Validation of uPA/SCID mouse with humanized liver as a human liver model: protein quantification of transporters, cytochromes P450, and UDP-glucuronosyltransferases by LC-MS/MS.
Chimeric mice with humanized liver (PXB mice) have been generated by transplantation of urokinase-type plasminogen activator/severe combined immunodeficiency mice with human hepatocytes. The purpose of the present study was to clarify the protein expression levels of metabolizing enzymes and transporters in humanized liver of PXB mice transplanted with hepatocytes from three different donors, and to compare their protein expressions with those of human livers to validate this human liver model. The protein expression levels of metabolizing enzymes and transporters were quantified in microsomal fraction and plasma membrane fraction, respectively, by means of liquid chromatography-tandem mass spectrometry. Protein expression levels of 12 human P450 enzymes, two human UDP-glucuronosyltransferases, eight human ATP binding cassette (ABC) transporters, and eight human solute carrier transporters were determined. The variances of protein expression levels among samples from mice humanized with hepatocytes from all donors were significantly greater than those from samples obtained from mice derived from each individual donor. Compared with the protein expression levels in human livers, all of the quantified metabolizing enzymes and transporters were within a range of 4-fold difference, except for CYP2A6, CYP4A11, bile salt export pump (BSEP), and multidrug resistance protein 3 (MDR3), which showed 4- to 5-fold differences between PXB mouse and human livers. The present study indicates that humanized liver of PXB mice is a useful model of human liver from the viewpoint of protein expression of metabolizing enzymes and transporters, but the results are influenced by the characteristics of the human hepatocyte donor.
Department of Pharmaceutical Microbiology, Faculty of Life Sciences, Kumamoto University, Kumamoto, Japan (S.O., K.N.); Laboratory of Veterinary Pathology, School of Veterinary Medicine, Azabu University, Kanagawa, Japan (H.K., J.K.); Division of Membrane Transport and Drug Targeting, Graduate School of Pharmaceutical Sciences, Tohoku University, Sendai, Japan (T.I, Y.K., W.O., Y.U., T.T.); PhoenixBio, Co., Ltd., Hiroshima, Japan (C.T.); DeThree Research Laboratory Inc., Ibaraki, Japan (T.H.).
Sumio
S
Ohtsuki
Hirotaka
H
Kawakami
Tae
T
Inoue
Kenji
K
Nakamura
Chise
C
Tateno
Yuki
Y
Katsukura
Wataru
W
Obuchi
Yasuo
Y
Uchida
Junichi
J
Kamiie
Toru
T
Horie
Tetsuya
T
Terasaki
D018528:ATP-Binding Cassette Transporters / Q000032:analysis / Q000096:biosynthesis*; D000818:Animals; D002648:Child; D002675:Child, Preschool; D002678:Chimera; D002853:Chromatography, Liquid / Q000379:methods; D003577:Cytochrome P-450 Enzyme System / Q000032:analysis / Q000096:biosynthesis*; D005260:Female; D014453:Glucuronosyltransferase / Q000032:analysis / Q000096:biosynthesis*; D022781:Hepatocytes / Q000737:chemistry / Q000378:metabolism; D006801:Humans; D008099:Liver / Q000737:chemistry / Q000378:metabolism*; D008297:Male; D051379:Mice; D016513:Mice, SCID; D053719:Tandem Mass Spectrometry / Q000379:methods
D018528:ATP-Binding Cassette Transporters; D003577:Cytochrome P-450 Enzyme System; D014453:Glucuronosyltransferase