It is established that the common myelomonocyte progenitor cell can differentiate into CD11b+CD11c+ mDC or CD11b+CD11c− OCP depending on the cytokine milieu. Previous research demonstrated that elevated levels of IFN-α influence myelomonocyte development into mDC, and that mDC are a major cellular mediator of SLE pathology (13
). It has also been shown that inflammatory erosive arthritis can be marked by an elevation in the CD11b+CD11c− population. While it is known that JA is typically non-erosive, and that IFN-α is considered the dysregulated cytokine in SLE, knowledge of a potential link between these two factors and myelopoiesis is scarce. Using the SIA model in conjunction with the NZBxNZW F1 model, we were able to add to the existing literature on SLE, myelopoiesis, and bone, by studying the effect of SLE disease and the effect of IFN-α in particular on the development of bone erosion in inflammatory arthritis.
The current etiologic model for the role of dysregulated IFN-α in SLE suggests that pathologically elevated levels of IFN-α are released from pDC after stimulation by an unknown factor (13
). Though early evidence of this came from detection of elevated serum levels of IFN-α, ELISA for IFN-α has been reported not to be very sensitive or reliable for measuring naturally elevated IFN-α levels because the range of detection antibodies would need to account for the approximately 14 known IFN-α isoforms. A more widely accepted method of determining IFN-α levels in SLE is to measure the levels of IFN-α-inducible genes since all IFN-α isoforms bind the same receptor and induce the same signaling cascade leading to gene expression (34
). Though many IFN-α-inducible genes have been identified as correlating with SLE pathophysiology, the Ifi202
gene has been best characterized as a susceptibility gene in the NZBxNZW F1 model we used in this study (24
). In NZB mice, natural levels of serum IFN-α were undetectable by ELISA, and CpG ODN injection was needed to see significant serum IFN-α levels detectable by ELISA compared to Balb/c controls (35
). Despite this, NZB mice have elevated expression of Ifi202
compared to Balb/c and C57Bl/6 mice (24
). The NZBxNZW F1 mice in our study exhibited elevated levels of Ifi202
mRNA expression compared to NZW controls at all ages. This is in accordance with previous literature that showed this gene not to be elevated in NZW mice (24
). Interestingly, the Ifi202
gene expression level was elevated even at 2 months of age though anti-dsDNA antibodies and proteinuria were not significantly different at this age from that seen in NZW mice. This may be explained by a gradual skewing of the monocyte differentiation program toward mDC with the age-related accumulation of IFN-α, and increasing numbers of circulating mDC presenting autoantigens, thereby breaking self-tolerance over time. Additionally, variables such as estrogen status and environmental factors likely contribute to the timing of autoantibody formation which precedes disease onset. Nevertheless, in these pre-autoimmune mice, induction of systemically elevated IFN-α via injection of Ad-IFN-α resulted in SLE disease features suggesting that innate barriers to a break in tolerance can be overcome if these mice exceed threshold levels of IFN-α.
As seen in human disease, the presence of SLE protected NZBxNZW F1 mice from developing bone erosions in the setting of inflammatory arthritis. When these mice were at an age where they did not exhibit SLE disease features (anti-dsDNA autoantibodies or proteinuria), they displayed a similar susceptible to SIA bone erosions as the non-SLE NZW mice were at all ages. To explain this, we examined osteoclastogenesis in these animals. We observed that NZBxNZW F1 mice had lower numbers of OC in vivo at the site of resorption compared to NZW mice. This could be the result of decreased circulating OCP, and/or expression of local factors at the site of resorption that inhibit osteoclastogenesis in mice with SLE-like disease. To address this, we also examined the ex vivo osteoclastogenic potential of circulating OCP from NZBxNZW F1 mice. Our results show that although SIA is capable of inducing osteoclastogenesis in both NZW and NZBxNZW F1 strains, it is more effective in mice without SLE. These data demonstrate that the reduced degree of bone erosion seen in inflammatory arthritis in NZBxNZW F1 mice is the result of a more global anti-osteoclastogenic process and not solely a local effect. Furthermore, our finding that NZBxNZW F1 mice with SLE-like disease had a greater percentage of circulating mDCP compared to the percentage of circulating OCP supports the idea that the global deficit in osteoclastogenesis seen in these mice is from a skewing of myelomonocytic differentiation.
The osteoclastogenesis, bone erosion, and mDCP frequency findings in the NZBxNZW F1 mice could be explained by events associated with SLE, or could represent another direct effect of IFN-α in SLE pathophysiology. The injection of Ad-IFN-α induced Ifi202 gene expression and the development of SLE disease markers in the NZW mice whereas Ad-Null and uninjected mice did not show these findings. Our results show that the presence of elevated IFN-α levels was able to mitigate the erosive nature of inflammatory arthritis by globally reducing the number of OC. These findings were explained by an elevation in the relative proportion of mDCP to OCP in the blood. Thus, the artificial induction of elevated systemic IFN-α is capable of replicating the findings we saw in NZBxNZW F1 mice with naturally high levels of IFN-α-inducible Ifi202 and SLE disease markers. This supports a direct effect of IFN-α on OCP frequency and bone erosion in inflammatory arthritis rather than general events related to SLE.
It is important to note that most of the research on interactions between SLE disease processes and bone pathology has focused on steroid-induced osteoporosis and avascular necrosis of bone, which are prominent side-effects of effective SLE therapy (20
). The mechanisms responsible have largely been attributed to glucocorticoid activation of osteoclastogenesis, direct inhibition of osteoblasts, and prolonged suppression of osteogenesis (36
). Since glucocorticoid therapy reverses the elevation in the IFN-α transcriptome (18
), it consequently mitigates the associated protection against bone erosions we have described here. Thus, the studies on steroid-induced bone loss in SLE patients do not conflict with our findings. However, steroid-independent osteopenia is also known to occur in SLE patients, which is somewhat inconsistent with our mouse models. Thus, the complexities of this disease may have dual inhibitor effects on both osteoclasts that results in decrease erosions, and osteoblasts that results in osteopenia, of which the later warrants further investigation.
Taken together, our findings support a new mechanism for the non-erosive nature of inflammatory JA in SLE. We have shown that a naturally elevated or experimentally induced IFN-α transcriptome directly correlates with protection from bone erosions in inflammatory arthritis as a result of biased myelopoiesis toward mDC and away from OC in a mutually exclusive manner. Further research to uncover how IFN-α influences the molecular interactions that promote this skewed differentiation program will help in identifying therapeutic targets for RA and other inflammatory erosive arthritides.