Control of differentiation in a rectal adenocarcinoma cell line: the role of diffusable and cell-associated factors

The rectal adenocarcinoma cell line, HRA-19a1.1, was cultured in a three-dimensional type I collagen gel and then xenografted in nu/nu mice to determine whether the in vivo environment could induce further morphological differentiation of the in vitro collagen gel cultures. Three phenotypic changes were observed. Type I collagen induces glandular differentiation in vitro in which well polarized cells are organized around a central lumen, as has been previously reported. Seven days after xenografting this structure in nu/nu mice, the glandular structures appeared to have 'ballooned' forming cyst-like structures lined by a monolayer of flattened cells. There were no stromal cells associated with the graft at this stage, but with time stromal cells invaded the collagen. At points where these cells were closely associated with the HRA-19 cells there was a marked phenotypic change, with the flattened lining cells seen at day 7 becoming columnar. By 21 days the stromal cells had replaced the collagen and the histology of the graft now resembled that of an adenocarcinoma. Placing this cell-collagen culture in a Millipore chamber prior to grafting resulted in cyst-like structures only. Here we provide conclusive evidence that heterologous connective tissue cells can induce differentiation of a rectal adenocarcinoma cell line by a non- or poorly diffusible factor(s). Furthermore, we show that this cell-collagen xenograft method has certain advantages over conventional xenograft methods: notably, a consistent 100 per cent take rate; considerably fewer cells are required to form a tumour; and the time taken to form a tumour is dramatically reduced.