title: Heme modulates Trypanosoma cruzi bioenergetics inducing mitochondrial ROS production.
abstract: Trypanosoma cruzi is the causative agent of Chagas disease and has a single mitochondrion, an organelle responsible for ATP production and the main site for the formation of reactive oxygen species (ROS). T. cruzi is an obligate intracellular parasite with a complex life cycle that alternates between vertebrate and invertebrate hosts, therefore the development of survival strategies and morphogenetic adaptations to deal with the various environments is mandatory. Over the years our group has been studying the vector-parasite interactions using heme as a physiological oxidant molecule that triggered epimastigote proliferation however, the source of ROS induced by heme remained unknown. In the present study we demonstrate the involvement of heme in the parasite mitochondrial metabolism, decreasing oxygen consumption leading to increased mitochondrial ROS and membrane potential. First, we incubated epimastigotes with carbonyl cyanide p-(trifluoromethoxy) phenylhydrazone (FCCP), an uncoupler of oxidative phosphorylation, which led to decreased ROS formation and parasite proliferation, even in the presence of heme, correlating mitochondrial ROS and T. cruzi survival. This hypothesis was confirmed after the mitochondria-targeted antioxidant ((2-(2,2,6,6 Tetramethylpiperidin-1-oxyl-4-ylamino)-2-oxoethyl) triphenylphosphonium chloride (MitoTEMPO) decreased both heme-induced ROS and epimastigote proliferation. Furthermore, heme increased the percentage of tetramethylrhodamine methyl ester (TMRM) positive parasites tremendously-indicating the hyperpolarization and increase of potential of the mitochondrial membrane (ΔΨm). Assessing the mitochondrial functional metabolism, we observed that in comparison to untreated parasites, heme-treated epimastigotes decreased their oxygen consumption, and increased the complex II-III activity. These changes allowed the electron flow into the electron transport system, even though the complex IV (cytochrome c oxidase) activity decreased significantly, showing that heme-induced mitochondrial ROS appears to be a consequence of the enhanced mitochondrial physiological modulation. Finally, the parasites that were submitted to high concentrations of heme presented no alterations in the ultrastructure. Consequently, our results suggest that heme released by the insect vector after the blood meal, modify epimastigote mitochondrial physiology to increase ROS as a metabolic mechanism to maintain epimastigote survival and proliferation.
authors.affiliation: Laboratório de Interação de Tripanossomatídeos e Vetores, Departamento de Bioquímica, Instituto de Biologia Roberto Alcantara Gomes, Universidade do Estado do Rio de Janeiro, RJ, Brazil; Instituto Nacional de Ciência e Tecnologia - Entomologia Molecular (INCT-EM), Brazil.
authors.forename: Natália P
authors.initials: NP
authors.lastname: Nogueira
authors.affiliation: Laboratório de Interação de Tripanossomatídeos e Vetores, Departamento de Bioquímica, Instituto de Biologia Roberto Alcantara Gomes, Universidade do Estado do Rio de Janeiro, RJ, Brazil.
authors.forename: Francis M S
authors.initials: FMS
authors.lastname: Saraiva
authors.affiliation: Laboratório de Bioquímica de Resposta ao Estresse, Instituto de Bioquímica Médica Leopoldo de Meis, Universidade Federal do Rio de Janeiro, RJ, Brazil.
authors.forename: Matheus P
authors.initials: MP
authors.lastname: Oliveira
authors.affiliation: Laboratório de Bioquímica de Resposta ao Estresse, Instituto de Bioquímica Médica Leopoldo de Meis, Universidade Federal do Rio de Janeiro, RJ, Brazil.
authors.forename: Ana Paula M
authors.initials: APM
authors.lastname: Mendonça
authors.affiliation: Laboratório de Bioquímica de Tripanossomatídeos, Instituto Oswaldo Cruz, Fundação Oswaldo Cruz, Manguinhos, RJ, Brazil.
authors.forename: Job D F
authors.initials: JDF
authors.lastname: Inacio
authors.affiliation: Laboratório de Bioquímica de Tripanossomatídeos, Instituto Oswaldo Cruz, Fundação Oswaldo Cruz, Manguinhos, RJ, Brazil.
authors.forename: Elmo E
authors.initials: EE
authors.lastname: Almeida-Amaral
authors.affiliation: Laboratório de Biologia Celular, Instituto Oswaldo Cruz, Fundação Oswaldo Cruz, RJ, Brazil.
authors.forename: Rubem F
authors.initials: RF
authors.lastname: Menna-Barreto
authors.affiliation: Laboratório de Interação de Tripanossomatídeos e Vetores, Departamento de Bioquímica, Instituto de Biologia Roberto Alcantara Gomes, Universidade do Estado do Rio de Janeiro, RJ, Brazil.
authors.forename: Gustavo A T
authors.initials: GAT
authors.lastname: Laranja
authors.affiliation: Laboratório de Helmintologia Romero Lascasas Porto, Faculdade de Ciências Médicas, Universidade do Estado do Rio de Janeiro, RJ, Brazil.
authors.forename: Eduardo J Lopes
authors.initials: EJL
authors.lastname: Torres
authors.affiliation: Laboratório de Bioquímica de Resposta ao Estresse, Instituto de Bioquímica Médica Leopoldo de Meis, Universidade Federal do Rio de Janeiro, RJ, Brazil.
authors.forename: Marcus F
authors.initials: MF
authors.lastname: Oliveira
authors.affiliation: Laboratório de Interação de Tripanossomatídeos e Vetores, Departamento de Bioquímica, Instituto de Biologia Roberto Alcantara Gomes, Universidade do Estado do Rio de Janeiro, RJ, Brazil; Instituto Nacional de Ciência e Tecnologia - Entomologia Molecular (INCT-EM), Brazil.
authors.forename: Marcia C
authors.initials: MC
authors.lastname: Paes
mesh_terms: D000818:Animals; D002259:Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone / Q000378:metabolism; D048708:Cell Growth Processes; D002478:Cells, Cultured; D014355:Chagas Disease / Q000276:immunology*; D004579:Electron Transport; D003576:Electron Transport Complex IV / Q000378:metabolism; D004734:Energy Metabolism; D006418:Heme / Q000378:metabolism*; D006801:Humans; D008018:Life Cycle Stages; D053078:Membrane Potential, Mitochondrial; D008928:Mitochondria / Q000378:metabolism*; D009943:Organophosphorus Compounds / Q000378:metabolism; D010101:Oxygen Consumption; D010880:Piperidines / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism; D012235:Rhodamines / Q000378:metabolism; D014349:Trypanosoma cruzi / Q000502:physiology*
chemical_list: C555916:MitoTEMPO; D009943:Organophosphorus Compounds; D010880:Piperidines; D017382:Reactive Oxygen Species; D012235:Rhodamines; C401833:tetramethylrhodamine methyl ester; D002259:Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone; D006418:Heme; D003576:Electron Transport Complex IV
keywords: Bioenergetics; Heme; Mitochondrion; Oxygen consumption; ROS; Trypanosoma cruzi