Immunochemical, biomolecular and biochemical characterization of bovine epithelial intestinal primocultures
1 Department of Biochemistry and General Physiology, University of Liege, Institute of Chemistry B6C, B-4000 Liege (Sart-Tilman), Belgium
2 Department of Biology, University of Namur (FUNDP), Rue de Bruxelles, 61, B-5000 Namur, Belgium
3 Department of Infectious and Parasitic Diseases/Bacteriology, Faculty of Veterinary Medicine, University of Liege, Boulevard de Colonster B43, B-4000 Liege (Sart-Tilman), Belgium
BMC Cell Biology 2005, 6:42 doi:10.1186/1471-2121-6-42Published: 1 December 2005
Cultures of enterocytes and colonocytes represent valuable tools to study growth and differentiation of epithelial cells. In vitro models may be used to evaluate passage or toxicity of drugs, interactions of enteropathogenes bacteria strains with intestinal epithelium and other physiologic or pathologic phenomenon involving the digestive tract.
Cultures of bovine colonocytes and jejunocytes were obtained from organoid-enriched preparations, using a combination of enzymatic and mechanical disruption of the intestine epithelium, followed by an isopicnic centrifugation discarding most single cells.
Confluent cell monolayers arising from plated organoids exhibited epithelium typical features, such as the pavement-like structure, the presence of apical microvilli and tight junctions. Accordingly, cells expressed several markers of enterocyte brush border (i.e. maltase, alkaline phosphatase and fatty acid binding protein) as well as an epithelial cytoskeleton component (cytokeratin 18). However, enterocyte primocultures were also positive for the vimentin immunostaining (mesenchyme marker). Vimentin expression studies showed that this gene is constitutively expressed in bovine enterocytes. Comparison of the vimentin expression profile with the pattern of brush border enzymes activities, suggested that the decrease of cell differentiation level observed during the enterocyte isolation procedure and early passages of the primoculture could result from a post-transcriptional de-repression of vimentin synthesis. The low differentiation level of bovine enterocytes in vitro could partly be counteracted adding butyrate (1–2 mM) or using a glucose-deprived culture medium.
The present study describes several complementary approaches to characterize bovine primary cultures of intestinal cells. Cultured cells kept their morphologic and functional characteristics during several generations.