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Françoise Brignole, Pierre-Jean Pisella, Magda De Saint Jean, Marie Goldschild, Alain Goguel, Christophe Baudouin; Flow Cytometric Analysis of Inflammatory Markers in KCS: 6-Month Treatment with Topical Cyclosporin A. Invest. Ophthalmol. Vis. Sci. 2001;42(1):90-95.
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purpose. Immune-based inflammation has been observed as a common mechanism of
keratoconjunctivitis sicca (KCS). In KCS-affected eyes, upregulated
expression of HLA DR and various immune- or apoptosis-related markers
by conjunctival epithelial cells has been demonstrated in an earlier
study, by a technique of flow cytometry in impression cytology (IC)
specimens. The purpose of this study was to monitor the effects of
topical cyclosporin A on the expression of these markers throughout a
6-month period of treatment.
methods. Patients with moderate to severe KCS included in a large European
multicenter clinical trial (Cyclosporin Dry Eye Study, Allergan,
Irvine, CA) underwent collection of IC specimens at baseline, month 3,
and month 6. For 6 months, they randomly received 0.05% or 0.1%
cyclosporin A or vehicle. Specimens were processed and analyzed in a
masked manner by flow cytometry, using monoclonal antibodies directed
to HLA DR, CD40, CD40 ligand, Fas, and the apoptotic marker APO2.7.
Percentages of positive cells were calculated and levels of expression
quantified after conversion into standardized units of fluorescence.
results. One hundred fifty-eight patients had at least two IC specimens
available for flow cytometry analysis. HLA DR expression, both in
percentage of positive cells and level of expression, was highly
significantly reduced after 0.05% and 0.1% cyclosporin A treatment at
months 3 and 6 compared with baseline values, whereas vehicle did not
induce any change in HLA DR expression over time. The 0.05% and 0.1%
cyclosporin emulsions were significantly more effective than the
vehicle in reducing HLA DR at months 3 and 6 (0.05%), and at month 6
(0.1%). CD40 expression was significantly reduced at month 3 and
partially at month 6, compared with baseline, with no reduction in
patients who received the vehicle. CD40 ligand expression also
decreased at months 3 and 6 in patients taking both concentrations of
cyclosporin A. APO2.7 expression was significantly increased in all
three groups, whereas percentage of Fas-positive cells decreased only
in patients treated with 0.05% cyclosporin A at months 3 and 6.
conclusions. Flow cytometry provided an objective technique to monitor the effects
of topical cyclosporin A on immune- and apoptosis-related markers in
the conjunctival epithelium of patients with KCS enrolled in a large
multicenter trial. Topical cyclosporin A strikingly reduced HLA DR and
to a lesser extent, other inflammatory and apoptotic markers, whereas
the vehicle, used as a control tear substitute, had almost no effect.
This study confirms that cyclosporin A may be efficient in reducing
conjunctival inflammation in moderate to severe KCS and is consistent
with clinical results in this indication.
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