TABLE 3 

Protein abundance of hepatic drug transporters in sandwich cultures of UHH and PHH

Concentrations based on selected signature peptides are given as total amount of plasma membrane protein (femtomoles per microgram plasma membrane protein) and are expressed as the mean ± S.E.M. of triplicates or quadruplicates (n = 34 MRM transitions) of a single experiment.

ProteinSCUHHSCHH
Donor 151-03Donor 653-03Donor 10-03Donor Hu1601Donor HC3-31Donor 371
fmol/µg
OATP1B1<LOQ (0.338)0.376 ± 0.063<LOQ (0.338)4.81 ± 0.2519.4 ± 1.088.73 ± 0.74
OATP1B3NQNQNQ1.96 ± 0.731.93 ± 0.482.15 ± 0.49
OATP2B11.29 ± 0.130.849 ± 0.0640.802 ± 0.1334.51 ± 0.304.18 ± 0.344.70 ± 0.64
OCT11.79 ± 0.533.20 ± 0.472.28 ± 0.26>ULQ (100)81.7 ± 16.547.7 ± 10.6
NTCP1.95 ± 0.821.19 ± 0.181.79 ± 0.475.11 ± 0.414.00 ± 0.604.12 ± 0.79
MDR17.28 ± 0.956.83 ± 0.7116.8 ± 4.47.70 ± 0.8711.5 ± 0.75.65 ± 0.59
MRP24.00 ± 0.211.43 ± 0.121.96 ± 0.0819.7 ± 2.945.19 ± 0.283.67 ± 0.31
BCRP1.63 ± 1.061.78 ± 0.401.81 ± 0.601.74 ± 0.373.02 ± 0.852.21 ± 0.53
BSEP<LOQ (0.467)0.570 ± 0.0360.523 ± 0.0493.22 ± 0.822.47 ± 0.143.15 ± 0.14
MATE1<LOQ (0.334)0.352 ± 0.066<LOQ (0.334)0.525 ± 0.1380.614 ± 0.0450.566 ± 0.078
Na/K-ATPase6.45 ± 0.5424.5 ± 1.822.7 ± 1.433.2 ± 3.5223.6 ± 3.4422.7 ± 1.68
γ-GTP12.2 ± 2.315.9 ± 2.415.5 ± 0.642.8 ± 2.621.5 ± 0.732.2 ± 4.3
  • Abundance of surrogate peptides for canalicular and sinusoidal transporter proteins was quantified from digested plasma membrane extractions derived from sandwich-cultured upcyte and primary hepatocytes (SCUHH and SCHH), respectively. In cases in which concentrations were found below 1 fmol/µg protein and the corresponding peak area of detected signals of at least two MRM transitions was below 5000 counts, the LOQ was defined as described by Uchida et al. (2011). In brief, the protein level of a signature peptide equivalent to a peak area of 5000 counts was calculated based on the linear regression equations obtained from calibrations curves (determination coefficient r2 > 0.99; Supplemental Table 4). If signal peaks were not detectable for two or more MRM transitions of a signature peptide, the corresponding transporter protein was defined as not quantifiable and was considered not to be present in the corresponding UHH culture, applying the quantitative targeted absolute proteomics method described herein. LOQ and ULQ values were as follows: LOQMATE1 = 0.334 fmol/µg plasma membrane protein; LOQOATP1B1 = 0.338 fmol/µg plasma membrane protein; LOQBSEP = 0.467 fmol/µg plasma membrane protein; and ULQOCT1 = 100 fmol/µg plasma membrane protein. NQ, not quantifiable; ULQ, upper limit of quantification.