PURPOSE: It was determined in cultured human retinal pigment epithelial (HRPE) cells whether there is gene expression for insulin, insulin-like growth factors (IGFs), insulin and IGF receptors, and IGF-binding proteins (IGFBPs); if these peptides are secreted by the HRPE cells, and whether they have mitogenic effects on these cells; and if IGF-related peptides bind to HRPE cells. METHODS: Reverse transcriptase-polymerase chain reaction (RT-PCR), nucleotide sequencing, and Southern blot analyses were done to identify gene expression. The presence of IGFs in the tissue culture medium was detected by radioimmunoassay. For determination of mitogenic effects and receptor binding characterization, [3H]-thymidine incorporation and radioreceptor binding measurements were performed. RESULTS: The genes for IGF-I and IGF-II, IGFBP-2, insulin receptor, and type I and II IGF receptors were detected. In the tissue culture supernatant, there was immunoreactivity for IGF-I and IGF-II. Insulin, IGF-I, and IGF-II stimulated DNA synthesis with EC50s of 3, 10, and 30 to 100 nM, respectively. Scatchard analyses of [125I]-IGF-I binding and [125I]-insulin binding to the cells showed that the cells have relatively abundant insulin and IGF-I binding sites. CONCLUSIONS: HRPE cells express genes for IGF-I and IGF-II, and conditioned medium from these cells is immunoreactive to their protein products. The cells express genes for insulin receptor, type I and II IGF receptors, and IGFBP-2. IGF-I, IGF-II, and insulin are all mitogenic, possibly as a result of their interactions with either the insulin or type I IGF receptor. The cells bind insulin and IGF-I with high affinity. These results suggest that IGF-I and IGF-II production by HRPE cells may be essential for autocrine/paracrine-mediated regulation of proliferation. The presence of insulin receptor suggests that insulin has a role in the regulation of HRPE function.
PURPOSE: It was determined in cultured human retinal pigment epithelial (HRPE) cells whether there is gene expression for insulin, insulin-like growth factors (IGFs), insulin and IGF receptors, and IGF-binding proteins (IGFBPs); if these peptides are secreted by the HRPE cells, and whether they have mitogenic effects on these cells; and if IGF-related peptides bind to HRPE cells. METHODS: Reverse transcriptase-polymerase chain reaction (RT-PCR), nucleotide sequencing, and Southern blot analyses were done to identify gene expression. The presence of IGFs in the tissue culture medium was detected by radioimmunoassay. For determination of mitogenic effects and receptor binding characterization, [3H]-thymidine incorporation and radioreceptor binding measurements were performed. RESULTS: The genes for IGF-I and IGF-II, IGFBP-2, insulin receptor, and type I and II IGF receptors were detected. In the tissue culture supernatant, there was immunoreactivity for IGF-I and IGF-II. Insulin, IGF-I, and IGF-II stimulated DNA synthesis with EC50s of 3, 10, and 30 to 100 nM, respectively. Scatchard analyses of [125I]-IGF-I binding and [125I]-insulin binding to the cells showed that the cells have relatively abundant insulin and IGF-I binding sites. CONCLUSIONS: HRPE cells express genes for IGF-I and IGF-II, and conditioned medium from these cells is immunoreactive to their protein products. The cells express genes for insulin receptor, type I and II IGF receptors, and IGFBP-2. IGF-I, IGF-II, and insulin are all mitogenic, possibly as a result of their interactions with either the insulin or type I IGF receptor. The cells bind insulin and IGF-I with high affinity. These results suggest that IGF-I and IGF-II production by HRPE cells may be essential for autocrine/paracrine-mediated regulation of proliferation. The presence of insulin receptor suggests that insulin has a role in the regulation of HRPE function.
Authors: Shervi Lie; Bárbara R Vieira; Sigrid Arruda; Milena Simões; Liam M Ashander; João M Furtado; Justine R Smith Journal: Microorganisms Date: 2019-09-29