Development of catecholamine producing cells by the transfection of CV40 TS gene and its application to intracranial neural transplantation.

CV40 TS基因转染儿茶酚胺产生细胞的研制及其在颅内神经移植中的应用

基本信息

  • 批准号:
    04557005
  • 负责人:
  • 金额:
    $ 12.99万
  • 依托单位:
  • 依托单位国家:
    日本
  • 项目类别:
    Grant-in-Aid for Developmental Scientific Research (B)
  • 财政年份:
    1992
  • 资助国家:
    日本
  • 起止时间:
    1992 至 1993
  • 项目状态:
    已结题

项目摘要

1. Fetal or neonatal mesencephalic (MS) cells, striatal (STR) cells and adrenal (AD) cells were transfected with SV40T/TS gene by culturing on the confluent 2 cells or culturing in the conditioned medium of 2 cells that have retroviral particles containing SV40T/TS.2. Transfected cells grew better when EGF, FGF or conditioned medium of striatal cells were added in the culture medium.3. After the growing period for 4 weeks in the standard or enriched medium, the transfected cells were cultured in the medium that contains G418 and eight G418-resistant cells were selected (MS-1, 2, 3 ; STR-1, 2 ; AD-1, 2, 3).4. They grew under the temperature of 33。C and stopped growing under 39。C, thus they had thermosensitive character. The doubling time was approximately 4 weeks.5. Only MS-1 and AD-1 were immunoreactive for tyrosine hydroxylase (TH). MS-2, 3 and STR-1, 2 were immunoreactive for both neurofilament and GFAP.6. After growing in vitro, MS-1 and AD-1 were transplanted in the striatum of an animal model of Parkinson's disease, and STR-1, 2 were transplanted in the striatum of an animal model of brain ischemia. Each cells survived in the host brain for two weeks. However, after 4 weeks they became unhealthy and finally disappeared. Thus, they could not survive for a long perid that enables the functional restoration.The reason why the grafted cells can not survive for a long period in the host brain is not clear yet. Various micro-environmental factors modulate the survival of the grafted cells. The characteristic of thermosensitivity might be a reason for that the cells cannot survive for a long period in the brain.Thus, it is an urgent problem to find an endogenous promoter that enables the expression of transfected genes for a long period.
1.将SV40T/TS基因导入胎儿或新生儿中脑(MS)细胞、纹状体(STR)细胞和肾上腺(AD)细胞,在融合的2个细胞上培养或在含有SV40T/TS逆转录病毒颗粒的2个细胞条件培养液中培养。在培养液中加入EGF、成纤维细胞生长因子或纹状体细胞条件培养液时,转基因细胞生长较好。在标准或浓缩培养液中培养4周后,在含有G418的培养液中培养,筛选出8个G418抗性细胞(MS-1、2、3;STR-1、2;AD-1、2、3)。它们在33℃的温度下生长。C并在39岁以下停止生长。C,具有热敏特性。倍增时间约为4周。仅MS-1和AD-1对酪氨酸羟化酶(TH)呈阳性反应。MS-2、3和STR-1、2对神经丝和GFAP均呈免疫阳性反应。在体外生长后,将MS-1和AD-1移植到帕金森病动物模型的纹状体内,将STR-1、2移植到脑缺血动物模型的纹状体内。每个细胞在宿主大脑中存活了两周。然而,4周后,他们变得不健康,最终消失了。因此,它们不能长期存活,从而能够进行功能恢复。移植的细胞为什么不能在宿主大脑中长期存活的原因尚不清楚。各种微环境因素调节着移植细胞的存活。热敏感的特性可能是细胞不能在脑内长期存活的原因之一,因此寻找一种内源性启动子使转基因基因能够长期表达是迫切需要解决的问题。

