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Ultrastructural Expansion Microscopy in Three In Vitro Life Cycle Stages of Trypanosoma cruzi
Published on: May 12, 2023
Study of Trypanosoma cruzi epimastigote cell death by NMR-visible mobile lipid analysis
Diego Benitez1, Horacio Pezaroglo, Verónica Martínez
1Grupo de Química Medicinal, Laboratorio de Química Orgánica, Universidad de la República, Montevideo, Uruguay.
Abstract:
Cell death mechanisms in Trypanosoma cruzi have not been disclosed in detail though different conventional techniques have been used in the classification of parasite-cell death type. Nuclear magnetic resonance (NMR) has successfully been used as a tool to evaluate the onset of apoptosis in a number of higher eukaryote-cell models analysing the ratio of CH(2)/CH(3) integration from the visible mobile lipids (VML). Surprisingly, this versatile non-invasive spectroscopy technique has never been employed with this purpose in T. cruzi. In the present study it is shown that under different parasite death-conditions the ratio CH(2)/CH(3) varied drastically. Thus, T. cruzi epimastigotes in apoptotic conditions increase significantly this ratio while in necrotic as well as in autophagic situations the parasites maintain the VML, CH(2)/CH(3) ratio, in normal values. Additionally, other VML markers commonly used in these studies, such as the change in the region of methyl-choline moiety, -N(+)(CH(3))(3), exhibited different particular patterns according to the type of cell death. Our results suggest that the (1)H NMR-VML technique is an adequate tool to discriminate different T. cruzi death pathways.
Insights
Nuclear magnetic resonance (NMR) spectroscopy can now differentiate cell death in Trypanosoma cruzi. This technique analyzes visible mobile lipids (VML) to distinguish apoptosis from necrosis and autophagy in parasites.
Area of Science:
- Parasitology
- Biochemistry
- Spectroscopy
Background:
- Cell death mechanisms in Trypanosoma cruzi remain poorly understood.
- Conventional methods for classifying parasite cell death lack detail.
- Nuclear magnetic resonance (NMR) spectroscopy is established for apoptosis detection in eukaryotes.
Purpose of the Study:
- To investigate the utility of (1)H NMR-VML technique for discriminating cell death pathways in T. cruzi.
- To establish NMR as a tool for analyzing T. cruzi cell death mechanisms.
Main Methods:
- Analysis of the CH(2)/CH(3) ratio from visible mobile lipids (VML) using (1)H NMR.
- Application of the technique to T. cruzi epimastigotes under different cell death conditions (apoptosis, necrosis, autophagy).
- Evaluation of other VML markers, including the methyl-choline moiety.
Main Results:
- The CH(2)/CH(3) ratio within VML varied significantly across different T. cruzi death pathways.
- Apoptotic conditions led to a significant increase in the CH(2)/CH(3) ratio.
- Necrotic and autophagic conditions maintained normal CH(2)/CH(3) ratios, with distinct patterns observed in other VML markers.
Conclusions:
- The (1)H NMR-VML technique effectively distinguishes between apoptotic, necrotic, and autophagic cell death in T. cruzi.
- NMR spectroscopy provides a non-invasive and versatile method for studying T. cruzi cell death.
- This approach offers a novel tool for understanding parasite viability and drug development.
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