Primary cultures of fully differentiated and pure human intestinal epithelial cells

The epithelium of the small intestinal mucosa is a highly dynamic system pa rticularly well suited for analyzing key biological phenomena such as cell differentiation and migration, cell-matrix interactions, and apoptosis. How ever, in vitro models of fully differentiated normal human enterocytes are still lacking. The objective of the present study was to investigate the po ssibility of generating such differentiated intestinal cell cultures from t he fetal small intestine. For this purpose, various dissociation methods we re tested in order to obtain pure, viable, and functional enterocytes. One of these methods, based on the procedure to recover epithelial cells grown on Matrigel and involving the use of Matrisperse, a nonenzymatic dissociati ng solution, was found to allow the isolation of the integral epithelial li ning from the mesenchyme. In culture, these epithelial fractions plated on collagen I spread rapidly and reached confluence after 3-4 days. When teste d after 5-7 days, these primary cultures of differentiated enterocytes (PCD E) remained well preserved. Both goblet and absorptive cells exhibit all th e main characteristics of intact villus intestinal cells as assessed by ele ctron microscopy. Indirect immunofluorescence and Western blot analyses con firmed the purity of PCDE. The functional status of these cells was demonst rated by the presence of uniformly distributed tight junction, zonula adher ens, and desmosomal components at the region of cell to cell contact as wel l as expression of various brush border enzymes, namely sucrase-isomaltase and lactase, and goblet cell mucins. As expected, cell proliferation was fo und to be negligible as assessed by DNA synthesis. Taken together, these da ta show that primary cultures of pure and viable differentiated enterocytes can be generated from the human fetal small intestine. (C) 1998 Academic P ress.