Characterization of hepatic cellular uptake of α1-acid glycoprotein (AGP), part 2: involvement of hemoglobin β-chain on plasma membranes in the uptake of human AGP by liver parenchymal cells.
Human α(1) -acid glycoprotein (AGP), a lipocalin family member, serves as a carrier for basic drugs and endogenous hormones. It is mainly distributed in the liver and also has anti-inflammatory effects. We previously discovered a protein in liver parenchymal cells that interacts with AGP and it was identified as hemoglobin β-chain (HBB). The purpose of this study was to clarify the role of HBB in the hepatic cellular uptake of AGP. Ligand blotting experiments showed that the interaction of (125) I-AGP with hemoglobin was saturable and was significantly suppressed in the presence of excess unlabeled AGP. In addition, the cellular uptake of fluorescein isothiocianate-AGP by HepG2 cells was saturable and temperature dependent. This uptake was inhibited by fillipin and methyl-β-cyclodextrin, but not chlorpromazine, suggesting that AGP is taken up via caveolae/lipid rafts endocytic pathway. Immunostaining showed that HBB and caveolin-1, exclusively expressed in caveolae, were partially colocalized on the plasma membranes of HepG2 cells. HBB knockdown with siRNA decreased the uptake of AGP by HepG2 cells by 40%, and exogenous hemoglobin inhibited the uptake by 40%-50%. These findings indicate that HBB is located on the liver plasma membrane and that it contributes to the intracellular uptake of AGP.
Department of Biopharmaceutics, Graduate School of Pharmaceutical Sciences, Kumamoto University, Kumamoto, Japan.
Hisakazu
H
Komori
Koji
K
Nishi
Nao
N
Uehara
Hiroshi
H
Watanabe
Tsuyoshi
T
Shuto
Ayaka
A
Suenaga
Toru
T
Maruyama
Masaki
M
Otagiri
D021941:Caveolae / Q000502:physiology; D002462:Cell Membrane / Q000378:metabolism*; D004705:Endocytosis; D006454:Hemoglobins / Q000378:metabolism*; D056945:Hep G2 Cells; D006801:Humans; D008099:Liver / Q000378:metabolism*; D021962:Membrane Microdomains / Q000502:physiology; D009961:Orosomucoid / Q000737:chemistry / Q000493:pharmacokinetics*
D006454:Hemoglobins; D009961:Orosomucoid