Aberrant expression of cyclin D1 in pulmonary proliferative lesions induced by high doses of urethane in transgenic mice carrying the human prototype c-H-ras gene.

In our previous study, when rasH2 mice and non-transgenic (non-Tg) littermates were injected intraperitoneally with 1,000 mg/kg of urethane once or three times at two-day intervals, the incidence of lung proliferative lesions in rasH2 mice given triple doses of urethane was significantly increased, compared to that in rasH2 mice given a single dose of urethane, and the mutation frequency of the transgene in lung tumors in rasH2 mice given triple doses was lower than that in rasH2 mice given a single dose of urethane. In the present study, differential immunohistochemical expressions of Cyclin D1 and PCNA, that lead to abnormal cell proliferation and tumor development due to uncontrolled G1-S transition in the cell cycle, as well as p53 tumor suppressor gene in pulmonary proliferative lesions obtained from our previous study were investigated. Over-expression of Cyclin D1 in hyperplasias in rasH2 mice given triple doses was significantly increased, compared to that in the single-injection group, but no significant differences in Cyclin D1 between the single and triple injection groups were observed in hyperplasias in non-Tg mice or lung tumors in either rasH2 or non-Tg mice. There were no differences in the PCNA labeling index of hyperplasias in rasH2 or non-Tg mice between the triple-injection and single-injection groups, while the PCNA labeling index tended to be increased in the tumor, compared with that in hyperplasias. There was neither mutation of p53 nor an increase in immunoreactivity of wild type p53 in these proliferative lesions. These results suggest that cyclin D1 over-expression in alveolar/bronchiolar hyperplasias in rasH2 mice in the triple-injection group is not only indicative of a high cell proliferation rate but also of an important role in the process of malignant transformation.

Carcinogen dose-dependent variation in the transgene mutation spectrum in urethane-induced lung tumors in transgenic mice carrying the human prototype c-Ha-ras gene.

Urethane-induced lung tumors and their genetic changes were investigated in transgenic (Tg) mice carrying a human prototype c-Ha-ras gene (rasH2 mice). Male and female rasH2 mice and non-transgenic (non-Tg) littermates were injected intraperitoneally with 1000 mg/kg of urethane once or three times at 2-day intervals. Hyperplasias and adenomas of the lung were observed in all animals of each group from week 10, and carcinomas were observed in male and female rasH2 mice of the triple injection group from week 10 and female non-Tg mice of the single injection group at 15/20 weeks. The multiplicities of lung proliferative lesions including hyperplasias, adenomas and carcinomas, in treated rasH2 mice were significantly higher than those in treated non-Tg mice. CAG to CTG transversions were observed in the c-Ha-ras gene in these lung proliferative lesions of rasH2 mice of the single injection group at high incidence (male: 58.3%, female: 62.5%), but no mutations of the mouse c-Ki-ras gene were evident in either rasH2 or non-Tg mice. In the triple injection group, transgene mutations were detected at a relatively low incidence, and mouse c-Ki-ras gene mutations(CAA to CGA) were observed in both rasH2 and non-Tg mice. These results suggest that the variation of the lesions induced by different doses of urethane was not the cause of the variation of the mutation spectrum and mutations of both transgene and mouse c-K-ras gene are not principal genetic events in urethane-induced lung proliferative lesions in rasH2 mice.