Gene expression of human hepatocytes isolated from chimeric mice with humanized liver

Chimeric mice with humanized livers are considered a useful animal model for predicting human drug metabolism and toxicity. In this study, the characteristics of fresh h-hepatocytes (cFHHs, PXB-cells®) isolated from chimeric mice (PXB-mice®) were evaluated in vitro to confirm their utility for drug development. The cFHHs cultured at high density (2.13 × 10^5 cells/cm2) displayed stable production of human albumin and cytochrome P450 (CYP) 3A activities for at least 21 days. The mRNA expression levels of 10 of 13 CYPs, UDP-glucuronosyltransferase (UGP), and transporters were maintained at >10% of the levels of freshly isolated cFHHs after 21 days. From 7-days cultured cFHHs at high density, many bile canaliculi were observed between cFHHs, and the accumulation of multidrug resistance-associated protein (MRP2) and bile salt export pump (BSEP) substrates in these bile canaliculi was clearly inhibited by cyclosporin A. We used microarrays to elucidate global gene expressions underlying this higher hepatic functions of high density cultured cFHHs (PXB-cells). Microarray analysis revealed that high density cultured cFHHs maintained high expressions of some transcription factors (HNF4α, PXR, and FXR) perhaps involved in the high CYP, UGT and transporter gene expressions of cFHHs. To identify genes related with the characterization of the cFHHs, we carried out microarray analysis using Affymetrix technolgy with total RNA extracted from high density (2.13 × 10^5 cells/cm2) cultured cFHHs, low density (0.53 x 105 cells/cm2) cultured cFHHs (Day 7) and cFHHs without culture (Day 0) as a reference group.