Prevention of Carcinoma In Situ of Human Papillomavirus Type 16-Immortalized Human Endocervical Cells by Retinoic Acid in Organotypic Raft Culture

Abstract

Objective: To determine the effect of retinoic acid on the development of severe dysplasia or carcinoma in situ from endocervical cells containing human papillornavirus (HPV) type 16. Methods: Two independent lines of HPV 16-immortalized endocervical cells were reconstructed into two squamous epithelial tissues using the organotypic raft culture system to examine the differentiated phenotype. The effect of retinoic acid on dysplastic morphology of differentiation of the epithelia was examined by light microscopy of stained sections and electron microscopy. The endocervical cell type cytokeratin expression pattern was determined by indirect immunofluorescence using specific monoclonal antibodies. Ribonucleic acid expression of the HPV 16 E7 oncogene was examined by in situ hybridization. Results: Untreated HPV 16-immortalized endocervical cells were reconstructed into squamous dysplastic lesions resembling carcinoma in situ observed in women. Retinoic acid-treated rafts formed epithelia composed of two to three cell layers of columnar-like cells resembling simple epithelium of the endocervix. Electron microscopy and cytokeratin expression patterns confirmed the histology of a differentiated endocervical phenotype after treatment with retinoic acid. Expression of HPV 16 E7 was modestly lower in treated epithelia, preferentially in basal cells. Conclusion: Retinoic acid prevents the histology and cytokeratin differentiation markers of carcinoma in situ of HPV 16-immortalized endocervical cells. Because the epithelia closely mimic HPV 16-containing severe dysplasias and native endocervical epithelium in women, this immortalized endocervical cell-raft system may be useful as a model to assess the efficacy of agents such as retinoic acid for preventing progression of these lesions to malignant cervical carcinoma.