克彦, 加納 聖, 前田 貞俊
     
    NFkB抑制核タンパク質MAILを欠損する新規マウスを用いて新しいアトピー性皮膚炎モデル動物として確立するとともに、これを材料としてアトピー性皮膚炎の発症機構を解析した。新規MAIL欠損マウスにおける皮膚炎病態を解析してその発症メカニズムを考察し、MAILと機能的に相互作用する遺伝子を検索して胎生期致死や皮膚炎の原因を探った。また、MAIL欠損マウスにおける胎生期致死の問題を克服してマウスを作出する効率を上昇させることを試みた。
  • 文部科学省:科学研究費補助金(萌芽研究)
    研究期間 : 2005年 -2006年 
    代表者 : 森松 正美
     
    研究代表者らは炎症刺激で誘導される遺伝子をスクリーニングする過程で機能未知の遺伝子を同定し、MAILと命名した。ジーンターゲティングによりこの遺伝子を破壊したマウスを作製したところ、ホモ型欠損マウスの顔面を中心にアトピー性皮膚炎様の病変が認められた。本研究の第1の目的は、申請者が開発したMAIL破壊マウスを用いてMAILの機能を解析するとともに、これを疾患モデル動物として確立することである。第2の目的は、このモデルマウスの原因遺伝子が、ヒトでもアトピー性皮膚炎と関係があるか否かを探ることである。1.マウスの交配と維持:MAILのホモ欠損型が胎生期致死となることが原因でマウスの数が不足している点が研究を進める上で支障となっていた。交配規模を拡大し,産子の遺伝型をPCRにより判定して欠損マウスを得た。2.マウスの病態解析:MAILがケラチノサイトの初代培養や株化細胞で無刺激の状態でも発現していることを明らかにした。この発現には転写因子NF-kappaBが重要な役割を果たすことがわかった。この発現は,炎症性サイトカインによって上昇することが判明し,MAILが皮膚炎病態に関わっていることが示唆された。さらに皮膚炎を発症しているMAIL破壊マウスの皮膚組織を検索したところ,アトピー性皮膚炎に関与すると考えられているTh2系サイトカインの産生が認められた。3.ヒトの遺伝子解析:ヒトの皮膚生検試料におけるMAILの発現をin situ hybridizationで検索したが,はっきりした発現は認められなかった。今後,検出感度を上昇させるなどの方法の改善が望まれる。さらに,ヒトMAIL遺伝子の多型と皮膚炎との関係を調べるために,MAILのcDNAをRT-PCRで増幅するためのプライマーを設計し,疫学的解析を実施するための基盤を築いた。
  • 文部科学省:科学研究費補助金(基盤研究(B))
    研究期間 : 2003年 -2005年 
    代表者 : 森松 正美, 吉松 組子, 山本 欣郎
     
    自然免疫(innate immunity)は、各種病原体に共通して存在する構造物(リポ多糖体(LPS)など)を認識する生体防御反応であり、これらの構造物がToll様受容体(TLR)を介してNF-kBを活性化する細胞内情報伝達系により支えられている。最近、研究代表者らはLPS刺激によって発現誘導される新規分子MAILを発見した。この分子を中心に研究を実施した。1.MAIL破壊マウスの解析MAIL破壊マウスの約9割は胎生期に死亡することが判明した。胎子を摘出して調べたところ、肝臓の形成不全が認められ、これが胎生期致死の主な要因であると考えられた。MAIL破壊マウスの約1割は出生したが、これらはすべてアトピー性皮膚炎様の病態を示した。2.MAIL遺伝子発現調節の解析MAIL遺伝子のプロモータをクローニングしてそれより各種変異体を作製して機能解析を行った。NF-kBがMAIL遺伝子の発現を促進し、特に転写開始点近傍に存在するkBモチーフ配列が重要であることが判明した。MAIL遺伝子はNF-kBによって発現誘導され、転写翻訳によりできたMAILタンパク質はNF-kBをフィードバック制御することが示唆された。3.BRCA2遺伝子産物の解析NF-kB標的分子のひとつであるBRCA2は、相同組換えによるDNA損傷修復や細胞周期の調節を通じて細胞増殖や胎子発生に重要な役割を果たしている。BRCA2がC末端で組換え酵素Rad51と結合することを確認した。イヌのBRCA2の核移行シグナル(NLS)に多型を発見した。このNLS多型は、核移行効率や腫瘍発症頻度と相関している可能性が示された。
  • 文部科学省:科学研究費補助金(萌芽研究)
    研究期間 : 2003年 -2004年 
    代表者 : 森松 正美
     
