Candrian U, You M, Goodrow T, Maronpot R R, Reynolds S H, Anderson M W
Laboratory of Molecular Toxicology, National Institute of Environmental Health Sciences, Research Triangle Park, North Carolina 27709.
Cancer Res. 1991 Feb 15;51(4):1148-53.
The C57BL/6 x C3H F1 (hereafter called B6C3F1) mouse is an important animal model for long-term carcinogenesis studies. Maintained under normal laboratory conditions, these mice develop various types of spontaneous tumors during their lifetime. Activated Ha-ras genes have been detected in 66% of spontaneous hepatocellular tumors in the B6C3F1 mouse [Reynolds et al., Science (Washington DC), 237:1309, 1988]. In this study 49 spontaneous non-liver tumors were investigated for oncogene activation by DNA transfection techniques. Of the 49 tumor DNAs analyzed, only 5 yielded multiple foci in the NIH 3T3 focus assay: 2 of 10 pulmonary adenocarcinomas; 0 of 25 lymphomas; 2 of 2 Harderian gland adenomas; 0 of 1 adenocarcinoma of the small intestine; 1 of 6 malignant skin tumors; 0 of 4 hemangiosarcomas; and 0 of 1 lung metastasis of a hepatocellular carcinoma. DNA from six lymphomas which were negative in the NIH 3T3 focus assay were further analyzed for transforming genes by the nude mouse tumorigenicity assay. One of the five lymphomas tested positive with this assay. Southern blot analysis identified five activated ras genes: H-ras in two Harderian gland adenomas; K-ras in one pulmonary adenocarcinoma and in one s.c. adenocarcinoma; and N-ras in one lymphoma. The mutations involved were CG to AT and AT to TA in codon 61 of the Ha-ras genes, GC to AT or TA in codon 12 of the K-ras genes, and a GC to AT mutation in codon 12 of the N-ras gene. Transformant DNA from a pulmonary adenocarcinoma which yielded multiple foci in the transfection assay did not hybridize to DNA probes specific for the K-, H-, and N-ras, raf, neu, and met genes. Thirteen additional tumor DNAs yielded a single focus in the NIH 3T3 transfection assay. The transformant DNAs retransmitted in a second cycle transfection assay. Rearranged and/or amplified raf genes were detected in six of the transformant DNAs. At present we do not know whether these activated raf genes were present in the original tumor DNA. The other seven transformant DNAs did not hybridize with any of the above mentioned specific DNA probes utilized in Southern blot analysis. Unlike liver tumors, the activation of ras protooncogenes is not a frequent event in the development of spontaneous non-liver tumors of the B6C3F1 mouse. The results from this study should aid in understanding the neoplastic development associated with exposure to chemical carcinogens in the B6C3F1 mouse.
C57BL/6×C3H F1（以下简称B6C3F1）小鼠是长期致癌研究的重要动物模型。在正常实验室条件下饲养时，这些小鼠在其一生中会发生各种类型的自发性肿瘤。在B6C3F1小鼠66%的自发性肝细胞瘤中检测到激活的Ha-ras基因[雷诺兹等人，《科学》（华盛顿特区），237:1309，1988]。在本研究中，通过DNA转染技术对49个自发性非肝肿瘤进行了癌基因激活研究。在分析的49个肿瘤DNA中，只有5个在NIH 3T3焦点试验中产生了多个病灶：10个肺腺癌中的2个；25个淋巴瘤中的0个；2个哈氏腺腺瘤中的2个；1个小肠腺癌中的0个；6个恶性皮肤肿瘤中的1个；4个血管肉瘤中的0个；以及1个肝细胞癌肺转移瘤中的0个。对在NIH 3T3焦点试验中呈阴性的6个淋巴瘤的DNA，通过裸鼠致瘤性试验进一步分析其转化基因。所检测的5个淋巴瘤中有1个在此试验中呈阳性。Southern印迹分析鉴定出5个激活的ras基因：2个哈氏腺腺瘤中的H-ras；1个肺腺癌和1个皮下腺癌中的K-ras；以及1个淋巴瘤中的N-ras。所涉及的突变分别为Ha-ras基因第61密码子中的CG突变为AT和AT突变为TA，K-ras基因第12密码子中的GC突变为AT或TA，以及N-ras基因第12密码子中的GC突变为AT。在转染试验中产生多个病灶的肺腺癌的转化体DNA，与针对K-、H-和N-ras、raf、neu和met基因的DNA探针不杂交。另外13个肿瘤DNA在NIH 3T3转染试验中产生了单个病灶。这些转化体DNA在第二轮转染试验中再次传递。在6个转化体DNA中检测到重排和/或扩增的raf基因。目前我们不知道这些激活的raf基因是否存在于原始肿瘤DNA中。其他7个转化体DNA与Southern印迹分析中使用的上述任何特异性DNA探针均不杂交。与肝肿瘤不同，ras原癌基因的激活在B6C3F1小鼠自发性非肝肿瘤的发生过程中并非常见事件。本研究结果应有助于理解B6C3F1小鼠接触化学致癌物相关的肿瘤发生发展过程。
