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J. H. Hou, Y. B. Shui, S. S. Shetty, C. J. Siegfried, N. M. Holekamp, D. C. Beebe; The Biochemistry of Ascorbate-Dependent O2 Consumption in Human Vitreous. Invest. Ophthalmol. Vis. Sci. 2009;50(13):2113.
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Human vitreous plays a novel physiologic role in the maintenance of a hypoxic intraocular environment by consuming O2 in an ascorbate (AsA)-dependent manner (Shui et al. Arch Ophthalmol. In press). Destruction of the vitreous gel by vitrectomy or age-related liquefaction increases intraocular pO2 and the risk of nuclear cataract. Another report found increased risk of primary open angle glaucoma (POAG) following vitrectomy and cataract surgery, presumably due to a secondary rise in pO2 in the anterior chamber (Chang. Am J Ophthalmol. 141 1033). Manipulation of the anti-oxidative potential of the vitreous and aqueous humors is a promising avenue for therapeutic interventions aimed at reducing the incidence of cataracts and POAG. However, little is known about the mechanism of O2 metabolism in the vitreous. This study evaluated factors in the vitreous that affect the rate of O2 consumption.
O2 consumption was measured with a fiberoptic probe in cadaver vitreous. Ultrafiltration and chelating agents were used to assess the size and nature of the molecule(s) that catalyze the reaction between AsA and O2. Vitreous was treated by ultrafiltration, boiling, or addition of 3-aminotriazole (3-AT) to remove, denature, or inhibit catalase activity, respectively. Western blotting was used to quantify catalase levels.
Vitreous O2 consumption was only slightly inhibited by treatment with deferoxamine, an iron chelator. The catalytic activity readily passed through a 10 kDa cutoff filter. The 10 kDa filtrate showed a 2-fold increase in the rate of O2 consumption, compared to unfiltered vitreous. Addition of 3-AT to unfiltered vitreous caused an increase in the rate of O2 consumption, similar to that observed after ultrafiltration. Addition of 3-AT to the filtered vitreous caused only minimal change in O2 consumption. Boiling vitreous resulted in a ~50% loss of AsA in the treated samples, but treated samples consumed O2 at rates similar to untreated vitreous. Catalase was surprisingly abundant in human vitreous, ~ 0.1 - 1.0 µg/ml in initial assays.
Our data suggest that catalase in human vitreous substantially slows the rate of vitreal O2 consumption, probably by releasing O2 from the H2O2 produced during AsA oxidation. Catalase may play an important role in protecting the eye from peroxide generated in the reaction between AsA and O2. A proper balance between the levels of AsA, O2, catalyst(s), and catalase is likely to be critical for protecting the lens and trabecular meshwork from oxidative damage. Measurement of these variables in vitreous and aqueous samples obtained during retina, cataract and glaucoma surgery is in progress.
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