    最近、ある機能未知の遺伝子を同定して機能解析のためにジーンターゲティングによりこの遺伝子を破壊したマウスを作製したところ、ホモ型欠損マウス(仮にAllergic Dermatitis、ALDマウスと呼ぶ)の顔面を中心にアトピー性皮膚炎様の病変が認められた。本研究の目的は、外見的に皮膚炎を認めるALDマウスについてその病態を解析し、これを疾患モデル動物として確立することである。1.マウスの交配と維持:研究の途中で岩手大学から北海道大学に拠点を移したが、いずれの場所でも問題なく微生物学的に清浄な環境でマウスを飼育した。繁殖規模を拡大すると同時に近交系への戻し交配を進めた。また、万が一の事故に備えて受精卵を凍結保存させた。2.病態の解析:皮膚病変部位の組織学的検索を行い、白血球系細胞の細胞浸潤による炎症反応を認めた。皮膚病変部では、ケモカインであるTARCとeotaxin、ケモカイン受容体であるCCR3の増加が認められた。これらはヒトのアトピー性皮膚炎とよく似た所見であり、MAIL欠損マウスが良いモデルとなる可能性を示すものである。正常マウスでのMAILの発現を免疫組織化学およびリアルタイムPCRで検索したところ、表皮ケラチノサイトでの発現を認めた。この結果は、ケラチノサイトのMAILが皮膚の恒常性に重要であり、これが欠損すると皮膚炎を発症することを示唆している。マウスの系統によって免疫応答に差があることが報告されているので、Th1優勢のC57BL/6、およびTh2優勢のBALB/cへの戻し交配を進めたが、いずれの系統でもMAIL欠損マウスで皮膚炎が認められた。以上より、本遺伝子操作マウスを疾患モデル動物として応用するための基盤の整備を進めることができた。
  • 文部科学省:科学研究費補助金(基盤研究(B))
    研究期間 : 1999年 -2001年 
    代表者 : 森松 正美, 冨澤 伸行, 津田 修治, 首藤 文榮, 西森 克彦
     
    本研究の目的は,Brca2を中心とした癌関連遺伝子について、生理機能を分子,細胞,個体レベルで解明して,乳癌の診断,予防,治療技術の発展に貢献することである。特にBrca2とRad51との相互作用およびその意義の解明、Brca2解析の獣医学領域への展開,および新規lkB分子MAILの生理機能の解明を目標として研究した。1.イヌBrca2のクローニングと乳腺腫瘍症例における解析イヌBrca2のcDNAめ全長をクローニングして塩基配列を決定した。また、症例におけるBrca2の変異解析を行うため,10kbにおよぶcDNAの全長を安定して増幅させるPCR法を開発した。さらに、Brca2遺伝子で最大であるエキソン11について、in vitro転写翻訳系を用いたProtein trancation testの検査系の開発に成功し、症例における変異解析を行った。2.ネコのBrca2のクローニングと発現解析ネコBrca2のcDNAの全長をクローニングして塩基配列を決定し、データベースに登録した。また、RT-PCR法により組織分布を調べ、乳腺等の広範な組織での発現を確認した。3.Brca2とRad51との会合イヌのBrca2とRad51について行った。イヌBrca2はそのC末端領域でRad51と会合することが判明したことに加え、エキソン11にコードされる中央部に転写活性化能をもつ領域が存在することが示唆された。4.新規lkB分子MAILの機能解析新規lkB分子MAILのゲノムをクローニングしてその構造を明らかにした。また、この分子の発現調節機構を解析した。
  • 文部科学省:科学研究費補助金(基盤研究(B))
    研究期間 : 1999年 -2001年 
    代表者 : 森松 正美, 吉松 組子, 小川 和重, 首藤 文榮, 吉村 佳典, 三好 一郎
     
