Elsevier

Experimental Eye Research

Volume 79, Issue 3, September 2004, Pages 357-365
Experimental Eye Research

Etanercept treatment in the endotoxin-induced uveitis of rats

https://doi.org/10.1016/j.exer.2004.06.001Get rights and content

Abstract

This study was conducted to investigate therapeutic value of a soluble tumor necrosis factor-α (TNF-α) receptor, etanercept, in a rat model of endotoxin-induced uveitis (EIU). Forty-two inbred male Lewis rats were divided into seven equal groups. 200 μg of Escherichia coli 055:B55 lipopolysaccharide (LPS) was injected in one hind footpad of the Groups 2, 3, 4, 5, 6, and 7 rats. Group 5, 6, and 7 rats also received subcutaneous etanercept 24 hr prior to LPS injection at a dose of 0·4 mg kg−1. Group 1 rats were used as controls. Eight, 24, and 48 hr after treatment clinical uvetis scores (miosis, iris hyperemia, and hypopyon) were assessed by a masked observer and the rats were euthanized. Neutrophil leukocytes, CD8+, CD4+, and CD45RO+ cells in the anterior uveal tissue were counted either after hematoxylin–eosin or monoclonal antibody staining. TNF-α levels were also measured in the aqueous humor samples by an ELISA method. Etanercept treatment significantly improved clinical uveitis scores at all examination points compared to the LPS injected animals. The improvement was almost complete expect for the miosis score, since no significant difference was detected between the controls and LPS+Etanercept treated animals at all examination points. Cell counts were also at significantly lower levels in LPS+Etanercept treated animals at all examination points, except for CD8+ and CD45RO+ cell counts at 24 hr examination point. There was no significant difference between the controls and LPS+Etanercept treated animals at all examination points as with CD4+ and CD45RO+ cell counts at 48 hr. Our data showed that etanercept had a definite effect on the treatment of EIU. Further studies should clarify its efficacy on clinical uveitis conditions.

Introduction

A single, low-dose injection of endotoxin, the lipopolysaccharide (LPS) component of gram-negative bacterial cell walls, at a site distant from the eye, such as the footpad, induces anterior uveitis in susceptible species and strains (Rosenbaum et al., 1980, Bhattacherjee et al., 1983, Kogiso et al., 1992). This phenomenon is called endotoxin-induced uveitis (EIU). Four hours after LPS injection, the breakdown of the blood-aqueous barrier can be detected, but cell infiltrates of the anterior segment are maximal at 24 hr (Okumura and Mochizuki, 1982, Ruiz-Moreno et al., 1992). The predominant infiltrating cell in EIU is the neutrophil, but some data support the fundamental role for T cells especially CD4+ T cells (Kogiso et al., 1992, McMenamin and Crewe, 1995). Several studies have indicated that tumor necrosis factor-α (TNF-α) was also involved in the development of EIU (de Vos et al., 1992, Yoshida et al., 1994, Rosenbaum and Boney, 1993, de Kozak et al., 1994). Evidence for a direct role of TNF-α in EIU came from the finding that intraocular injection of TNF-α into rat eyes induced acute uveitis that closely resembled the response to LPS (de Vos et al., 1995a). Thus, it is logical to think that TNF-α antagonists would have beneficial effect on EIU. Indeed, Koizumi et al. (2003) have recently demonstrated that administration of etanercept reduced leukocyte rolling and adhesion significantly in their rat model of EIU. EIU is characterized by an increased leukocyte adhesion and rolling, predominantly in veins (Miyamoto et al., 1996, Miyamoto et al., 1998, Becker et al., 2001). Etanercept is a soluble TNF-α receptor which binds to TNF-α and prevents its binding to TNF-α receptor, hence blocks its action. If there is any beneficial effect of a TNF-α antagonist on EIU this effect should most probably occur early in the course of EIU, since TNF-α plays a crucial role in the early phases of EIU (Yoshida et al., 1994, de Vos et al., 1994). Thus, to investigate the value of etanercept in the treatment of EIU, we treated some rats 24 hr prior to induction of EIU. The clinical uveitis scores of animals and immunohistopathological findings were compared to sham injected controls and rats with EIU without any treatment.

Section snippets

Material and methods

Forty-two inbred male Lewis rats, weighting 150–200 g (6–8 weeks old) were randomly divided into seven groups. Each group contained six rats. This study was carried out at the Selcuk University Experimental Medicine Research and Application Center (SUDAM). All experiments were approved by the SUDAM Ethics Committee and adhered to the Association for Research in Vision and Ophthalmology resolution on the use of animals in research. Animals were housed at 21°C and 50% humidity in a 12-hr light

Clinical uveitis scores

The control animals did not show any clinical signs of uveitis.

ANOVA test revealed statistically significant differences between groups at all different time points (8, 24, and 48 hr; all p=0·001). There was no statistically significant difference between LPS injected and LPS+ETA treated animals at 8 hr in terms of miosis, but both LPS injected and LPS+ETA treated animals showed significantly higher miosis scores compared to the control animals (both p=0·001). We did not detect any significant

Discussion

In the present study, we clearly showed that the TNF-α receptor fusion protein etanercept had a beneficial effect on both clinical parameters and cellular infiltration in the rat model of endotoxin-induced uveitis. The beneficial effect of etanercept treatment resulted in complete regression of some parameters of EIU, since no significant difference was present between the controls and LPS+ETA treated experimental animals (i.e. iris hyperemia and hypopyon scores at all examinations, neutrophil

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    Contributed equally to the work and therefore should be considered equivalent authors.

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