DCA delays the onset of CIA and at the same time alleviates the progress of CIA. Interestingly, this outcome is observed in female mice only. Female DBA/1 mice treated with DCA had a significant slower onset and less severe arthritis compared with water-treated controls. Importantly, the destructive action of inflammation on bone was almost totally inhibited in mice provided with DCA. The results of this study reveal for the first time that destructive arthritis can be inhibited by DCA administration. Our results also suggest that estrogen plays an important role in the beneficial effect of DCA.
DCA is a well-established drug used for the treatment of lactic acidosis. It also exhibits efficient anti-tumor properties due to its pro-apoptotic and anti-proliferative effects without visibly affecting non-cancerous cells or eliciting systemic toxicity [1
]. RA, like a malignant tumor, is also characterized by increased cell proliferation. We therefore wanted to evaluate whether DCA could prevent the development of arthritis in a model of RA. To this end, we treated CIA in DBA/1 mice with DCA. We found that DCA can ameliorate arthritis potently, but only in female mice.
How does DCA abolish the development of chronic destructive inflammation in CIA? B cells are important in the pathogenesis of RA by producing auto-antibodies and in T-cell activation [27
]. Antibodies to CII have been detected in serum and synovial fluid of patients with RA [28
]. As in RA, anti-CII antibody production contributes to the development of CIA [30
]. The beneficial effect of DCA on arthritis is likely due to the down-regulation of B cells producing anti-CII antibodies because DCA-treated mice displayed significantly lower levels of anti-CII antibodies (Figure ). This may be a direct effect of DCA or mediated via reduction of pro-inflammatory cytokines. IL-6 is a pro-arthritogenic cytokine that affects B cells by promoting plasma cell differentiation, antibody production, and class switch [32
]. A tendency toward lower serum levels of IL-6 was observed in DCA-treated female mice compared with water-treated mice (Figure ), which thus reflects a possible role of IL-6 in DCA-mediated inhibition of anti-CII antibody production. We could not show any effect of DCA on T cell-mediated inflammation (DTH) or granulocyte-mediated inflammation (olive oil-induced), suggesting that the observed beneficial effect is through humoral immunity, as indicated by the effect on anti-CII antibody production.
Because only female DBA/1 mice displayed amelioration of arthritis in response to DCA treatment, female hormones such as estrogen may play a role. A role of estrogens in RA is suggested by the therapeutic effect of estradiol in menopausal women with RA, a group of patients characterized by low levels of estrogens and high incidence of RA [3
]. In addition, estradiol has both prophylactic and therapeutic effects on arthritis development in CIA [37
]. To test the hypothesis, endogenous estrogens were removed by OVX. The OVX group treated with DCA had an earlier onset of disease, more frequent and more severe arthritis, and more synovitis and bone destruction compared with the sham-operated group treated with DCA, which had intact estrogen production. A confounding factor is the fact that the OVX treatment in itself may worsen CIA [36
], which could possibly mask anti-inflammatory effects of DCA. But the net effect of DCA on sham versus OVX mice is of several magnitudes greater than the effect of OVX on arthritis. DCA ameliorates arthritis by decreasing the frequency of arthritis by at least 40% in sham versus OVX mice (Figure , day 38) and the severity by at least 65% (Figure , days 37 and 38), whereas the earlier reported aggravating effect of OVX on arthritis severity is 20% and the effect on arthritis frequency is minimal or non-existent [36
]. This suggests that the observed difference between mice unable to produce estrogens and water controls is indeed due to the DCA treatment. An ameliorating effect of estrogens on arthritis has been demonstrated in both mice and humans [3
]. However, the effect of DCA is probably not via increased production of estrogens as DCA treatment did not cause increased uterus weight. Rather, DCA elicits its effect by affecting estrogen signaling.
The beneficial effect of DCA on CIA may not be entirely dependent on estrogens. This is supported by the fact that DCA-treated mice unable to produce estrogen (OVX mice) had a lower frequency of arthritis (61.3%) than water-treated mice (86.4%) (Figures and , respectively). As only female mice benefited from DCA treatment, this observation indicates that DCA has some estrogen-independent but gender-dependent effects on inflammation. Also, the difference in response to DCA could be due to the fact that male hormones may inhibit the DCA effect.
Previous studies of DCA in inflammation have not focused on the effect on bone density. Our results clearly indicate that DCA can prevent cortical bone mineral loss in female mice in CIA (Figure ) as a result of increased cortical thickness (Figure ). This is in line with a recent study showing that down-regulation of arthritis severity will lead to not only absence of local erosion (that is, in cartilage and subchondral bone) but also systemic effects on BMD [36
]. We believe that the beneficial effect of DCA on bone is mediated by a combination of estrogen-dependent effects and the decrease of the inflammatory response manifested by a reduced level of anti-CII antibodies. First, estrogen is important for bone maintenance and may provide protection from bone destruction in arthritis [3
]. The role of estrogen in the DCA-mediated effect was demonstrated by the fact that female mice, but not male or OVX-treated female mice, benefited from DCA treatment. Second, the lower levels of anti-CII antibodies found in DCA-treated animals may also prevent bone destruction as anti-CII antibodies induce bone erosions in the CIA model [38
]. Such bone erosions may be mediated via anti-CII antibody-dependent C3 recruitment to the cartilage surface, which will initiate an immunological attack, eventually leading to bone destruction [39
]. The DCA-mediated reduction of the pro-inflammatory cytokine IL-6 may also contribute to bone protection as IL-6 has potent effects on cartilage and bone destruction [40
The effect of DCA on cortical bone may also be indirect. As animals have less arthritis, they are more prone to physical activity, which may account for the difference in cortical BMD [41
] between treated and non-treated animals (Figure ). This notion is furthermore in line with the observation that the effect of DCA on bone was mainly on cortical bone, and not on trabecular bone, which is less affected by physical training [42