    本研究の目的は,マウスのジーンターゲティングにおいて,ノックインにより哺乳動物に存在しない特異的な目印(エピトープタグ)を標的分子に導入し,生体反応におけるその動向を抗エピトープタグ抗体で調べる方法を開発するための基礎研究を行うことである。それに向けて、以下のごとく数種類のモデル遺伝子を標的とすべく基礎研究を行った。また、導入するエピトープタグについての解析を行った。1.内毒素誘導性新規遺伝子(MAIL)を標的とした検討MAILは、内毒素による顕著な発現調節を受け`遺伝子産物は速やかに核に移行するため、標的に適している。この遺伝子のゲノムDNAを分離し,塩基配列を決定してその構造を明らかにした。これを用いてジーンターゲティングを行った。2.キチナーゼ遺伝子を標的とした検討キチナーゼは特徴的な組織特異的発現調節を示す遺伝子であるため、これを標的とすることを試みた。この遺伝子のクローニング、構造解所、および発現解析を行い、ターゲティングを実施するための基盤を築いた。3.他の標的遺伝子候補の検索上記の他誘導性遺伝子であるB-13や、組織特異的遺伝子であるGlucose transporter、Leptin等、疾病に関わると考えられる遺伝子を選んで標的の候補として検索した。4.エピトープタグの解析新規エピトープの候補であるハンタウイルス由来タンパク質について、抗体の反応性や、細胞に与える影響の解析を行った。5.新規ノックイン法の開発に関する基礎的研究二段階置換法によるジーンターゲティングを計画した。ES細胞にベクターを導入して第一段階目の置換を施した後、第二段階目の置換を試みたが、目的とする相同組換え体を得ることができなかった。その原因の解明と問題の克服は今後の課題である。
  • 文部科学省:科学研究費補助金(萌芽的研究)
    研究期間 : 1999年 -1999年 
    代表者 : 森松 正美, 冨澤 伸行
     
    脳の多彩な精神活動は,神経細胞によるエネルギー代謝とそれにともなう酸素の消費によって支えられている.脳活動が変動すると脳の血流量や血液内ヘモグロビン(Hb)の酸素化度が変化するため,これらを測定することは,動物の精神活動を探る糸口となることが期待される.本研究ではイヌの精神活動の分析に,近赤外分光法による脳Hb酸素化度の測定が利用可能か否かを検討した.1.麻酔下のイヌによる脳血流測定の基礎的解析脳血流量を測定する際に筋血流量の影響が障害となる可能性があるため,麻酔下のイヌを用いて検討した.プローブを外科的処置により露出させた頭蓋骨表面に装着した場合と,皮膚表面からあてた場合とで測定値を比較し,測定における筋血流量の影響を調べた.呼気中酸素分圧や頚動脈の開閉等を行って検討した.プローブを頭部正中に装着すれば,筋血流量の変化の影響を抑えて脳血流量を測定できる可能性が高いことが示された.さらに,脳特異的血流変化が検出できるかどうかを検討した.脳は二酸化炭素に感受性が高いため,これを吸入させてその影響を調べたところ,脳に特徴的な変化が観察された.2.無侵襲・無麻酔・無拘束のイヌにおける脳血流測定イヌの情動変化を近赤外分光法で捉えることができるか否かを検討するため,イヌにとって望まれる状況と望まれない状況を設定し脳血流の変化を観察した.餌を与えて喜ばせた場合,Hb酸素化度の低下が認められた.別離不安の見られるイヌについて,飼い主がイヌから離れて隠れたところ,Hb酸素化度の一過性の上昇とそれに続く比較的長時間の低下が観察された
  • 文部科学省:科学研究費補助金(奨励研究(A))
    研究期間 : 1997年 -1998年 
    代表者 : 森松 正美
     
