Ascorbic acid accumulation and transport in human fibroblasts.
As the initial step in the use of fibroblasts as a model system for 'in situ kinetics', ascorbic acid (vitamin C) accumulation in normal human fibroblasts was investigated for the first time. Ascorbic acid was transported into fibroblasts and accumulated against a concentration gradient up to 20-fold, as measured by h.p.l.c. with coulometric electrochemical detection. Ascorbic acid accumulation was mediated by two concentration-dependent transport activities. The first was a high-affinity activity with an apparent Km of 6 microM and an apparent Vmax. of 203 microM/h, and the second was a low-affinity activity with an apparent Km of 5 mM and an apparent Vmax. of 1.8 mM/h. Both activities were inhibited by metabolic inhibitors and inhibitors of ascorbic acid transport in human neutrophils. The low-affinity transporter could not be accounted for by diffusion. Although the high-affinity transport activity was comparable with that described for human neutrophils, the low-affinity transporter was different. These data provide the first evidence that two-component ascorbic acid transport may be a generalized mechanism for accumulation of this vitamin in humans.
Laboratory of Cell Biology and Genetics, National Institute of Diabetes, Digestive, and Kidney Diseases, National Institutes of Health, Bethesda, MD 20892.
R W
RW
Welch
P
P
Bergsten
J D
JD
Butler
M
M
Levine
D019297:2,4-Dinitrophenol; D001205:Ascorbic Acid / Q000378:metabolism*; D001692:Biological Transport; D002259:Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone / Q000494:pharmacology; D002352:Carrier Proteins / Q000378:metabolism; D002460:Cell Line; D002851:Chromatography, High Pressure Liquid; D003571:Cytochalasin B / Q000494:pharmacology; D004140:Dinitrophenols / Q000494:pharmacology; D005347:Fibroblasts / Q000378:metabolism*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D009504:Neutrophils / Q000378:metabolism; D010693:Phloretin / Q000494:pharmacology; D011190:Potassium Cyanide / Q000494:pharmacology; D012964:Sodium / Q000494:pharmacology
D002352:Carrier Proteins; D004140:Dinitrophenols; D002259:Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone; D003571:Cytochalasin B; D012964:Sodium; D011190:Potassium Cyanide; D001205:Ascorbic Acid; D019297:2,4-Dinitrophenol; D010693:Phloretin