Reduced functional activity of natural killer cells in spondyloarthritis patients may be related to the expression of killer immunoglobulin-like receptor 1 (KIR3DL1)

Background: Inflammatory mediators in spondyloarthritis (SpA) comprise components of both innate and adaptive immune system (1). In this respect, interesting are the findings that natural killer (NK) cells possess killer immunoglobulin-like receptors (KIRs) that bind HLA-B27 homodimers, abnormal molecules lacking beta2-microglobulin: the interaction between these molecules seems to be relevant in the pathogenesis of SpA in humans. Objectives: To determine the cytotoxic activity of NK cells bearing the inhibitory receptor KIR3DL1 in SpA patients. Methods: After informed consent was obtained, we enrolled twenty-one outpatients (14 males, 7 females; mean age 49.2 years, range 39-68; mean disease duration 129 months, range 12-264): among these, eleven fulfilled the modified New York criteria for ankylosing spondylitis and ten were classified as having psoriatic arthritis, characterized predominantly by axial involvement, according to the Moll and Wright criteria modified by Helliwell. All patients showed active disease, as evidenced by BASDAI score (median 6.5, standard deviation ± 2.2). Twenty-one healthy volunteers matched for sex and age were also included. Peripheral blood mononuclear cells (PBMCs) from patients and controls were isolated from heparinized blood by Lymphoprep gradient centrifugation. Immunofluorescence and flow cytometry analysis were used to evaluate the expression of KIR3DL1 on NK and T lymphocytes by sequential gating on forward, side scatter and fluorescence parameters. Flow cytometry was also used to detect NK cell cytotoxicity through a degranulation assay which measures cell-surface expression of the lysosomal protein CD107a (LAMP-1) in PBMCs incubated with either K562, an erythroleukemia cell line, or the mouse mastocytoma cell line P815 pre-incubated with antibodies directed against the NK cell activating receptors CD16 and NKG2D (CD314). The significance of the differences was determined using the nonparametric Mann-Whitney test for unpaired samples and the Wilcoxon's test for matched samples; p values less than 0.05 were deemed statistically significant. Results: A relevant increase in the expression of KIR3DL1 receptor on peripheral blood NK cells was found in SpA patients respect to healthy controls (p=0.024). Among patients, a significant increase in the KIR3DL1 expression on NK cells was found in respect to CD4+ (p=0.0001) and CD8+ T lymphocytes (p=0.0004). When we evaluated the cytotoxicity through the expression of CD107a on NK cells, no difference between patients and controls was found using either K562 or mAb-coated P815 cells as target, even if controls tended to show higher CD107a expression. In SpA patients a negative correlation between degranulation activity and BASDAI was shown (p=0.016, r=-0.639). Conclusion: The increased expression of KIR3DL1 on NK cells from SpA patients respect to controls may be responsible for the dampened cytotoxic activity in our patients. Indeed, the NK cell population was largely activated, as indicated by high level of CD69 and HLA-DR expression. This observation supports the ''intrinsic'' reduced activity of NK cells from SpA patients, rather than the possibility that these cells may have exhausted their classical activity in vivo. References: Spondyloarthropathy: disease at the crossroads of immunity. FitzGerald O et al. Best Pract Res Clin Rheumatol 2006; 20: 949-67.