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K. Kahook, A. M. Tawfik, R. Mussell, V. Sarthy, M. A. Al-Shabrawey; Role of 12-Lipoxygenase in Pathological Retinal Neovascularization. Invest. Ophthalmol. Vis. Sci. 2009;50(13):5691.
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Pathological angiogenesis is a common and potentially vision threatening complication of eye diseases such as diabetic retinopathy and retinopathy of prematurity. Angiogenesis, a growth of new blood vessels from preexisting vessels is controlled by a large number of modulating factors, such as VEGF and PEDF. These pro-and antiangiogenic factors are both produced by Müller cells and regulated by hypoxia. Disruption of the balance which is normally present between the level of VEGF and PEDF is crucial in mediating retinal neovascularization. Arachidonic acid is metabolized by 12-lipoxygenase (12-LO) to 12- hydroxyeicosatetraenoic acid (12-HETE). 12- HETE has been reported to play a significant role in tumor angiogenesis and promotes endothelial cell proliferation and migration. However, the specific role of 12-HETE in retinal angiogenesis has not been yet identified. The goal of this study was to investigate whether 12-LO plays a role in the pathological retinal neovascularization by targeting VEGF and PEDF expression.
Oxygen-induced retinopathy (OIR) was developed by exposing neonatal mice to 75% oxygen (P7-P12) followed by normoxia (P12-P17). Baicalein, a 12- lipoxygenase inhibitor, was administered intraperitoneally (20mg/kg) into one group on P12, P14 and P16. The mice then were sacrificed on P17 and one eye was processed for morphological study and the other for biochemical assay. Labeling of retinal flat mounts with Isolectin B4 was used to assess retinal neovascularization. 12-lipoxygenase expression and localization in retina was demonstrated by Immunofluroscence. Hypoxic areas in the retina were identified by a specific hypoxia marker, pimonidazole HCl. For in vitro study, rat Müller cells (rMC) treated with or without 12-HETE (0.5 µM and 1 µM ) were used. This was followed by analyzing VEGF and PEDF production using ELISA and Western blotting respectively.
OIR was associated with increased retinal expression of 12-LO and patchy areas of hypoxia localized particularly in Müller cells. Retinal neovascularization was markedly abrogated in baicalein treated mice compared with the control (3.1+0.3 vs 1.3+0.01). rMC treated with 0.5 µM or 1.0 µM 12-HETE showed increases in VEGF production (367+18 and 343+74 respectively) compared with the control (143+25) and decreases in PEDF expression (0.1+0.01 and 0.2 +0.03) compared with the control (0.3+0.06).
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