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Molecular investigation on the fulminant hepatitis-resistant model animals

暴发性肝炎耐药模型动物的分子研究

基本信息

批准号：
10470253
负责人：
MIURA Naoyuki
金额：
$ 8.38万
依托单位：
HAMAMATSU UNIVERSITY SCHOOL OF MEDICINE (1999-2000)Akita University (1998)
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B).
财政年份：
1998
资助国家：
日本
起止时间：
1998 至 2000
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-10470253/
关键词：
Rb protein Transgenic mice Apoptosis Fulminant hepatitis Fas antigen TNFα Hepatocellular carcinoma Nodule トランスジェニックマウス HNF1 化学発癌 GOT 肝細胞変性出血トランスジーン

项目摘要

Two lines of the transgenic mice in which the human Rb cDNA was controlled under the) kbp of rat hepatocyte nuclear factor-1 (HNF-1) promoter/enhancer were generated. Transgenic mice, line A (TGA) had about 11 copies of the transgenes per haploid and line B (TGB) had about 4 copies of the transgenes. By Western blot analysis, we observed that a large amount of Rb protein was expressed in the liver of TGA mice and a small amount of Rb protein was expressed in that of TGB mice. In control mice, injection of anti-Fas antibody and TNFα induced increase of GOT and GPT in serum, hemorrhages and hepatocyte apoptosis in the histology. However, in TGA mice, the increase in GOT and GPT was marginal and no hemorrhages and apoptosis were detected In TGB mice, the increase in GOT and GPT was moderate and apoptosis of the substantial number of hepatocytes was observed, but no hemorrhages. In order to investigate the molecular mechanism of anti-apoptotic condition, we did the western blot analysis of … More apoptosis-related proteins. Fas, Bcl-2, Bcl-X, Bid, Bad, ICE, CPP32, E2F1, E2F2, E2F3, E2F4, E2F5 and p53 proteins had no differences between control mice and Rb transgenic mice. However, the Bax protein was decreased in Rb transgenic mice compared to control mice. This suggests that the Bax protein is on e of the contributing proteins for the anti-apoptotic condition.Next we investigated whether the Rb transgenic mice showed resistance to chemical carcinogenesis. We inject diethylnitrosamine into the peritoneal cavity at the age of 6 weeks and treated phenobarbital in drinking water for 35 weeks. After the experiment, the mice were sacrificed to make histological sections for counting the numbers of hepatocellular carcinoma and hepatic nodule. In control mice, a large number of nodules and several hepatocellular carcinoma were developed. In contrast, the number of nodules was greatly reduced and no hepatocellular carcinomas were detected in Rb transgenic mice. These results indicate that the Rb protein act as an anti apoptotic agent and an anti-tumorigenic agent in the liver in vivo. Less

在大鼠肝细胞核因子-1 （HNF-1）启动子/增强子的kbp范围内，制备了2株人Rb cDNA的转基因小鼠。转基因小鼠A系（TGA）每单倍体约有11个转基因拷贝，B系（TGB）每单倍体约有4个转基因拷贝。通过Western blot分析，我们发现TGA小鼠肝脏中有大量Rb蛋白表达，TGB小鼠肝脏中有少量Rb蛋白表达。在对照组小鼠中，注射抗fas抗体和TNFα可引起血清中GOT和GPT含量升高，组织学上出现出血和肝细胞凋亡。而在TGA小鼠中，GOT和GPT的增加是边缘性的，未见出血和凋亡。在TGB小鼠中，GOT和GPT的增加是中度的，有大量肝细胞凋亡，但未见出血。为了探讨抗凋亡的分子机制，我们对更多的凋亡相关蛋白进行了western blot分析。Fas、Bcl-2、Bcl-X、Bid、Bad、ICE、CPP32、E2F1、E2F2、E2F3、E2F4、E2F5和p53蛋白在对照小鼠和Rb转基因小鼠之间无差异。然而，与对照小鼠相比，转Rb基因小鼠的Bax蛋白含量降低。这表明Bax蛋白是抗凋亡条件的贡献蛋白之一。接下来，我们研究了Rb转基因小鼠是否表现出对化学致癌的抗性。我们在6周龄时腹腔注射二乙基亚硝胺，并在饮用水中使用苯巴比妥治疗35周。实验结束后处死小鼠，进行肝细胞癌和肝结节组织切片计数。对照组小鼠出现大量结节和几种肝细胞癌。相比之下，Rb转基因小鼠的肝结节数量大大减少，未发现肝细胞癌。这些结果表明Rb蛋白在体内肝脏中具有抗凋亡和抗肿瘤作用。少