    生体が、感染、炎症、ストレス等による侵襲を受けると、肝臓から血液中に特定の一群の生体防御関連タンパク質、すなわち急性相タンパク質が分泌される。この研究の目的は、急性相タンパク質の測定による動物の健康管理を新規家畜や野生動物に応用することにより、基礎研究及び関連産業の発展への貢献をはかることである。本研究では、クマおよびシカの急性相タンパク質に焦点をあてて、免疫測定法を確立するとともに飼育個体や野生個体における変動を解析して,これらの新規家畜や野生動物の健康管浬に利用することを計画した。平成10年度は,急性相タンパク質測定法の確立,試料の収集と測定の実施を中心に以下のごとく検討した.クマの急性相タンパク質:クマの急性相タンパク質のうち,ハプトグロビン(Hp),α2-マクログロブリン(α2M),α1-アンチトリプシン(α1-AT)およびC-反応性タンパク質(CRP)について単純放射免疫拡散法(SRID法)あるいはウエスタンプロット法による測定を行った。野生個体の捕獲例では,外見的に正常な個体に比べ外傷を負った個体で顕著なハブトグロビンの増加が認められた。健康な飼育個体で冬眠との関係に着目して調べたところ,冬眠に伴ってHpとα2Mが増加したが,α1-ATとCRPは変化しなかった。クマの健康管理に急性相タンパク質の測定が有用であると思われるが,冬眠など生理的な変化の影響について留意する必要がある。シカの急性相タンパク質:シカの血清をポリアクリルアミドゲル電気泳動により分析したところ,健康状態によって顕著に変化するタンパク質を見いだした。これを分離して分析し,シカHpと同定した。クマの例と同様に特異的な免疫測定法を確立して調べた結果,シカでHpが炎症性の刺激により急激に増加する良いマーカーとなりうろことが示された。
  • 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 : 森松 正美
     
    急性相タンパク質は、炎症性サイトカインが肝細胞を刺激することにより合成・分泌される血清タンパク質の総称であり、生体の防御・免疫機能に関与している。申請者はこれまでウシの急性相タンパク質の血中レベルの変動等について研究を進め、ラット等の実験動物やヒトでこれまで報告されてきたものとは異なるいくつかの特徴的な現象を見いだしてきた。本研究ではこれを更に発展させ,この現象の根幹となる遺伝子発現調節機構をin vitroの肝細胞培養実験系を用いて解明することを試み,いくつかの新知見を得ることができた。[1]ウシ急性相タンパク質の型別(I型とII型)の検討近年、ラットやヒト由来の材料を用いた研究結果に基づき、急性相タンパク質は作用するサイトカインの種類によりI型(インターロイキン(IL)-1と腫瘍壊死因子(TNF)が例外無くともに作用し,IL-6も効果があることがある)とII型(IL-6でのみ誘導される)に分類されるのが一般的となってきた。申請者は,これがウシの急性相タンパク質にもあてはまるか否かを明らかにするため,ウシのハプトグロビン(Hp),C-反応性タンパク質(CRP),alpha 1-酸性糖タンパク質(al-AG),alpha 2-マクログロブリン(a2-MG)のcDNAをそれぞれクローニングしてこれをプローブとし,サイトカインで刺激したウシの初代培養肝細胞におけるこれら急性相タンパク質のmRNA発現量をNorthernハイブリダイゼーション法で解析した。その結果,ラットやヒトでI型急性相タンパク質と考えられているHp,CRP,al-AGがいずれもTNFとIL-6で誘導されるがIL-1では誘導されないことを明らかにした.すなわち,これまでラットやヒトで提唱されているI型とII型の分類はウシにあてはまらないことを発見し,ウシに特有の急性相タンパク質合成調節機構が存在することが示唆された。[2]ウシ特有の上記機構解明の試み上記(1)についてHp,CRP,al-AGがラットやヒトでI型であることを考えると、ウシでもこれらが基本的にはI型類似だが、何らかの因子の欠陥によりIL-1に不応答であると予想できる。このウシ肝細胞に欠けている因子を遺伝子相補により単離することを最終目標としてそれに不可欠なウシ肝細胞株の樹立を試みた。ウシ初代培養肝細胞にSV40ウイルス由来の大型T抗原(LT)を導入して不死化することを試みた。哺乳動物細胞発現ベクターあるいはアデノウイルスに組み換えたLTを導入したところ,長期培養に耐える細胞がいくつか得られたが,残念ながらこれらでは急性相タンパク質遺伝子の発現を認めなかった。現在,不死化の方法を変更してさらにウシ肝細胞株樹立を試みている。
  • 文部科学省:科学研究費補助金(奨励研究(A))
    研究期間 : 1994年 -1994年 
    代表者 : 森松 正美
     
