The Significance of Ascorbate in the Aqueous Humour Protection Against UV-A and UV-B

doi:10.1006/exer.1996.0031

Experimental Eye Research

Volume 62, Issue 3, March 1996, Pages 261-264

https://doi.org/10.1006/exer.1996.0031 Get rights and content

Abstract

It has previously been suggested that the aqueous humour acts as a UV-filter protecting the structures behind. This hypothesis has here been further evaluated by use of spectrophotometry and spectrofluorimetry. Principally three different aspects of the protecting mechanism have been unveiled: absorption, fluorescence quenching and wavelength transformation. The extremely high ascorbate values obviously play a key role in all of them. Thus, aqueous ascorbate both increased absorption and suppressed fluorescence of radiation below roughly 310nm wavelength. In addition, as a consequence of ascorbate quenching, fluorescence emission to the UV-A range (320–400nm) is substantially reduced.

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In addition, two important non-enzymatic antioxidants exist in the cornea; ascorbic acid and reduced glutathione (GSH). Ascorbic acid (Vitamin C) is found in high concentrations in the corneal epithelium (Brubaker et al., 2000), where it acts to absorb UV radiation (Ringvold, 1996). While ascorbic acid is found at higher levels in the epithelium compared to GSH (Brubaker et al., 2000; Riley and Yates, 1977), GSH is essential for reducing oxidised ascorbic acid to ascorbic acid to maintain ascorbic acid levels (Wu et al., 1998).
In this study we have sought to complete the identification and localisation of uptake pathways involved in accumulating precursor amino acids involved in GSH synthesis in the rat cornea. To do this, we performed reverse transcription PCR (RT-PCR) to identify the Excitatory Amino Acid Transporters (EAAT 1–5) responsible for glutamate uptake, and glycine transporters (GLYT 1–2) at the transcript level. Western blotting was used to verify protein expression, while immunolabelling of sagittal sections was used to localise transporters to the different layers of the cornea. Immunolabelling of en face sections was used to examine the subcellular distribution of proteins in the corneal endothelium. Our findings revealed EAAT 1–5 and GLYT 1–2 to be expressed at the transcript and protein level in the rat cornea. Immunohistochemistry revealed all amino acid transporters to be localised to the epithelium. In the majority of cases, labelling was restricted to the epithelium, and labelling absent from the stroma or endothelium. However, EAAT 4 and GLYT 2 labelling was detected in the stroma with EAAT 4 labelling also present in the endothelium. Overall, the identification of amino acid transporters strongly supports the existence of an intracellular GSH synthesis pathway in the rat corneal epithelium. This suggests that regional differences in GSH accumulation pathways exist, with direct uptake of GSH and intracellular synthesis of GSH restricted to the endothelial and epithelial cell layers, respectively. This information is important in the design of targeted strategies to enhance GSH levels in specific layers of the cornea to prevent against oxidative damage, corneal swelling and loss of corneal transparency.
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Selenium and pseudoexfoliation syndrome
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Citation Excerpt :
It is for this purpose that aqueous humor contains a series of molecules such as ascorbic acid, urate, L-tyrosine, L-cysteine, and glutathione (GSH) that protect the eye against free radical damage. Ascorbic acid is the most important of these molecules in terms of ocular antioxidant defense.19–21 A nonenzymatic extracellular oxido-reduction system has been postulated as a major mechanism for protecting the anterior eye from free radical damage and H2O2 toxicity.22
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^f1: Address for correspondence: Eye Department, Rikshospitalet, Pilestr. 32, N-0027 Oslo, Norway

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Regular articleThe Significance of Ascorbate in the Aqueous Humour Protection Against UV-A and UV-B

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Regular article
The Significance of Ascorbate in the Aqueous Humour Protection Against UV-A and UV-B