Experimental study of heterotopic bone formation by in vivo and in vitro gene transfer of bone morphogenetic protein(BMP) and sonic hedgehog(SHH) in the maxillofacial resions.

颌面部骨形态发生蛋白(BMP)和音刺猬(SHH)体内外基因转移异位骨形成的实验研究。

基本信息

  • 批准号:
    13470430
  • 负责人:
  • 金额:
    $ 6.14万
  • 依托单位:
  • 依托单位国家:
    日本
  • 项目类别:
    Grant-in-Aid for Scientific Research (B)
  • 财政年份:
    2001
  • 资助国家:
    日本
  • 起止时间:
    2001 至 2003
  • 项目状态:
    已结题

项目摘要

cDNA encoding the NH2-terminal domain of human and mouse sonic hedgehog(Shh) was amplified by PCR from the full length human and mouse Shh cDNA. It was inserted into the multiple cloning site of the pCR3.1(+) expression vector. The complete coding resion of human and mouse BMP-2 was inserted into the multiple cloning site of the pCR3.1(+) expression vector as well as BMP-4. Human and mouse fibroblasts derived from oral tissue were transfected with human and mouse BMP-2, BMP-4, and Shh cDNA plasmid vector each by calcium phosphate precipitation. Selected storongly positive clones were mouse fibroblast-clones by transfected with mouse BMP-2, BMP-4, and Shh cDNA plasmid vector each. Selected positive clone was grown to confluence in large scale flasks and cells were enclosed in gelatin capsules and implanted into the thigh muscles of nude mice. 14days after implantation, a radiopaque area was observed in cells-implanted area on the soft X-ray films and light microscopic examination disclo … More sed new bone formation by endochondral ossification. with toluidine bulue staining.Vascularized muscle flaps in mice cervical, shoulder and limbs were elevated to examine longterm-viability and the following results were obtained that ectopectoralis flap, superficial gluteal muscle flap and biceps femoris muscle flap were useful for gene-transfer and cells-implant experiments.In the A9 cells, C3H10T1/2 cells, KMS-6 cells and fibroblasts derived from mouse oral tissues transfected with mouse BMP-2, BMP-4 and Shh cDNA plasmid vector each, BMP and Shh-positive cells were detected in C3H10T1/2 cells and fibroblasts derived from mouse oral tissues. Though BMP and Shh protein secretion was stable by 12 passages, these protein secretions and gene expressions were not detected soon and therefore, these gene expressions were transient as we, had expected.Storongly positive cells transfected with mouse BMP-2, BMP4, and Shh cDNA plasmid vector each were enclosed in gelatin capsules and implanted into ectopectoralis flap, superficial gluteal muscle flap and biceps femoris muscle flap and a radiopaque area was observed in only cells-implanted area on the soft X-ray films and light microscopic examination disclosed new bone formation.Optison solution(200μl) containing mouse BMP-2, BMP-4 and Shh cDNA plasmid vector each were injected into ectopectoralis flap, superficial gluteal muscles flap and biceps femoris muscle flap with ultrasound for 2 minutes(2.5 W/cm2). 14days after injection, a small radiopaque area was observed in only solution-injected area on the soft X-ray films and light microscopic examination disclosed new bone formation. Less
通过PCR从全长人和小鼠Shh cDNA扩增编码人和小鼠音刺猬(Shh)的NH 2-末端结构域的cDNA。将其插入pCR3.1(+)表达载体的多克隆位点。将人和小鼠BMP-2的完整编码区插入pCR3.1(+)表达载体的多克隆位点,以及BMP-4。通过磷酸钙沉淀,用人和小鼠BMP-2、BMP-4和Shh cDNA质粒载体分别转染源自口腔组织的人和小鼠成纤维细胞。筛选出的强阳性克隆为分别用小鼠BMP-2、BMP-4和Shh cDNA质粒载体转染的小鼠成纤维细胞克隆。将选择的阳性克隆在大规模烧瓶中生长至汇合,并将细胞封闭在明胶胶囊中并植入裸鼠的大腿肌肉中。植入14天后,软X线片及光镜检查可见细胞植入区有不透射线区, ...更多信息 通过软骨内骨化形成新骨。将带血管蒂的小鼠颈、肩、四肢肌瓣移植于体外,观察其长期存活情况,结果表明:胸大肌瓣、臀浅肌瓣和股二头肌瓣可用于基因转移和细胞移植实验,A9细胞、C3 H10 T1/2细胞、分别用小鼠BMP-2、BMP-4和Shh cDNA质粒载体转染KMS-6细胞和小鼠口腔组织来源的成纤维细胞,在C3 H10 T1/2细胞和小鼠口腔组织来源的成纤维细胞中检测到BMP和Shh阳性细胞。虽然BMP和Shh蛋白的分泌在12代时是稳定的,但这些蛋白的分泌和基因的表达并没有很快被检测到,因此,这些基因的表达是短暂的,正如我们所预期的。软X线片和光镜检查显示,仅在细胞植入区域观察到不透射线区域,并发现新骨形成。含小鼠BMP-2的Optison溶液(200μl),将BMP-4和Shh cDNA质粒载体分别注入胸大肌皮瓣、臀浅肌皮瓣和股二头肌肌皮瓣,超声(2.5W/cm 2)2 min。注射后14天,软X线片上仅在溶液注射区观察到一个小的不透射线区域,光镜检查显示新骨形成。少

