mTOR INHIBITION IN trypanosoma cruzi INFECTED MACROPHAGES ACTIVES NLRP3 AND INDUCES MITOCHONDRIAL ROS PRODUCTION THAT REGULATE PARASITE SURVIVAL.

ROJAS MÁRQUEZ, JORGE DAVID; ANA, YAMILE; STEMPIN, CINTHIA C.; CERBAN, FABIO M.

Buenos Aires

Congreso; Reunión conjunta de sociedades de biociencia; 2017

Sociedad Argentina de Inmunología

We have previously shown that Trypanosoma cruzi infection inmacrophages (Mo) activates mTOR pathway and its inhibition de-582 MEDICINA - Volumen 77 - (Supl. I), 2017creased parasite replication. However, in rapamycin (Rap) pretreatedand T. cruzi infected Mo, we observed reduced nitric oxide productionand iNOS expression compared to control cells. Therefore,the aim of this work was to determine alternative activated mechanismsinvolved in controlling parasite during mTOR inhibition. In thissense, we found that cytoplasmic ROS (cROS) as well as IDO activitywere not involved as inflammatory mechanisms. Consequently,to study possible mediators involved in parasite killing, we obtainedbone marrow derived Mo (BMDM) from different KO mice pretreatedthem with Rap and then infected. Parasite load was evaluated 72hpost infection (pi) by immunofluorescencia (IF). The results showeda significant increase in parasite load in BMDM from IL-6-KO, TNFaR-KOand NLRP3-KO compared to WT BMDM. However, parasitenumber stands out in BMDM from NLRP3 KO (p<0.05). Taking intoaccount that NLRP3 is a key component of the inflammasome, ournext aim was to determine whether this pathway is involved. Weobserved that Rap-pretreated and infected BMDM showed a significantincrease in NLRP3 and IL-1β expression at 6h pi (p<0.05).Besides, TNF-a, IL-6, and IL12 expression were also increase at 12,18 and 24h pi (p<0.05). It has been shown that inflammasome usesthe mitochondria as platform of assembly. We found a significantlyincrease in mitochondrial ROS (mtROS) production at 3 and 6h pi(p<0.05). To evaluate the relevance of mtROS, BMDM were treatedwith DPI (NADPH oxidase inhibitor), then infected and parasite loadwas studied by IF. We observed that Rap+DPI pretreated BMDMhad significantly higher parasite load compared to Rap pretreatedBMDM (p<0.05). Our findings strongly support that mTOR inhibitionduring T. cruzi infection in Mo induced inflammasome activation thatleads to mtROS production controlling parasite survival.