背景:HRASKO/NRASKO双基因敲除小鼠表现出极高的围产期致死率,这是由于显著的肺成熟延迟引起的呼吸衰竭。少数成年动物的寿命正常,但目前肺不张区域与肺气肿和正常组织混合。
方法:使用微X射线计算机断层扫描和小动物生理监测系统分析了八只双敲除小鼠和八只对照小鼠。使用标准组织学和分子生物学方法分析来自这些小鼠的组织和样品,并使用学生T检验分析结果的显著性。
结果:存活到成年期的极少数双基因敲除小鼠表现出明显的颅面异常,让人联想到在RASopathy小鼠模型中看到的异常,以及血小板减少症,出血异常,并降低凝血酶诱导的血小板活化。这些存活的小鼠还出现心脏和脾脏增生,脾脏中骨髓来源的抑制细胞数量增加。机械上,我们观察到这些表型改变伴随着心脏中KRAS-GTP水平的升高,这些动物的血小板和原代小鼠胚胎成纤维细胞。
结论:我们的数据揭示了一个新的,由于联合去除HRAS和NRAS,能够在小鼠中触发RASopathy表型的先前未识别的机制。
BACKGROUND: HRASKO/NRASKO double knockout mice exhibit exceedingly high rates of perinatal lethality due to respiratory failure caused by a significant lung maturation delay. The few animals that reach adulthood have a normal lifespan, but present areas of atelectasis mixed with patches of emphysema and normal tissue in the lung.
METHODS: Eight double knockout and eight control mice were analyzed using micro-X-ray computerized tomography and a Small Animal Physiological Monitoring system. Tissues and samples from these mice were analyzed using standard histological and Molecular Biology methods and the significance of the results analyzed using a Student´s T-test.
RESULTS: The very few double knockout mice surviving up to adulthood display clear craniofacial abnormalities reminiscent of those seen in
RASopathy mouse models, as well as thrombocytopenia, bleeding anomalies, and reduced platelet activation induced by thrombin. These surviving mice also present heart and spleen hyperplasia, and elevated numbers of myeloid-derived suppressor cells in the spleen. Mechanistically, we observed that these phenotypic alterations are accompanied by increased KRAS-GTP levels in heart, platelets and primary mouse embryonic fibroblasts from these animals.
CONCLUSIONS: Our data uncovers a new, previously unidentified mechanism capable of triggering a
RASopathy phenotype in mice as a result of the combined removal of HRAS and NRAS.