small-scale-colorimetric-assays-intracellular-lactate-pyruvate

Small-Scale Colorimetric Assays of Intracellular Lactate and Pyruvate in the Nematode <em>Caenorhabditis elegans</em>

Lactate and pyruvate are key intermediates of intracellular energy metabolic pathways. Monitoring the lactate/pyruvate ratio in cells helps to determine ...

Daito Bunka University

Tokai University

The hormetic effect, which extends the lifespan by various stressors, has been confirmed in Caenorhabditis elegans (C. elegans). We have previously reported that oxidative stress resistance in a long-lived mutant age-1 is associated with the hormesis. In the age-1 allele, which activates an insulin/insulin-like growth factor-1 (Ins/IGF-1) signaling pathway, the superoxide dismutase (SOD) and catalase activities increased during normal aging. We now demonstrate changes in the mitochondrial superoxide radical (*O(2)(-)) levels of the hormetic conditioned age-related strains. The *O(2)(-) levels in age-1 strain significantly decreased after intermittent hyperoxia exposure. On the other hand, this phenomenon was not observed in a daf-16 null mutant. This hormesis-dependent reduction of the *O(2)(-) levels was observed even if the mitochondrial Mn-SOD was experimentally reduced. Therefore, it is indicated that the hormesis is mediated by events that suppress the mitochondrial *O(2)(-) production. Moreover, some SOD gene expressions in the hormetic conditioned age-1 mutant were induced over steady state mRNA levels. These data suggest that oxidative stress-inducible hormesis is associated with a reduction of the mitochondrial *O(2)(-) production by activation of the antioxidant system via the Ins/IGF-1 signaling pathway.

Hyperoxia exposure induced hormesis decreases mitochondrial superoxide radical levels via Ins/IGF-1 signaling pathway in a long-lived age-1 mutant of Caenorhabditis elegans.

Superoxide dismutase (SOD) is an enzyme that catalytically removes the superoxide radical (*O2-) and protects organisms from oxidative damage during normal aging. We demonstrate that not only the cytosolic *O2- level but also the mitochondrial *O2- level increases in the deletion mutants of sod-1 gene encoding Cu/Zn SOD in Caenorhabditis elegans (C. elegans). Interestingly, this suggests that the activity of SOD-1, which so far has been thought to act mainly in cytoplasm, helps to control the detoxification of *O2- also in the mitochondria. We also found functional compensation by other SODs, especially the sod-5 gene, which was induced several fold in the mutants. Therefore, the possibility exists that the compensative expression of sod-5 gene in the sod-1 deficit is associated with the insulin/insulin-like growth factor-1 (Ins/IGF-1) signaling pathway, which regulates longevity and stress resistance of C. elegans because the sod-5 gene may be a target of the pathway.

SOD-1 deletions in Caenorhabditis elegans alter the localization of intracellular reactive oxygen species and show molecular compensation.

Astaxanthin (AX), which is produced by some marine animals, is a type of carotenoid that has antioxidative properties. In this study, we initially examined the effects of AX on the aging of a model organism C. elegans that has the conserved intracellular pathways related to mammalian longevity. The continuous treatments with AX (0.1 to 1 mM) from both the prereproductive and young adult stages extended the mean lifespans by about 16-30% in the wild-type and long-lived mutant age-1 of C. elegans. In contrast, the AX-dependent lifespan extension was not observed even in a daf-16 null mutant. Especially, the expression of genes encoding superoxide dismutases and catalases increased in two weeks after hatching, and the DAF-16 protein was translocated to the nucleus in the AX-exposed wild type. These results suggest that AX protects the cell organelle mitochondria and nucleus of the nematode, resulting in a lifespan extension via an Ins/IGF-1 signaling pathway during normal aging, at least in part.

Supplemental cellular protection by a carotenoid extends lifespan via Ins/IGF-1 signaling in Caenorhabditis elegans.

In the nematode Caenorhabditis elegans, the mammalian tumor suppressor p53 ortholog CEP-1 mediates the stress response, activates germ line apoptosis and regulates meiotic chromosome segregation. A reduction in its expression, which frequently occurs in mammalian cancer cells, extends lifespan and induces an adaptive response in C. elegans. However, these effects do not involve an increase in oxidative stress resistance. Here, we showed that intermittent exposure to hyperoxia, which induces oxidative stress resistance and lowers the production of ROS derived from mitochondrial respiration in C. elegans, slightly improved the lifespan extension of cep-1 mutant. Interestingly, ATP levels were increased without an increase in oxygen consumption in cep-1 mutant during aging. In the wild-type, lactate levels and consequentially the lactate/pyruvate ratio decreased during aging in adults. Furthermore, the expression levels of mitochondrial respiration-related sco-1, which is a target of p53/CEP-1, as well as those of gluconeogenesis regulation and mammalian sirtuin ortholog genes, were also increased in the aged and adaptive conditioned wild-type animals. In contrast, the lactate/pyruvate ratio increased in cells of the cep-1 mutant and was amplified by intermittent hyperoxia. These results suggest that impaired p53/CEP-1 leads to an imbalance in the age-related energy metabolic alteration between mitochondrial oxidative phosphorylation and aerobic glycolysis and plays an important role in the extension of both intact and adaptive lifespans.

Sumino Yanase

Department of Health Science,
School of Sports & Health Science,
Department of Health Science, School of Sports & Health Science

Daito Bunka University

Small-Scale Colorimetric Assays of Intracellular Lactate and Pyruvate in the Nematode Caenorhabditis elegans

Sumino Yanase

Department of Health Science,School of Sports & Health Science,Department of Health Science, School of Sports & Health Science

Daito Bunka University

Small-Scale Colorimetric Assays of Intracellular Lactate and Pyruvate in the Nematode Caenorhabditis elegans

Department of Health Science,
School of Sports & Health Science,
Department of Health Science, School of Sports & Health Science