Fibroblasts and transforming growth factor beta induce organization and differentiation of T84 human epithelial

Background & Aims: The gut epithelium in the cryptvillus axis represents a continuous developmental system in which the role of fibroblast-epithelial interactions is obvious. The aim of this study was to establish an in vitro method whereby fibroblast-guided differentiation of crypt-like gut epithelial cells can be studied. Methods: Intestinal epithelial cells (T84 and HT-29) were cultured within type I collagen gel together with fibroblasts without cell-to-cell contact. T84 cells were also grown in the presence of transforming growth factor beta and hepatocyte growth factor. The gels were studied using light and electron microscopy and histochemical and immunohistochemical methods. Results: The epithelial cells formed unorganized cell clusters within the gels, but when given fibroblast support, 76% of the T84 cell colonies (not HT-29) organized into luminal formations, and basement membranes including laminin were well deposited. The cells in the columnar single cell-layer luminal formations (49% of all colonies) were differentiated, showing microvilli, up-regulated alkaline phosphatase brush border activity, and mucin profiles typical for small intestine. This fibroblast-induced organization and differentiation was induced by transforming growth factor beta. Conclusions: Crypt-like T84 epithelial cells are able to differentiate when grown three-dimensionally together with fibroblasts or transforming growth factor beta. This method may be used for mesenchymal-epithelial cell cross-talk studies.