Enterocytes differentiation

The main reasons for the popularity of the Caco-2 cell line are that the cells are easy to maintain in culture, and they develop unusually high degree of differentiation spontaneously under standard culture conditions. In fact, Caco-2 is the only human intestinal cell line that has been found so far spontaneously to undergo functional enterocytic differentiation. The cells exhibit a good reproducibility, robustness and functional properties of human intestinal epithelial cells. The model has proved capable of predicting the oral absorption of a variety of drug compounds [see references in 10]. 

E., Brattain, M. G., Zweibaum, A., Epithelial polarity, villin expression, and enterocytic differentiation of cultured human colon carcinoma cells a survey of twenty cell lines, Cancer Res. 1988, 48, 1936-1942. 

There are several cell monolayer models that are frequently used for the evaluation of drug permeability and absorption potential (Table 18.1). For a more detailed discussion, please refer to Chap. 8. Caco-2 cells (adenocarcinoma cells derived from colon) are the most extensively characterized and frequently used of the available cell lines [5-9], A unique feature of Caco-2 cells is that they undergo spontaneous enterocyte differentiation in cell culture. Unlike intestinal enterocytes, Caco-2 cells are immortalized and replicate rapidly into confluent monolayers. When the cells reach confluency during culture on a semi-porous membrane, they start to polarize and form tight junctions, creating an ideal system for permeability and transport studies. During the past decade, use of... 

Hypocalcemia directly increases PTH synthesis and release and inhibits calcitonin release. PTH in turn restores plasma calcium by initially stimulating transport of free or labile calcium from bone into the blood. PTH also increases renal 1,25-dihydroxycholecalciferol (1,25-(0H)2D3) production, which is the most active form of D3. 1,25-(0H)2D3 induces enterocyte differentiation in the intestine, which in turn results in increased absorption of calcium. Finally, during long periods of hypocalcemia, PTH can mobilize more stable calcium deep in the hydroxyapatite of bone by activating deep osteoclasts. 

Carriere V, Lessufleur T, Barbat A et al. (1994) Expression of cy-tochrom P450 3A in HT-29-MTX cells and CACO-2 clone TC7. FEBS Letter 355 247-250 Chantret I, Barbat A, Dussaulx E et al. (1988) Epithelial polarity, villin expression, and enterocytic differentiation of cultured human colon carcinoma cells a survey of 20 cell-lines. Cancer Res 48 1936-1942 Cordon-Cardo C, O Brien JP, Boccia J et al. (1990) Expression of the multidrug resistance product (P-glycoprotein) in human normal and tumor tissues. J Histochem Cytochem 38 1277-1287... 

Jumarie Cand Malo C (1991) CACO-2 cells cultured in serum-free medium as a model for the study of enterocytic differentiation in vitro. J Cell Physiol 149 24-33 Karlsson J, Ungell AL, Artursson P (1994) Effect of an oral rehydration solution on paracellular drug transport in intestinal epithelial cells and tissues assessment of charge and tissue selectivity. Pharm Res 11 248... 

Zweibaum A, Traidou N, Kedinger M et al. (1983) Sucrase-isomaltase a marker of foetal and malignant epithelial cells of the human colon. Int J Cancer 32 407 112 Zweibaum A, Pinto M, Chevalier G et al. (1985) Enterocytic differentiation of a subpopulation of the human colon tumor cell line HT-29 selected for growth in sugar-free medium and its inhibition by glucose. J Cell Physiol 122 21-29... 

M Pinto, MD Appay, P Simon-Assman, G Chevalier, N Dracopoli, J Fogh, A Zweibaum. Enterocytic differentiation of cultured human cancer cells by replacement of glucose by galactose in the medium. Biol Cell 44 193—196,1982. 

Changes in the expression and activity ofC2GnTl occur upon activation of lymphocytes with interleukin-2/ and after enterocytic differentiation of Caco-2 cells. This indicates a central role of C2GnT in the biological responses of cells. 

Adenocarcinoma Primary EM20 Hypertetraploid exhibits enterocyte differentiation tumorigenic... 

Reynier, M, Sari, H, d Anglebermes, M, Ah Kye, E and Pasero, L (1991) Differences in lipid characteristics of undifferentiated and enterocytic differentiated HT human colonic cells. 

