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Emergence of permanently differentiated cell clones in a human colonic cancer cell line in culture after treatment with sodium butyrate.

The human colonic cancer cell line HT29 is undifferentiated in standard culture conditions (Dulbecco's medium:10% fetal bovine serum). These cells were cultured in 5 mM sodium butyrate for 9 days; then they were trypsinized and subcultured in sodium butyrate for an additional 14 days. Multinucleation occurred during this second phase of the treatment. The cells were then transferred to standard medium and multinucleation disappeared. Morphological changes appeared 10 to 12 days after return to standard culture conditions; some cells flattened and became more adherent to the bottom of the flasks. These altered cells divided actively and formed "flat foci" interspersed among the densely packed undifferentiated HT29 cells. This altered phenotype persists after more than 24 months of culture in standard medium. Clonal cell lines were established from these flat foci-forming cells and characterized. These clonal lines exhibited morphological cell polarity defined by an apical cell surface separated by junctional complexes from the basolateral cell surface. Functional differentiation did also occur since some clonal lines formed domes representing active transepithelial transport, and others exhibited massive mucus secretion. In conclusion, our findings indicate that permanently differentiated cell populations emerged in a colonic cancer cell line after sodium butyrate treatment. These new clonal lines will be useful in future models for the study of differentiation programs of both normal and cancerous colonic cells.