Utility of B-13 progenitor-derived hepatocytes in hepatotoxicity and genotoxicity studies.
AR42J-B-13 (B-13) cells form hepatocyte-like (B-13/H) cells in response to glucocorticoid treatment. To establish its utility in toxicity and genotoxicity screening, cytochrome P450 (CYP) induction, susceptibility to toxins, and transporter gene expression were examined. Conversion to B-13/H cells resulted in expression of male-specific CYP2C11 and sensitivity to methapyrilene. B-13/H cells constitutively expressed CYP1A, induced expression in response to an aryl hydrocarbon receptor agonist, and activated benzo[α]pyrene to a DNA-damaging species. Functional CYP1A2 was not expressed due to deletions in the Cyp1a2 gene. A B-13 cell line stably expressing the human CYP1A2 was therefore engineered (B-13(-TR/h1A2)) and the derived B-13/H cells expressed metabolically functional CYP1A2. Treatment with the cooked food mutagen 2-amino-1-methyl-6-phenylimidazo(4,5-b)pyridine resulted in a dose-dependent increase in DNA damage. B-13/H cells expressed constitutive androstane receptor (CAR) and induced CYP2B1 mRNA levels in response to classical CAR activators. However, translation to functional CYP2B1 protein was low and increased minimally by CAR activator treatment. B-13/H cells expressed high levels of pregnane X-receptor (PXR) and induced CYP3A1 in response to classical PXR activators. CYP3A genes were inducible, functional, and activated aflatoxin B1 to a DNA-damaging species. All 23 major hepatic transporters were induced when B-13 cells were converted to B-13/H cells, although in many cases, levels remained below those present in adult rat liver. However, bile salt export pump, Abcb1b, multidrug resistance-associated protein, and breast cancer resistance protein transporters were functional in B-13/H cells. These data demonstrate that the B-13 cell generates hepatocyte-like cells with functional drug metabolism and transporter activities, which can alone--or in a humanized form--be used to screen for hepatotoxic and genotoxic endpoints in vitro.
* Institute of Cellular Medicine.
Philip M E
PM
Probert
Git W
GW
Chung
Simon J
SJ
Cockell
Loranne
L
Agius
Pasquale
P
Mosesso
Steven A
SA
White
Fiona
F
Oakley
Colin D A
CD
Brown
Matthew C
MC
Wright
D000826:Animal Testing Alternatives; D000818:Animals; D018929:Cell Culture Techniques; D002454:Cell Differentiation / Q000187:drug effects; D002460:Cell Line; D049109:Cell Proliferation / Q000187:drug effects; D003577:Cytochrome P-450 Enzyme System / Q000096:biosynthesis / Q000235:genetics; D004249:DNA Damage; D003907:Dexamethasone / Q000494:pharmacology; D053595:Embryonic Stem Cells / Q000166:cytology* / Q000187:drug effects / Q000201:enzymology; D004790:Enzyme Induction; D022781:Hepatocytes / Q000166:cytology* / Q000187:drug effects / Q000201:enzymology; D008099:Liver / Q000187:drug effects* / Q000201:enzymology / Q000473:pathology; D008297:Male; D009152:Mutagenicity Tests / Q000379:methods; D009153:Mutagens / Q000633:toxicity; D051381:Rats; D012727:Sex Characteristics; D018675:Toxicity Tests / Q000379:methods; D014162:Transfection
D009153:Mutagens; D003907:Dexamethasone; D003577:Cytochrome P-450 Enzyme System
cytochrome P450; liver; stem cell; transporters.; xenobiotic metabolism