Superfused microcarrier cultures of choriocarcinoma cells: a dynamic model for studying transport of glucose and amino acids

Abstract

We here describe a novel procedure for culturing BeWo and JAr choriocarcinoma cells on solid microcarrier beads. The regime developed to stir the beads resulted, after about 10 days, in small aggregates of two to six beads covered with a layer of differentiated cells. Bead aggregates were packed into small columns and superfused, providing a dynamic in vitro system for studying the uptake mechanisms for sugars and amino acids. The rapid, unidirectional uptake of tritiated l-phenylalanine, l-serine, l-arginine and d-glucose was determined, relative to an extracellular reference tracer, in cells superfused in 0.5 ml (range 0.3–0.8 ml) columns. Several sequential measurements could be made in the same column. Twenty-four hour pre-incubation with dexamethasone (0.25 μm) was found to increase the transport of d-glucose. Uptake of d-glucose was reduced by over 80 per cent following 20 min perfusion of the cells with 1 mM phloretin. Pre-incubation with growth hormone (0.2 μg/ml) decreased the transport of serine, whereas nicotine (0.5 μg/ml) decreased both serine and phenylalanine uptake. Atropine (1 mM) or 5-oxoproline (0.5 mM) had no short-term effects on amino acid uptake. Insulin (12.5 mIU/ml) had no effect on the transport of the amino acids but caused a small but significant increase in glucose transport ( P<0.05). This model allows characterization of human trophoblast function without the complications resulting from the presence of other cell types in placental slices or fragments.