研究内容（折野）

1. 哺乳動物の血清フェリチン測定とその臨床診断的意義

鉄貯蔵タンパク質は細胞内に存在しますが、血液にも微量に存在します（血清フェリチンの分泌機構は詳細には解明されておりません）。血清フェリチンレベルは体内鉄貯蔵量と正の相関をすることから、鉄欠乏性貧血および鉄過剰症の診断に用いられます。現在、ウマ、ウシ、イヌの血清フェリチン測定法を既に開発しております。また、血清フェリチンは炎症および腫瘍性疾患でも上昇することから、血清フェリチンは炎症および腫瘍マーカーに成り得ます。各種哺乳動物における血清フェリチン測定の臨床診断的意義について検討しております。

2. 脊椎動物のフェリチンの遺伝子配列決定とその発現

現在までウマ、ウシ、イヌ、ネコ、ニワトリのフェリチンの遺伝子配列を決定しております。最近ではイルカフェリチンの遺伝子配列決定を完了しました。また、フェリチンを遺伝子工学を用いて発現させ、フェリチンの機能および構造を調べております。

3. フェリチンの酸化ストレス防御効果

病気の90％は酸化ストレスが原因であると言われております。鉄は酸化ストレスを増悪させます。鉄は生命に必須な微量金属元素ですが、生体に利用されない鉄はフェリチンに貯蔵されます。さらにフェリチンの鉄貯蔵機能は、鉄の毒性を無毒化し、酸化ストレスを防護します。しかしながら、呼吸で酸素を取り入れてエネルギーを産生し、生命を維持するには酸化ストレスは避けることが出来ない問題です。現在、細胞内フェリチンの酸化ストレスに対する防御効果を分子生物学的および遺伝子的レベルで解明しようとしております。

4. フェリチン結合タンパク質に関する研究

細胞内外にはフェリチンと結合する多種多様なタンパク質が見出されております。現在まで血液、ミルク、腎臓細胞等にフェリチン結合タンパク質を存在することを明らかにしております。循環血中のフェリチン結合タンパク質はフェリチンのクリアランスに関与することが知られています。しかしながら、多くのフェリチン結合タンパク質の生理的意義は明らかにされておらず、その解明に取り組んでいます。

より詳しい内容を知りたい方は 獣医生化学研究室にお越しください。

研究業績

折野宏一　原著論文他（1989-2022）
【著書】

  1. 獣医生化学実験＜第３版＞　Q：薄層クロマトグラフィー　R：免疫化学実験法　　獣医生理学・生理化学教育懇談会　編著　分担執筆　チクサン出版社　pp. 181-188, 176-178.
  2. 獣医生化学　７章：糖質の代謝　　斉藤昌之，鈴木嘉彦，横田　博　編　分担執筆　朝倉書店

