title: Bioenergetic profiling of Trypanosoma cruzi life stages using Seahorse extracellular flux technology.
abstract: Energy metabolism is an attractive target for the development of new therapeutics against protozoan pathogens, including Trypanosoma cruzi, the causative agent of human Chagas disease. Despite emerging evidence that mitochondrial electron transport is essential for the growth of intracellular T. cruzi amastigotes in mammalian cells, fundamental knowledge of mitochondrial energy metabolism in this parasite life stage remains incomplete. The Clark-type electrode, which measures the rate of oxygen consumption, has served as the traditional tool to study mitochondrial energetics and has contributed to our understanding of it in T. cruzi. Here, we evaluate the Seahorse XF(e)24 extracellular flux platform as an alternative method to assess mitochondrial bioenergetics in isolated T. cruzi parasites. We report optimized assay conditions used to perform mitochondrial stress tests with replicative life cycle stages of T. cruzi using the XF(e)24 instrument, and discuss the advantages and potential limitations of this methodology, as applied to T. cruzi and other trypanosomatids.
authors.affiliation: Department of Immunology and Infectious Diseases, Harvard School of Public Health, 665 Huntington Ave, Boston, MA, 02115, USA.
authors.forename: Sheena
authors.initials: S
authors.lastname: Shah-Simpson
authors.affiliation: Instituto de Biofísica Carlos Chagas Filho, Universidade Federal do Rio de Janeiro Av, Carlos Chagas Filho, 373, Centro de Ciências da Saúde, Bloco G, Cidade Universitária, Rio de Janeiro, Brazil.
authors.forename: Camila F A
authors.initials: CF
authors.lastname: Pereira
authors.affiliation: Department of Immunology and Infectious Diseases, Harvard School of Public Health, 665 Huntington Ave, Boston, MA, 02115, USA.
authors.forename: Peter C
authors.initials: PC
authors.lastname: Dumoulin
authors.affiliation: Seahorse Bioscience, 16 Esquire Road, Billerica, MA, 01862, USA.
authors.forename: Kacey L
authors.initials: KL
authors.lastname: Caradonna
authors.affiliation: Department of Immunology and Infectious Diseases, Harvard School of Public Health, 665 Huntington Ave, Boston, MA, 02115, USA. Electronic address: bburleig@hsph.harvard.edu.
authors.forename: Barbara A
authors.initials: BA
authors.lastname: Burleigh
mesh_terms: D004734:Energy Metabolism; D008018:Life Cycle Stages; D055442:Metabolome; D055432:Metabolomics / Q000379:methods*; D008928:Mitochondria / Q000378:metabolism; D010085:Oxidative Phosphorylation; D014349:Trypanosoma cruzi / Q000254:growth & development* / Q000378:metabolism*
keywords: Glycolysis; Mitochondrial bioenergetics; Trypanosoma cruzi