The development of a cell culture maintenance system is important for alternatives to animal testing and experimentation for drug discovery. Generally, cryopreserved human hepatocyte viability and function are maintained for approximately 3 days after thawing in two-dimensional (2D) monolayer culture using a collagen-coated dish. In this study, we cultured human hepatocytes in the silicate fiber-based three-dimensional (3D) scaffold Cellbed®. As a result of observation with a scanning electron microscope (SEM), hepatocytes were cuboidal as compared with monolayer culture. This cell morphology seems to be appropriate for maintenance of cell viability and function of hepatocytes. Cell viability in Cellbed culture was maintained at a higher level and for a longer period (about 2 weeks) than that of monolayer culture. Levels of glutamic oxaloacetic transaminase (GOT) and glutamic pyruvic transaminase (GPT) leakage were also lower in Cellbed culture than in monolayer culture. Based on gene expression analysis, CYP2C19 and CYP3A4 were expressed in Cellbed culture at levels more than 3.0 times higher than those of monolayer culture. Moreover, SLC22A1, the gene for bile acid transporter, was expressed in Cellbed culture at a level more than 2.9 times higher than that in monolayer. This culture system may be applied to hepatic metabolism studies and long-term in vitro liver toxicity testing.
Drugs are exposed to drastic pH changes following oral administration, for instance, from pH 1 in the stomach to pH 6 in the small intestine for healthy subjects. In the case of poorly soluble basic drugs, which exhibit pH-dependent solubility in the range of pH 1–6, supersaturation may occur, inducing higher absorption. Therefore, it is preferable to simulate the effect of various pH changes in the gastrointestinal tract for oral formulation studies. The results of in vivo studies with humans or animals may not accurately show how pH changes can effect drug dissolution and absorption in reality. In the present study, we constructed a new in vitro experimental system that can reflect the changes induced by dramatic shifts in pH. Albendazole (ABZ) was selected as a lipophilic weak basic model drug. Supersaturation of ABZ was induced by pH changes in a dynamic condition, which led to a higher permeation than that observed under the constant pH condition. This changeable pH dissolution/permeation system can simulate the effects of drastic pH shifts without the need for humans or experimental animals.