 
【原著論文】

  1. Pandey, G. S., Shuimizu, K., Orino, K. and Schneebeli, M. 1989. Preliminary observations on ovine paratuberculosis (Johne’s disease) in Zambia. Rev. Vet. Pays Trop. 42: 515-516.
  2. Orino, K. and Naiki, M. 1990. Two kinds of P-fimbrial variants of uropathogenic Escherichia coli recognizing Forssman glycosphingolipid. Microviol. Immunol. 34: 607-615.
  3. 折野宏一，渡辺清隆，山本晋二． ウマフェリチンの免疫測定に及ぼす血清および血漿の阻害効果. 家畜生化学研究会報. 29：37-44.
  4. Orino, K., Saji, M., Ozaki, Y., Ohya, T., Yamamoto, S. and Watanabe, K. 1993. Inhibitory effects of horse serum on immunoassay of horse ferritin. J. Vet. Med. Sci. 55: 45-49.
  5. Orino, K., Yamamoto S, Watanabe K. 1993. Fibrinogen as a ferritin-binding protein in horse plasma. J. Vet. Med. Sci. 55: 785-787.
  6. Watanabe, K., Sakashita, Y., Orino, K. and Yamamoto S. 1994. Identification of bovine serum transferrin phenotypes by polyacrylamide gel isoelectric focusing (PAGIEF). J. Vet. Med. Sci. 56: 421-423.
  7. Watanabe, K., Yahikozawa, S., Orino, K. and Yamamoto, S. 1995. Immunological cross-reaction between lactoferrin and transferrin. J. Vet. Med. Sci. 57: 519-521.
  8. 山田真理子、坂田美和子、坪内亜矢子、折野宏一、山本晋二、渡辺清隆. 1995. 牛血漿および好中球ラクトフェリン含量. 家畜生化学. 32：25-30.
  9. 角田健一、折野宏一、山本晋二、渡辺清隆. 1996. 牛胎仔血清のフェリチンレベル 家畜生化学. 33:25-30.
  10. Kakuta, K., Orino, K., Yamamoto, S. and Watanabe, K. 1997. High levels of ferritin and its iron in fetal bovine serum. Comp. Biochem. Physiol. 118A:165-169.
  11. 折野宏一、江口邦明、山本晋二、渡辺清隆. 牛フェリチンL鎖cDNAの塩基配列. 家畜生化学. 33:31-38.
  12. 折野宏一、中山貴雄、江口邦明、渡辺清隆、山本晋二. 1997. 牛フェリチンおよびL鎖cDNAのクローニング、シークエンシングおよび発現.　家畜生化学.　34：27-34.
  13. Orino, K., Eguchi, K., Nakayama, T., Yamamoto, S. and Watanabe, K. 1997. Sequencing of cDNA clones that encode bovine ferritin H and L chains. Comp. Biochem. Physiol. 118B: 667-673.
  14. Watanabe, K., Ozawa, M., Ochiai, H., Kamohara, H., Iijima, N., Negita, H., Orino, K. and Yamamoto S. 1998. Changes in iron and ferritin in anemic calves infected with Theileria sergenti. J. Vet. Med. Sci. 60: 943-947.
  15. Orino, K., Tsuji, Y., Torti, F. M. and Torti, S. V. 1999. Adenovirus E1A blocks oxidant-dependent ferritin induction and sensitizes cells to pro-oxidant cytotoxicity. FEBS Lett. 46: 334-338.
  16. Watanabe, K., Muranishi, N., Murata, Y., Orino, K., Okano, S. and Yamamoto S. 2000. Biochemical properties of canine serum ferritin: iron content and nonbinding to concanavalin A. Biometals 13: 319-324.
  17. Orino, K., Lehman, L., Tsuji, Y., Ayaki, H., Torti, S. V. and Torti, F. M. 2001. Ferritin and the response to oxidatice stress. Biochem. J. 357: 241-247.
  18. Watanabe, K., Yamashita, Y., Ohgawara, H., Sekiguchi, M., Satake, N., Orino, K., and Yamamoto S. 2001. Iron content of rat serum ferritin. J. Vet. Med. Sci. 63: 587-589.
  19. Inagaki, M., Kikuchi, M., Orino, K., Ohnami, Y. and Watanabe K. 2002. Purification and quantification of lactoferrin in equine seminal plasma. J. Vet. Med. Sci. 64: 75-77.
  20. Orino, K., Kamura, S., Natsuhori, M., Yamamoto, S. and Watanabe K. 2002. Two pathways of iron uptake in bovine spleen apoferritin dependent on iron concentration. Biometals 15: 59-63.
  21. Orino, K., Higa, M., Fujikawa, M., Takehara, K., Okano, S., Yamamoto, S. and Watanabe, K. 2002. Characterization of autoantibodies to ferritin in canine serum. Biometals 15: 161-165.
  22. 荻野心太郎，折野宏一，渡辺清隆． 食餌鉄によるラット体内貯蔵鉄の制御．獣医生化学 39: 19-24.
  23. K., Terada, A., Yoshida, A., Okano, S., Higuchi, S., Natsuhori, M. and Watanabe, K. 2003. Clearance rates of biotinylated ferritins in canine: A possible physiological role of canine serum ferritin as an iron transporter. Biometals 16: 341-347.
  24. Kikuchi, M., Mizoroki, S., Kubo, T., Ohiwa, Y., Kubota, M., Yamada, N., Orino, K., Ohnami, Y. and Watanabe, K. 2003. Seminal plasma lactoferrin but not transferrin reflects gonadal function in dogs. J. Vet. Med. Sci. 65: 679-684.
