Cellular uptake of conjugated bilirubin and sulfobromophthalein (BSP) by the human hepatoma cell line Hep G2 is mediated by a membrane BSP/bilirubin binding protein.
Cellular influx kinetics of 4-50 microM bilirubin diglucuronide and sulfobromophthalein (BSP) by the human hepatoma cell line Hep G2 was examined at 37 degrees C. In confluent monolayer cultures, cellular influx of increasing concentrations of conjugated bilirubin and BSP revealed similar saturation kinetics with Km values of 9.9 and 12.1 microM, and Vmax values of 0.512 and 0.473 nmol.mg cell protein-1.min-1, respectively. Uptake of [3H]bilirubin diglucuronide was competitively inhibited by unlabeled BSP, and was temperature dependent with maximal cellular influx rates at 37 degrees C. When the confluent monolayer cultures were pretreated with a monospecific antibody to the rat liver BSP/bilirubin binding membrane protein, initial uptake rates of conjugated and unconjugated bilirubin as well as of BSP were significantly inhibited, whereas uptake of oleate was not affected. Furthermore, immunoblot analysis of the homogenate of Hep G2 cells with the same antibody revealed predominant reactivity with a 55 kDa protein. These data suggest that cellular uptake of bilirubin and related cholephilic organic anions by the human hepatoma cell line Hep G2 is mediated by a specific 55 kDa membrane BSP/bilirubin binding protein.
Stremmel
W
W
Department of Internal Medicine, Heinrich Heine University, Düsseldorf, Federal Republic of Germany.
Diede
H E
HE
D001663:Bilirubin / Q000378:metabolism*; D001667:Binding, Competitive; D006528:Carcinoma, Hepatocellular / Q000378:metabolism* / Q000473:pathology; D002352:Carrier Proteins / Q000378:metabolism*; D006801:Humans; D015151:Immunoblotting; D007158:Immunologic Techniques; D008113:Liver Neoplasms / Q000378:metabolism* / Q000473:pathology; D013448:Sulfobromophthalein / Q000378:metabolism*; D013696:Temperature; D013997:Time Factors; D014407:Tumor Cells, Cultured
D002352:Carrier Proteins; C029783:bilirubin binding protein, human; C488927:bilirubin binding protein, rat; D013448:Sulfobromophthalein; D001663:Bilirubin