PARP1-cGAS-NF-κB pathway of proinflammatory macrophage activation by extracellular vesicles released during Trypanosoma cruzi infection and Chagas disease

Author summary Chagas disease, caused by Trypanosoma cruzi (T. cruzi) parasite, is a life-threatening debilitating illness. Tissue inflammation is a hallmark of clinical form of Chagas dilated cardiomyopathy. Recent studies show that cell membranes shed extracellular vesicles (Ev) that carry DNA, proteins, and lipids of host and pathogen origin. In this study, we show that Ev released by T. cruzi, T. cruzi-infected cells and in plasma of Chagas mice (vs. controls) carry parasite and host DNA that is oxidized. When captured by macrophages, the Ev-carried oxidized DNA molecules were recognized by cytosolic DNA sensors cGAS and PARP1, and subsequently, signaled NF-kappa B-mediated proinflammatory cytokine production. Chemical or genetic inhibition of PARP1 resulted in significant decrease in Ev-induced inflammatory response of macrophages and cardiac inflammation in Chagas mice. We propose that chemical inhibitors of PARP1 offer a potential therapeutic target in arresting chronic inflammation in Chagas disease through modulation of the macrophage proinflammatory signaling. Trypanosoma cruzi (T. cruzi) is the etiological agent of Chagas cardiomyopathy. In the present study, we investigated the role of extracellular vesicles (Ev) in shaping the macrophage (M phi) response in progressive Chagas disease (CD). We purified T. cruzi Ev (TcEv) from axenic parasite cultures, and T. cruzi-induced Ev (TEv) from the supernatants of infected cells and plasma of acutely and chronically infected wild-type and Parp1(-/-) mice. Cultured (Raw 264.7) and bone-marrow M phi responded to TcEV and TEv with a profound increase in the expression and release of TNF-alpha, IL-6, and IL-1 beta cytokines. TEv produced by both immune (M phi) and non-immune (muscle) cells were proinflammatory. Chemical inhibition or genetic deletion of PARP1 (a DNA repair enzyme) significantly depressed the TEv-induced transcriptional and translational activation of proinflammatory M phi response. Oxidized DNA encapsulated by TEv was necessary for PARP1-dependent proinflammatory M phi response. Inhibition studies suggested that DNA-sensing innate immune receptors (cGAS>>TLR9) synergized with PARP1 in signaling the NF kappa B activation, and inhibition of PARP1 and cGAS resulted in >80% inhibition of TEv-induced NF kappa B activity. Histochemical studies showed intense inflammatory infiltrate associated with profound increase in CD11b(+)CD68(+)TNF-alpha(+) M phi in the myocardium of CD wild-type mice. In comparison, chronically infected Parp1(-/-) mice exhibited low-to-moderate tissue inflammation, >80% decline in myocardial infiltration of TNF-alpha(+) M phi, and no change in immunoregulatory IL-10(+) M phi. We conclude that oxidized DNA released with TEv signal the PARP1-cGAS-NF-kappa B pathway of proinflammatory M phi activation and worsens the chronic inflammatory pathology in CD. Small molecule antagonists of PARP1-cGAS signaling pathway would potentially be useful in reprogramming the M phi activation and controlling the chronic inflammation in CD.