  25. Kikuchi, M., Takao, Y., Tokuda, N., Ohnami, Y., Orino, K. and Watanabe, K. 2003. Relationship between seminal plasma lactoferrin and gonadal function in horses. J. Vet. Med. Sci. 65: 1273-1274.
  26. Orino, K., Ishiji, T., Yamamoto, S. and Watanabe, K. 2004. Characterization of bovine serum ferritin-binding proteins. Comp. Biochem. Physiol. 137A: 375-381.
  27. Orino, K., Harada, S., Natsuhori, M., Takehara, K. and Watanabe, K. 2004. Kinetic analysis of bovine spleen apoferritin and recombinant H and L chain homopolymers: iron uptake suggests early stage H chain ferroxidase activity and second stage L chain cooperation. Biometals 17: 129-134.
  28. Orino, K., Miura, T., Muto, S. and Watanabe, K. 2005. Sequence analysis of canine and equine ferritin H and L subunit cDNAs. DNA Seq. 16:58-64.
  29. Orino, K., Hirata, A. and Watanabe, K. 2005. Sequence analysis of feline ferritin H and L subunit cDNAs. Vet. Biochem. 42: 7-11.
  30. Orino, K., Uehara, M., Okano, S. and Watanabe, K. 2006. Purification and characterization of canine serum ferritin-binding proteins. Biometals 19: 315-322.
  31. Orino, K., Watanabe, S., Ohtsuka, H., Kohiruimaki, M. and Watanabe, K. 2006. Measurement of ferritin in bovine milk and its clinical significance. J. Dairy Sci. 89: 3842-3845.
  32. Abe, T., Kinda, T., Takano, Y., Chikazawa, S., Higuchi, M., Kawasaki, N., Orino, K. and Watanabe, K. 2006. Relationship between body iron stores and diquat toxicity in male Fischer-344 rats. Biometals 19: 651-657.
  33. Orino, K., Hamada, S., Hashimoto, M., Nambo, Y., Kondo, T. and Watanabe, K. 2006. Identification of horse anti-ferritin autoantibody. J. Equine Sci. 17: 113-119.
  34. Higuchi, M., Kobayashi, S., Kawasaki, N., Hamaoka, K., Watabiki, S,, Orino, K. and Watanabe, K. 2007. Protective effects of wheat bran against diquat toxicity in male Fischer-344 rats. Biosci. Biotechnol. Biochem. 71: 1621-1625.
  35. Seki, T., Kunichika, T., Watanabe, K. and ★Orino, K. Apolipoprotein B binds ferritin by hemin-mediated binding: evidence of direct binding of apolipoprotein B and ferritin to hemin. Biometals 21: 61-69.
  36. Takaesu, A., Watanabe, K., Takai, S., Sasaki, Y. and ★Orino, K. Sequence analysis of dolphin ferritin H and L subunits and possible iron-dependent translational control of dolphin ferritin gene. Acta Vet. Scand. 50: 42.
  37. Sugawara, G., Inoue, R., Watanabe, K., Ohtsuka, H. and ★Orino, K. Bovine α-casein is a ferritin-binding protein and inhibitory factor of milk ferritin immunoassay. J. Dairy Sci. 92: 3810-3814.
  38. Sakamoto, H., Kuboi, T., Nagakura, T., Hayashi, S., Hoshi, F., Mutoh, K., Watanabe, K. and ★Orino K. 2009. Characterization of feline serum ferritin-binding proteins: the presence of a novel ferritin-binding protein as an inhibitory factor in feline ferritin immunoassay. Biometals 22: 793-802.
  39. Naoi, T., Shibuya, N., Inoue, H., Mita, S., Kobayashi, S., Watanabe, K. and ★Orino, K. The effect of tert-butylhydroquinone-induced oxidative stress in MDBK cells using XTT assay: Implication of tert-butylhydroquinone-induced NADPH generating enzymes. J. Vet. Med. Sci. 72: 321-326.
  40. Higuchi, M., Oshida, J., Orino, K. and Watanabe, K. 2011. Wheat bran protects Fischer-344 rats from diquat-induced oxidative stress by activating antioxidant system: selenium as an antioxidant. Biosci. Biotechnol. Biochem. 75: 496-499.
  41. Hashimoto, M., Nambo, Y., Kondo, T., Watanabe, K. and ★Orino, K. A study on the presence of ferritin-binding proteins in fetal horse plasma. J. Equine Sci. 22: 1-7.
