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B. D. Quill, E. M. McElnea, N. G. Docherty, M. Irnaten, R. C. Barry, A. F. Clark, C. J. O’Brien; Elevated Intracellular Free Calcium in Glaucoma Derived Lamina Cribrosa Cells. Invest. Ophthalmol. Vis. Sci. 2009;50(13):2790.
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At confluence, LC cells from ‘normal’ and glaucoma donors were serum starved for 24 hours then exposed to isotonic or hypotonic solution to create hypotonic stretch. Cellular protein was harvested between 1 and 30 minutes to measure phosphorylated PKC- by Western blotting. Intracellular free calcium was measured before and during hyotonic stretch using FURA 2AM spectrofluorescence ratios. In addition, cDNA from both cell types was used to run Taqman low density array analysis of twenty genes involved in calcium homeostasis.
In glaucomatous LC cells, basal and hypotonic stretch induced intracellular calcium levels are significantly higher (0.71 ± 0.03 v 0.66 ± 0.03, 0.94 ± 0.03 v 0.74 ± 0.02; p < 0.05). Glaucomatous LC cells also showed increased basal and induced levels of activated PKC alpha and significant upregulation of the endoplasmic reticulum Ca2+ pump SERCA3 mRNA(6 fold, p<0.05). In contrast the store-operated channel TRPC1 mRNA was 70% lower in glaucomatous LC cells (-0.7 fold, p<0.05).
Glaucomatous LC cells have elevated intracellular calcium and increased PKC alpha signalling. Therefore, regulating intracellular Ca2+ levels and associated downstream signalling may represent a potential therapeutic target in glaucoma.
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