Adenosine abolishes MTX-induced suppression of osteoclastogenesis and inflammatory bone destruction in adjuvant-induced arthritis

Abstract

Methotrexate (MTX) is widely utilized for the treatment of patients with rheumatoid arthritis (RA); however, recent observation of the MTX-resistant patients proposed some difficulty in MTX-dependent therapeutic approach for RA. To access cellular events related to MTX resistance in RA in respect to inflammatory bone destruction, we investigated on an involvement of the potent inflammatory mediator adenosine in the regulation of osteoclastogenesis and inflammatory bone destruction. In rats with adjuvant-induced arthritis (AA rats), MTX efficiently suppressed bone destruction when it was administrated within 3 days after adjuvant injection, while it could not suppress inflammatory bone destruction if MTX was injected at the time of onset of inflammation (at day 10 after adjuvant injection). Time-course change in the level of plasma adenosine of AA rats was estimated by use of high-performance liquid chromatography and elucidated that adenosine level was markedly elevated till 10 days after adjuvant injection. In vitro bone marrow culture system for evaluating osteoclastogenesis, MTX markedly suppressed osteoclastogenesis in a stromal cell-dependent manner. This MTX-induced suppression of osteoclastogenesis was abrogated by the addition of adenosine. MTX suppressed the expression of mRNA for the receptor activator NF-kB ligand (RANKL), but it did not suppress the expression of osteoprotegerin (OPG). The addition of MTX and adenosine together markedly suppressed the level of OPG expression. Abolishment of MTX action by adenosine was significantly blocked by MRS1754, a highly selective antagonist for the A 2b adenosine receptor (A 2b AR), but not by caffeine, an antagonist for A 1 , A 2a , A 3 AR (A 1 AR, A 2a AR, and A 3 AR), which suggests that adenosine acts through A 2b AR. Immunohistochemical studies showed abundant expression of A 2b AR in cells localized in the bone-bone marrow boundary of the distal tibia in AA rats but not in control rats. When adenosine was injected in the ankle joints of MTX-treated AA rats, the suppressive effects of MTX on bone destruction was abolished. The current data therefore suggest that upregulation of adenosine production abolished the suppressive effect of MTX on osteoclastic bone destruction. Involvement of the adenosine-A 2b AR system may explain MTX resistance in RA.

Adenosine A1 receptor regulates osteoclast formation by altering TRAF6/TAK1 signaling

“…Moreover, we have observed dose-dependent inhibition of RANKL-induced osteoclast formation and expression of osteoclast markers by the A 2b R specific agonist BAY 60-6,583 in murine BMMs (supplemental Figure 1). These observations contrast, at least at a superficial level, with those of Teramachi et al [49] who reported that adenosine abolished MTX-induced osteoclast inhibition via stimulation of A 2b receptors. However, the effects described by Teramachi and colleagues are most likely mediated by A 2b receptors on osteoblasts since, as noted in their work, the primary reason for inhibition of osteoclast formation in these bone marrow cultures was inhibition of RANKL expression without diminution of osteoprotegerin expression.…”

Adenosine and adenosine receptors in the pathogenesis and treatment of rheumatic diseases

“…As noted above, A 2b promotes a phenotypical switch from M1 to M2 macrophages, 75-77 but also stimulates the differentiation of dendritic cells, which can promote type 17 T helper cells (T H 17) immune responses via production of IL-6 87-89 . Indeed, in in vivo studies high concentrations of adenosine exacerbated arthritis and tissue destruction in rats via an A 2b mechanism 90 .…”

Regulation of bone and cartilage by adenosine signaling

“…For example, while some have found the application of an A2BR-specific agonist, BAY60-6583 in murine bone marrow macrophages inhibits RANKL-induced osteoclast formation in a dose-dependent manner [53], this contrasts with the findings of Termachi et al who reported that adenosine blocked the MTX-induced osteoclast inhibition via A2BR activation [63]. These effects may be secondary to involvement of the A2BR receptor on osteoblasts, which play a crucial role in stimulating osteoclast differentiation via RANKL.…”