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W. Lu, C. Rasmussen, B. Gabelt, B. Hennes, P. Kaufman, A. M. Laties, C. H. Mitchell; Upregulation of NTPDase 1 in an Experimental Monkey Glaucoma Model. Invest. Ophthalmol. Vis. Sci. 2007;48(13):4804.
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ATP is released in numerous tissues to signal a change in flow, pressure or other mechanical perturbation. The increased intraocular pressure that can accompany glaucoma may also trigger the release of ATP. Retinal ganglion cells die in glaucoma, and stimulation of the P2X7 receptors for ATP can kill ganglion cells. Elevated pressure increases the concentration of ATP in the vitreous chamber of the bovine eyecup. Direct measurements of ATP levels in the extracellular space of the retina are not presently possible. However, we have recently found that the enzyme ecto-nucleoside triphosphate diphosphohydrolase 1 (NTPDase1; aka CD39) is upregulated after prolonged exposure to extracellular ATP. This study asked whether expression of the NTPDase1 was increased in the retinas of primates with elevated intraocular pressure (IOP).
Elevation of IOP was induced in one eye of 15 monkeys by laser photocoagulation of the trabecular meshwork. The IOP was monitored weekly; IOP values are the mean measurement throughout the experiment. For immunoblot experiments, eyes were fast frozen, retinal proteins purified using standard techniques and run on a SDS-PAGE. Gels were blotted with antibodies to NTPDase1(BU61) and staining quantified with Image Pro Plus software. For immunohistochemical studies, tissue was perfused with 2% paraformaldehyde, sectioned at 12 µm and processed using the same antibody as above.
The IOP of the lasered eyes was significantly higher than the non-lasered controls, at 21.8 ±1.2 mm Hg vs. 15.6 ±0.4 mm Hg, respectively (p<0.01). Expression of NTPDase1 was also increased in the lasered eyes. Western blot analysis gave the mean ratio of protein in lasered vs non-lasered (L/NL) eye as 2.00 ± 0.28 (Mean ±SE). There was a significant correlation between the IOP and the increase in NTPDase1 protein in the glaucomatous retina (r2 = 0.714). Immunohistochemical analysis indicated that the upregulation of NTPDase1 occurred both in the inner nuclear layer and the optic nerve.
We have demonstrated that NTPDase1 levels are increased in the inner nuclear layer and optic nerve head of eyes with increased intraocular pressure. This result provides indirect evidence for chronic exposure to excess extracellular ATP in experimental glaucoma.
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