Type I insulin-like growth factor receptors in the BeWo choriocarcinoma cell (b30 clone) during cell differentiation

Abstract

The expression of insulin-like growth factor (IGF) receptors in the differentiating human trophoblast was studied using the b30 clone of the BeWo choriocarcinoma cell line (BeWo b30) as a model system. This clonally derived cell line differentiates over 48–72 h, in culture, to form syncytiotrophoblasts when intracellular cAMP levels are elevated by exposure to 100 μ m forskolin (FSK). IGF receptors were studied at various times during the differentiation process by measuring the specific binding of [ 125I]-IGF-I and [ 125I]-IGF-II to attached cells. First, [ 125I]-IGF-I bound to a single class of binding sites in the untreated cells ( K D ≈ 1−2 × 10 −10 m) that exhibited binding specificity characteristic of the type I IGF receptor (IGF-I ≥ IGF-II ⪢ Insulin FSK treatment resulted in a two- to threefold increase in the number of these binding sites. Increased receptor expression was observed as early as 24 h after FSK treatment and remained elevated for at least 72 h. Next, [ 125I]-IGF-II bound to two classes of binding sites in the untreated cells, a high-affinity ( K D 2.5 x 10 −10 m), low-capacity site and a low-affinity ( K D ≈ 6 × 10 −9 m), high-capacity site. The B max and K D of the high-affinity site suggested that it represented the type I IGF receptor. Competition studies revealed that 15–20 per cent of total [ 125I]-IGF-II binding only was sensitive to IGF-I competition in the untreated cells. After FSK treatment, however, unlabelled IGF-I inhibited 60–70 per cent of specific [ 125I]-IGF-II binding. Scatchard analysis revealed a two- to fourfold increase in the number of both binding sites with no change in their respective binding affinities. Cross-linking analysis demonstrated that [ 125I]-IGF-II bound to two structurally distinct binding sites in the untreated BeWo b30 cell consistent with both the type I and II IGF receptors. After FSK treatment, however, there was an increase in the relative amount of [ 125I]-IGF-II associated with the higher affinity type I IGF receptor. The BeWo b30 cells expressed no insulin receptors at any stage of differentiation. These data demonstrate that the BeWob3o choriocarcinoma cell line expresses both type I and II IGF receptors. Induction of cell differentiation is associated with an increase in type I IGF receptors expressed at the cell surface. These receptors bind IGF-II with high-affinity, providing additional binding capacity for locally available IGF-II. These data are consistent with specific roles for the type I IGF receptor in regulating differentiated trophoblast cell function. Furthermore, the early rise in type I IGF receptor number suggests they may play a regulatory role in the differentiation process itself.