Cells of different tissues for in vitro and in vivo studies in toxicology: Compilation of isolation methods

An advantage of using freshly isolated intact cells of different organs in toxicology is that they reflect more closely the in vivo situation than do long-term cultures. In vitro, primary cells provide the possibility of determining cell-specific xenobiotic metabolism, in the absence of artificial extracellular activation systems, which may result in cytotoxic and genotoxic effects. After in vivo exposure of animals to xenobiotics, isolated primary cells can be studied to elucidate toxicokinetic effects. In the review presented here, selected methods are described for isolating cells with high viability from pig liver and avian embryonic liver, and from the nasal cavity, lungs, kidneys, gastro-intestinal tract, urinary bladder, testes and thymus of the rat. Two techniques for preparing rat lymphocytes are also described. Cell isolation may be initiated with an in situ perfusion to clear the organ of blood. Steps to loosen cell-to-cell contacts and to digest the intercellular connective material may then follow. Also, in situ digestion may be performed, as described for the epithelial cells from different mucosal tissues. Following initial digestion, a single-cell suspension is prepared by tissue mincing and a second digestive step with proteolytic enzymes. Frequently used digestive enzymes are collagenase (types I, IV and P; from Clostridium histolyticum), trypsin and proteinase K. Follow-up filtration is usually required to remove undigested material. The quantities and viabilities of the harvested cells vary with the organ of choice and the procedure used; the values obtained are stated.