Abstract
Purpose: :
Dry eye disease can be associated with increased tear film osmolarity leading to elevated inflammatory cytokine and matrix metalloproteinases levels in ocular surface epithelial cells. As peroxisome proliferator-activated receptor γ (PPARγ) gene overexpression is implicated in inhibiting injury-induced corneal inflammation in mice, we determined if changes in the PPARγ activation status of human corneal epithelial cells (HCEC) affect the increases in proinflammatory cytokine release induced by a hypertonic challenge.
Methods: :
Immunocytochemistry probed for PPARγ expression in SV40-immortalized human corneal epithelial cells (HCEC). To monitor changes in IL-8 release in triplicate, ELISA was used. HCEC were preexposed for 30 min to either a PPARγ agonist or an antagonist, (1 µM ciglitazone or 1 µM GW9662) under isotonic conditions. Subsequently, they were maintained for an additional 20 h in sucrose supplemented hypertonic (450 mOsm) DMEM/F12 medium, respectively. In the other three groups, HCEC were maintained in an isotonic DMEM/F12 medium (i.e.300 mOsm) throughout and were exposed to either: a) ciglitazone; b) GW9662. Supernatants were collected and centrifuged at 1,000 rpm for 5 min to remove cell debris. The amount of IL-8 in the culture medium was normalized to the total amount of lysed cellular protein.
Results: :
PPARγ expression was detected in the nuclear membrane. In isotonic medium (300 mOsm), PPARγ activation by ciglitazone decreased IL-8 release by 26% from its isotonic level of 2053 ± 67 pg/mg protein to 1522±52 pg/mg protein whereas PPARγ inhibition by GW9662 elevated this control IL-8 level by 51% to 3107±178 pg/mg protein. Under the hypertonic condition (450 mOsm), IL-8 release instead increased 3.83-fold from the isotonic control to 9911± 52 pg/mg protein. On the other hand, ciglitazone preexposure suppressed this rise by 81%. IL-8 release declined from 9911± 52 to 1929± 58 pg/mg protein. Conversely, GW9662 preexposure enhanced the hypertonicity-induced IL-8 increase by 68% to 16647 ± 453 pg/mg protein.
Conclusions: :
Ciglitazone blocked essentially a large amount of the rise in IL-8 release induced by hypertonicity whereas GW9662 enhanced the increase in IL-8 induced by this challenge. These opposing effects on PPARγ activation point to the possibility that ligand-induced PPARγ activation in a clinical setting may reduce in some cases anterior ocular surface inflammation.
Keywords: cornea: epithelium • cytokines/chemokines • inflammation