Expression of the myeloid inhibitory receptor CLEC12A correlates with disease activity and cytokines in early rheumatoid arthritis

Abstract

The myeloid inhibitory receptor CLEC12A negatively regulates inflammation. Reduced CLEC12A expression enhances inflammation in CLEC12A knock-out mice with collagen antibody-induced arthritis. Moreover, CLEC12A internalisation augments human neutrophil activation. We thus postulated that CLEC12A expression on circulating myeloid cells of rheumatoid arthritis patients is associated with disease manifestations. Cell-surface, CLEC12A receptor expression was determined on circulating neutrophils and monocytes of eRA patients and of healthy donors. Generalized estimating equations model, Student's t-test and Spearman's correlations were performed to compare CLEC12A expression between groups and test its association with disease activity and clinical parameters. Plasma cytokines were measured by multiplex immunoassay. Patients with reduced neutrophil or monocyte CLEC12A expression at baseline and at 3 months have an increased simple disease activity index. Low baseline CLEC12A expression also correlates with a higher SDAI at 6 months. In contrast, positive correlations were observed between baseline CLEC12A expression and several cytokines. Moreover, neutrophil and monocyte CLEC12A expression is significantly higher in early rheumatoid arthritis patients at baseline than healthy controls. Circulating neutrophil and monocyte CLEC12A expression correlates with disease activity at baseline and is predictive of SDAI at later stages of the disease indicative of a regulatory role for CLEC12A in RA.

Figure 1

Neutrophil CLEC12A expression in eRA patients correlates with SDAI. Cell-surface, CLEC12A expression was determined by flow cytometry on circulating neutrophils of eRA patients. Cross-sectional correlations between neutrophil CLEC12A expression and SDAI score in eRA patients at (A) baseline, and (B) three months. Correlations between baseline, neutrophil CLEC12A expression and SDAI at (C) three and (D) six months. Statistical analysis: Spearman’s rank correlation coefficient and p value are shown in each panel. Symbols represent individual patients.

Figure 2

Monocyte CLEC12A expression in eRA patients correlates with SDAI. Cell-surface, CLEC12A expression was determined by flow cytometry on peripheral monocytes. Cross-sectional correlations between monocyte CLEC12A expression in eRA patients and SDAI score at (A) baseline, and (B) three months. Correlations between baseline, monocyte CLEC12A expression and SDAI at (C) three months and (D) six months. Statistical analysis: Spearman’s rank correlation coefficient and p value are shown in each panel. Symbols represent individual patients.

Figure 3

Neutrophil and monocyte CLEC12A expression diminishes with treatment in eRA patients. Cell-surface, CLEC12A expression was determined by flow cytometry on peripheral neutrophils and monocytes and analyzed with GEE models and Bonferroni correction. (A) Analysis of neutrophil CLEC12A expression revealed significant differences between groups (p = 0.017). Pairwise significant difference p values are 0.0007 for baseline (BL) and 18 months, 0.018 for 3 and 12 months and < 0.0001 for 3 and 18 months. (B) Analysis of monocyte CLEC12A expression did not reveal significant differences between groups (p = 0.081). Symbols represent individual patients at baseline (BL) and after 3, 6, 12 and 18 months of treatment. Data show the mean ± SEM. *** < 0.001 ** < 0.01 * < 0.05.

Figure 4

CLEC12A is differentially expressed on eRA neutrophils and monocytes compared to healthy donors. CLEC12A expression was measured by flow cytometry on peripheral neutrophils and monocytes of eRA patients and healthy donors. (A) Comparison of neutrophil CLEC12A expression of healthy donors and eRA patients at baseline (Student’s t-test, p = 0.014). (B) Comparison of monocyte CLEC12A expression of healthy donors and eRA patients at baseline (Student’s t test, p = 0.026). Symbols represent individual subjects. Data show the mean ± SEM. *** < 0.001 ** < 0.01 * < 0.05.