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Journal Français d'Ophtalmologie
Vol 26, N° 8  - octobre 2003
pp. 782-789
Doi : JFO-10-2003-26-8-0181-5512-101019-ART1
Épithélium conjonctival et glaucome : analyse par cytofluorimétrie en flux de l'expression des marqueurs inflammatoires HLA-DR, IL-6 et IL-8 chez les patients traités
 

L. Bensoussan [1], C. Blondin [1], C. Baudouin [1 et 2], P. Hamard [1], G. Sabeh Afaki [3], C. Creuzot-Garcher [4], J.-M. Warnet [1 et 3], F. Brignole [3]
[1]  Centre Hospitalier National d'Ophtalmologie des Quinze-Vingts, EA 3123 Université Pierre et Marie Curie Paris 6.
[2]  Hôpital Ambroise Paré, AP-HP, Boulogne.
[3]  Laboratoire de Toxicologie, Faculté des Sciences Pharmaceutiques et Biologiques, Université René Descartes Paris 5.
[4]  Centre Hospitalo-Universitaire de Dijon.

Tirés à part : C. Baudouin [5]

[5]  Centre Hospitalier National d'Ophtalmologie des Quinze-Vingts, service III, 28 rue de Charenton, 75012, Paris.

Abstract
Flow cytometric analysis of HLA-DR, IL-6 and IL-8 expression by conjunctival epithelial cells from patients with prolonged topical antiglaucoma treatments

Aim of the study: To investigate the expression of three inflammation markers, HLA-DR, IL-6, and IL-8, by conjunctival epithelial cells obtained using impression cytology (IC) samples from long-term treated glaucoma patients.

Patients and methods: IC samples were obtained from the 60 following individuals: 45 patients suffering from primary open-angle glaucoma and receiving topical treatments for at least 1 year and 15 subjects with no ophthalmological disease (controls). Membrane expression of HLA-DR and intracellular expression of IL-6 and IL-8 were quantified, respectively, by indirect and direct immunofluorescence techniques. Fluorescence levels were quantified using calibrated fluorescent beads.

Results: The percentage of HLA-DR-positive cells was significantly higher on IC samples from multitreated glaucoma patients and from patients treated by either preserved betablocker or preserved prostaglandin analogue than on IC samples from control individuals. Interestingly, the percentage of HLA-DR-positive cells was not significantly increased upon treatment with unpreserved betablocker. However, the percentage of IL-6- and IL-8-positive cells as well as IL-6 and IL-8 expression levels was significantly higher in patients than in controls, regardless of the treatment type and the presence of preservative. A significant positive correlation was found between HLA-DR and cytoplasmic IL-6 expression, between HLA-DR and cytoplasmic IL-8 as well as between IL-6 and IL-8 cytoplasmic expressions.

Conclusion: The present study confirms that there is an increased expression of HLA-DR in treated glaucoma patients compared to controls and demonstrates that antiglaucoma treatments lead to increased IL-6 and IL-8 expressions. It also indicates that benzalkonium-preserved eyedrops and preserved multitherapy may induce stronger inflammatory responses than do unpreserved eyedrops, although further studies are needed to determine the respective inflammatory role of preservative and therapeutic molecules. In this study, flow cytometry was used to detect the intracellular pro-inflammatory cytokines in conjunctival cells obtained by impression cytology. This standardized and reliable technique was a useful tool to assess inflammatory and allergic ocular surface disorders.


Mots clés : Collyres antiglaucomateux , épithélium conjonctival , inflammation , cytokines , cytométrie en flux

Keywords: Antiglaucoma drugs , conjunctiva , inflammation , cytokines , flow cytometry


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