Caco-2 cell cultures in the assessment of intestinal absorption: Effects of some co-administered drugs and natural compounds in biological matrices

Several different in vitro absorption models are used in the screening of new drug candidates. One of them is the Caco-2 model, a widely used in vitro model for small intestinal absorption. Caco-2 cells, which originate from a colon carcinoma, differentiate spontaneously to cells that resemble mature small intestinal enterocytes and express carrier proteins similar to the small intestine, and can therefore be used for the assessment of active transport processes during intestinal absorption. Due to variation in cell line differentiation and selection of subpopulations, permeability data obtained from different laboratories is seldom directly comparable. Hence, the use of several drugs or compounds with known permeability characteristics are recommended for model standardisation and the use of internal standards in every experiment could offer a solution to the problem. In this study, the simultaneous use of several drugs as internal standards was evaluated. Drugs with different permeability characteristics (high and low permeability, passive and active transport and active efflux) were used to detect their possible effects on cell viability, monolayer integrity and possible interactions during active transport processes. Five to ten drugs in one experiment at individual 50 μM concentrations did not cause problems in cell viability (MTT test) or monolayer integrity (transepithelial electrical resistance (TEER) measurements, and mannitol diffusion test). Drugs with passive permeability can be included in cocktails; no interactions between them are expected. Drugs, which occupy the same binding sites of a transport protein, cannot be used simultaneously. After validation of the use of several drugs simultaneously as internal standards, the usefulness of the method was further probed by testing the possible interactions during absorption between drugs and plant extracts that are used as food supplements, functional foods, or natural laxatives. These extracts contain several active compounds, such as flavonoids, alkyl gallates, or anthraquinones, which are able to partition into the cell membranes and thus affect e.g. the fluidity of the membranes, or diffuse across the cell monolayers, either by using active transport mechanisms or passively. The effects of different concentrated plant extracts, which may contain high concentrations of several different active compounds, are difficult to predict, because their interactions may be mediated via several active transport proteins, such as OCT, MDR1 and different MRP ́s, or additionally via the effects on the rigidity of cell membranes. Whereas several food-drug interactions have often been attributed to the inhibition of drug metabolizing enzymes, information regarding the effects of food components on transporters during absorption, distribution and excretion of drugs is still limited. Caco-2 cell monolayers, when expressing active transport and efflux proteins, are therefore suitable as in vitro model for this type of studies. _____________________________________________________________________________ TABLE OF CONTENTS TABLE OF CONTENTS . I LIST OF ABBREVIATIONS AND ACRONYMS . II LIST OF ORIGINAL PUBLICATIONS . III