Autocrine effects of IGF-I-induced VEGF and IGFBP-3 secretion in retinal pigment epithelial cell line ARPE-19

Abstract

Hypoxia-induced physiological stress plays a central role in various neovascular diseases of the eye. Increased expression of hypoxia-inducible factor 1alpha (HIF-1alpha) and subsequent formation of HIF-1 dimers active at the vascular endothelial growth factor (VEGF) promoter lead to expression of this potent angiogenic factor in the retina, including retinal pigment epithelial (RPE) cells. We previously demonstrated that insulin-like growth factor I (IGF-I) stimulates VEGF and IGF binding protein (IGFBP)-3 secretion in RPE cells. In this study we examined IGF-I-induced HIF-1alpha expression, VEGF and IGFBP-3 secretion, and the autocrine actions of VEGF and IGFBP-3 on these processes in the spontaneously transformed RPE cell line ARPE-19. Cells were treated with CoCl(2), IGF-I, recombinant human (rh)IGFBP-3, and rhVEGF. Immunoblot analysis revealed IGF-I-induced upregulation of total HIF-1alpha protein, whereas luciferase reporter assays of HIF-1 transcriptional activity demonstrated accumulation of HIF-1alpha correlated with the formation of functional HIF-1 heterodimers. Western and ligand blot analyses of RPE cell conditioned medium confirmed that IGF-I stimulated VEGF and IGFBP-3 secretion. rhVEGF stimulated IGFBP-3 secretion in an IGF-I- and HIF-1alpha-independent manner, whereas rhIGFBP-3 attenuated IGF-I-induced VEGF secretion. These findings demonstrate the multifaceted autocrine regulation of IGF-I-induced VEGF secretion by IGFBP-3 secreted in response to both IGF-I and, to a lesser extent, VEGF. These results provide evidence for HIF-1-dependent and -independent mechanisms by which IGF-I regulates VEGF and IGFBP-3 secretion.

Hypoxia-Inducible Factor-1-Dependent and -Independent Regulation of Insulin-Like Growth Factor-1-Stimulated Vascular Endothelial Growth Factor Secretion

“…Abrogation of Hif-1␣ degradation resulting from physiologic or chemical hypoxia (CoCl 2 ) affects Hif-1␣ protein accumulation (Fig. 3A) (Slomiany and Rosenzweig, 2004a;Brahimi-Horn et al, 2005;Slomiany et al, 2006). Consistent with this observation, CoCl 2 treatment led to a more significant increase in HRE-dependent luciferase activity at the enolase promoter than IGF-1 treatment (Fig.…”

“…CsA addition abrogated CoCl 2 -stimulated VEGF secretion, jpet.aspetjournals.org while only partially (ϳ20%) reducing IGF-1-stimulated VEGF secretion. ARPE-19 cells secrete a single species of IGFBP, IGFBP-3, which resolves into four differentially glycosylated species ranging from 28 to 45 kDa in size on SDS gels (Slomiany and Rosenzweig, 2004a). Treatment of ARPE-19 cells with IGF-1 and, to a lesser extent, with CoCl 2 increased IGFBP-3 secretion (Fig.…”

Octreotide Inhibits Choroidal Neovascularization in Rats

“…IGF-1, which is secreted by the RPE cells under autocrine regulation [40,41] , selectively stimulates these cells to release VEGF and upregulates the transcription of this factor through the secondary messenger Ca 2+ , thus inducing CNV [26] . IGF-1 has been identified as an important angiogenic factor in the pathogenesis of CNV [25][26][27] .…”

Inhibition of proliferation, migration and tube formation of choroidal microvascular endothelial cells by targeting HIF-1α with short hairpin RNA-expressing plasmid DNA in human RPE cells in a coculture system

“…Even though the chemical hypoxia method was not identical to physical hypoxia, it was easy and manipulable to mimic the hypoxic condition of solo RPE cells, and could induce the expression of HIF-1α and the downstream transcript, VEGF [17,18]. However, physical hypoxia seemed chronic compared with hypoxia induced by CoCl 2.…”