Phenotype-associated changes in retinal pigment epithelial cell expression of insulin-like growth factor binding...

The objectives of this study were to evaluate retinal pigment epithelial (RPE) cells as a source of insulin-like growth factor binding proteins (IGFBPs) and to characterize biosynthetic changes associated with the cell phenotype and vitreous growth factor stimuli known to be present in fibrocontractive diseases.

Early culture-associated changes in RPE phenotype were characterized by indirect immunofluorescence localization and Western blot analysis of cell lysates. IGFBP expression was evaluated by RT-PCR and Northern blot analysis of total RNA preparations.

Normal unperturbed RPE are immunoreactive for cytokeratin 18 and negative for cytokeratin 19, vimentin, and alpha-smooth muscle actin (alphaSMA). Early reactive RPE (7 days in culture) express cytokeratin 18, cytokeratin 19, and vimentin. Myofibroblastic RPE (35 days in culture) express cytokeratin 19, vimentin, and alphaSMA. RT-PCR studies revealed that normal RPE can produce IGFBP-2, -3, -4, -5, and -6 but not IGFBP-1. Early reactive and myofibroblastic RPE have detectable levels of message for IGFBP-3, -5, and -6. However, Northern blot analysis suggests that IGFBP-5 is the predominant binding protein produced. Finally, stimulation with biologically relevant quantities of IGF-I and IGF-II had no detectable effects on IGFBP expression.

Changes in RPE phenotype are accompanied by dramatic changes in IGFBP expression profile, with IGFBP-5 the predominant binding protein produced by myofibroblastic RPE cells.