The results presented in this study demonstrate for the first time that daily administration of exogenous IFN-β starting at the onset of disease in the murine CIA model reduces synovial inflammation and protects against cartilage and bone destruction. The fact that clinical effects, but not histologic changes, were detected at the lowest dosage might be explained by the relative lack of sensitivity to change of semiquantitative histologic analysis. IFN-β treatment also resulted in a reduction in proinflammatory cytokine expression by synovial cells, which could be explained in part by inhibition of NF-κB activity.
Of importance, histologic examination revealed a reduction in the number of osteoclasts in the animals treated with IFN-β, correlating with a reduction in cartilage and bone destruction, suggesting that osteoclastogenesis is inhibited by the presence of IFN-β. Recent studies by Takayanagi and colleagues have described an essential role for IFN-β in the negative regulation of RANKL- and c-Fos-dependent osteoclast differentiation [14
]. In agreement with their predictions, decreases in osteoclastogenesis observed in arthritic mice treated with IFN-β were paralleled by a decrease in the number of RANKL- and c-Fos-positive cells. In addition to effects on osteoclastogenesis, IFN-β might also affect osteoclast activity directly.
Src tyrosine kinase function (and two of its targets, Ras and Cbl) are required for bone resorption by osteoclasts [28
]. It was previously reported that IFN-β has negative effects on tyrosine kinase signalling pathways in HL-60 cells [31
]. Thus, IFN-β treatment may represent a potentially therapeutic strategy in inhibiting bone degradation in arthritis.
Further experiments were carried out to determine the effect of IFN-β on the total numbers of T cells, B cells, macrophages, and granulocytes. We found no clear differences in the total numbers of T cells and B cells in the animals treated with IFN-β in comparison with controls as assessed by immunohistochemistry. Recent studies have proposed that endogenous IFN-β production in RA might theoretically promote inflammation, as IFN-β can promote T-cell and neutrophil survival in vitro
]. While our studies did not specifically address the effects of exogenous IFN-β on T-cell survival in the synovial joint, we did not observe an increase in T-cell numbers, and any potential anti-apoptotic effect did not prevent a beneficial therapeutic effect of IFN-β in vivo
In contrast to the lack of effect on T- and B-cell infiltration, morphological analysis revealed a tendency towards a decreased number of macrophages and a significant reduction in the number of granulocytes in animals treated with IFN-β. Although one study has reported that IFN-β induces apoptosis in a monocytic leukaemia cell line [33
], on the basis of that study, it is unclear whether IFN-β is influencing monocyte recruitment, survival, and/or retention in the synovial joint.
Alternatively, IFN-β may inhibit inflammatory cell infiltration indirectly, via suppression of FLS and/or monocyte activation. Our data demonstrated statistically significant modulation of proinflammatory and anti-inflammatory cytokine production in animals treated with IFN-β. We found a statistically significant reduction of TNF-α and IL-6 expression and an increase in IL-10 production in the animals treated with the highest dose IFN-β. TNF-α plays an important role in the pathogenesis of CIA as well as RA. Treatment with TNF-α blockade has been shown to be effective in both CIA and RA [34
]. The reduced expression of IL-6 could also be beneficial, as IL-6 may inhibit bone formation and induce bone resorption through its stimulatory effects in osteoclasts, and it is known that IL-6 knockout mice do not develop bone erosions. Moreover, it has been suggested that treatment with anti-IL-6 receptor antibody may be effective in RA patients [37
]. Of interest, IL-10 production was increased in the IFN-β-treated animals. IL-10 may be a potent anti-inflammatory cytokine, achieving the effect through suppression of TNF-α, IL-6, and IL-1 production by activated macrophages [38
]. A trend towards clinical improvement has been suggested in RA patients treated with recombinant human IL-10 [39
It has been shown previously that IFN-β has an inhibitory effect on the production of TNF-α by lipopolysaccharide-stimulated macrophages from mouse bone marrow [11
]. In this study, we show for the first time that IFN-β can decrease the production of IL-6, IL-8, and GM-CSF by stimulated FLS from RA and OA patients. Although the molecular signalling mechanism underlying this inhibitory effect requires further elucidation, preliminary evidence presented here suggests that IFN-β acts at least in part via inhibition of NF-κB activity induced by IL-1.
The potential effects of IFN-β were previously investigated in CIA in mice by IFN-β gene therapy [11
]. Fibroblasts from DBA/1 mice were infected with a retrovirus expressing murine IFN-β and were injected intraperitoneally into CIA mice before and after the onset of arthritis, leading to continuous IFN-β delivery. A significant decrease in inflammation was observed after IFN-β gene therapy both before and after the onset of disease. At present, viral and nonviral vectors that are used for gene therapy have limited applications for use in humans. Therefore, our approach using daily injections with murine IFN-β could have the advantage of easily translating results into RA patients. However, it remains to be shown whether the exciting biological effects described in the present study can be achieved in RA patients if IFN-β is administered only three times weekly, in accordance with the regular treatment regimen in patients with multiple sclerosis. A recent pilot study, which was not designed to evaluate clinical efficacy, did not suggest clinical improvement after IFN-β treatment three times weekly [40
]. It is conceivable that more frequent injections as in the present study, higher doses, or the use of compounds with a longer half-life is required to induce clinically meaningful effects in RA patients. Obviously, although animal arthritis models are very useful for screening interesting compounds, the results are not necessary identical to those obtained in RA patients.