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Institute of Nutritional Sciences, Laboratory for Nutrition in Prevention and Therapy, Biomedical Research Center Seltersberg (BFS), Justus Liebig University Giessen, Schubertstrasse 81, 35392 Giessen, Germany.
Published: May 2020
AI Article Synopsis
Article Abstract
Introduction: Mitochondria supply cellular energy and are key regulators of intrinsic cell death and consequently affect longevity. The nematode is frequently used for lifespan assays. Using paraquat (PQ) as a generator of reactive oxygen species, we here describe its effects on the acceleration of aging and the associated dysfunctions at the level of mitochondria.
Methods: Nematodes were incubated with various concentrations of paraquat in a heat-stress resistance assay (37°C) using nucleic staining. The most effective concentration was validated under physiological conditions, and chemotaxis was assayed. Mitochondrial membrane potential (m) was measured using rhodamine 123, and activity of respiratory chain complexes determined using a Clark-type electrode in isolated mitochondria. Energetic metabolites in the form of pyruvate, lactate, and ATP were determined using commercial kits. Mitochondrial integrity and structure was investigated using transmission electron microscopy. Live imaging after staining with fluorescent dyes was used to measure mitochondrial and cytosolic ROS. Expression of longevity- and mitogenesis-related genes were evaluated using qRT-PCR.
Results: PQ (5 mM) significantly increased ROS formation in nematodes and reduced the chemotaxis, the physiological lifespan, and the survival in assays for heat-stress resistance. The number of fragmented mitochondria significantly increased. The ∆m, the activities of complexes I-IV of the mitochondrial respiratory chain, and the levels of pyruvate and lactate were significantly reduced, whereas ATP production was not affected. Transcript levels of genetic marker genes, , , , and , were significantly upregulated after PQ incubation, which implicates a close connection between mitochondrial dysfunction and oxidative stress response. Expression levels of and were unchanged.
Conclusion: Using paraquat as a stressor, we here describe the association of oxidative stress, restricted energy metabolism, and reduced stress resistance and longevity in the nematode making it a readily accessible model for mitochondrial dysfunction.
