Intracellular adenosine released from THP-1 differentiated human macrophages is involved in an autocrine control of Leishmania parasitic burden, mediated by adenosine A2A and A2B receptors

Abstract

Leishmania infected macrophages have conditions to produce adenosine. Despite its known immunosuppressive effects, no studies have yet established whether adenosine alter Leishmania parasitic burden upon macrophage infection. This work aimed at investigating whether endogenous adenosine exerts an autocrine modulation of macrophage response towards Leishmania infection, identifying its origin and potential pharmacological targets for visceral leishmaniasis (VL), using THP-1 differentiated macrophages. Adenosine deaminase treatment of infected THP-1 cells reduced the parasitic burden (29.1 ± 2.2%, P < 0.05). Adenosine A_2A and A_2B receptor subtypes expression was confirmed by RT-qPCR and by immunocytochemistry and their blockade with selective adenosine A_2A and A_2B antagonists reduced the parasitic burden [14.5 ± 3.1% (P < 0.05) and 12.3 ± 3.1% (P < 0.05), respectively; and 24.9 ± 2.8% (P < 0.05), by the combination of the two antagonists)], suggesting that adenosine A₂ receptors are tonically activated in infected THP-1 differentiated macrophages. The tonic activation of adenosine A₂ receptors was dependent on the release of intracellular adenosine through equilibrative nucleoside transporters (ENT1/ENT2): NBTI or dipyridamole reduced (~25%) whereas, when ENTs were blocked, adenosine A₂ receptor antagonists failed to reduce and A₂ agonists increase parasitic burden. Effects of adenosine A₂ receptors antagonists and ENT1/2 inhibitor were prevented by L-NAME, indicating that nitric oxide production inhibition prevents adenosine from increasing parasitic burden. Results suggest that intracellular adenosine, released through ENTs, elicits an autocrine increase in parasitic burden in THP-1 macrophages, through adenosine A₂ receptors activation. These observations open the possibility to use well-established ENT inhibitors or adenosine A₂ receptor antagonists as new therapeutic approaches in VL.

Keywords: Adenosine; Immunoregulation; Leishmania infantum; Macrophage; Visceral leishmaniasis.