In culture, the human colon carcinoma cell hne Caco-2 spontaneously differentiates at confluency into polarized cells with enterocyte-like characteristics. The principle of this approach consists of following the passage of the compound of interest from the apical or lumen-like sides to the basolateral or lymph-hke sides of Caco-2 cells, thus following the absorption of the compound per se. One obhgate step for fat-soluble nutrients such as carotenoids to cross the intestinal barrier is their incorporation into CMs assembled in the enterocytes. Under normal cell culture conditions, Caco-2 cells are unable to form CMs. When supplemented with taurocholate and oleic acid, Caco-2 cells were reported to assemble and secrete CMs. ... 

The Caco-2 cell line was isolated from a human colon carcinoma, and has been characterized as one of the best in vitro models of intestinal epithelium. Indeed, in contrast to other intestinal cell lines, Caco-2 cells are able to constitute a homogenous monolayer and to spontaneously differentiate into polarized cells, highly similar to human mature enterocytes, after approximately 2 weeks of culture. Furthermore, the Caco-2 cells present microvillosities at the apical side and have a high transmembrane resistivity, which confirms the fact that the cells are confluent and link to one another via gap junctions. Finally, they can absorb different compounds, express many enzymes involved in intestinal metabolic pathways (Pinto et al. 1983, Musto et al. 1995, Salvini et al. 2002), and give reproducible in vitro results consistent with results obtained in in vivo studies (Artursson and Karlsson 1991). 

Pinto, M. et al. (1983). Enterocyte-like differentiation an polarizationof the human colon carcinoma cell line Caco-2 in culture. Biol. Cell 47 323-330. 

The situation prevailing in the crypt cell at the beginning of its differentiation into an enterocyte and before it has begun to climb towards the villus is shown in the lower panel. The cell s iron requirements are supplied by receptor-mediated diferric transferrin uptake from the basolateral membrane. The TfR in turn is involved in an interaction with the HFE protein, which decreases the affinity of TfR for diferric transferrin. The level of transferrin saturation, or some other factor, determines the amount of iron taken up, and presets the IRP system at a level that corresponds to the iron requirements of the organism. 

The enthusiasm for using Caco-2 cells and other epithelial cell cultures in studies of drug transport processes has been explained by the ease with which new information can be derived from these fairly simple in vitro models [7]. For instance, drug transport studies in Caco-2 cells grown on permeable supports are easy to perform under controlled conditions. This makes it possible to extract information about specific transport processes that would be difficult to obtain in more complex models such as those based on whole tissues from experimental animals. Much of our knowledge about active and passive transport mechanisms in epithelia has therefore been obtained from Caco-2 cells and other epithelial cell cultures [10-15]. This has been possible since Caco-2 cells are unusually well differentiated. In many respects they are therefore functionally similar to the human small intestinal enterocyte, despite the fact that they originate from a human colorectal carcinoma [16, 17]. 

Paul, E. C. A., J. Hochman, and A. Quaroni. Conditionally immortalized intestinal epithelial cells, novel approach for study of differentiated enterocytes., Am. J. Physiol. 1993, 265, C266-C278... 

M. D., Kedinger, M., Triadou, N., Dussaulx, E., Lacroix, B., Simon-Assman, P., Happen, K., Fogh, J., Zweibaum, A., Enterocyte-like differentiation and polarization of the human colon carcinoma cell line Caco-2 in culture, Biol. Cell 1983, 47, 323-330. 

Cell monolayers grown on permeable culture inserts form confluent mono-layers with barrier properties and can be used for drug absorption experiments. The most well-known cell line for the in vitro determination of intestinal drug permeability is the human colon adenocarcinoma Caco-2 [20, 21], The utility of the Caco-2 cell line is due to its spontaneous differentiation to enterocytes under conventional cell culture conditions upon reaching confluency on a porous membrane to resemble the intestinal epithelium. This cell model displays small intestinal carriers, brush borders, villous cell model, tight junctions, and high resistance [22], Caco-2 cells express active transport systems, brush border enzymes, and phase I and II enzymes [22-24], Permeability models... 

Pine, L., Kathariou, S., Quinn, F., George, V., Wenger, J. D., and Weaver, R. E. (1991). Cytopathogenic effects in enterocyte-like Caco-2 cells differentiate virulent from aviru-lent Listeria strains. /. Clin. Microbiol. 29,990-996. 

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