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Paula Urquhart, Jenny Wang, David F. Woodward, Anna Nicolaou; Analysis of Prostamides E2 and F2α Produced in Rabbit Cornea. Invest. Ophthalmol. Vis. Sci. 2012;53(14):4207.
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One main function of the cornea is to protect the intraocular structures of the eye. It does this partly by responding to intraocular pressure. Corneal injury resulting from surgery or infections, can cause pain and result in tissue scarring. Injury stimulates arachidonic acid release, cyclo-oxygenase-2 (COX-2) up-regulation and, consequently, formation of bioactive lipid mediators including prostaglandins (PG). PGE2 suppresses the mitogenic response to epithelial growth factor, thus regulating cell proliferation, whilst PGF2α is a well established anti-hypertensive agent. Stable analogues of prostaglandins such as bimatoprost, an ethanolamide derivative of PGF2α, are widely used in the treatment of glaucoma and management of ocular hypertension. Here, we report the identification and quantitation of prostaglandin ethanolamides (prostamides; PG-EA) PGE2-EA and PGF2α-EA in rabbit corneal tissue.
Lipids were extracted with chloroform : methanol (2:1, v/v). Lipid extracts were analysed by reverse phase chromatography using gradient elution with acetonitrile: water: acetic acid, pH 3.0. Quantitation was achieved by positive ion tandem electrospray ionisation mass spectrometry using deuterated arachidonoyl ethanolamine (anandamide; A-EA-d8) as internal standard. Each prostamide was identified by three separate transition ions: PGE2-EA, m/z 378>360, m/z 378>342 and m/z 378>62; PGF2α-EA, m/z 380>344, m/z 380>283 and m/z 380>62.
The concentrations of PGE2-EA and PGF2α-EA were 0.02 and <0.01 pg/mg tissue, respectively. Furthermore, A-EA, their biochemical precursor, was also identified in the rabbit cornea at a concentration of 0.55 pg/mg. The ability of the corneal tissue to metabolise A-EA to prostamides via COX-2, was confirmed using tissue homogenate incubated with external A-EA (10μM). This generated PGE2-EA and PGF2α-EA at a ratio of 3:1. Profiling of corneal prostaglandins revealed production of PGE2 and PGF2α at the same ratio, suggesting that the profile of prostamides in the cornea could be determined by the expression of the corresponding prostaglandin synthases.
Overall, the identification of prostamides in corneal tissue confirms the presence of this family of lipid mediators in the eye and may, in part, explain the potent pharmacological activity of prostaglandin derivatives used in the treatment of ocular conditions.
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