Diadenosine tetraphosphate improves adrenergic anti-glaucomatous drug delivery and efficiency

The effect of the dinucleotide P-1, P-4-Di (adenosine-5') tetraphosphate (ANA) in improving adrenergic anti-glaucomatous delivery by modifying the tight junction proteins of the corneal epithelium was evaluated. Stratified human corneal epithelial cells (HCLE) were treated with Ap(4)A (100 mu M) for 5 min and TJ protein levels and barrier function were analysed by western blotting and transepithelial electrical resistance (TEER), respectively. Western blot experiments showed a significant reduction at 2 h (45% reduction of ZO-1 and 65% reduction of occludin protein levels) as compared to non-treated (control) cells. Two hours after ANA treatment, TEER values were significantly reduced (65% as compared to control levels (p < 0.001)), indicating an increase in corneal barrier permeability. Topical application of ANA in New Zealand white rabbits two hours before the instillation of the hypotensor compounds (the alpha 2-adrenergic receptor agonist, brimonidine and the beta-adrenergic receptor antagonist, timolol), improved the delivery of these compounds to the anterior chamber as well as their hypotensive action on the intraocular pressure. The results obtained showed that, when ANA was topically applied two hours before the adrenergic compounds, the concentration of brimonidine in the aqueous humour increased from 643 +/- 5.3 nM to 240.6 +/- 8.6 nM and from 58.9 +/- 9.2 nM to 183.7 +/- 6.8 nM in the case of timolol, which also produces a more profound effect on IOP. Therefore, ANA treatment results in a better entrance of adrenergic anti-glaucomatous compounds within the eye and consequently improved therapeutic efficiency by increasing corneal epithelial barrier permeability. (C) 2015 Elsevier Ltd. All rights reserved.