Identification and characterization of an ascorbic acid transporter in human granulosa-lutein cells

Ascorbic acid serves a vital role as a pre-eminent antioxidant. in animals, it has been shown to be concentrated in granulosa and theca cells of the f ollicle, in luteal cells of the corpus luteum, and in the peripheral cytopl asm of the oocyte. We have previously identified hormonally-regulated ascor bic acid transporters in rat granulosa and luteal cells, and herein present preliminary evidence for the presence of a transporter for ascorbic acid i n human granulosa-lutein cells. Granulosa-lutein cells were obtained from t he follicular fluid of patients undergoing in-vitro fertilization. Followin g an overnight incubation, the cells were incubated with [C-14]-ascorbic ac id (0.15 mu Ci; 150 mu MM) and ascorbic acid uptake was determined. The upt ake of ascorbic acid was saturable with a Michaeli's constant (K-m) and max imum velocity (V-max) of 21 mu M and 3 pmol/10(6) cells/min respectively. O uabain, low Na+ medium, and dinitrophenol significantly inhibited ascorbic acid uptake (P < 0.05). Neither the presence of insulin, human chorionic go nadotrophin (HCG), insulinlike growth factor (IGF)-I, nor IGF-II affected t he uptake of ascorbic acid in a statistically significant fashion. Followin g saturation of cellular uptake, the ascorbic acid level was estimated to b e 1.04 pmoles/10(6) cells or similar to 1 mM, a high concentration similar to that seen in rat luteal cells. Active ascorbic acid transport in human g ranulosa-lutein cells appears to occur via a Na+- and energy-dependent tran sporter, with high levels of ascorbic acid being accumulated in these cells .