Generation of model mice for brain tumors using conditional knockout technique

使用条件敲除技术产生脑肿瘤模型小鼠

基本信息

批准号：
17390395
负责人：
USUI Hiroshi
金额：
$ 10.52万
依托单位：
Niigata University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
2005
资助国家：
日本
起止时间：
2005 至 2007
项目状态：
已结题

项目摘要

The purpose of this research is generating model mice for brain tumors, using conditional knockout technique to destroy the functions of tumor-suppressor genes selectively in brain cells. We produced five lines of genetically modified mice, by use of the homologous recombination vector DNAs and mouse ES cell line RENKA. The five lines included 2 kinds of "flox" mice in which frame-shift exon of tumor-suppressor gene Ptc or Nf1 was flanked with recombination target sequence loxP, and 2 kinds of "knock-in" mice in which Cre recombinase was introduced in the translation initiation site of Nestin or Tubb3 gene, marker genes for neural cells. We also produced Nf1-flox; P53-null mice in which a tumor suppressor gene P53 located nearly in Nf1 gene was destroyed. Crossbreeding of the "flox" mice and "knock-in" mice successfully generated the mice in which tumor suppressor gene was destroyed selectively in brain cells by the Cre-loxP recombination system.Among the mice obtained, brain tumors which arose from the cells in cerebellar cortex were observed with frequency of 50% (2/4)in Ptc-flox;Tubb3-Cre hetero mice before 5 months of age. The tumor cells consisted of undifferentiated cells which hardly expressed mature marker proteins and resembled those of human medulloblastomas in histology. On the other hand, no brain tumor was observed in Nf1-flox;Neatin-Cre hetero nor Nf1-flox;P53-null;Neatin-Cre hetero mice within 6 months of age, suggesting the necessity of generating homo knockout mice for the Nf1 and P53 gene.These results indicate that mutation of Ptc gene is one of the causative events of medulloblastoma, and, therefore, we were able to contribute to clarification of etiology of the medulloblastoma. In addition, the Ptc-flox;Tubb3-Cre hetero mice may work as model mice of medulloblastoma which is useful to develop new therapeutic drugs or protocols.

这项研究的目的是使用条件敲除技术为脑肿瘤生成模型小鼠，以选择性地破坏脑细胞中肿瘤抑制基因的功能。我们通过使用同源重组载体DNA和小鼠ES细胞系Renka产生了五行转基因的小鼠。这五行包括两种“氟氧杆”小鼠，其中肿瘤 - 抑制剂基因PTC或NF1的框架偏移外显子与重组靶序列LOXP侧面，以及2种“敲入”小鼠，其中CRE重组酶在Nestin或Tubb3 Gene，Marker neural neural neural neural in Neural of the Neftiation intriation of介绍中。我们还产生了NF1-flox； p53-null小鼠，其中几乎位于NF1基因中的肿瘤抑制基因p53被破坏。 “ Flox”小鼠和“敲入”小鼠的杂交成功产生了小鼠，其中抑制肿瘤抑制基因在脑细胞中被CRE-LOXP重组系统选择性破坏。年龄。肿瘤细胞由未分化的细胞组成，这些细胞几乎没有表达成熟的标记蛋白，并且类似于组织学中人类髓母细胞瘤的细胞。 On the other hand, no brain tumor was observed in Nf1-flox;Neatin-Cre hetero nor Nf1-flox;P53-null;Neatin-Cre hetero mice within 6 months of age, suggesting the necessity of generating homo knockout mice for the Nf1 and P53 gene.These results indicate that mutation of Ptc gene is one of the causative events of medulloblastoma, and, therefore,我们能够促进髓母细胞瘤病因的澄清。此外，PTC-Flox; tubb3-cre Hetero小鼠可以用作髓母细胞瘤的模型小鼠，这对于开发新的治疗药物或方案很有用。