title: Quantification of polarized trafficking of transferrin and comparison with bulk membrane transport in hepatic cells.
abstract: Transport of the recycling marker transferrin was analysed in polarized hepatic HepG2 cells using quantitative fluorescence microscopy and mathematical modelling. A detailed map and kinetic model for transport of transferrin in hepatic cells was developed. Fluorescent transferrin was found to be transported sequentially through basolateral SE (sorting endosomes) to a SAC/ARC (subapical compartment/apical recycling compartment). DiI (di-indocarbocyanine) lipid probes of different acyl chain length (DiIC12 and DiIC16) co-localized with transferrin in basolateral SE and in the SAC/ARC. By kinetic comparison of hepatic transport of transferrin and labelled HDL (high-density lipoprotein), it is shown that transport of transferrin from SE to the SAC/ARC follows a default pathway together with HDL. Kinetic modelling of fluorescence data provides an identical half-time for SE-to-SAC/ARC transport of transferrin and fluorescent HDL (t(1/2)=4.2 min). Fluorescent transferrin was found to recycle with a half-time of t(1/2)=12.9 min from the SAC/ARC to the basolateral cell surface of HepG2 cells. In contrast with HDL, targeting of labelled transferrin from the SAC/ARC to the apical biliary canaliculus was negligible. The results indicate that transport from basolateral hepatic SE to the SAC/ARC represents a bulk flow process and that polarized sorting occurs mainly at the level of the SAC/ARC.
authors.affiliation: Department of Biochemistry and Molecular Biology, University of Southern Denmark, Campusvej 55, DK-5230 Odense M, Denmark. wuestner@bmb.sdu.dk
authors.forename: Daniel
authors.initials: D
authors.lastname: Wüstner
mesh_terms: D001692:Biological Transport; D002232:Carbocyanines / Q000378:metabolism; D045744:Cell Line, Tumor; D002462:Cell Membrane / Q000378:metabolism; D016764:Cell Polarity / Q000502:physiology; D011992:Endosomes / Q000378:metabolism; D005456:Fluorescent Dyes / Q000378:metabolism; D022781:Hepatocytes / Q000378:metabolism* / Q000502:physiology; D006801:Humans; D006834:Hydrazines / Q000378:metabolism; D007700:Kinetics; D008075:Lipoproteins, HDL / Q000378:metabolism; D008113:Liver Neoplasms; D008856:Microscopy, Fluorescence; D008954:Models, Biological; D014168:Transferrin / Q000378:metabolism*
chemical_list: C417561:Alexa 488 hydrazide; D002232:Carbocyanines; D005456:Fluorescent Dyes; D006834:Hydrazines; D008075:Lipoproteins, HDL; D014168:Transferrin