项目成果

期刊论文数量(61)
专著数量(0)
科研奖励数量(0)
会议论文数量(0)
专利数量(0)
Furuyama,F.: "Genetic development of an inbred rat strain with increased resistance adaptation to a hot environment" Am.J.Physiol.265. R957-R962 (1993)
Furuyama, F.:“近交系大鼠品系的遗传发育对炎热环境的抵抗力增强”Am.J.Physiol.265。
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  • 期刊:
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    0
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  • 通讯作者:
Hashimoto,N.: "Dopamine has inhibitory and accelerating effects on ischemia-induced neuronal cell damage in the hot striatum" Brain Res.Bull.33. 281-288 (1994)
Hashimoto,N.:“多巴胺对热纹状体中缺血引起的神经元细胞损伤具有抑制和加速作用”Brain Res.Bull.33。
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    0
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Hattori,S.et al.: "Treadmill running test for evaluating locomotor activity after 6-OHDA lesions and depaminergic cell grafts in the rat" Brain Res.Bull.31. 433-435 (1993)
Hattori,S.et al.:“用于评估大鼠 6-OHDA 损伤和去氨能细胞移植后运动活性的跑步机跑步测试”Brain Res.Bull.31。
  • DOI:
  • 发表时间:
  • 期刊:
  • 影响因子:
    0
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  • 通讯作者:
Czurko,A.et al.: "Appearance of immunoglobulin G and complement factor C3 in the striatum after transient focal ischemia in the rat" Neurosci.Lett.166. 51-54 (1994)
Czurko,A.et al.:“大鼠短暂局部缺血后纹状体中免疫球蛋白 G 和补体因子 C3 的出现”Neurosci.Lett.166。
  • DOI:
  • 发表时间:
  • 期刊:
  • 影响因子:
    0
  • 作者:
  • 通讯作者:
Karadi, Z., Oomura, Y., Nishino, H., Scott, T.R., Lenard, L.and Aou, S.: "Responses of lateral hypothalamic glucose-sensitive and glucose-insensitive neurons to chemical stimuli in behaving rhesus monkeys." J.Neurophysiol.76. 389-400 (1992)
Karadi, Z.、Oomura, Y.、Nishino, H.、Scott, T.R.、Lenard, L. 和 Aou, S.:“行为恒河猴中外侧下丘脑葡萄糖敏感和葡萄糖不敏感神经元对化学刺激的反应。
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    0
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NISHINO Hitoo其他文献

NISHINO Hitoo的其他文献

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{{ truncateString('NISHINO Hitoo', 18)}}的其他基金

Development of stem (neural stem-, embryonic stem-, medullary stem) cells to dopaminergic neurons and their application to reconstruction medicine.
干细胞(神经干细胞、胚胎干细胞、髓质干细胞)发育为多巴胺能神经元及其在重建医学中的应用。
  • 批准号:
    15200026
  • 财政年份:
    2003
  • 资助金额:
    $ 12.99万
  • 项目类别:
    Grant-in-Aid for Scientific Research (A)
Striatal vulnerability and functional repair by neural grafts
神经移植物的纹状体脆弱性和功能修复
  • 批准号:
    10044311
  • 财政年份:
    1998
  • 资助金额:
    $ 12.99万
  • 项目类别:
    Grant-in-Aid for Scientific Research (B).
Development of Huntington's model animals and protection of the symptom by antisense strategy
亨廷顿舞蹈症模型动物的开发和反义策略对症状的保护
  • 批准号:
    09558104
  • 财政年份:
    1997
  • 资助金额:
    $ 12.99万
  • 项目类别:
    Grant-in-Aid for Scientific Research (B)
Mechanisms of cell in following brain ischemia and reconstruction of disturbed function by neural graft
脑缺血后的细胞机制及神经移植重建受损功能
  • 批准号:
    07458223
  • 财政年份:
    1995
  • 资助金额:
    $ 12.99万
  • 项目类别:
    Grant-in-Aid for Scientific Research (B)
Identification of dopaminergic neuron trophic factor using subtraction hybridization and bioassay
使用消减杂交和生物测定法鉴定多巴胺能神经元营养因子
  • 批准号:
    03454133
  • 财政年份:
    1991
  • 资助金额:
    $ 12.99万
  • 项目类别:
    Grant-in-Aid for General Scientific Research (B)
Phenotypic Plasticity of Grafted Catecholaminergic Neurons and its Regulatory Mechanisms.
嫁接儿茶酚胺能神经元的表型可塑性及其调节机制。
  • 批准号:
    01480127
  • 财政年份:
    1989
  • 资助金额:
    $ 12.99万
  • 项目类别:
    Grant-in-Aid for General Scientific Research (B)
The mechanism of differentiation of transplanted chromaffin cells into dopaminergic neurons.
移植的嗜铬细胞分化为多巴胺能神经元的机制。
  • 批准号:
    62570054
  • 财政年份:
    1987
  • 资助金额:
    $ 12.99万
  • 项目类别:
    Grant-in-Aid for General Scientific Research (C)

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微重力场中髓核细胞诱导骨髓间充质干细胞分化及永生化
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