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Estrone‐3‐Sulfate is a substrate to verify functionality of uptake transporters in primary hepatocytes
Author(s) -
Keiser Markus,
Radebold Jennifer,
Ullrich Anett,
Damm Georg,
Nüssler Andreas,
Siegmund Werner,
Runge Dieter
Publication year - 2012
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.26.1_supplement.274.3
Subject(s) - transporter , hepatocyte , estrone sulfate , efflux , in vitro , substrate (aquarium) , chemistry , metabolism , drug metabolism , estrone , toxicity , biochemistry , pharmacology , biology , hormone , ecology , gene , organic chemistry
Background Primary mammalian hepatocytes are used for several in vitro applications like testing of drug metabolism, toxicity and transporter assays. However, little is known about species specific differences or similarities in the activity of uptake and efflux transporter. Therefore, we started a species specific characterization and comparison of uptake transporters functionalities in relevant species used for drug testing. Here, we report the uptake of estrone‐3‐sulfate (E 3 S) in primary hepatocyte cultures of human, rat and monkey. Methods Hepatocytes were incubated in serum free media. Two ‐ four days after cell isolation a time and concentration dependent uptake of [ 3 H]‐E 3 S was measured using liquid scintillation counting. Rifampicin was added in various concentrations to measure its effects on E 3 S uptake. Results All hepatocytes showed at 37 °C a time‐dependent and saturable increase in E 3 S uptake compared to the uptake at 4 °C. E 3 S revealed a high affinity to human ( K m = 12.9 ± 10.1 μmol/l; V max = 84.2 ± 30.3 pmol/mg × min) and monkey ( K m = 8.2 ± 2.3 μmol/l; V max = 31.3 ± 3.6 pmol/mg × min) but not to rat hepatocytes. Rifampicin clearly inhibited the E 3 S uptake in monkey hepatocytes while rat hepatocytes were not influenced. Conclusion Our data suggest that [ 3 H]‐E 3 S is a suitable substrate to verify the functionality of uptake transporters in primary hepatocytes of humanoids but not of rats.