  42. Nakai, M., Murata, N., Yoda, T., Yoshikawa, Y., Watanabe, K. and ★Orino, K. Binding of mammalian and avian ferritins with biotinylated hemin: demonstration of preferential binding of the H subunit to heme. J. Vet. Med. Sci. 73: 313-318.
  43. Watanabe, M., Yuge, M,, Uda, A., Yoshikawa, Y., Watanabe, K. and ★Orino, K. Structural and functional analyses of chicken liver ferritin. Poultry Sci. 90: 1489-1495.
  44. Higuchi, M., Yoshikawa, Y., Orino, K., Watanabe, K. 2011. Effect of diquat-induced oxidative stress on iron metabolism in male Fischer-344 rats. Biometals 24: 1123-1131.
  45. Usami, A., Tanaka, M., Yoshikawa, Y., Watanabe, K., Ohtsuka, H. and ★Orino, K. Heme-mediated binding of α-casein to ferritin: evidence for preferential α-casein binding to ferrous iron. Biometals 24: 1217-1224.
  46. Ohya, T., Kondo, T., Yoshikawa, Y., Watanabe, K. and ★Orino, K. Change of ferritin-binding activity in the serum of foal after birth. J. Equine Sci. 22: 73-76.
  47. Yoshikawa, Y., Morimatsu, M., Ochiai, K., Okuda, K,, Taoda, T., Chikazawa, S., Shimamura, A., Omi, T., Bonkobara, M., Orino, K. and Watanabe, K. 2012. Establishment of a PCR analysis method for canine BRCA2. BMC Res. Notes 5: 173.
  48. Shibuya, N., Kobayashi, S., Yoshikawa, Y., Watanabe, K. and ★Orino, K. Effects of oxidative stress caused by tert-butylhydroquinone on cytotoxicity in MDCK cells. J. Vet. Med. Sci. 74: 583-589.
  49. Shibuya, N., Yoshikawa, Y., Watanabe, K., Ohtsuka, H. and ★Orino, K. Iron-dependent binding of bovine milk α-casein with holo-lactoferrin, but not holo-transferrin. Biometals 25: 1083-1088.
  50. Yoshikawa, Y., Ochiai, K., Morimatsu, M., Suzuki, Y., Wada, S., Taoda, T., Iwai, S., Chikazawa, S., Orino, K. and Watanabe, K. 2012. Effects of the missense mutations in canine BRCA2 on BRC repeat 3 functions and comparative analyses between canine and human BRC repeat 3. PLOS ONE 7: e45833.
  51. Chikazawa, S., Hori, Y., Hoshi, F., Kanai, K., Ito, N., Sato, J., Orino, K. and Watanabe, K, Higuchi S. 2013. Development of a sandwich enzyme-linked immunosorbent assay to detect and measure serum levels of canine ferritin. J. Vet. Med. Sci. 75: 515-517.
  52. Kanno, Y., Ohtsuka, H., Yoshikawa, Y., Watanabe, K. and ★Orino, K. Measurement of ferritin and anti-ferritin autoantibodies in serum and colostrum of Holstein and Japanese Black cows. Anim. Sci. J. 84: 556-561.
  53. Mieno, A., Yamamoto, Y., Yoshikawa, Y., Watanabe, K., Mukai, T. and ★Orino, K. Binding analysis of ferritin with heme using α-casein and biotinylated-hemin: detection of heme-binding capacity of Dpr derived from heme synthesis-deficient Streptococcus mutans. J. Vet. Med. Sci. 75: 1101-1105.
  54. Takahashi, K., Kondo, T., Yoshikawa, Y., Watanabe, K. and ★Orino K. The presence of heat-labile factors interfering with binding analysis of fibrinogen with ferritin in horse plasma. Acta Vet. Scand. 55: 70.
  55. Chikazawa, S., Nakazawa, T., Hori, Y., Hoshi, F., Kanai, K., Ito, N., Orino, K., Watanabe, K. and Higuchi, S. 2013. Change in serum ferritin concentration in experimentally induced anemia of chronic inflammation in dogs. J. Vet. Med. Sci. 75: 1419-1426.
  56. Chikazawa, S., Hori, Y., Kanai, K., Ito, N., Hoshi, F., Orino, K. and Watanabe, K, Higuchi S. 2013. Factors influencing measurement of serum iron concentration in dogs: diurnal variation and hyperferritinemia. J. Vet. Med. Sci. 75: 1615-1618.
  57. Numata, M., Kondo, T., Nambo, Y., Yoshikawa, Y., Watanabe, K. and ★Orino, K. Change of antibody levels to ferritin in the sera of foals after birth: possible transfer of maternal anti-ferritin autoantibody via colostrum and age-related anti-ferritin autoantibody production. Anim. Sci. J. 84: 782-789.
  58. ★Orino, K. Functional binding analysis of human fibrinogen as an iron- and heme-binding protein. Biometals 26:789-794.
