Adenosine regulates cytotoxic CD4 T lymphocytes in Trypanosoma cruzi infection

BERGERO, GASTÓN; MAZZOCO YANINA; EBERHARDT, NATALIA; AOKI, MARIA PILAR

Congreso; 54th Annual Meeting; 2021

Chagas disease, a major public health problem in Latin America, is caused by the infection with the protozoan parasite Trypanosoma (T.) cruzi and has been spread to non-endemic regions by migratory movements. Chagas cardiomyopathy represents the most severe complication of the infection. Parasite persistence within the cardiac tissue is central to the etiology of the cardiomyopathy. The immune regulatory pathways that allow the chronic parasite persistence for such a long time remain mostly undefined. In recent years, the purinergic system has taken a robust significance as a central signaling pathway driving the outcome of the immune response. Expression of the CD39 and CD73 ectonucleotidases mediate the catabolism of extracellular ATP, which promotes pro-inflammatory responses, towards the anti-inflammatory mediator adenosine (ADO). We have reported that CD73-generated ADO dampens cardiac microbicidal responses and increases local T. cruzi load in acutely infected mice. Recently, a new CD4 T cell population with cytotoxic effector functions (CD4 CTL) has been reported. At steady state, CD4 CTL only resides in the intestine, whereas under inflammatory conditions, including viral infections, autoimmune disorders, and in response to tumor antigens, it expands in the peripheral blood and tissues. Their differentiation and participation in different adaptive immune responses, both beneficial and pathogenic, have not been completely understood yet. In the present work, we evaluated the role of ADO on the differentiation and functional capacity of the CD4 CTL population in T. cruzi infected mice.To this aim, we explored the effect of CD73 activity abrogation (CD73KO) on immune cells infiltrating target tissues during the acute phase of T. cruzi infection. We found that CD73KO cardiac tissue presented a higher frequency of CD4 T cells expressing the degranulation marker CD107a at 21 days post-infection (dpi), compared to the wild-type C57BL/6 (WT) counterpart (p<0.05). Furthermore, CD4 CTL (CD3+ CD4+ Granzyme B+ Perforin+) spleen population was significantly increased in CD73KO mice in comparison with WT mice at 14 dpi (p<0.005). The functional profile shows a higher frequency of CD4 T cells co-expressing two (TNF-α+ Perforin+ or IFN-γ+ TNF-α+) and three effector molecules (TNF-α+ Perforin+ Granzyme B+ or IFN-γ+ TNF-α+ Granzyme B+) and increased percentages of CD107a+ CD4 T cells in CD73KO cells compared to WT cells (CD73KO vs. WT: p <0.05). The frequency of CD107a+ CD4 T cells and CD4 T cells producing IFN-γ increased in CD73KO splenocytes culture compared to the WT counterpart (CD73KO vs. WT: p <0.0005 and p <0.005 respectively). Moreover, both populations from WT spleens had increased frequency after re-stimulation with T. cruzi lysate (WT Lysate vs. medium: p <0.05), while no differences were found in CD73KO spleen cells subjected or not to re-stimulation. On the other hand, CD73-deficient cells cultivated in an ADO-enriched medium decreased the frequency of CD4 CTL (CD73KO vs CD73KO+ADO: p <0,05). Remarkably, ADO stimulation significantly reduced the multifunctional capacity (IFN-γ+ TNF-α+ Granzyme B+ or IFN-γ+ TNF-α+ Perforin+) in CD4 T cells from CD73KO infected spleens (CD73KO vs CD73KO+ADO: p <0.05). To mimic CD73-deficient environment, WT cells were cultivated in an ATP-enriched medium. As expected, an increase in the multifunctional capacity (IFN-γ+ TNF-α+ Perforin+ Granzyme B+) of CD4 T cells was observed (WT vs. WT+ATP: p<0.05) reaching similar frequency observed for these populations in CD73KO cells.These data suggest that CD73 activity and ADO balance frankly influence the functionality of CD4 CTL during the early immune response against T. cruzi infection. Further experiments are needed to discover the role of this cell population on parasite persistence.