CD14 bright CD16+ intermediate monocytes are induced by interleukin-10 and positively correlate with disease activity in rheumatoid arthritis

In this study, we demonstrate that circulating CD14^brightCD16+ monocytes are increased in patients with RA in the active phase and decrease after MTX treatment in a manner that correlates with decreasing disease activity. Moreover, this monocyte subset is associated with expression of inflammatory cytokines in peripheral blood, and the cytokine IL-10, which is increased in patients with RA, induces CD16 expression on monocytes. These results suggest that CD14^brightCD16+ monocytes play a role in the pathogenesis of RA, and that IL-10 is a key cytokine in the regulation of CD16 expression.

Although an increase in CD14^brightCD16+ monocytes in patients with RA has been reported [14], the possibility of correlation between CD14^brightCD16+ monocytes and cytokines in untreated patients with active RA has not yet been investigated. Previous studies have not ruled out any influence of treatment on cytokine and disease activity, because the patients with RA in these studies were not treatment-naïve. Our results support previous observations [14] and show that the proportion of CD14^brightCD16+ monocytes is increased in treatment-naïve patients with active RA compared to healthy controls, and is positively correlated with disease activity in these patients.

We further showed that the CD14^brightCD16- population was negatively correlated with RA disease activity. Notably, the proportion of CD14^brightCD16+ monocytes decreased and that of CD14^brightCD16- monocytes increased when the patients received MTX treatment.

Though the function of CD14^brightCD16+ monocytes in RA is not still clarified, this population may be involved in the pathogenesis of RA in accordance with our findings that CD14^brightCD16+ monocytes decreased after MTX treatment. It has been shown that MTX inhibits inflammatory cytokine production [19] and cell proliferation in vitro, and to induce apoptosis of immune cells [20], but it has not been found to inhibit a specific subset of monocytes. One of the reasons for the decreased proportion of CD14^brightCD16+ monocytes after MTX treatment was probably secondary to a phenomenon derived from the improvement in RA disease activity.

It is reported that M-CSF plays an important role in the introduction of CD14^brightCD16+ monocytes. Anti-M-CSF antibody caused a decrease in circulating CD14^brightCD16+ and CD14^dimCD16+ monocytes in a clinical trial in two patients with active RA [17]. Moreover, M-CSF and IFN-? therapy has been found to induce CD16 expression on circulating monocytes in patients with cancer or lymphoma [21]. CD16 expression has been shown to be induced in monocytes with culture of whole PBMCs with M-CSF or IL-10 on CD14^brightCD16- monocytes in vitro [12]. Although this report may indicate a possible direct role for cytokines, it did not exclude interactions with other cell subsets among the PBMCs. In our study, using highly purified CD14^brightCD16- monocytes, we clearly showed that IL-10, but not M-CSF, directly induces CD16 expression in CD14^brightCD16- monocytes. In addition, we proved that the enhancement of CD16 expression on CD14^brightCD16- monocytes required the interaction with IL-10 and IL-10 receptor by a neutralization assay with anti-IL-10 receptor antibody.

It is well-known that IL-10 plays a crucial role, such as anti-inflammatory and/or pro-inflammatory roles in the pathogenesis of RA. IL-10 has been shown to inhibit production of IL-6, TNF-?, and GM-CSF from immune cells [22], and to enhance B cell differentiation to cells secreting IgG, IgM, and IgA [23, 24], resulting in increased RF and IgG-RF production by B cells in peripheral blood. Moreover, IL-10 is localized to the synovial membrane lining layer, the site of monocyte migration, and inhibits pro-inflammatory cytokines in RA [25]. In this study, we demonstrated that serum IL-10 in patients with RA was significantly elevated compared with healthy volunteers, and was correlated with disease activity.

IL-10 is secreted by many kinds of cells such as T-cells, B-cells, macrophages, dendritic cells, natural killer cells, and monocytes themselves [26–28]. It is reported that CD16 expression on monocytes is maintained by IL-10 production by human naïve CD4+ T cells [29]. The function of CD14^brightCD16+ monocytes may be regulated by these cells producing IL-10. IL-10 tended to decrease in patients with RA with decreasing CD14^brightCD16+ monocytes after 12 weeks of treatment. IL-10 may play a role in the induction of CD16 on monocytes in patients with RA.

We note two limitations to our study. First, the number of patients was relatively small, albeit large enough to provide statistically significant data. Second, we did not show that CD14^brightCD16+ monocytes are directly associated with inflammatory cytokines in RA in vivo. Production of IL-6 and TNF-? in CD14^brightCD16+ monocytes was not higher than that in CD14^brightCD16-monocytes (Additional file 1). We thought that CD14^brightCD16+ monocytes could exert both inflammatory and anti-inflammatory effects, and which effect’s dominance would depend on cells producing IL-10. The functions of these monocytes in RA will need to be clarified in future studies.