Development of a serum-free co-culture of human intestinal epithelium cell-lines (Caco-2/HT29-5M21).

BACKGROUND: The absorptive and goblet cells are the main cellular types encountered in the intestine epithelium. The cell lineage Caco-2 is a model commonly used to reproduce the features of the bowel epithelium. However, there is a strong debate regarding the value of Caco-2 cell culture to mimick in vivo situation. Indeed, some authors report in Caco-2 a low paracellular permeability and an ease of access of highly diffusible small molecules to the microvilli, due to an almost complete lack of mucus. The HT29-5M21 intestinal cell lineage is a mucin-secreting cellular population. A co-culture system carried out in a serum-free medium and comprising both Caco-2 and HT29-5M21 cells was developed. The systematic use of a co-culture system requires the characterization of the monolayer under a given experimental procedure. RESULTS: In this study, we investigated the activity and localization of the alkaline phosphatase and the expression of IAP and MUC5AC genes to determine a correlation between these markers and the cellular composition of a differentiated monolayer obtained from a mixture of Caco-2 and HT29-5M21 cells. We observed that the culture conditions used (serum-free medium) did not change the phenotype of each cell type, and produced a reproducible model. The alkaline phosphatase expression characterizing Caco-2 cells was influenced by the presence of HT29-5M21 cells. CONCLUSION: The culture formed by 75% Caco-2 and 25% HT29-5M21 produce a monolayer containing the two main cell types of human intestinal epithelium and characterized by a reduced permeability to macromolecules.

Intestinal effects of long-lasting spermine ingestion by suckling rats.

Spermine ingestion induces the precocious maturation of the small intestine in suckling rats. Previous observations suggest that spermine-induced intestinal maturation is a two-step phenomenon. The first step is the elimination of immature enterocytes (4-10 h post spermine ingestion) and the second step is the replacement of previous immature cells by adult-type enterocytes (2-3 days post initial spermine administration). The spermine-induced maturation is reversible when spermine administration is stopped. This work was undertaken in order to check whether the extension of polyamine administration (for 3-7 days) after the appearance of spermine-induced maturation can retain the mature state of the small intestine. Our results indicate that extension of spermine administration does not prevent some parameters (sucrase and maltase specific activities) reverting to a typical 'immature' value while others remain at a typical 'mature' level (mucosal weight and lactase specific activity). Our results show that there are at least two different mechanisms in required for the control of spermine-induced maturation of the small intestine.

Short-term effects of spermine ingestion on the small intestine: a comparison of suckling and weaned rats.

We have previously shown that spermine, shortly after its ingestion, can induce the alteration of the morphology of the small intestine of suckling rats. It was proposed that this alteration is due to polyamine accumulation inside the epithelial cells. This could also be related to the fact that the intestine of the suckling rat is in an immature state. To shed light on this issue, disaccharidase and alkaline phosphatase activity assays, protein, DNA and RNA content measurements and polyamine concentration analysis were performed on the small intestine of suckling and weaned Wistar rats treated with spermine. Spermine did not induce the same intestinal alterations in weaned rats compared to suckling animals. Indeed, in sucklings, spermine administration induced a decrease of the protein, DNA, putrescine and spermidine intestinal content, suggesting a cell loss. The cell loss impaired the activity of intestinal enzymes: lactase, maltase and alkaline phosphatase. In weaned rats, the same treatment did not alter these parameters. Exogenous spermine by itself is not sufficient to induce the alterations described here and previously. The maturity degree of the small intestine could be the basis of this process.