项目成果

期刊论文数量(66)
专著数量(0)
科研奖励数量(0)
会议论文数量(0)
专利数量(0)
Ueda M: "Clinical application of tissue engineering concept andits industrialization"Surgery Frontier. Vol.10(3). 49-54 (2003)
上田男:《组织工程概念的临床应用及其产业化》外科前沿。
  • DOI:
  • 发表时间:
  • 期刊:
  • 影响因子:
    0
  • 作者:
  • 通讯作者:
Hattori H, Mizutani H, Ueda M: "Sonic Hedgehog"CLINICAL CALCIUM. 12・2. 55-59 (2002)
服部 H、水谷 H、上田 M:“音速刺猬”临床钙 12・2(2002 年)。
  • DOI:
  • 发表时间:
  • 期刊:
  • 影响因子:
    0
  • 作者:
  • 通讯作者:
上田実: "再生医療の実用化と産業移転"Surgery Frontier. 10・3. 49-54 (2003)
上田稔:“再生医学的实际应用和产业转移”《外科前沿》10・3(2003)。
  • DOI:
  • 发表时间:
  • 期刊:
  • 影响因子:
    0
  • 作者:
  • 通讯作者:
服部 宇: "Sonic hedgehog"CLINICAL CALCIUM. 12・2. 233-237 (2002)
服部 U:“音速刺猬”临床钙 12・2(2002 年)。
  • DOI:
  • 发表时间:
  • 期刊:
  • 影响因子:
    0
  • 作者:
  • 通讯作者:
Ueda M: "Maxillofacial bone regeneration using tissue engineering concepts."Dentistry in Japan. 39. 199-207 (2003)
Ueda M:“利用组织工程概念进行颌面骨再生。”日本牙科。
  • DOI:
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  • 影响因子:
    0
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UEDA Minoru其他文献

Improvement of High Temperature Fatigue Properties of TiAl Alloys Fabricated by Electron Beam Melting Through Hot Isostatic Pressing Process
热等静压电子束熔炼TiAl合金高温疲劳性能的改善
  • DOI:
    10.7791/jspmee.9.180
  • 发表时间:
    2020
  • 期刊:
  • 影响因子:
    0
  • 作者:
    CHO Ken;YASUDA Hiroyuki;TODAI Mitsuharu;UEDA Minoru;TAKEYAMA Masao;NAKANO Takayoshi
  • 通讯作者:
    NAKANO Takayoshi
高クロム系多合金白鋳鉄の高温エロージョン摩耗特性に及ぼすチタン添加の影響
添加钛对高铬多元合金白口铸铁高温冲蚀磨损特性的影响
  • DOI:
  • 发表时间:
    2022
  • 期刊:
  • 影响因子:
    0
  • 作者:
    CHO Ken;YASUDA Hiroyuki;TODAI Mitsuharu;UEDA Minoru;TAKEYAMA Masao;NAKANO Takayoshi;門間雄大,清水一道,楠本賢太
  • 通讯作者:
    門間雄大,清水一道,楠本賢太

