APPLICATION OF GENE MANIPULATION TECHNOLOGY TO GENERATION OF MODEL ANIMALS FOR ENVIRONMENTAL ASSESSMENT

1.AhR高表达小鼠的建立。携带由角蛋白14启动子驱动的AhR基因的小鼠表现出皮肤的炎症和过度角化。这些小鼠现在正在接受检查，看它们是否更容易受到化学致癌物的影响。一代人AhR(HAHR)敲入小鼠。在ES细胞中通过同源重组将人AhR基因替换为小鼠AhR基因。通过将ES细胞注射到胚泡中，我们获得了表达人AhR分子的HAHR-敲入小鼠，而不是小鼠AhR。当暴露在3-甲基胆蒽中时，HAHR基因敲除小鼠表现出与对照动物相似的I相酶基因诱导。当使用TCDD时，小鼠的HAHR-爆震反应明显弱于对照组。我们怀疑，在HAHR-敲击小鼠中观察到的反应与人类的反应相似。该小鼠将作为环境化学物质毒性评价的动物模型。NRF2基因缺陷小鼠对环境化学物质的反应分析。Nrf2基因缺陷小鼠被几种环境化学物质挑战，以检查Nrf2在体内的贡献的意义。由于活性代谢中间产物水平的增加，NRF2缺陷小鼠对扑热息痛的肝毒性更加敏感。在暴露在柴油废气中的Nrf2缺失突变小鼠中，观察到DNA加合物水平增加。口服苯并[a]芘导致Nrf2基因缺陷小鼠比对照小鼠患上更严重的胃癌。这些结果表明，Nrt2是对环境化学物质毒性的保护性反应的关键调节因子。

1. Generation of AhR-overexpressing mouse. Mice carrying AhR cDNA driven by keratin14 promoter displayed inflammation and hyperkeratinization of the skin. These mice are now under examination whether they are more vulnerable to chemical carcinogens.2. Generation of human AhR (hAhR)-knock-in mice. The mouse AhR gene was replaced with human AhR cDNA by homologous recombination in ES cells. By injecting the ES cells into blastcysts, we generated hAhR-knock-in mouse expressing human AhR molecule instead of mouse AhR. When exposed to 3-methylcholanthrene, hAhR-knock-in mice displayed the comparable induction of the phase I enzyme genes with the control animals. When TCDD was used, hAhR-knock in mice showed much weaker response than the control mice. We suspect that the response observed in hAhR-knock in mouse mimics that of the humans. This mouse will serve as an animal model for the assessment of the toxicity of environmental chemicals.3. Analysis on the responses to environmental chemicals of Nrf2-deficient mice. Nrf2-deficient mice were challenged by several environmental chemicals to examine the significance of Nrf2 contribution in vivo. Nrf2-deficient mice were more sensitive to acetaminophen hepatotoxicity due to the increased level of active metabolic intermediates. Increased levels of DNA adducts was observed in Nrf2-null mutant mice after the exposure to diesel exhaust. Oral administration of benzo[a]pyrene caused severer gastric cancer in Nrf2-deficient mice than the control animals. These results indicate that Nrt2 is a key regulator of the protective responses against the toxicity of environmental chemicals.