Transforming growth factor beta1 induces differentiation in human papillomavirus-positive keratinocytes

Human papillomaviruses (HPVs) are implicated in the etiology of anogenital cancers. Expression of the HPV E6 and E7 oncoproteins is believed to contribute to the carcinogenic process. Progressive loss of the ability to differentiate and resistance to the growth-inhibitory effects of endogenous signals also appear important in multistep tumorigenesis. Transforming growth factor beta1 (TGF-beta1) is a potent growth inhibitor for a variety of cultured cells. There have been conflicting reports on the ability of TGF-beta1 to inhibit the growth of HPV-positive keratinocytes in monolayer cultures. We have employed the organotypic (raft) tissue culture system, which more accurately mimics the in vivo cellular environment and architecture. We have investigated the TGF-beta1 response of HPV-positive keratinocytes derived from neoplastic cervical biopsies. Growth of these cell lines as raft tissues showed that many were altered in the ability to stratify and synthesize differentiation-specific proteins. When the organotypic tissues were treated with TGF-beta1, a more complete differentiation of the keratinocytes was induced. Treatment with 12-0-tetradecanoylphorbol-13-acetate gave similar results. TGF-beta1 treatment of HPV-positive raft epithelia led to a dose-dependent increase in E7 RNA expression in contrast to results from previous studies with monolayer cultures. Furthermore, TGF-beta1 interfered with the proliferation of HPV-positive cell lines grown in monolayer cultures. Our results suggest that loss of the ability to express markers of differentiation, a characteristic of malignancy, is a two-step process. The first step is reversible; the second is irreversible.