    ハプトグロビン(Hp)は,肝臓で合成・分泌される血清タンパク質であり,急性炎症時に血中レベルが上昇して生体の防御・免疫機能に関与する急性相タンパク質のひとつである.本研究では,ウシHpに着目し,その遺伝子進化の背景をさぐるとともに,特徴的な遺伝子転写調節機構について,いくつかの新知見を得ることに成功した.1.ウシHp遺伝子の進化申請者はこれまでの研究成果から,ウシのHpは特徴的な遺伝子重複による分子進化を経て形成されたと予想した.本研究では,ウシゲノムDNAをサザンブロット法で解析して制限酵素地図を作製したところ,ゲノム上に単一コピーで存在することが明らかとなり,重複前の遺伝子は著しく変異したか,あるいはすでに欠失したと予想した.さらに,他の哺乳動物のHpをタンパク質レベルで調べた結果から,ウシで発見した遺伝子重複による分子進化は,広く反芻動物に認められることを明らかにした.なお,当初の実験計画では,ウシHpのゲノムDNAをクローニングして全塩基配列を決定する計画であったが,残念ながらまだクローニングには成功しておらずこの実験は継続中である.2.ウシHp遺伝子の転写調節ウシ肝臓におけるmRNA量を調べたところ,Hpの発現は,正常では極めて低いが炎症性疾患で急激に上昇することを明らかにした.さらに,ウシ初代培養肝細胞実験系で調べたところ,炎症性サイトカインのうち,インターロイキン(IL)-6と腫瘍壊死因子(TNF)が,Hp遺伝子の転写を誘導したが,IL-1は効果がなかった.このことは,これまでに報告された急性相タンパク質では,TNFで誘導されるものは例外なくIL-1でも誘導され,両者が共通の細胞内情報伝達系を活性化するとされてきたことと極めて対照的な新知見である.また,泌乳ホルモンであるプロラクチンがウシHpの遺伝子発現を誘導することを,あらゆる急性相タンパク質の中で初めて発見した.
  • 文部科学省:科学研究費補助金(奨励研究(A))
    研究期間 : 1993年 -1993年 
    代表者 : 森松 正美
     
