Human skin fibroblasts induce anchorage‐independent growth of HPV‐16‐DNA‐immortalized cervical epithelial cells

Abstract

Immortalized non-malignant cells do not grow in soft agarose. We found, however, that HPV-16-DNA-immortalized human uterine exocervical epithelial cells (HCE16/3 cell line) formed colonies when co-cultured with human embryonic skin fibroblasts. This appeared to be mediated by diffusible growth factors, and direct cell-cell contacts were not required. HCE16/3 cells from colonies remained unable to grow alone in soft agarose like the parental HCE16/3 cells, indicating that only transient phenotypic changes without genetic alterations had occurred in the co-cultures. In a modified soft-agarose assay, HCE16/3 cells grown as an underlying monolayer did not produce enough (transforming) growth factors which could confer an anchorage-independent phenotype to non-transformed fibroblasts. Lethally irradiated fibroblasts were able to stimulate DNA synthesis and proliferation of HCE16/3 cells. Northern-blot analysis showed that HPV-16-DNA expression of the immortalized cells was not altered in the presence of fibroblasts. The expression of the IL-I gene in the immortalized cells, however, was augmented by the co-cultured fibroblasts compared with immortalized cells alone. In organotypic collagen raft co-cultures, in which epithelial cells were grown on top of a collagen gel containing fibroblasts, HCE16/3 cells appeared to be chemotactic to fibroblasts and fibroblasts stimulated the growth of the immortalized epithelial cells. Our results suggest that fibroblasts may contribute to human epithelial carcinogenesis in vivo.