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Developmental and molecular cell biological study of the mechanisms of limb differentiation

Research Project

Project/Area Number 10470004
Research Category

Grant-in-Aid for Scientific Research (B).

Allocation TypeSingle-year Grants
Section一般
Research Field General anatomy (including Histology/Embryology)
Research InstitutionKYOTO UNIVERSITY

Principal Investigator

SHIOTA Kohei  Kyoto Univ., Anatomy, Professor, 医学研究科, 教授 (80109529)

Co-Investigator(Kenkyū-buntansha) MORIMOTO Takeshi  Kyoto Univ., Anatomy, Instructor, 医学研究科, 助手 (00311746)
TAKIGAWA Toshiya  Kyoto Univ., Anatomy, Instructor, 医学研究科, 助手 (90263095)
ISHIBASHI Makoto  Kyoto Univ., Anatomy, Lecturer, 医学研究科, 講師 (30232341)
MIURA Takashi  Kyoto Univ., Anatomy, Instructor, 医学研究科, 助手 (10324617)
成瀬 一郎  京都大学, 大学院・医学研究科, 助教授 (20113326)
森 千里  京都大学, 大学院・医学研究科, 助教授 (90174375)
Project Period (FY) 1998 – 2000
Project Status Completed (Fiscal Year 2000)
Budget Amount *help
¥10,100,000 (Direct Cost: ¥10,100,000)
Fiscal Year 2000: ¥2,700,000 (Direct Cost: ¥2,700,000)
Fiscal Year 1999: ¥2,300,000 (Direct Cost: ¥2,300,000)
Fiscal Year 1998: ¥5,100,000 (Direct Cost: ¥5,100,000)
KeywordsLimb bud / Programmed cell death / Vascular remodelling / Matrix metalloproteinase / Limb anomalies / Mouse fetus / 細胞死 / メタロプロテアーゼ / カスパーゼ / マウス / Hammer-toe / 分化 / 形態形成 / 指奇形 / アポトーシス / リモデリング
Research Abstract

We have reported that so-called "programmed cell death" (PCD) in the interdigital tissue of the fetal mouse limb is apoptosis with DNA fragmentation and that the regional heterogeneity and temporal sequence of cell death in the developing limb play essential roles in patterning the limb. However, the cells which undergo apoptosiss were not identified nor was the process of how the ECM is disrupted during digit separation. We examined the remodeling of the microvascular network in the interdigital tissue of mouse fetuses and found that cell death occurs in PECAM1/C031-positive endothelial cells. Whole-mount in situ hybridization and Northern blot analysis revealed that gelatinase/MMP2 was highly expressed in the areas where vascular remodeling was occurring. In the limb of Hammertoe mutant (Hm/Hm) mouse fetuses in which interdigital webbing persists, neither apoptosis nor the regression of microvascular plexus in the interdigital tissue were observed to occur normally. Our study has provided the first evidence that apoptotic cell death occurs in endothelial cells of the interdigital microvascular plexus and that vascular remodeling and gelatinase/MMP2 expression may play essential roles in the separation of mammalian digits.
Next, to analyze the mechanisms of chondrogenic pattern formation in the developing limb, we utilized a micromass culture system of limb mesenchymal cells, and examined the chondrogenic pattern formation in vitro. The periodic nature of limb chondrogenesis was retained in micromass culture. We developed a novel method to automatically measure the periodicity of the pattern and demonstrated that the reaction-diffusion model is the most plausible for explaining the periodic chondrogenic pattern formation. We also demonstrated that TGFβ2 acts as an "activator"-like molecule in chondrogenic pattern formation and is possibly responsible for the cell sorting phenomenon.

Report

(4 results)
  • 2000 Annual Research Report   Final Research Report Summary
  • 1999 Annual Research Report
  • 1998 Annual Research Report
  • Research Products

    (27 results)

All Other

All Publications (27 results)