  59. Masuda, K., Haruta, S., Orino, K., Kawaminami, M. and Kurusu, S. 2013. Autotaxin as a novel, tissue-remodeling-related factor in regressing corpora lutea of cycling rats. FEBS J. 280: 6600-6612.
  60. Higashi, S., Nagasawa, K., Yoshikawa, Y., Watanabe, K. and ★Orino, K. Characterization analysis of human anti-ferritin autoantibodies. Antibodies 3: 169-181.
  61. Schnell, S. A., Ohtsuka, H., Kakinuma, S., Yoshikawa, Y., Watanabe, K. and ★Orino, K. Iron and ferritin levels in the serum and milk of bovine leukemia virus-infected dairy cows. Front. Vet. 2:1-5.
  62. Okada, A., Yoshikawa, Y., Watanabe, K. and ★Orino, K. Analysis of the binding of bovine and human fibrinogen to ferritin: evidence that fibrinogen is a common ferritin-binding protein in mammals. Biometals 28: 679-685.
  63. Yoshikawa, Y., Morimatsu, M., Ochiai, K., Ishiguro-Oonuma, T., Wada, S., Orino, K., Watanabe, K. Reduced canine BRCA2 expression levels in mammary gland tumors. BMC Vet. Res. 11:159.
  64. Yamanaka, Y., Matsugano, S., Yoshikawa, Y. and ★Orino, K. Binding analysis of human immunoglobulin G as a zinc-binding protein. Antibodies 5: 13. doi: 10.3390/antib5020013.
  65. ★Orino, K. 2017. Inhibitory effect of transferrin on serum zinc measurement. Biometals 30:615-621. doi: 10.1007/s10534-017-0032-z
  66. Hasegawa, R., Kiyota, H., Kurusu, S., Yoshikawa, Y. and ★Orino, K. Changes in anti-Ferritin autoantibody during pregnancy in rats. EMS Vet Sci J 1(1):1.
  67. Saito, N., Iio, T., Ohtsuka, H., Yoshikawa, Y. and ★Orino, K. Heme-binding of bovine lactoferrin: the potential presence of a heme-binding capacity in an ansestral transferrin gene. Biometals 31(1):131-138. doi: 10.1007/s10534-017-0075-1
  68. ★Orino, K. Binding of immunoglobulin G to protoporphyrin IX and its derivatives: evidence the Fab domain recognizes the protoporphyrin ring. 2019. Antibodies 8(1):6. doi: https://doi.org/10.3390/antib8010006
  69. Naito K., Iio, T., Katagi, M., Yoshikawa, Y., Ohtsuka, H. and ★Orino, K. Binding analysis of bovine milk proteins, especially casein interactions and the interaction between α–casein and lactoferrin, using beads immobilized with zinc ion and poly-L-lysine or α-casein. International Dairy Journal 105 104690 doi: https://doi.org/10.1016/j.indairyj.2020.104990
  70. Yoshikawa Y, Kozuma H, Morimatsu M, Sugawara K, Orino K. Reduced translation efficiency due to novel splicing variants in 5’untranslated region and identification of novel cis-regulatory elements in canine and human BRCA2. BMC Mol Cell Biol 22: 2 doi:https://doi.org/10.1186./s12860-020-0336-4
  71. Yoshikawa Y, Morimatsu M, Ochiai K, Ishiguro-Oonuma T, Morioka R, Okuda K, Orino K. Identification of the core motif of the BRCA2 C-terminal RAD51-binding domain by comparing canine and human BRCA2. J Vet Med Sci, 83(5):759-766.
  72. Yoshikawa, Y., Morimatsu, M., Ochiai, K., Ishiguro-Oonuma, T., Morioka, R., Okuda, K., Orino, K. Identification of the core motif of the BRCA2 C-terminal RAD51-binding domain by comparing canine and human BRCA2. J Vet Med Sci, 83(5):759-766. doi: https://doi.og/10.1186/s12860-020-00336-4
  73. Zhu, Z., Kitano, T., Morimatsu, M., Tanaka., A, Morioka, R., Lin, X., Orino, K., Y. 2022. BRCA2 C-terminal RAD51-binding domain core confers resistance to DNA-damaging agents. Int J Mol Sci 23:4060. doi: https://doi.org/10.3390/ijms23074060.
  74. Yoshikawa, Y., Kimura, S., Soga, A., Sugiyama, M., Ueno, A., Konco, H., Zhu, Z., Ochiai, K., Nakayama, K., Hakozaki, J., Kusakisako, K., Haraguchi, A., Kitano, T., Orino, K., Fukumoto, S., Ikadai, K. 2022. Plasemodium berghei Braca2 is required fro normal develoment and diffenciation in mice and mosquitoes. Parasites Vectors 15:244. doi: https://doi.org/10.1186/s13071-022-05357-w.

