Co-Exposure of Cardiomyocytes to IFN-γ and TNF-α Induces Mitochondrial Dysfunction and Nitro-Oxidative Stress: Implications for the Pathogenesis of Chronic Chagas Disease Cardiomyopathy

Abstract

Infection by the protozoan Trypanosoma cruzi causes Chagas disease cardiomyopathy (CCC) and can lead to arrhythmia, heart failure and death. Chagas disease affects 8 million people worldwide, and chronic production of the cytokines IFN-γ and TNF-α by T cells together with mitochondrial dysfunction are important players for the poor prognosis of the disease. Mitochondria occupy 40% of the cardiomyocytes volume and produce 95% of cellular ATP that sustain the life-long cycles of heart contraction. As IFN-γ and TNF-α have been described to affect mitochondrial function, we hypothesized that IFN-γ and TNF-α are involved in the myocardial mitochondrial dysfunction observed in CCC patients. In this study, we quantified markers of mitochondrial dysfunction and nitro-oxidative stress in CCC heart tissue and in IFN-γ/TNF-α-stimulated AC-16 human cardiomyocytes. We found that CCC myocardium displayed increased levels of nitro-oxidative stress and reduced mitochondrial DNA as compared with myocardial tissue from patients with dilated cardiomyopathy (DCM). IFN-γ/TNF-α treatment of AC-16 cardiomyocytes induced increased nitro-oxidative stress and decreased the mitochondrial membrane potential (ΔΨm). We found that the STAT1/NF-κB/NOS2 axis is involved in the IFN-γ/TNF-α-induced decrease of ΔΨm in AC-16 cardiomyocytes. Furthermore, treatment with mitochondria-sparing agonists of AMPK, NRF2 and SIRT1 rescues ΔΨm in IFN-γ/TNF-α-stimulated cells. Proteomic and gene expression analyses revealed that IFN-γ/TNF-α-treated cells corroborate mitochondrial dysfunction, transmembrane potential of mitochondria, altered fatty acid metabolism and cardiac necrosis/cell death. Functional assays conducted on Seahorse respirometer showed that cytokine-stimulated cells display decreased glycolytic and mitochondrial ATP production, dependency of fatty acid oxidation as well as increased proton leak and non-mitochondrial oxygen consumption. Together, our results suggest that IFN-γ and TNF-α cause direct damage to cardiomyocytes’ mitochondria by promoting oxidative and nitrosative stress and impairing energy production pathways. We hypothesize that treatment with agonists of AMPK, NRF2 and SIRT1 might be an approach to ameliorate the progression of Chagas disease cardiomyopathy.

Chagas Disease Megaesophagus Patients Carrying Variant MRPS18B P260A Display Nitro-Oxidative Stress and Mitochondrial Dysfunction in Response to IFN-γ Stimulus

“…Chronic immune response is also a key factor associated with worse progression of patients with Chagas disease [ 44 ]. The pro-inflammatory cytokines IFN-γ and TNF-α are increased in patients with CCC and digestive disease compared to patients with dilated cardiomyopathy [ 21 , 45 ] and these stimuli are reported to be mitochondrial dysfunction inducers by causing damage to cardiac tissue [ 3 , 24 , 27 ] and, in in vitro cardiomyocytes [ 46 ], promoting an increase in the production of reactive oxygen species (ROS) and nitrogen (RNS) [ 45 , 47 , 48 , 49 ]. Considering that the trigger shared by patients with megaesophagus and Chagas cardiomyopathy is infection with T. cruzi , our hypothesis is that these CME patients may also have mitochondrial/energy dysfunction associated with pathology plus mutation in the mitochondrial gene.…”

Targeting the Macrophage-Ferroptosis Crosstalk: A Novel Insight into Tumor Immunotherapy

“…Then the STAT1 homodimers are translocated into the nucleus where they could bind to the transcription region of SLC7A11 and suppressing system Xc − . Furthermore, IFN-γ can lead to high mitochondrial oxidative status and increased mitochondrial-derived ROS, implying that IFN-γ may disrupt the mitochondrial metabolism [31]. This is probably caused by IFN-γ-induced nitrooxidative stress via STAT1/NF-κB/NOS2 axis, though the exact mechanism still remains unknown [31,32].…”

“…Furthermore, IFN-γ can lead to high mitochondrial oxidative status and increased mitochondrial-derived ROS, implying that IFN-γ may disrupt the mitochondrial metabolism [31]. This is probably caused by IFN-γ-induced nitrooxidative stress via STAT1/NF-κB/NOS2 axis, though the exact mechanism still remains unknown [31,32]. Apart from this mechanism, Liao's team discovered that IFNγ stimulates ACSL4 to promote incorporation of arachidonic acid into into C16 and C18 acyl chain-containing phospholipids (Fig.…”

Cellular Stress and Senescence Induction during Trypanosoma cruzi Infection

“…Particularly, we found that cellular stress elicited by T. cruzi infection in the early stages of infection inhibits fibroblast proliferation and induces a senescent phenotype [55]. Furthermore, experimental evidence demonstrated that cells under senescence are apoptosis-resistant [56,57], and our findings suggest that senescent fibroblasts act as a long-term reservoir of parasites, allowing T. cruzi to replicate and establish a chronic infection [55]. Although our group correlated for the first time the early induction of senescence in T. cruzi infected fibroblasts, previous studies have reported the induction of immunosenescence on T cell subsets in both chronically infected human subjects [58,59] and mouse models of T. cruzi infection [8].…”