Enhanced tumorigenesis and reduced transforming growth factor-beta type IIreceptor in lung tumors from mice with reduced gene dosage of transforminggrowth factor-beta 1

To elucidate the role of transforming growth factor-beta1 (TGF-beta1) and t he TGF-beta type II receptor (TGF-beta RII)as tumor-suppressor genes in lun g carcinogenesis, we mated C57BL/6 mice heterozygous (HT) for deletion of t he TGF-beta1 gene with A/J mice to produce AJBL6 TGF-beta1 HT progeny and t heir wild-type (WT) littermates. Immunohistochemical staining, in situ hybr idization, and northern blot analyses showed lower staining and hybridizati on for TGF-beta1 protein and mRNA, respectively, in the lungs of normal HT mice versus WT mice. Competitive reverse transcription-polymerase chain rea ction (CRT-PCR) amplification showed the level of TGF-beta1 mRNA in the lun gs of HT mice to be fourfold lower than the level in WT lung. When challeng ed with ethyl carbamate, lung adenomas were detected in 55% of HT mice by 4 mo but only in 25% of WT littermates at this time. Whereas all HT mice had adenomas by 6 mo, it was not until 10 mo before all WT mice had adenomas. After 12 mo, the average number of adenomas was fivefold higher in HT lungs than in WT lungs. Most dramatic was the appearance of lung carcinomas in H T mice 8 mo before they were visible in WT mice. Thus, the AJBL6 TGF-beta1 HT mouse provides an excellent model system to examine carcinogen-induced l ung tumorigenesis by increasing progressive lesion incidence and multiplici ty relative to their WT littermates. Immunohistochemical staining showed ex pression of the TGF-beta type I receptor (TGF-beta RI) at moderate to stron g levels in lung adenomas and carcinomas in HT and WT mice. In contrast, wh ereas weak immunostaining for TGF-beta RII was detected in 67% of HT carcin omas at 12 mo, only 22% of WT carcinomas showed weak staining for this prot ein. individual lung carcinomas showing reduced TGF-beta RII expression and adjacent normal bronchioles were excised from HT lungs using laser capture microdissection, and CRT-PCR amplification of the extracted RNA showed 12- fold less TGF-beta RII mRNA in these carcinomas compared with bronchioles. Decreasing TGF-beta RII mRNA levels occurred with increasing tumorigenesis in lung hyperplasias, adenomas, and carcinomas, with carcinomas having four fold and sevenfold lower levels of TGF-beta RII mRNA than adenomas and hype rplasias, respectively. These data show enhanced ethyl carbamate-induced lu ng tumorigenesis in AJBL6 HT mice compared with WT mice, suggesting that bo th TGF-beta1 alleles are necessary for tumor-suppressor activity. Reduction of TGF-beta RII mRNA expression in progressive stages of lung tumorigenesi s in HT mice suggests that loss of TGF-beta RII may play an important role in the promotion of lung carcinogenesis in mice with reduced TGF-beta1 gene dosage when challenged with carcinogen. Mol. Carcinog. 29:112-126, 2000. P ublished 2000 Wiley-Liss. Inc.(dagger).