Elsevier

Experimental Eye Research

Volume 91, Issue 3, September 2010, Pages 456-461
Experimental Eye Research

Stromal interleukin-1 expression in the cornea after haze-associated injury

https://doi.org/10.1016/j.exer.2010.06.023Get rights and content

Abstract

The purpose of this study was to determine whether myofibroblasts or other cells in the stroma in the cornea produce interleukin (IL)-1α or IL-1β that could modulate myofibroblast viability in corneas with haze after photorefractive keratectomy (PRK). Twenty-four female rabbits had haze-generating PRK for 9 diopters of myopia and were sacrificed at 1 week, 2 weeks, 3 weeks or 4 weeks after surgery. Corneal rims were removed, frozen in OCT at −80 °C, and analyzed by immunocytochemistry using primary antibodies to IL-1α, IL-1β and alpha smooth muscle actin (SMA). Double immunostaining was performed for the co-localization of SMA with IL-1α or IL-1β. Central dense haze and peripheral slight haze regions of each cornea were analyzed. SMA+ cells that expressed IL-1α protein were detected in both regions of the corneas at most time points following PRK. However, in the haze region at the 1, 3 and 4 week time points, significantly more (p < 0.01) SMA+ cells did not express IL-1α. Also, in the haze region at all three time points, significantly more (p < 0.01) SMA- cells than SMA+ cells expressed interleukin-1α protein. IL-1β expression patterns in SMA+ and SMA- stromal cells was similar to that of IL-1α after PRK. Previous studies have demonstrated that IL-1α or IL-1β triggers myofibroblast apoptosis in vitro, depending on the available concentration of apoptosis-suppressive TGFβ. This study demonstrates that SMA- cells such as corneal fibroblasts, keratocytes, or inflammatory cells may produce IL-1α and/or IL-1β that could act in paracrine fashion to regulate myofibroblast apoptosis—especially in the region where there is haze in the cornea after PRK was performed and SMA+ myofibroblasts are present at higher density. However, some SMA+ myofibroblasts themselves produce IL-1α and/or IL-1β, suggesting that myofibroblast viability could also be regulated via autocrine mechanisms.

Introduction

Corneal injury, including surgery or infection, may trigger a loss of corneal transparency associated with the generation of stromal myofibroblasts that produce disordered matrix components such as collagen and glycosaminoglycans (Masur et al., 1996, Jester et al., 1999a, Wilson et al., 1999, Netto et al., 2006). TGFβ and PDGF have been shown to have important roles in modulating the development of corneal myofibroblasts from precursor cells (Masur et al., 1996, Jester et al., 1999b, Jester et al., 2002, Kaur et al., 2009b). The appearance and persistence of the myofibroblasts has been hypothesized to occur when structural and/or functional defects in the regenerated epithelial basement membrane allows penetration of TGFβ and PDGF from the epithelium into the stroma at sufficient levels required for receptor activation (Netto et al., 2006, Kaur et al., 2009b).

Over a period of years following surgery, many cases of corneal haze that occur after photorefractive keratectomy resolve spontaneously (Rajan et al., 2004). Recent work demonstrated that myofibroblast apoptosis has an important role in the resolution of corneal haze through the removal of the cellular contribution to the opacity (Wilson et al., 2007). Remaining anomalous extracellular matrix is subsequently removed by keratocytes that repopulate the anterior stroma. In a recent in vitro study, Kaur et al. (2009a) demonstrated that exogenous interleukin (IL)-1 alpha or IL-1 beta triggers apoptosis of corneal myofibroblasts and that TGFβ blocks this effect. The current study examines potential in situ sources of IL-1 alpha or IL-1 beta in the corneal stroma following haze-generating photorefractive keratectomy in a rabbit model.

Section snippets

Animals and surgery

All animals were treated in accordance with the tenets of the ARVO Statement for the Use of Animals in Ophthalmic and Vision Research. The Animal Control Committee at the Cleveland Clinic approved the animal studies included in this work. Anaesthesia was obtained by intramuscular injection of ketamine hydrochloride (30 mg/kg) and xylazine hydrochloride (5 mg/kg). In addition, topical proparacaine hydrochloride 1% (Alcon, Ft. Worth, TX, USA) was applied to each eye at the time of photorefractive

Results

In response to -9 diopter PRK in the rabbit, little haze can be noted in corneas 1 week or 2 weeks after PRK (not shown). At week 3, a significant haze response to the PRK is present (not shown). By 4 weeks after the PRK there is dense haze that obscures iris details (Fig. 1).

Immunocytochemistry (Fig. 2) and cell counts performed on double-stained slides (Table) from the areas of the stroma with dense haze in all of the corneas treated with PRK revealed the presence of SMA+ and SMA- stromal

Discussion

Severe haze (or opacity) sometimes develops in the corneal stroma after photorefractive keratectomy (PRK) and other surgeries or injuries. Myofibroblasts have an important role in the development of haze after they are generated following injury to some corneas. Large numbers of these cells result in altered transparency through the production of disordered collagen and other extra cellular matrix materials produced by these cells and the opacity of the cells themselves (Jester et al., 1999a,

Acknowledgements

This study was supported by EY10056, EY015638, and Research to Prevent Blindness, New York, NY.

References (24)

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