Regulation of Pigment Epithelium–Derived Factor Production and Release by Retinal Glial (Müller) Cells under Hypoxia

PURPOSE. To assess the regulation of pigment epithelium- derived factor (PEDF) production by retinal Miiller glial cells, especially under ischemic or hypoxic conditions. METHODS. PEDF was determined in surgically excised retinal tissue originating from patients with ischemic diabetic retinopathy and in primary guinea pig Miiller cell cultures exposed to the protein synthesis inhibitor cycloheximide (CHX) and to hypoxia. PEDF production and secretion were studied by immunohistochemistry, quantitative RT-PCR, ELISA, fluorescence-activated cell sorter analysis, and Western blotting. RESULTS. Gliotic Miiller cells displayed decreased PEDF immunoreactivity in fibrovascular tissue from patients with diabetes compared with tissue from subjects with pathologic myopia. In Muller cell cultures, CHX treatment resulted in an increase, whereas mild hypoxia (2.5%-10% O 2 ) induced a decrease, of PEDF mRNA and protein levels. However, strong hypoxia (0.2% O 2 ) induced an upregulation of PEDF mRNA expression and resulted in only slightly reduced PEDF levels after 24 hours, detected as either a released, soluble, or cell surface-linked protein. CONCLUSIONS. These results suggest that under certain conditions including mild hypoxia, Miiller cells synthesize a protein factor that downregulates PEDF expression or its turnover. Generally, the cells appear to generate a biphasic response to hypoxia. In moderate hypoxia, PEDF is downregulated such that the VEGF-to-PEDF ratio increases (and angiogenesis is facilitated). During severe (or chronic) oxygen deficiency, however, the PEDF decline is arrested or even reversed; thus, the neurotrophic effects of PEDF remain available.