Immunological analyses and gene therapy in chronic colitis of IL-12p40 transgenic mice

IL-12p40转基因小鼠慢性结肠炎的免疫学分析和基因治疗

基本信息

  • 批准号:
    09670504
  • 负责人:
  • 金额:
    $ 2.3万
  • 依托单位:
  • 依托单位国家:
    日本
  • 项目类别:
    Grant-in-Aid for Scientific Research (C)
  • 财政年份:
    1997
  • 资助国家:
    日本
  • 起止时间:
    1997 至 1998
  • 项目状态:
    已结题

项目摘要

(Aims) : To investigate the roles of IL-12p40 in chronic intestinal inflammation, we generated IL-12p40 transgenic mice using T3^b promoter and determined the restricted expression of IL-12p40 to intestinal epithelial cells. (Methods & Results) : T3^b-IL-12p40 trans genic mice were produced by DNA microinjection method. Transgene contains T3^b promoter, rabbit B globin exon and intron, mouse IL-12p40 cDNA and rabbit beta globin polyA.As a result of screenings by PCR, six founders carrying this transgene were produced. Their phenotypes were almost normal. These mice were backcrossed with C57BL/6 mice, and next generations were obtained in four founders, line #9, #13, #20 and #24. The copy number of the trans gene in mouse genome was determined by Southern blot analysis. As judged from hybridizing intensity of bands, the four transgenic mice carry less than 10 copies of the T3^b-IL-12p40 construct. Northern blot analysis and RT-PCR were performed to examine mRNA expression derived from trangene. IL-12p40 mRNA was detected in large and small intestine of the transgenic mice. Except for gastrointestinal tract, mRNA derived from transgene was not detected in major organs including thymus. And immunohistochemical analysis of the large intestine derived from transgenic mouse and negative littermate were conducted using IL-12 (p40/p70) monoclonal antibody. The epithelial cells of the large intestine in transgenic mice was mainly stained. Colitis was induced in T3^b-IL-12p40 transgenic mice and negative littermate by 1.5% dextran sulfate sodium. The differences of colitis score between transgenic mice and negative littermate were not significant.
(目的):为了研究IL-12 p40在慢性肠道炎症中的作用,我们用T3 ^b启动子构建了IL-12 p40转基因小鼠,并测定了IL-12 p40在肠上皮细胞中的限制性表达。(方法与结果):采用DNA显微注射法制备T3 β-IL-12 p40转基因小鼠。转基因含有T3 ^B启动子、兔B珠蛋白外显子和内含子、小鼠IL-12 p40 cDNA和兔β珠蛋白polyA。他们的表型几乎正常。将这些小鼠与C57 BL/6小鼠回交,并在四个建立者(品系#9、#13、#20和#24)中获得下一代。用Southern印迹分析法测定转基因小鼠基因组的拷贝数。从条带的杂交强度判断,四只转基因小鼠携带少于10个拷贝的T3 β b-IL-12 p40构建体。通过北方印迹分析和RT-PCR检测来自转基因的mRNA表达。IL-12 p40 mRNA在转基因小鼠的大肠和小肠中均有表达。除胃肠道外,胸腺等主要器官均未检测到转基因mRNA。用IL-12(p40/p70)单克隆抗体对转基因小鼠和阴性同窝小鼠的大肠组织进行免疫组化分析。转基因小鼠大肠上皮细胞主要染色。用1.5%葡聚糖硫酸钠在T3 β b-IL-12 p40转基因小鼠和阴性同窝小鼠中诱导结肠炎。转基因小鼠与阴性同窝小鼠结肠炎评分差异不显著。

项目成果

期刊论文数量(0)
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Aihara H,Hiwatashi N,et al: "T3^b promoter directs specific expression on intestinal cells in trans genic mice" Gastroenterology. 116 (5). (1999)
Aihara H、Hiwatashi N 等人:“T3^b 启动子指导转基因小鼠肠细胞上的特异性表达”胃肠病学。
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H Aihara, N.Hiwatashi.et al: "T3^b promotor directs specific expression on intestinal epithelial cells in transgenic mice" Gastroenterology. 116・5. (1999)
H Aihara、N.Hiwatashi.et al:“T3^b 启动子指导转基因小鼠肠上皮细胞的特异性表达”Gastroenterology 116·5。
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相原裕之, 樋渡信夫, 他: "腸管特異的なプロモーターを用いたIL-12p40トランスジェニックマウスの作製" 日本消化器病学会誌. 96. (1999)
Hiroyuki Aihara、Nobuo Hiwatari 等人:“使用肠道特异性启动子创建 IL-12p40 转基因小鼠”,日本胃肠病学会杂志 96。(1999 年)
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H.Aihara, N.Hiwatashi, et al: "T3^b promotor dlrects specific expression on intestinal epithelial cells in transgenic mice." Gastroenterology. 116・5. (1999)
H.Aihara、N.Hiwatashi 等人:“T3^b 启动子指导转基因小鼠肠上皮细胞的特异性表达。胃肠病学”116·5。
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相原裕之、樋渡信夫、他: "腸管特異的なプロモーターを用いたIL-12p40トランスジェニックマウスの作製" 日本消化器病学会誌. 96. (1999)
Hiroyuki Aihara、Nobuo Hiwatari 等人:“使用肠道特异性启动子创建 IL-12p40 转基因小鼠”,日本胃肠病学会杂志 96。(1999 年)
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HIWATASHI Nobuo其他文献

HIWATASHI Nobuo的其他文献

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{{ truncateString('HIWATASHI Nobuo', 18)}}的其他基金

Regulation of TNFalpha and TNFbeta gene expression in inflammatory bowel disease
炎症性肠病中 TNFα 和 TNFβ 基因表达的调节
  • 批准号:
    03670345
  • 财政年份:
    1991
  • 资助金额:
    $ 2.3万
  • 项目类别:
    Grant-in-Aid for General Scientific Research (C)

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