Human cervical cells immortalized in vitro with oncogenic human papillomavirus DNA differentiate dysplastically in Vivo

Abstract

Human papillomavirus (HPV) types 16 and 18 are associated with cervical dysplasia and carcinoma. In vitro integration of HPV-16 or HPV-18 DNA into cultured human cervical cells results in their immortalization. In this study, in vivo differentiation of human cervical cells immortalized with recombinant HPV-16 or HPV-18 DNA was studied by implanting confluent monolayers of HPV-16 and HPV-18 DNA immortalized cell lines under dorsal skin flaps of immunodeficient mice. Grafts of a human cervical cancer cell line (C4-1) and cultured normal cervical cells served as controls. Histologic analysis 2 to 3 weeks after grafting demonstrated dysplastic differentiation of the HPV-16 and HPV-18 cell lines as characterized by abnormal mitotic figures, nuclear pleomorphism, and loss of basal polarity. RNA in situ hybridization confirmed the presence of viral transcripts in the dysplastic grafts. G-banded chromosome analysis showed that both the HPV-16 and HPV-18 cell lines were aneuploid. Even after prolonged periods of implantation tumors did not form and invasion of underlying mouse stroma did not occur. Normal cervical cells differentiated normally and the C4-1 cell line formed tumors and demonstrated more pronounced nuclear abnormalities than the HPV immortalized grafts. Integration of HPV-16 and HPV-18 DNA into cervical cells leads to an indefinite growth capacity. These aneuploid cells differentiate abnormally in vivo, but do not form tumors or invade host tissue, consistent with a multistage theory of cervical carcinogenesis.