UEDA Minoru的其他文献

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{{ truncateString('UEDA Minoru', 18)}}的其他基金

Regulation of plant hormone receptor using stereochemical isomers of a natural product
利用天然产物的立体化学异构体调节植物激素受体
  • 批准号:
    18KK0162
  • 财政年份:
    2019
  • 资助金额:
    $ 6.14万
  • 项目类别:
    Fund for the Promotion of Joint International Research (Fostering Joint International Research (B))
Molecular mechanism of trap-snapping movement of carnivorous plant
食虫植物捕捉运动的分子机制
  • 批准号:
    24651236
  • 财政年份:
    2012
  • 资助金额:
    $ 6.14万
  • 项目类别:
    Grant-in-Aid for Challenging Exploratory Research
Development of new regenerative therapy for central nerve system using the conditioned media derived from human dental pulp stem cells
使用源自人牙髓干细胞的条件培养基开发新的中枢神经系统再生疗法
  • 批准号:
    23659914
  • 财政年份:
    2011
  • 资助金额:
    $ 6.14万
  • 项目类别:
    Grant-in-Aid for Challenging Exploratory Research
Intracellular glycoside signal participating in the regulation of biological phenomena
细胞内糖苷信号参与生物现象的调节
  • 批准号:
    23310147
  • 财政年份:
    2011
  • 资助金额:
    $ 6.14万
  • 项目类别:
    Grant-in-Aid for Scientific Research (B)
Analysis of benefits of dental pulp stem cells from permanent and milk teeth for the treatment of the intractable diseases and of their banking system.
分析恒牙和乳牙牙髓干细胞治疗疑难杂症及其银行系统的益处。
  • 批准号:
    21390524
  • 财政年份:
    2009
  • 资助金额:
    $ 6.14万
  • 项目类别:
    Grant-in-Aid for Scientific Research (B)
Bioorganic studies on membrane protein involved in the regulation of plant ion channels
参与植物离子通道调节的膜蛋白的生物有机研究
  • 批准号:
    20310128
  • 财政年份:
    2008
  • 资助金额:
    $ 6.14万
  • 项目类别:
    Grant-in-Aid for Scientific Research (B)
Periodontal tissue regeneration method by regenerative medicine with stem cells-multi-center study for practical use
干细胞再生医学牙周组织再生方法-多中心实用化研究
  • 批准号:
    16390583
  • 财政年份:
    2004
  • 资助金额:
    $ 6.14万
  • 项目类别:
    Grant-in-Aid for Scientific Research (B)
Study of cultured bone or cartilage regeneration using human mesenchymal stem cells
利用人间充质干细胞培养骨或软骨再生的研究
  • 批准号:
    12794015
  • 财政年份:
    2000
  • 资助金额:
    $ 6.14万
  • 项目类别:
    Grant-in-Aid for University and Society Collaboration
Bioorganic studies on nyctinastic leaf-movement controlled by a biological clock
生物钟控制的夜间叶运动的生物有机研究
  • 批准号:
    12680598
  • 财政年份:
    2000
  • 资助金额:
    $ 6.14万
  • 项目类别:
    Grant-in-Aid for Scientific Research (C)
Development technique of bone regeneration using Mesenchymal Stem Cell
利用间充质干细胞进行骨再生的开发技术
  • 批准号:
    11307048
  • 财政年份:
    1999
  • 资助金额:
    $ 6.14万
  • 项目类别:
    Grant-in-Aid for Scientific Research (A)

相似国自然基金

Shh(Sonic Hedgehog)基因在颅面发育过程中作用及其机制研究
  • 批准号:
    30171023
  • 批准年份:
    2001
  • 资助金额:
    18.0 万元
  • 项目类别:
    面上项目
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