  • [Publications] 木村澄子 他: "Palmar and plantar pads and flexion creases of Polydactyly Nagoya (Pnd) mice with polydactyly"Journal of Morphology. 239. 87-96 (1999)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2000 Final Research Report Summary
  • [Publications] Li,Z.L. 他: "Stage-specific vertebral homeotic transformations in mouse fetuses induced by brief hyperthermia"Developmental Dynamics. 216. 336-348 (1999)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2000 Final Research Report Summary
  • [Publications] 木村澄子 他: "Pads and flexion creases on the plantar surface of Mammertoe mutant mouse (Hm)"Anatomical Record. 260. 26-32 (2000)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2000 Final Research Report Summary
  • [Publications] 三浦岳 他: "Extracellular matrix environment influences chondrogenic pattern formation in limb bud culture"Anatomical Record. 258. 100-107 (2000)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2000 Final Research Report Summary
  • [Publications] 三浦岳 他: "TGF-β2 acts as an "activator in the reaction-diffusion model and is involved in cell sorting phenomenon in mouse limb bud culture"Developmental Dynamics. 217. 241-249 (2000)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2000 Final Research Report Summary
  • [Publications] 三浦岳 他: "A novel method for analysis of the peridicity of chondrogenic patterns in limb bud cell culture"Anatomy and Embryology. 201. 419-428 (2000)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2000 Final Research Report Summary
  • [Publications] Kimura, S.et al.: "Palmar and plantar pads and flexion creases of Polydactyly Nagoya (Pdn) mice with polydactyly."Journal of Morphology. 239. 87-96 (1999)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2000 Final Research Report Summary
  • [Publications] Li, ZL.et al.: "Stage-specific vertebral homeotic trnasformations in mouse fetuses induced by brief hyperthermia"Development Dynamics. 216. 336-348 (1999)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2000 Final Research Report Summary
  • [Publications] Kimura, S.et al.: "Pads and flexion creases on the plantar surface of Hammertoe mutant mice."Anatomical Record. 260. 26-32 (2000)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2000 Final Research Report Summary
  • [Publications] Miura, T.et al.: "Extracellular matrix environment influences chondrogenic pattern formation in limb bud culture."Anatomical Record. 258. 100-107 (2000)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2000 Final Research Report Summary
  • [Publications] Miura, T.et al.: "TGF-β2 acts as an "activator" molecule in the reaction-diffusion model and is involved in cell sorting phenomenon in mouse limb bud micromass culture"Developmental Dynamics. 217. 241-249 (2000)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2000 Final Research Report Summary
  • [Publications] Miura, T.et al.: "A novel method for analysis of the periodicity of chondrogenic patterns in limb bud cell culture"Anatomy and Embryology. 201. 419-428 (2000)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2000 Final Research Report Summary
  • [Publications] 三浦岳 他: "Extracellular matrix environment influences chondrogenic pattern formation in limb bud culture"Anatomical Record. 258(1). 100-107 (2000)

    • Related Report
      2000 Annual Research Report
  • [Publications] 三浦岳 他: "TGF-β2 acts as an "activator" molecule in the reaction-diffusion model and is involved in cell sorting phenomenon"Developmental Dynamics. 217. 241-249 (2000)

    • Related Report
      2000 Annual Research Report
  • [Publications] 三浦岳 他: "A novel method for analysis of the periodicity of chondrogenic patterns in limb bud cell culture"Anatomy and Embryology. 201(5). 419-428 (2000)

    • Related Report
      2000 Annual Research Report
  • [Publications] 木村澄子 他: "Pads and flexion creases on the palmar surface of Hammetoe mutant mouse (Hm)."Anatomical Record. 259(1). 26-32 (2000)

    • Related Report
      2000 Annual Research Report
  • [Publications] 木村澄子 他: "Comparative investigations of human and rat dermatoglyphics : Palmar and digital pads, and flexion creases."Acta Anatomica Nipponica. (印刷中). (2001)

    • Related Report
      2000 Annual Research Report
  • [Publications] 塩田浩平 他: "Color Atlas of Clinical Embryology"Saunders (Philadelphia, USA). 294 (2000)

    • Related Report
      2000 Annual Research Report
  • [Publications] Naruse I.et al.: "Palar and plantar pads and flexion creases of genetic polydactyly mice (pdn)"Journal of Morphology. 239(1). 87-96 (1999)

    • Related Report
      1999 Annual Research Report
  • [Publications] Takigawa T.et al.: "Study of the vascular remodelling and programmed cell death during the separation of digits in the mouse limb"Acta Anatomica Nipponica. 75(1). 67 (2000)

    • Related Report
      1999 Annual Research Report
  • [Publications] Takigawa T.et al.: "Study of the differentiation of the medial edge epithelium in the fetal mouse palate in organ culture"Acta Anatomica Nipponica. 75(1). 60 (2000)

    • Related Report
      1999 Annual Research Report
  • [Publications] Shiota,K.and Nakatsu,T.: "Normal and abnormal neurulation in human embryos: Implications for the pathogenesis of neural tube defects." Neuropathology. 18. 415-418 (1998)

    • Related Report
      1998 Annual Research Report
  • [Publications] Peeters,M.C.E.他: "Differences in axial curvature correlate with species-specific rate of neural tube closure in the embryo of rabbit,rat,chick and human." Anatomy and Embryology. 198(3). 185-194 (1998)

    • Related Report
      1998 Annual Research Report
  • [Publications] 水関健司 他: "SoxD: an essential mediator of induction of anterior neural tissues in Xenopus embryos." Neuron. 21(1). 77-85 (1998)

    • Related Report
      1998 Annual Research Report
  • [Publications] 山本淑子 他: "Effects of amphetamine on rat embryos developing in vitro" Reproductive Toxicology. 12(2). 133-137 (1998)

    • Related Report
      1998 Annual Research Report
  • [Publications] 木村澄子 他: "Palmar and plantar pads and fiexion creases of Polydactyly Nagoya (Pdn) mice with polydactyly" J.Morphol.,. (in press). (1999)

    • Related Report
      1998 Annual Research Report
  • [Publications] Shin,H.-J.他: "Involvement of germ cell apoptosis in the induction of testicular toxicity following hydroxyurea treatment." Toxicol.Appl.Pharmacol.155. 139-149 (1999)

    • Related Report
      1998 Annual Research Report

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Published: 1998-04-01   Modified: 2016-04-21  

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