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Insulin‐like growth factor (IGF) binding to human fibroblast and glioblastoma cells: The modulating effect of cell released IGF binding proteins (IGFBPs)
Author(s) -
McCusker R. H.,
CamachoHubner C.,
Clemmons D. R.,
Bayne M. L.,
Cascieri M. A.
Publication year - 1990
Publication title -
journal of cellular physiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.529
H-Index - 174
eISSN - 1097-4652
pISSN - 0021-9541
DOI - 10.1002/jcp.1041440210
Subject(s) - receptor , cycloheximide , growth factor , binding site , cell culture , cell , plasma protein binding , biology , cell surface receptor , endocrinology , chemistry , medicine , biochemistry , genetics
The cell surface of human fibroblasts contains not only type IIGF receptors but at least two forms of IGFBPs. Studies were undertaken to analyze the mechanisms by which these IGFBPs alter IGF‐I‐cell surface interactions. Human fetal fibroblasts (GM10) and a human glioblastoma cell line (1690) were chosen for analysis. During assays to quantify [ 125 l]‐IGF‐l binding, both cell lines were shown to release IGFBPs into the binding assay buffer. Under equilibrium conditions, [ 125 I]‐IGF‐I preferentially associates with IGFBPs in the assay buffer (up to 40% of the [ 125 I]‐IGF‐I added) since they have a higher affinity than type IIGF receptors or IGFBPs associated with the cell surface. Likewise the addition of increasing concentrations of unlabeled IGF‐I results in preferential competition for binding to assay buffer IGFBPs. This results in a repartitioning of the [ 125 I]‐IGF‐I that is bound to assay buffer IGFBPs onto cell surface binding sites. The degree of repartitioning is quantitatively related to the amount of [ 125 I]‐IGF‐I bound to released IGFBPs. When cultures are exposed to cycloheximide before the binding assay, both the amount of IGFBPs that are released into the assay buffer and the amount of [ 125 I]‐IGF‐l that is repartitioned are decreased. In contrast when [Gln 3 , Ala 4 , Tyr 15 , Leu 16 ]‐IGF‐l (IQAYLj‐IGF‐I, an IGF analog that has unaltered affinity for type I IGF receptors) is iodinated and tested, the competition curve with unlabeled IGF‐I shows no repartitioning effect. This form of IGF can be used to quantify type I receptor number independent of the presence of IGFBPs. IGF‐I and the [QAYL]‐IGF‐I compete equally with the [ 125 I]‐[QAYL]‐IGF‐I for binding to cell surfaces, whereas unlabeled [QAYL]‐IGF‐I is > 25‐fold less potent compared to IGF‐I in competing with [ 125 I]‐IGF‐I for cell surface binding. Specific binding of [ 125 I]‐[QAYL]‐IGF‐I to GM10 and 1690 cell surfaces is < 20% of [ 125 I]‐IGF‐I binding. These findings suggest that IGFBPs that are present on human fibroblast surfaces represent a large portion of the IGF binding sites. We conclude that the amount of IGFBPs released into assay buffer is a major determinant of the repartitioning of [ 125 I]‐IGF‐I to cell surface binding sites and that both cell surface and assay buffer IGFBPs modulate type I IGF receptor binding.