title: Riboflavin uptake in human trophoblast-derived BeWo cell monolayers: cellular translocation and regulatory mechanisms.
abstract: Riboflavin (vitamin B2) is essential for fetal development and must be acquired from maternal sources. The uptake mechanism of riboflavin and the major regulatory pathways involved were characterized in a model for the placental barrier, the human choriocarcinoma cell line, BeWo. Uptake of [3H]riboflavin was saturable (Kt = 1.32 +/- 0.68 nM, Jmax = 266.63 +/- 26.89 fmol/mg of protein/20 min), and was significantly reduced at low temperature and in the presence of metabolic inhibitors (azide, 2-deoxyglucose) or structural analogs. Ouabain, amiloride, sodium-free buffers, and medium with pH values ranging from 3 to 8 did not affect uptake of [3H]riboflavin. In contrast, substitution of chloride with other monovalent anions significantly inhibited its uptake. Induced differentiation of BeWo cells into syncytiotrophoblasts by forskolin or 8-bromo-cyclic adenosine monophosphate introduced a time-dependent decrease of riboflavin uptake. Preincubation with activators of cyclic nucleotide-dependent protein kinase pathways (3-isobutyl-1-methylxanthine and p-chlorophenylthio-cyclic guanosine monophosphate) and calmodulin antagonists (calmidazolium and W-13) resulted in a concentration-dependent reduction of [3H]riboflavin uptake, whereas specific modulators of protein kinase C pathways did not have significant effects. 3-Isobutyl-1-methylxanthine exerted its regulatory effect on riboflavin uptake via decreasing both Kt and Jmax of the riboflavin uptake process (Kt = 6.32 +/- 1.29 nM, Jmax = 135.57 +/- 10.42 fmol/mg of protein/20 min). In summary, we report the presence of high- affinity riboflavin transporter(s) on the microvillous membrane of BeWo cells that appears to be modulated by cellular cyclic nucleotide levels and calmodulin.
authors.affiliation: Division of Pharmaceutics, College of Pharmacy, The Ohio State University, Columbus, Ohio, USA.
authors.forename: S N
authors.initials: SN
authors.lastname: Huang
authors.forename: P W
authors.initials: PW
authors.lastname: Swaan
mesh_terms: D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid / Q000494:pharmacology; D000818:Animals; D001693:Biological Transport, Active / Q000187:drug effects / Q000502:physiology; D018118:Chloride Channels / Q000187:drug effects / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D006863:Hydrogen-Ion Concentration; D017136:Ion Transport / Q000187:drug effects / Q000502:physiology; D010920:Placenta / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D051381:Rats; D012256:Riboflavin / Q000031:analogs & derivatives / Q000493:pharmacokinetics*; D014327:Trophoblasts / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D014407:Tumor Cells, Cultured / Q000378:metabolism
chemical_list: D018118:Chloride Channels; D004791:Enzyme Inhibitors; D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid; D012256:Riboflavin