MMP-9 Mediates IL-2 Receptor Alpha Chain (CD25) Loss in Murine Corneal and Conjunctival Epithelia in Desiccating Stress

Abstract

Purpose:: IL-2 is a pleiotropic cytokine that has been identified to play a pivotal role in regulating the adaptive immune response. We propose to investigate the expression of interleukin-2 (IL-2) receptor alpha (CD25) and beta (CD122) chains by the ocular surface epithelia in experimental murine dry eye.

Methods:: Experimental dry eye (EDE) was created by subjecting C57BL/6 and MMP-9 knockout (BKO) mice and their wild-type littermates (WT) mice to desiccating environmental stress for 5 or 10 days. A separate group of C57BL/6 mice was subjected to EDE and treatment with topical 0.025% doxycycline, administered QID. The expression of CD25 and CD122 was evaluated in cryosections by dual-label laser scanning confocal microscopy. Western-blot was used to measure relative levels of CD25 in epithelial lysates. Gelatinase activity was evaluated by in situ zymography. Soluble CD25 in tear fluid was measured by an immunobead assay.

Results:: CD25 and CD122 were found in the corneal (all layers) and conjunctival (apical and subapical layers) epithelia of untreated C57BL/6, BKO and WT mice. Desiccating stress caused a marked decrease in CD25 immunoreactivity in the corneal and conjunctival epithelium of C57BL/6 and WT mice, while CD122 expression remained unchanged. The decrease in CD25 expression corresponded to an increase in gelatinase activity. Treatment of EDE with doxycycline abolished EDE induced gelatinase activity and preserved CD25. EDE had no effect on CD25 expression in the MMP-9 knockout mice. Soluble CD25 protein was detected in higher levels in tears of dry eye C57BL/6 mice than controls.

Conclusions:: Desiccating stress promotes CD25 loss from the ocular surface epithelia in C57BL/6 mice and WT mice, and this was accompanied by increased gelatinase activity and higher soluble CD25. Preservation of membrane CD25 with doxycycline and the resistance of MMP-9 knock-out mice to loss of CD25 immunoreactivity in response to desiccating stress suggests that CD25 is proteolytically cleaved by MMP-9 from the ocular surface epithelia into the tears in dry eye.

Keywords: cornea: tears/tear film/dry eye • cytokines/chemokines