Influence of Antiglaucoma Compounds on Ocular Surface Corneal and Conjunctival 2 Adrenergic Receptor Expression in vitro

Abstract

Purpose: : Brimonidine, an adrenergic receptor (₂AR) agonist used for the treatment of glaucoma, elicits conjunctivitis in 15% of patients by a mechanism that still remains elusive. Our group has reported the presence of ₂AR in human conjunctiva and their up-regulation in cells exposed to inflammatory cytokines. To start investigating whether this drug could exert its mechanism of action by altering the expression of ₂AR at the ocular surface, we studied the expression levels of ₂AR in conjunctival and corneal cells treated with brimonidine.

Methods: : Human conjunctival (IOBA-NHC) and corneal (HCE) cell lines were cultured for 24h and 48h under 3 different conditions: 1) control: supplemented culture medium (SCM); 2) SCM and brimonidine, 3) SCM and brimonidine plus timolol. The mRNA and protein expression of _2A, _2B and _2CAR was analyzed by real time RT-PCR, Western blotting, and flow cytometry. Cell viability, apoptosis, and cell cycle stage were also studied by flow cytometry.

Results: : The _2AAR gene and protein expression was up-regulated in both cell lines after 48h with either treatment. The mRNA expression of _2BAR and the gene and protein expression of _2cAR were down-regulated after 48h with either treatment and in both cell lines. The _2BAR protein expression was not altered. Both treatments affected viability, apoptosis, and cell cycle stage.

Conclusions: : Both brimonidine and brimonidine plus timolol were able to alter the level of expression of ₂AR in conjunctival and corneal cells. Whether this in vitro effect will have a clinical relevance remains to be confirmed.

Keywords: conjunctiva • cornea: basic science • gene/expression