CD56+ CD3+CD8bright Natural Killer T–Cells Appear to Correlate With Inflammatory Activity in Behçet’s Disease

Abstract

Purpose: : To analyze a unique population of CD56⁺CD3⁺CD8^bright cells in patients with Behçet’s disease which appears to correlate with disease activity and proliferate in response to ConA and cytokines.

Methods: : Peripheral blood mononuclear cells (PBMCs) were isolated from 5 patients with Behçet’s disease (BD) and 5 normal donors. Following carboxyfluorescein diacetate succinimidyl ester (CFSE) labeling, cells were cultured and stimulated for 5 days using ConA and cytokines. Cells were then stained with antibodies to CD3, CD8 and CD56, and analyzed using flow cytometry. Whole blood phenotyping was performed on the patients and normal donors prior to CFSE labeling and proliferation. Multiplex cytokine array analysis was performed in cultured CD56⁺CD3⁺CD8^bright cells.

Results: : Lymphocytes from peripheral blood sample of one patient with active BD was found to have a significantly higher proportion of CD56⁺CD3⁺CD8^bright cells compared to other BD patients and normal donors. This patient was followed over the course of 4 months, during which disease activity, measured by vitreous haze, fluctuated between 1⁺ to 3⁺. Evaluation of the CD56⁺CD3⁺CD8^bright population during this time indicates that quantitative changes in this population seem to correlate with disease activity, ranging from 8% to 27% of total PBMCs, compared to an average of 3.8% in normal donors. Anterior chamber activity did not appear to correlate with fluctuations in CD56⁺CD3⁺CD8^bright cells. Furthermore, this population of cells proliferates in response to ConA, IL–2, IL12, and IL15. The proliferative response was greatest with ConA, IL2, and IL15. Cytokine array analysis suggests that these CD56⁺CD3⁺CD8^bright cells secrete pro–inflammatory cytokines/chemokines.

Conclusions: : Our data supports previous observations that CD56⁺CD8^bright cells are increased in Behçet’s disease, though this was observed in 1 of 5 patients in our study. The proportion of CD56⁺CD3⁺CD8^bright cells in one patient with BD appears to correlate with severity of disease activity. Our data showing that these CD56⁺CD3⁺CD8^bright cells proliferate in response to ConA and cytokines, and themselves secrete various pro–inflammatory cytokines, suggests that they may contribute to pathogenesis in BD.

Keywords: uveitis-clinical/animal model • flow cytometry • cytokines/chemokines