 
　　　★：Corresponding Author
 
【総説】

  1. ★折野宏一，渡辺清隆，山本晋二． フェリチンの分子性状および生理学的意義．獣医生化学 37: 1-10.
  2. ★Orino, K. and Watanabe, K. 2007. Anti-ferritin autoantibody as mammalian common ferritin-binding protein. Vet. Biochem. 44: 1-8.
  3. ★Orino, K. and Watanabe, K. 2008. Molecular, physiological and clinical aspects of the iron storage protein ferritin. Vet. J. 178: 191-201.
  4. ★Orino K. 2016. Physiological implications of mammalian ferritin-binding proteins interacting with circulating ferritin and a new aspect of ferritin- and zinc-binding proteins. Biometals 29: 15-24.
  5. ★Orino K. 2018. Physiological implications of the interaction of mammalian fibrinogens with metal ions, heme, ferritin, and anti-fibrinogen antibody. EMS Vet Sci J 1(1):4.
  6. ★Orino K. 2020. Heme-binding ability of bovine milk proteins, BioMetals 33(6):287-291.

 
 
 
★：Corresponding Author
 
 
【その他の印刷物】

  1. Orino, K., Hasebe, F., Kitada, R., Miyazaki, F., Mubiana, M., Sato, T. and Naiki M. 1994. Prevalence of bovine brucellosis of Zambia. UNZA Vet. 5:5-7.

 
 
 
 

模式図：たくさんの鉄原子を蓄えたフェリチンタンパク質