Abstract
BackgroundThe integrin VLA-4 (α4β1) plays an important role in leukocyte trafficking. This study investigated the efficacy of a novel topical α4β1 integrin inhibitor (GW559090, GW) in a mouse model for non-infectious posterior uveitis (experimental autoimmune uveitis; EAU) and its effect on intraocular leukocyte subsets.MethodsMice (female; B10.RIII or C57Bl/6; aged 6–8 weeks) were immunized with specific interphotoreceptor retinoid-binding protein (IRBP) peptides to induce EAU. Topically administered GW (3, 10, and 30 mg/ml) were given twice daily either therapeutically once disease was evident, or prophylactically, and compared with vehicle-treated (Veh) and 0.1% dexamethasone-treated (Dex) controls. Mice were sacrificed at peak disease. The retinal T cell subsets were investigated by immunohistochemistry and immunofluorescence staining. The immune cells within the retina, blood, and draining lymph nodes (dLNs) were phenotyped by flow cytometry. The effect of GW559090 on non-adherent, adherent, and migrated CD4+ T cell subsets across a central nervous system (CNS) endothelium was further assayed in vitro and quantitated by flow cytometry.ResultsThere was a significant reduction in clinical and histological scores in GW10- and Dex-treated groups as compared to controls either administered therapeutically or prophylactically. There were fewer CD45+ leukocytes infiltrating the retinae and vitreous fluids in the treated GW10 group (P < 0.05). Immunofluorescence staining and flow cytometry data identified decreased levels of retinal Th17 cells (P ≤ 0.001) in the GW10-treated eyes, leaving systemic T cell subsets unaffected. In addition, fewer Ly6C+ inflammatory monocyte/macrophages (P = 0.002) and dendritic cells (P = 0.017) crossed the BRB following GW10 treatment. In vitro migration assays confirmed that Th17 cells were selectively suppressed by GW559090 in adhering to endothelial monolayers.ConclusionsThis α4β1 integrin inhibitor may exert a modulatory effect in EAU progression by selectively blocking Th17 cell migration across the blood-retinal barrier without affecting systemic CD4+ T cell subsets. Local α4β1 integrin-directed inhibition could be clinically relevant in treating a Th17-dominant form of uveitis.
Highlights
The integrin very late activation antigen-4 (VLA-4) (α4β1) plays an important role in leukocyte trafficking
It is considered to be an immunemediated disorder based on clinical treatment responses to corticosteroids and immunosuppressants, the ability of adoptively transferred retinal antigen-specific CD4+ T cells to induce an experimental model of autoimmune uveitis (EAU), and histology obtained from postmortem uveitis eyes [3, 4]
The process is accompanied by evidence of blood-retinal barrier (BRB) breakdown [5]; leukocytes infiltrating the anterior and/or posterior chamber and vitreous; immigration of retinal CD4+ T cells, mainly Th1 and Th17 [6]; and structural damage mediated via cytokines released by infiltrating leukocytes, including activated macrophages and neutrophils [7]
Summary
This study investigated the efficacy of a novel topical α4β1 integrin inhibitor (GW559090, GW) in a mouse model for non-infectious posterior uveitis (experimental autoimmune uveitis; EAU) and its effect on intraocular leukocyte subsets. Non-infectious posterior uveitis is an inflammatory disease targeting the uveal tract in the eye mediated by leukocytes that infiltrate the retinal tissues. The specific cell migration process is largely mediated by leukocyte functional antigen-l (LFA-1)/intercellular adhesion molecule-1 (ICAM-l) and very late activation antigen-4 (VLA-4)/vascular cell adhesion molecule-1 (VCAM-1) interactions [10, 11]. Targeting VLA-4 or LFA1 molecules systemically has previously been shown to barricade lymphocytes from crossing the BRB and ameliorate the clinical signs of EAU, indicating the importance of the VLA-4/VCAM-1 pathway in inflamed retinae [15,16,17,18,19,20]. Whether there is a preferential use of specific integrins by Th1 cells versus Th17 cells has not been addressed within the uveitogenic CD4+ T cell subsets
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