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E. Bodek, L. Geng, H. J. Kaplan, T. H. Tezel; Retinal Pigment Epithelium (RPE) Hemoglobin Production Increases With Hypoxia. Invest. Ophthalmol. Vis. Sci. 2008;49(13):3977.
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. To determine the effect of physiologic or pathological hypoxia on RPE hemoglobin (Hb) production.
. The effect of physiological hypoxia on RPE-Hb production was studied on 24 C57Bl/6 mice and 6 brown Norwegian rats that were kept 12/12 hours of dark/light cycles for one day. Animals were euthanized at the end of either dark or light cycle and RPE cells were harvested. Change in Hb expression was studied with RT-PCR and western blot analyses. In another set of experiments, 20 C57Bl/6 were kept in an hypoxic chamber (10% oxygen) for 12 hours. At the end of this period change in Hb, erythropoietin (EPO), erythropoietin-receptor (EPO-R), VEGF expression was determined with immunohistochemistry, RT-PCR and western blot analyses. Animals kept in normoxic (21% oxygen) conditions were taken as controls. Modulation of Hb production with EPO was studied in vitro with ARPE-19 cells. Confluent cultures were incubated with EPO (20 U/ml) for 72 hours. Cells incubated with human retina and rod outer segments (106/ml) were used as controls.Result. Keeping the animals in dark upregulated Hb mRNA expression 3.6 ± 0.8 times, HIF1α expression 2.6 ± 0.6 times and VEGF expression 1.4 ± 0.4 times. Similarly, hypoxia increased the expression of Hb 3.0 ± 1.1 times. Despite an increase in retinal and RPE EPO-R expression we did not observe an increase in EPO levels. Immunohistochemistry showed intense HIF1α staining in RPE and photoreceptor outer segments in control animals. With hypoxia signal intensity increased and involved photoreceptor inner segments. EPO expression was observed in photoreceptor outer segments and ganglion cells. EPO expression was upregulated with hypoxia. Supplementing the culture medium with EPO increased Hb production in ARPE-19 cells; however no significant change was observed with the presence of retina or rod outer segments in the culture medium.
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