Precision-cut tissue slices (PCTS) are viable ex vivo explants of tissue with a reproducible, well defined thickness. They represent a mini-model of the organ under study and contain all cells of the tissue in their natural environment, leaving intercellular and cell-matrix interactions intact, and are therefore highly appropriate for studying multicellular processes. PCTS are mainly used to study the metabolism and toxicity of xenobiotics, but they are suitable for many other purposes. Here we describe the protocols to prepare and incubate rat and human liver and intestinal slices. Slices are prepared from fresh liver by making a cylindrical core using a drill with a hollow bit, from which slices are cut with a specially designed tissue slicer. Intestinal tissue is embedded in cylinders of agarose before slicing. Slices remain viable for 24 h (intestine) and up to 96 h (liver) when incubated in 6- or 12-well plates under 95% O(2)/5% CO(2) atmosphere.

Microfluidic technology is providing new routes toward advanced cell and tissue culture models to better understand human biology and disease. Many advanced devices have been made from poly(dimethylsiloxane) (PDMS) to enable experiments, for example, to study drug metabolism by use of precision-cut liver slices, that are not possible with conventional systems. However, PDMS, a silicone rubber material, is very hydrophobic and tends to exhibit significant adsorption and absorption of hydrophobic drugs and their metabolites. Although glass could be used as an alternative, thermoplastics are better from a cost and fabrication perspective. Thermoplastic polymers (plastics) allow easy surface treatment and are generally transparent and biocompatible. This study focuses on the fabrication of biocompatible microfluidic devices with low adsorption properties from the thermoplastics poly(methyl methacrylate) (PMMA), polystyrene (PS), polycarbonate (PC), and cyclic olefin copolymer (COC) as alternatives for PDMS devices. Thermoplastic surfaces were oxidized using UV-generated ozone or oxygen plasma to reduce adsorption of hydrophobic compounds. Surface hydrophilicity was assessed over 4 weeks by measuring the contact angle of water on the surface. The adsorption of 7-ethoxycoumarin, testosterone, and their metabolites was also determined after UV-ozone treatment. Biocompatibility was assessed by culturing human hepatoma (HepG2) cells on treated surfaces. Comparison of the adsorption properties and biocompatibility of devices in different plastics revealed that only UV-ozone-treated PC and COC devices satisfied both criteria. This paper lays an important foundation that will help researchers make informed decisions with respect to the materials they select for microfluidic cell-based culture experiments.

Over the past two decades, it has become increasingly clear that the intestine, in addition to the liver, plays an important role in the metabolism of xenobiotics. Previously, we developed a microfluidic-based in vitro system for the perifusion of precision-cut liver slices for metabolism studies. In the present study, the applicability of this system for the perifusion of precision-cut intestinal slices, and for the sequential perifusion of intestinal and liver slices, all from rat, was tested to mimic the in vivo first pass situation. Intestinal and liver slices, exposed to the substrates 7-ethoxycoumarin (7-EC), 7-hydroxycoumarin (7-HC) and lidocaine (Li), exhibited similar metabolic rates in the biochip and in the well plates for periods of at least 3 h. The metabolic rate remained the same when two slices were placed in adjacent microchambers and perifused sequentially. In addition, the system has been adapted to sequentially perifuse intestinal and liver tissue slices in a two-compartment co-culture perfusion system with a continuous flow of medium. It becomes possible to direct metabolites or other excreted compounds formed by an intestinal slice in the first compartment to the second compartment containing a liver slice. The intestine does not influence liver metabolism for these substrates. The interplay between these two organs was demonstrated by exposing the slices to the primary bile acid, chenodeoxycholic acid (CDCA). CDCA induced the expression of fibroblast growth factor 15 (FGF15) in the intestinal slice, which resulted in a stronger down-regulation of the enzyme, cytochrome P450 7A1 (CYP7A1), in the liver slice in the second compartment than when the liver slice was exposed to CDCA in a single-microchamber biochip. We thus demonstrate in this paper that intestinal slices, in addition to liver slices, remain functional in the biochip under flow conditions, and that the two-microchamber biochip has great potential for the study of interorgan effects. This is the first example of the incorporation of both liver and intestinal slices in a microfluidic device. Use of this microfluidic system will improve our insight into interorgan interactions and elucidate as yet unknown mechanisms involved in toxicity, gene regulation and drug-drug interactions.

Early detection of kinetic, metabolic, and toxicity (ADME-Tox) profiles for new drug candidates is of crucial importance during drug development. This article describes a novel in vitro system for the incubation of precision-cut liver slices (PCLS) under flow conditions, based on a poly(dimethylsiloxane) (PDMS) device containing 25-microL microchambers for integration of the slices. The microdevice is coupled to a perifusion system, which enables a constant delivery of nutrients and oxygen and a continuous removal of waste products. Both a highly controlled incubation environment and high metabolite detection sensitivity could be achieved using microfluidics. Liver slices were viable for at least 24 h in the microdevice. The compound, 7-ethoxycoumarin (7-EC), was chosen to test metabolism, since its metabolism includes both phase I and phase II metabolism and when tested in the conventional well plate system, correlates well with the in vivo situation (De Kanter et al. 2004. Xenobiotica 34(3): 229-241.). The metabolic rate of 7-EC was found to be 214 +/- 5 pmol/min/mg protein in the microdevice, comparable to well plates, and was constant over time for at least 3 h. This perifusion system better mimics the in vivo situation, and has the potential to significantly contribute to drug metabolism and toxicology studies of novel chemical entities.