The results of the present study indicate that resolution of joint swelling in ZIA was delayed in leptin deficient mice. Accordingly, the acute phase response, as assessed by circulating levels of IL-6 and SAA, remained elevated for a longer period of time in ob/ob mice than in control, lean littermates. Furthermore, at late time points histological features of arthritis tended to be more severe in ob/ob mice. Similar results were obtained in db/db mice, suggesting that the observed changes in the course of ZIA were mediated by a lack of interaction of leptin with OB-Rb.
In contrast to these findings we previously observed a milder form of AIA in ob
mice as compared with their controls, with decreased synovial levels of IL-1β and tumour necrosis factor (TNF)-α, and a switch toward production of Th2 cytokines [14
]. These contrasting observations in AIA and ZIA further suggest a greater sensitivity to agents stimulating the innate immune responses in leptin or leptin signalling deficient animals, as opposed to attenuated inflammation in models involving T-cell responses, in particular Th1-mediated diseases [4
]. Indeed, both T and B lymphocytes participate in the mechanisms that lead to articular inflammation in AIA, whereas ZIA exclusively involves the innate immune response.
Leptin was previously reported to play an important role in T-cell-mediated immune responses. Evidence of defective cell-mediated immunity and lymphoid atrophy, analogous to those observed in chronic under-nutrition in humans, are detected in ob
]. Leptin stimulates the proliferation of CD4+
T cells and promotes Th1 responses [6
]. Congenital leptin deficiency in humans is associated with a decreased number of circulating CD4+
T cells, impaired T-cell proliferation and cytokine release, all of which could be reversed by the administration of recombinant leptin [26
]. In addition, the OB-Rb receptor is also expressed on B cells and may participate in the development of humoral responses [14
]. Consistent with these findings, leptin deficient mice are protected from inflammation mediated by T and B cells in different disease models, including AIA, experimental autoimmune encephalomyelitis, type 1 diabetes and experimental colitis [11
Our results in ZIA suggest that chronic leptin deficiency interferes with adequate control of the inflammatory reaction. Protective effects of leptin were previously observed in studies of other experimental models conducted to explore innate immune responses. Ob/ob
mice are significantly more susceptible to lipopolysaccharide (LPS)-induced death, and this feature can partly be reversed by administration of leptin [28
]. OB receptor deficient fa/fa
rats also exhibit enhanced LPS-induced hepatotoxicity [29
]. Similarly, ob
mice are more likely to succumb after administration of TNF-α. The protective role of leptin against TNF-α induced toxicity was further supported by the deleterious effect of neutralizing anti-leptin antibodies administered to TNF-α injected mice [30
]. The mechanisms underlying these protective effects of leptin are still unclear. Although thymic and circulating lymphocytes are reduced, a fourfold increase in the number of circulating monocytes was observed in leptin deficient mice, suggesting enhanced responses to monocyte activators [7
]. Furthermore, an imbalance between proinflammatory and anti-inflammatory monokines has been observed in ob
mice injected with LPS, with plasma levels of the anti-inflammatory cytokines IL-10 and IL-1 receptor antagonist being lower in leptin deficient than in normal mice [28
]. However, LPS or TNF-α mediated systemic inflammation is a complex syndrome, and susceptibility to these systemic stimuli might also be influenced by the effects of leptin on nervous, endocrine, or other responses, independent of the production of inflammatory mediators.
Intravenous injection of Staphylococcus aureus
results in a severe form of septic arthritis in mice, which is associated with decreased circulating levels of leptin. In this model, treatment with leptin significantly decreased the severity of septic arthritis without interfering with staphylococcal load in the joints [31
]. The levels of IL-6 were significantly lower in mice with septic arthritis after administration of leptin [31
]. Consistent with these findings, our results in ZIA indicate that IL-6 levels were higher in ob
mice than in controls. IL-6 plays an important role in turning acute inflammation into a chronic synovitis, as demonstrated by limited duration of ZIA in IL-6 deficient mice [32
]. In addition, IL-6 has also been shown to play a major role in other models of arthritis [33
]. Thus, control of IL-6 production may be one of the mechanisms by which leptin is involved in the control of the inflammatory response during ZIA.
It is noteworthy that anaesthesia, skin cut and intra-articular injection with saline slightly enhanced serum levels of IL-6 and SAA in ob/ob and +/? mice, although to a lesser extent than in animals infected with zymosan A. Interestingly, this small, zymosan A independent inflammatory response was also greater in ob/ob mice than in lean controls, further supporting the presence of an inappropriate control of inflammatory responses in leptin deficiency.
Corticosterone secretion is known to be elevated in all forms of leptin deficiency and in leptin insensitivity [21
]. However, despite the presence of elevated levels of glucocorticoids, ob
mice still exhibited a more pronounced acute phase response and longer lasting arthritis than did controls. Thus, it is conceivable that leptin deficiency could result in an even more severe form of arthritis in the absence of hypercorticosteronaemia.
The mRNA levels of different cytokines were determined in arthritic and control joints at two time points. Consistent with a previous report [17
], the levels of IL-1α, IL-1β and IL-6 mRNA were increased during ZIA. In addition, we also detected elevated levels of IL-1 receptor antagonist and MIF, which to the best of our knowledge have not previously been reported in the joint during ZIA. MIF is a broad-spectrum proinflammatory cytokine that is implicated both in animal models of immune-mediated arthritis and in human rheumatoid arthritis [35
]. The levels of mRNAs encoding these different cytokines, including IL-6, were not different between ob
mice and their lean controls. However, we cannot exclude variations in post-transcriptional regulation, which might still result in different levels of active proteins at the site of inflammation.