The antiinflammatory mechanism of methotrexate depends on extracellular conversion of adenine nucleotides to adenosine by ecto‐5′‐nucleotidase: Findings in a study of ecto‐5′‐nucleotidase gene–deficient mice

Abstract

Objective. Evidence from in vitro, in vivo, and clinical studies indicates that adenosine mediates, at least in part, the antiinflammatory effects of methotrexate (MTX), although the biochemical events involved have not been fully elucidated. This study was undertaken to investigate whether MTX exerts antiinflammatory effects in mice that lack ecto-5-nucleotidase (ecto-5-NT) (CD73) and are unable to convert AMP to adenosine extracellularly, in order to determine whether adenosine is generated intracellularly and transported into the extracellular space or is generated from the extracellular dephosphorylation of AMP to adenosine.Methods. Male CD73 gene-deficient mice and age-matched wild-type mice received intraperitoneal injections of saline or MTX (1 mg/kg/week) for 5 weeks. Air pouches were induced on the back by subcutaneous injection of air; 6 days later, inflammation was induced by injection of carrageenan.Results. Fewer leukocytes, but higher levels of tumor necrosis factor ␣ (TNF␣), accumulated in the air pouches of vehicle-treated CD73-deficient mice compared with those of wild-type mice. As expected, MTX treatment reduced the number of leukocytes and TNF␣ levels in the exudates and increased exudate adenosine concentrations in wild-type mice. In contrast, MTX did not reduce exudate leukocyte counts or TNF␣ levels or increase exudate adenosine levels in CD73-deficient mice.Conclusion. These results demonstrate that the antiinflammatory actions of MTX are mediated, at least in part, by increased release of adenine nucleotides that are hydrolyzed extracellularly to adenosine via an ecto-5-NT-dependent pathway.Low-dose weekly methotrexate (MTX) is a mainstay in the treatment of rheumatoid arthritis and other inflammatory diseases, with a relatively safe profile compared with other therapies (1).

Low expression of CD39 on regulatory T cells as a biomarker for resistance to methotrexate therapy in rheumatoid arthritis

“…Blockage of adenosine receptors reduces the anti-inflammatory effect of MTX in humans and in a murine model of arthritis (8,9). Furthermore, MTX failed to inhibit carrageenan-induced inflammation in CD73-deficient mice (10).…”

The effect of low-dose methotrexate on autologous fat graft survival

“…It also promotes extracellular adenosine accumulation at the inflammatory site, which inhibits some cytokines, superoxide, nitric oxide, and TNF-α (14,15). Adenosine is a cytoprotective molecule due to its inhibitor effect on toxic oxygen metabolites created from the adhesion of stimulated neutrophils to the endothelium (11,16). Decreasing the inflammation and apoptosis at the fat graft area is a recently investigated method to increase long- term fat graft survival (5).…”

Effects of low-dose methotrexate in spinal cord injury in rats

“…[5] Additionally, adenosine has cytoprotective effects resulting from inhibition of the toxic oxygen metabolites that are generated from the adhesion of stimulated neutrophils to the endothelium. [3,6] Based on all these results, this preliminary study was designed to investigate the possible neuroprotective effects of low-dose MTX in the secondary damage of SCI in rats.…”