Caco-2 versus Caco-2/HT29-MTX co-cultured cell lines: permeabilities via diffusion, inside- and outside-directed carrier-mediated transport.
OBJECTIVE
The objective of this study was a systematic characterization and evaluation of cell culture models based on mixtures of Caco-2/HT29-MTX co-cultures for their use in screening for drug absorption and intestinal permeability in comparison to the properties of the respective mono-cultures.


METHODS
Co-cultures of Caco-2 cells (absorptive-type) and HT29-MTX cells (goblet-type) were set up. Three different co-cultures (initial seeding ratios Caco-2/HT29-MTX: 90/10, 70/30, and 50/50) were grown on permeable filter supports, and monolayers were used for permeability studies with model compounds for paracellular absorption (atenolol, furosemide, H334/75, mannitol, terbutaline), transcellular absorption (antipyrine, ketoprofen, metoprolol, piroxicam), carrier-mediated absorption (D-glucose, Gly-Pro, and L-phenylalanine) as well as substrates for carrier-mediated secretion via P-glycoprotein (cimetidine and talinolol). Electrophysiological and microscopic controls were performed to characterize the cell cultures.


RESULTS
For compounds undergoing passive intestinal absorption permeabilities were generally higher in co-cultures than in Caco-2 monolayers, yielding highest values in pure HT29-MTX monolayers. This difference was most obvious for compounds transported via the paracellular pathway, where HT29-MTX cells may be up to 30 times more permeable than Caco-2 cells, whereas for lipophilic and highly permeable compounds, the difference in permeability values was less obvious. For drugs undergoing intestinal secretion mediated by P-glycoprotein, co-cultivation of Caco-2 cells with HT29-MTX cells led to increased apical to basolateral permeability which was decreased in the opposite direction, consistent with the fact that HT29-MTX cells do not express P-glycoprotein. When a carrier-mediated absorption mechanism is involved, the permeabilities observed were lower than the values reported for human small intestine and co-cultivation of HT29-MTX cells with Caco-2 cells resulted in even lower values as compared to the plain Caco-2 cultures.


CONCLUSIONS
Co-cultures of HT29-MTX and Caco-2 cells offer the opportunity of modifying the permeability barrier of the cell monolayers both with respect to paracellular resistance and secretory transport via P-gp. Thus, in special cases, they allow more flexibility in adapting the in vitro system to the in vivo situation as compared to the monocultures. Another advantage is the obvious robustness of the method with respect to the reproducibility of the results. A problem remaining, however, is the quantitative expression of carriers involved in intestinal uptake of many nutrients and drugs.
Hilgendorf
C
C
School of Pharmacy, Martin Luther-University, Wolfgang-Langenbeck-Strasse 4, 06120 Halle/Saale, Germany.
Spahn-Langguth
H
H
Regårdh
C G
CG
Lipka
E
E
Amidon
G L
GL
Langguth
P
P
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000378:metabolism; D000983:Antipyrine / Q000493:pharmacokinetics; D001262:Atenolol / Q000493:pharmacokinetics; D001693:Biological Transport, Active; D018938:Caco-2 Cells / Q000378:metabolism* / Q000502:physiology / Q000648:ultrastructure; D002352:Carrier Proteins / Q000378:metabolism*; D002463:Cell Membrane Permeability / Q000502:physiology; D016922:Cellular Senescence / Q000502:physiology; D018920:Coculture Techniques; D005665:Furosemide / Q000493:pharmacokinetics; D019073:HT29 Cells / Q000378:metabolism* / Q000502:physiology / Q000648:ultrastructure; D006801:Humans; D007408:Intestinal Absorption / Q000502:physiology*; D007413:Intestinal Mucosa / Q000378:metabolism; D007660:Ketoprofen / Q000493:pharmacokinetics; D008353:Mannitol / Q000493:pharmacokinetics; D008790:Metoprolol / Q000493:pharmacokinetics; D008854:Microscopy, Electron; D010894:Piroxicam / Q000493:pharmacokinetics; D015203:Reproducibility of Results; D013726:Terbutaline / Q000493:pharmacokinetics
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D002352:Carrier Proteins; D010894:Piroxicam; D008353:Mannitol; D001262:Atenolol; D005665:Furosemide; D007660:Ketoprofen; D008790:Metoprolol; D013726:Terbutaline; D000983:Antipyrine