    急性相タンパク質は、主に肝臓により合成・分泌され、急性炎症時に特徴的な血中動態を示しながら生体の防御・免疫機能に関与している。本研究では、ウシ肝細胞の初代培養系を確立して急性相タンパク質の合成調節機構を細胞・分子レベルで解明するための基礎を築くとともにいくつかの新知見を得ることに成功した。1.ウシ肝細胞の初代培養の確立(1)ウシから肝尾状葉を摘出後、コラゲナーゼを灌流して結合組織を消化し、肝細胞を分離する方法を確立した。肝摘出前にヘパリンをウシに注射して血液凝固を抑制することにより実験の再現性が向上した。また、灌流するコラゲナーゼの濃度は0.05%が、灌流時間は10〜15分間が適当であった。(2)分離した肝細胞を培養することにより、タンパク質合成とグリコゲン分解の活性が上昇した。培養条件はラットのものを応用したが、ラットの場合と同様に、肝特異的代謝活性は培養開始後一週間で再び低下した。2.初代培養肝細胞を用いた急性相タンパク質の合成調節機構の解析(1)まず、ウシの急性炎症で血中に最も急激に上昇するハプトグロビン(Hp)について調べた。肝細胞の培養上清に放出されたHpを特異抗体で免疫沈降して測定した。調べた炎症性サイトカインのうち、インターロイキン6(IL-6)と腫瘍壊死因子(TNF)はHpの合成を誘導したが、IL-1では変化がなかった。特に、TNFが誘導する点は、ラット等とは異なる点であり、in vivoにおける急激な上昇と関係があるものと推察できる。(2)急性炎症のみならず泌乳に関連して変動するC-反応性タンパク質(CRP)について、上記Hpと同様に免疫沈降実験を行った。しかし、現在までのところCRPの検出に成功していない。これについては、検出感度を上昇させるべく特異抗体の再調製を行うとともに、遺伝子発現レベルでの変化の解析を行っているところである。
  • 炎症反応の制御に関わる細胞情報伝達機構
  • 相同組換え反応の分子機構
  • 家畜の急性相タンパク質の合成・分泌調節機構
  • Signal transduction mechanisms in the inflammatory response
  • Molecular mechanisms of homologous recombination.
  • Mechanisms of the synthesis and secretion of acute-phase proteins in domestic animals.

教育活動情報

主要な担当授業

  • 研究倫理演習
    開講年度 : 2018年
    課程区分 : 博士後期課程
    開講学部 : 国際感染症学院
  • 動物福祉学
    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 : 動物福祉、伴侶動物、産業動物、実験動物、野生動物、展示動物
  • 専門獣医科学特論
    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
  • 実験動物学実習
    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 : 実験動物学、微生物的モニタリング、遺伝学的モニタリング 遺伝学、発生工学
  • 獣医科学基礎科目B 動物実験倫理特論
    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
  • 研究倫理演習
    開講年度 : 2018年
    課程区分 : 博士後期課程
    開講学部 : 獣医学院
  • 先端獣医科学科目 実験動物学特論
    開講年度 : 2018年
    課程区分 : 博士後期課程
    開講学部 : 獣医学院
  • 専門獣医科学特論
    開講年度 : 2018年
    課程区分 : 博士後期課程
    開講学部 : 獣医学院
  • 病態科学演習
    開講年度 : 2018年
    課程区分 : 学士課程
    開講学部 : 獣医学部
  • 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

大学運営

委員歴

  • 2020年 - 現在   国立大学法人動物実験施設協議会   学術情報・広報委員会
  • 2020年 - 現在   日本実験動物学会   動愛法等対策委員会
  • 2020年 - 現在   日本実験動物学会   実験動物管理者研修制度委員会
  • 2019年 - 現在   日本実験動物医学専門医協会   試験問題検討委員会
  • 2019年 - 現在   国立大学法人動物実験施設協議会   中型動物委員会
  • 2018年 - 現在   日本実験動物学会   動物福祉・倫理委員会
  • 2016年 - 現在   日本実験動物学会   定款、細則、規定等検討委員会
  • 2016年 - 現在   日本実験動物学会   人材育成委員会
  • 2014年 - 現在   日本実験動物医学会   理事会
  • 2018年 - 2019年   日本実験動物学会   理事会
  • 2012年 - 2015年   日本実験動物医学専門医協会   試験委員会/試験問題作成委員会委員
  • 2015年   日本実験動物医学会、日本実験動物医学専門医協会   「げっ歯類の胎児・新生児の鎮痛・麻酔および安楽死に関する声明」アドホック専門委員会

社会貢献活動

  • AAALAC International アドホック専門家
    期間 : 2017年09月01日
    役割 : 助言・指導

学術貢献活動

  • 日本実験動物学会外部検証専門員
    期間 : 2018年07月10日
    役割 : 学術調査立案・実施
    種別 : 学術調査


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