Macrophages are key elements in the inflammatory process, whereas depending on the micro-environmental stimulation they exhibit a pro-inflammatory (classical/M1) or an anti-inflammatory/reparatory (alternative/M2) phenotype. Extracellular
ATP can act as a danger signal whereas
adenosine generally serves as a negative feedback mechanism to limit
inflammation. The local increase in
nucleotides communication is controlled by ectonucleotidases, such as members of the ectonucleoside
triphosphate diphosphohydrolase (E-NTPDase) family and
ecto-5'-nucleotidase/CD73 (ecto-5'-NT). In the present work we evaluated the presence of these
enzymes in resident mice M1 (macrophages stimulated with LPS), and M2 (macrophages stimulated with IL-4) macrophages. Macrophages were collected by a lavage of the mice (6-8 weeks) peritoneal cavity and treated for 24 h with
IL-4 (10 ng/mL) or LPS (10 ng/mL).
Nitrite concentrations were measured using the Greiss reaction. Supernatants were harvested to determine
cytokines and the
ATPase,
ADPase and
AMPase activities were determined by the
malachite green method and HPLC analysis. The expression of selected
surface proteins was evaluated by flow cytometry. The results reveal that M1 macrophages presented a decreased
ATP and
AMP hydrolysis in agreement with a decrease in
NTPDase1, -3 and
ecto-5'-nucleotidase expression compared to M2. In contrast, M2 macrophages showed a higher
ATP and
AMP hydrolysis and increased
NTPDase1, -3 and
ecto-5'-nucleotidase expression compared to M1 macrophages. Therefore, macrophages of the M1 phenotype lead to an accumulation of
ATP while macrophages of the M2 phenotype may rapidly convert
ATP to
adenosine. The results also showed that P1 and P2 purinoreceptors present the same
mRNA profile in both phenotypes. In addition, M2 macrophages, which have a higher
ATPase activity, were less sensitive to cell death. In conclusion, these changes in ectoenzyme activities might allow macrophages to adjust the outcome of the extracellular purinergic cascade in order to fine-tune their functions during the inflammatory set.