Previous studies have demonstrated significant changes in many proinflammatory cytokines, especially IL-1, IL-6, TNF-α, and IFNs, in SLE patients [28, 29]. However, serum levels of most of the proinflammatory cytokines measured here were not associated with disease activity indices such as, SLEDAI and CLASI, or with serum immunological biomarkers, most likely because our study only included SLE patients with LDA.
We detected decreases in serum levels of TNF-α, IL-6, VEGF-A, IL-1ra, IL-2, MIP-1α, IL-8, IL-1β, S100A8, and S100A9 after add-on HCQ treatment. Our results differ from those of Monzavi et al., who reported no change in serum IL-8 levels after treatment of newly diagnosed SLE patients with HCQ [30]. This difference, together with the observed lack of response of other factors, such as MCP-1, to HCQ treatment, is likely to be due to the inclusion of only patients with LDA. Plasma and urinary IL-8 levels are reportedly associated with LN activity [8, 31–33], and serum IL-6, IL-8 and IL-18 levels have been proposed to be useful predictors of relapse in SLE [34]. We found that HCQ had a greater effect on serum IL-8 levels in patients with a history of renal involvement compared with patients with no history, suggesting that HCQ may help improve the prognosis and prevent relapse of LN. Of note, it is possible that even renal lesions considered to be in remission may express elevated levels of proinflammatory cytokines.
Despite its common use, the mechanism of action of HCQ in autoimmune diseases is unclear. HCQ is a weak base and is known to raise the pH of acidic intracellular vesicles and interfere with their physiological functions, including autophagy and antigen processing [35]. In addition, HCQ interferes with intracellular signaling, which may suppress the response to engagement of the innate Toll-like receptors (TLRs), thereby inhibiting the production and release of cytokines and promoting apoptosis in lymphocytes and endothelial cells [36, 37]. Thus, both acidification of endosomal vesicles and increased lymphocyte apoptosis following HCQ treatment may contribute to the decreased production of proinflammatory cytokines. In addition, HCQ-mediated inhibition of TLR activation suppresses the activity of plasmacytoid dendritic cells and autoreactive B cells in SLE patients [37], leading to a reduction in inflammation. Indeed, Sacre et al. demonstrated that HCQ treatment of SLE patients reduced the ability of plasmacytoid dendritic cells to produce IFN-α and TNF-α in response to TLR-9 and TLR-7 stimulation in vivo [38]. We believe that the mechanism of cytokine reduction by HCQ in the present study may be mediated by these effects.
We previously reported that HCQ modulates serum levels of S100A8 and S100A9 in SLE patients with LDA [21]. S100 proteins are components of neutrophil extracellular traps (NETs), which play an important role in the pathogenesis of SLE [39] [40]. In addition, chloroquine and HCQ have been reported to inhibit NETs in vivo and in vitro [41, 42], which may suggest a mechanism for the regulation of S100 proteins. S100A8 and S100A9 proteins upregulate the expression of proinflammatory cytokines such as IL-6 and IL-8 [43, 44]. Therefore, HCQ-mediated modulation of S100 proteins may also be involved in the suppression of proinflammatory cytokine expression in SLE patients.
There are several limitations to this study. First, we did not monitor HCQ adherence by measuring blood HCQ levels. Second, whether the change in proinflammatory cytokine levels were a direct result of add-on HCQ treatment is difficult to determine unequivocally because some patients were receiving other immunosuppressants. Third, the sample size was small due to the strict inclusion and exclusion criteria. Finally, serum IFN-α levels were un detectable in most patients. Nevertheless, our study has merit because it is the first to demonstrate the effect of add-on HCQ treatment on proinflammatory cytokines in SLE patients with LDA.