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C. Luo, X. Yang, B. Tezel, J. Cai, W. M. Pierce, G. Tezel; Retinal Hemoglobin (Hb) Expression and Its Differential Regulation During Glaucomatous Neurodegeneration: Insights Into Intracellular Oxygen Transport to Retinal Ganglion Cells (RGCs). Invest. Ophthalmol. Vis. Sci. 2008;49(13):5483. doi: https://doi.org/.
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© ARVO (1962-2015); The Authors (2016-present)
Proteomics has recently discovered Hb in the retinal proteome and this study aimed to further explore this oxygen-binding protein in normal and glaucomatous tissues.
Retinal protein expression was determined by LC-MS/MS analysis. To validate proteomic findings, Hb expression was also studied by RT-PCR and western blot analysis of mixed retinal and enriched RGC samples. In addition, the extent and cellular localization of Hb was determined in histological sections from ocular hypertensive and control rat eyes (n:15) as well as from human donor eyes with glaucoma (n:38) and age-matched controls (n:30). To define whether this protein is regulated by ambient oxygen concentrations, in vitro experiments utilized primary cell cultures incubated in the absence and presence of hypoxia. To determine the role of HIF-1α/erythropoietin (EPO) signaling in Hb induction and cellular survival, recombinant EPO treatment was applied, and EPO neutralization experiments were performed using rsEPO-R treatment of hypoxic cultures.
Proteomic analysis, gene analysis, and immunohistochemistry demonstrated Hb expression in RGCs and glia and its up-regulation in ocular hypertensive rat eyes and glaucomatous human eyes. Up-regulated expression of Hb was found parallel to increased glial expression of EPO and up-regulation of EPO-R on RGCs. In vitro experiments revealed that hypoxia boosts glial Hb expression through HIF1-α and a well-known target of the HIF-1α signaling, EPO, in an autocrine manner. Hypoxia also up-regulated Hb expression in RGCs through the HIF-1α/EPO signaling in a paracrine manner, thereby providing an intrinsic mechanism to increase RGC survival.
Findings of this study provide new insights into how oxygen homeostasis is controlled through regulated expression of Hb in RGCs and glia, and warrant further research. (1) Increased Hb expression in glaucoma appears to be a compensatory mechanism to facilitate intracellular oxygen transport, and perhaps to provide free radical scavenging. (2) Such an Hb-mediated protective mechanism is likely associated with the neuroprotective ability of EPO treatment in glaucoma. (3) Further studies should determine whether an insufficiency of this intrinsic mechanism against hypoxic injury accompanies neurodegeneration, which could open up a new perspective for "RGC hypoxia" even in the existence of sufficient vascular perfusion.
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